Structure-Based Design and Synthesis of Potent,\n                    Ethylenediamine-Based, Mammalian Farnesyltransferase Inhibitors as Anticancer\n                    Agents
A potent class of anticancer, human farnesyltransferase (hFTase) inhibitors has been identified by \xe2\x80\x9cpiggy-backing\xe2\x80\x9d on potent, antimalarial inhibitors of Plasmodium falciparum farnesyltransferase (PfFTase). On the basis of a 4-fold substituted ethylenediamine scaffold, the inhibitors are structurally simple and readily derivatized, facilitating the extensive structure\xe2\x80\x93activity relationship (SAR) study reported herein. Our most potent inhibitor is compound 1f, which exhibited an in vitro hFTase IC50 value of 25 nM and a whole cell H-Ras processing IC50 value of 90 nM. Moreover, it is noteworthy that several of our inhibitors proved highly selective for hFTase (up to 333-fold) over the related prenyltransferase enzyme geranylgeranyltransferase-I (GGTase-I). A crystal structure of inhibitor 1a co-crystallized with farnesyl pyrophosphate (FPP) in the active site of rat FTase illustrates that the para-benzonitrile moiety of 1a is stabilized by a \xcf\x80\xe2\x80\x93\xcf\x80 stacking interaction with the Y361\xce\xb2 residue, suggesting a structural explanation for the observed importance of this component of our inhibitors.
Introduction<!>Design<!>Crystal Structure<!>Chemistry<!>Sulfonyl R2 SAR<!>Sulfonamide R3 SAR<!>Aniline R4 SAR: The para-Position (X)<!>Aniline R4 SAR: The Aromatic Ring<!>Scaffold Optimization<!>Optimization of FTIs<!>Discussion of SAR Data in the Context of the Crystal Structure of 1a and the\n                    GOLD Docking of 1a<!>Conclusions<!>Ligand Docking Studies<!>Chemistry: General Methods<!>General Procedure A (Reaction of Amines with Sulfonyl Chlorides)<!>General Procedure B (Alkylation of Sulfonamides)<!>General Procedure C (Reductive Amination of Primary Amines)<!>General Procedure D (Arylation of Primary Amines)<!>General Procedure E (Alkylation of Secondary Anilines)<!>General Procedure F (Cyanation of Aryl Bromides)<!>4-[(2-Aminoethyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-benzonitrile\n                    (9)<!>N-Benzyl,\n                        N-{2-[(4-Cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}\n                    Propane-1-sulfonamide (1b)<!>N-Benzyl,\n                        N-{2-[(4-Cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}\n                    Cyclopropanesulfonamide (1c)<!>[N-(Furan-3-ylmethyl),\n                        N-{2-[(4-Cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (1ah, 4.38)<!>[N-(Thiophen-3-ylmethyl),\n                        N-{2-[(4-Cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (1ai, 4.39)<!>[N-(Furan-2-ylmethyl),\n                        N-{2-[(4-Cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (1aj, 4.41)<!>[N-(Thiophen-2-ylmethyl),\n                        N-{2-[(4-Cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (1ak, 4.42)<!>[N-(3,5-Dimethyl-isoxazol-4-ylmethyl),\n                        N-{2-[(4-Cyanophenyl)(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (1al, 4.36)<!>[N-(2,4-Dimethyl-thiazol-5-ylmethyl),\n                        N-{2-[(4-Cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (1am, 4.40)<!>[N-(1,3,5-Trimethyl-1H-pyrazol-4-ylmethyl),\n                        N-{2-[(4-Cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (1an, 4.49)<!>[N-(Cyclopropylmethyl),\n                        N-{2-[(4-Cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (1aw, 4.34)<!>tert-Butyl\n                        4-[{2-[benzyl-(1-methyl-1H-imidazole-4-sulfonyl)-amino]-ethyl}-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-benzoate\n                    (10)<!>4-[{2-[Benzyl-(1-methyl-1H-imidazole-4-sulfonyl)-amino]-ethyl}(3-methyl-3H-imidazol-4-ylmethyl)-amino]-benzoic\n                    Acid (12)<!>4-[{2-[Benzyl-(1-methyl-1H-imidazole-4-sulfonyl)-amino]-ethyl}(1-methyl-1H-imidazol-4-ylmethyl)-amino]-benzamide\n                    (13)<!>[N-Benzyl,\n                        N-{2-[(biphenyl-4-yl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}]\n                        1-methyl-1H-imidazole-4-sulfonamide (14)<!>N-(2-tert-Butoxycarbonylaminoethyl)\n                        1-Methyl-1H-imidazole-4-sulfonamide (15)<!>[N-(2-tert-Butoxycarbonylaminoethyl),\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (16)<!>[N-(2-Aminoethyl),\n                    N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (17)<!>[N-{2-(2-Chloro-4-cyanophenyl)-aminoethyl},\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (18a)<!>N-(2-(2-Cyanopyridin-5-ylamino)ethyl)-1-methyl-N-((1-(pyrimidin-2-yl)piperidin-4-yl)methyl)-1H-imidazole-4-sulfonamide\n                    (18b)<!>[N-{2-(5-Bromopyridin-2-yl)-aminoethyl},\n                        N-{N-(2-pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-methyl-1H-imidazole-4-sulfonamide (18c)<!>[N-{2-(5-Bromopyrimidin-2-yl)-aminoethyl},N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (18d)<!>[N-{2-(3-Fluoro-4-cyanophenyl)-aminoethyl},\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (18e)<!>[N-{2-(2-Fluoro-4-cyanophenyl)-aminoethyl},\n                        N-{N-(2-pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (18f)<!>[N-{2-(2,6-Difluoro-4-cyanophenyl)-aminoethyl},\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (18g)<!>[N-{2-(2,3,5,6-Tetrafluoro-4-cyanophenyl)-aminoethyl},N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (18h)<!>[N-{2-[(2-Chloro-4-cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl},\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (19a)<!>N-(2-((2-Cyanopyridin-5-yl)((1-methyl-1H-imidazol-5-yl)methyl)-amino)ethyl)-1-methyl-N-((1-(pyrimidin-2-yl)piperidin-4-yl)methyl)-1H-imidazole-4-sulfonamide\n                    (19b)<!>[N-{2-[(5-Cyanopyridin-2-yl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl},\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (19c)<!>[N-{2-[(5-Cyanopyrimidin-2-yl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl},\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (19d)<!>[N-{2-[(3-Fluoro-4-cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl},\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (19e)<!>[N-{2-[(2-Fluoro-4-cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl},\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (19f)<!>[N-{2-[(2,6-Difluoro-4-cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl},\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (19g)<!>[N-{2-[(2,3,5,6-Tetrafluoro-4-cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl},\n                        N-{N-(2-Pyrimidinyl)-piperidin-4-ylmethyl}]\n                        1-Methyl-1H-imidazole-4-sulfonamide (19h)<!>N-(2-Aminoethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide\n                    (21)<!>(2-((4-Cyanophenyl)((1-methyl-1H-imidazol-5-yl)methyl)amino)-ethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide\n                    (21a)<!>N-(2-((4-Cyano-2-fluorophenyl)((1-methyl-1H-imidazol-5-yl)-methyl)amino)ethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide\n                    (21b)<!>N-(2-((5-Cyanopyridin-2-yl)((1-methyl-1H-imidazol-5-yl)-methyl)amino)ethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide\n                    (21c)<!>In Vitro and Whole Cell Inhibition Assays of Protein Prenylation
<p>As the successful treatment of cancer remains a challenging goal, research
                into novel, selective, and less toxic chemotherapeutic agents is gathering
                    pace.1-3 Indeed, increasing understanding of the cellular processes that lead
                to cancer has identified additional targets for the design of such
                chemotherapeutics. Ras, the protein product of the ras oncogene, is
                a small GTPase that is important in signal transduction, cell growth, and cell
                    proliferation.4 Mutations in Ras, which
                cause the protein to persistently bind GTP and thus become constitutively active,
                can lead to unregulated cell division; such Ras mutants are found in approximately
                30% of human tumors.5,6 In the 1980s, it was reported that Ras required farnesylation
                to enhance its hydrophobicity and thereby facilitate its anchorage to the plasma
                membrane, a process necessary for its signaling function.7,8 Accordingly, it was
                envisioned that inhibition of the enzyme that performs this post-translational
                modification, protein farnesyltransferase (FTasea), would offer an indirect method of blocking the function of Ras
                oncoproteins. Indeed, in addition to inhibiting FTase in vitro,9-12 farnesyltransferase
                inhibitors (FTIs) have demonstrated anti-tumor activity in several animal
                    models.2,9 Clinically, however, the results are mixed. For example, a lack of
                activity was reported when Tipifarnib13
                (R115777) was used against advanced colorectal and pancreatic cancers.14,15 In
                contrast, extremely encouraging results were observed when Tipifarnib was used
                against breast cancer in combination with cytotoxic agents.16,17 In recent years,
                it has become clear that aberrant Ras activity is not the only target for FTIs, and
                it is likely that other FTase substrates, such as Rheb, are also involved in
                    oncogenesis.18-21 Nonetheless, despite the now-apparent complexity of this
                system and the unclear molecular mechanisms by which FTIs operate, the past decade
                has seen many FTIs established as antiproliferative agents of high efficacy and low
                toxicity, validating the continued research into more drug-like FTIs as alternative
                chemotherapeutics for cancer.1-3</p><p>The prenyltransferases are a family of zinc metalloenzymes that catalyze the
                prenylation (addition of a prenyl group through a thioether linkage) of a particular
                set of proteins, many of which are crucial to signal transduction pathways, causing
                their localization to the plasma membrane and other cellular compartments and so
                rendering them biologically active.22 There
                are three members of the prenyltransferase family: farnesyltransferase (FTase),
                geranygeranyltransferaseI (GGTase-I), and geranygeranyltransferase-II (GGTase-II).
                FTase catalyzes the transfer of a farnesyl (C15 isoprenoid) group from
                the cosubstrate farnesyl pyrophosphate (FPP) to the cysteine residue within the
                    C-terminus Ca1a2X tetra-peptide sequence
                of the target protein (including Ras and Rheb), where C = cysteine, a = an aliphatic
                amino acid, and X = methionine (M), serine (S), alanine (A), or glutamine (Q).23 Likewise, GGTase-I catalyzes the
                corresponding S-geranylgeranylation by accelerating the transfer of the
                geranylgeranyl group (C20 isoprenoid) from geranylgeranyl pyrophosphate
                (GGPP) to the cysteine within the C-terminus
                    Ca1a2X sequence of the substrate protein (including Rho,
                Rap, and Ral),24 where this time X is usually
                leucine (L), isoleucine (I), or phenylalanine (F).23 It is the identity of the X residue that dictates if a target protein
                is farnesylated or geranylgeranylated, and is so-called the specificity residue.
                Finally, in a similar fashion, GGTase-II transfers two geranylgeranyl groups to
                protein trafficking Rab proteins that contain Cys-CysorCys-Ala-Cyssequencesatthe
                    C-terminus.23</p><p>Previous research within our laboratories has focused on the design of
                peptidomimetic inhibitors of FTase based on the Ca1a2X
                tetrapeptide substrate.25-28 Herein we describe a novel series of
                ethylenediamine-based, mammalian FTase inhibitors as anticancer compounds that were
                discovered by a "piggy-back" approach after the success of the core
                scaffold in a series of antimalarial plasmodial FTase inhibitors.29 We present an extensive
                structure–activity relationship (SAR) study of these inhibitors with both in
                vitro and whole cell data, including relative activities against FTase and GGTase-I.
                Additionally, we discuss our efforts to improve inhibitor potency against mammalian
                FTase, and we present crystallographic data that reveals the actual binding mode of
                our inhibitors within the active site of the enzyme.</p><!><p>We have previously reported on the design and synthesis of inhibitors of
                        Plasmodium falciparum farnesyltransferase
                        (PfFTase).29,30 An homology model of the active site of
                        PfFTase suggests the presence of four sub-pockets.31 By employing the computational modeling
                    program GOLD,32 we identified that an
                    ethylenediamine scaffold with both nitrogens doubly substituted in order to gain
                    simultaneous access to these four subpockets might furnish inhibitors of
                        PfFTase. Indeed, compounds based on this scaffold,
                    including 1a (Figure 1), in
                    which the choice of the four substituents (para-benzonitrile,
                    imidazolylmethyl, arylmethyl, and heterocycle-substituted sulfonyl) was
                    influenced by the BMS series of tetrahydrobenzodiazepine-based33 and tetrahydroquinoline (THQ)-based34 FTase inhibitors, proved particularly
                    potent inhibitors of PfFTase, both in vitro and in infected
                        erythrocytes.29,30 Although PfFTase is significantly
                    larger than rat FTase (rFTase) in both the α- (472 vs
                    379 residues) and β-subunits (923 vs 437 residues), the
                    differences are mainly due to insertions in the PfFTase protein
                    sequence, and overall there is minimal difference in the residues that form the
                    active site.29,31 Of the sequence segments in the model of
                        PfFTase, there is 23% identity (53% similarity) for the
                        α-subunit between PfFTase and
                    rFTase isoforms and 37% (56%) for the β-subunit.
                    Therefore, we anticipated that our PfFTase inhibitors may also
                    inhibit mammalian FTase, binding in the same sub-domains but, in certain
                    instances, to differing active site residues (i.e.,
                        PfFTase31 →
                    rFTase (PDB ID: 1JCR35): S150 →
                        N165α, F151 → Y166α,
                    N317 → H201α), providing a route to inhibitor
                    selectivity for the mammalian isoform of the enzyme.</p><p>Specifically, as with an almost identical analogue of 1a in
                    the homology model of the active site of PfFTase (Figure 2 in ref 29b), we hypothesized that the more basic of the two
                    imidazoles, the 1-methyl-1H-imidazol-5-ylmethyl substituent of
                        1a, would bind the zinc ion (itself held in place by
                        D297β (D659), C661β (C299),
                    and H362β (H838), where the labels in parentheses
                    represent the corresponding residues in PfFTase31). In addition, we envisioned that the
                        para-benzonitrile moiety would bind in the mostly
                    hydrophobic pocket constructed from the tetramethylene portion of the side chain
                    of K164α (K149) and Y166α (F151)
                    and whose deepest point forms a hydrophilic domain
                        (H201α (N317) and N165α
                    (S150)). We considered that the benzyl moiety would bind in the hydrophobic
                    pocket comprising W102β (W452),
                        W106β (W456), and Y361β
                    (Y837), and finally that the sulfonylimidazole would bind in the hydrophilic
                    domain formed by R202β (R564) and three water molecules
                    participating in a hydrogen-bonded network between S99β
                    (S449) and Q167α (Q152).</p><p>To maintain consistency with the GOLD docking experiments of our
                    ethylenediamine-based inhibitors in the homology model of the active site of
                        PfFTase,29,30 the only constraint imposed in the
                    flexible ligand GOLD docking with compound 1a in the active site of
                    rFTase (PDB ID: 1JCR) was that the more basic, non-sulfonylated imidazole should
                    again bind the Zn2+ ion. Due to the flexible nature of the
                    ethylenediamine scaffold in 1a, we did not want to bias the docking
                    results any further and used no additional constraints. Moreover, despite the
                    similarities of the pendant groups in 1a and the former clinical
                    candidate
                        (R)-7-cyano-2,3,4,5-tetrahydro-1-(1H-imidazol-4-ylmethyl)-3-(phenylmethyl)-4-(2-thienylsulfonyl)-1H-1,4-benzodiazepine
                    (BMS-214662, 233), we
                    elected not to use the knowledge of the crystal structure of 236 given its more rigid
                    tetrahydrobenzodiazepine scaffold. As was anticipated owing to the similarities
                    in the active sites of PfFTase and rFTase, several low energy
                    GOLD docked poses of compound 1a in rFTase demonstrated an almost
                    identical binding mode to that observed with a very close analogue of
                        1a in PfFTase, with the biggest difference
                    being that the benzyl group, rather than binding in the
                        W102β, W106β,
                        Y361β sub-pocket, is engaged in a
                        π–π stacking
                    interaction with Y361β (compare Figure 2 in ref 29b
                    with Figure 1a above). Figure 1A illustrates one such high scoring
                    (low energy) docked pose of compound 1a in green and colored by
                    atom type, using the graphical representation (Connolly analytical surface,
                        PyMOL37) and orientation employed in
                    previous publications.29,30 The binding surface of rFTase shown
                    incorporates the cosubstrate farnesyl pyrophosphate (FPP: farnesyl, red;
                    pyrophosphate, blue). This binding mode of 1a overlays well with
                    the tetrapeptide inhibitor CVFM from the rFTase crystal structure as shown in
                        Figure 1B, in which we have used an
                    alternative graphical representation ("cartoon", PyMOL37) and orientation that have also been
                    presented by us recently.38 For
                    simplicity, the latter graphical representation shall be used throughout the
                    remainder of this manuscript. Given the highly flexible nature of the ligand,
                    coupled with the fact that the other high scoring poses from our studies (data
                    not shown) were generally those in which the scaffold projected functionalities
                    to positions similar to those seen in Figure
                        1, we feel that it is likely that the molecule, in solution, would
                    occupy pockets as previously predicted as part of an ensemble of binding
                    motifs.</p><p>Initially, we selected a focused set of our PfFTase
                    inhibitors and analyzed their abilities to inhibit human FTase (hFTase), as well
                    as GGTase-I, both in vitro and in whole cells (Table 1). The amino acid sequences of rat and human FTase are 95%
                    identical with complete sequence and structural conservation around the active
                        site,35 so it is reasonable to
                    analyze inhibition of hFTase with respect to docking studies in rFTase. Briefly,
                    in vitro inhibition assays for hFTase and GGTase-I were carried out by measuring
                    the incorporation of [3H]FPP and [3H]GGPP into recombinant
                    H-Ras-CVLS and H-Ras-CVLL, respectively, as described previously.39 Whole cell inhibition of farnesylation
                    and geranylgeranylation were determined based on the level of inhibition by
                    synthetic compounds of H-Ras and Rap1A processing, respectively.40 In all cases, IC50 data
                    represent the average of three independent assays (n = 3),
                    unless otherwise stated, and errors are given as standard deviations. The
                    importance of both the para-substitution of the aniline
                    component and Nπ-methylation (R1 =
                    Me) of the imidazole that we observed in the inhibition of
                        PfFTase29 was
                    reflected in the mammalian isoform of the enzyme. Unsubstituted aniline
                        3 exhibited little activity toward the inhibition of hFTase.
                    Incorporation of bromine into the para position of the aniline
                    ring, however, led to an order of magnitude increase in potency with in vitro
                        IC50s for hFTase improving from 6300 ± 360 nM for
                        3 to 730 ± 20 nM for 4. Additionally, H-Ras
                    processing IC50s of the FTIs were enhanced from > 10
                        μM to 5.7 ± 1.2 μM. A
                    further order of magnitude increase in FTase inhibition in vitro was achieved
                    upon Nπ-methylation of the imidazole, with
                        5 exhibiting an IC50 of 79 ± 30 nM, and an
                    associated increase in whole cell activity (H-Ras processing IC50 =
                    1.6 ± 1.3 μM) was also observed. Even greater
                    hFTase inhibitory activity was achieved by the replacement of bromine with cyano
                    in the para position of the aniline ring
                        (1a:IC50 =56 ± 29 nM). It is interesting to
                    note that a considerable improvement in selectivity for hFTase over GGTase-I was
                    also observed; 1a was approximately 7-fold more selective for
                    hFTase than was 5. In addition, the trends observed here in the
                    inhibition of the mammalian isoform of FTase are mirrored by those observed in
                    the disruption of the plasmodial isoform.29 Given these very encouraging data, we embarked on an SAR study of
                    our ethylenediamine-based inhibitors in order to optimize the inhibitory
                    activity of 1a against hFTase.</p><!><p>We recently reported the crystal structure of the
                    1a:FPP:rFTase ternary complex (Figure
                        2A; PDB ID: 3E32).38 As a
                    comparison, in Figure 2B we have
                    superimposed the crystal structure of 1a with that of the
                    tetrapep-tide substrate CVFM (PDB ID: 1JCR). Interestingly, the crystal
                    structure shows that the ethylenediamine-based FTIs actually adopt a different
                    binding mode to that predicted (Figure 1B).
                        Figure 2A illustrates that the
                        para-benzonitrile substituent of 1a is
                    oriented toward the product exit groove and is partially stabilized by a
                    stacking interaction with Y361β. The rings are
                    approximately 30° offset from parallel, with the distance between them
                    ranging from 3.8 to 4.9 Å. The binding pocket also consists of
                        F360β (not shown in Figure),
                        Y93β, L96β, and
                        W106β, creating a generally hydrophobic environment
                    for this moiety. Inhibitor 1a possesses two
                    N-methylimidazole groups: one coordinates the catalytic zinc
                    ion (2.0 Å distance) through its nonmethylated
                    τ-nitrogen in place of the Ca1a2X
                    box cysteine residue, while the second (at the sulfonamide position) is stacked
                    between the first N-methylimidazole (3.7 Å distance) and
                    the first isoprene of FPP (4.1 Å distance). The non-methylated
                        τ-nitrogen of the
                    1-methyl-1H-imidazole-4-sulfonyl group also makes a single,
                    weak polar contact (3.7 Å distance) to the side chain hydroxyl of
                        Y361β. The phenyl substituent of 1a
                    binds in a pocket formed by W106β,
                        W102β, and L96β, essentially
                    the a2 residue binding site of the Ca1a2X
                    substrate, and occupying almost exactly the same space as the phenylalanine side
                    chain of CVFM (Figure 2B).</p><p>Several similarities exist between the chemical substituents and binding
                    mode of the ethylenediamine inhibitor 1a with
                    tetrahydrobenzodiazepine 2; their crystal structures are overlaid in Figure 3. The binding modes of the
                        para-benzonitrile group and the non-sulfonylated
                        N-methylimidazole group of the ethylenediamine inhibitor
                        1a are nearly indistinguishable from the similar substituents
                    in 2, with the imidazole of 2 coordinating the
                    catalytic zinc ion like the non-sulfonylated N-methylimidazole
                    in 1a. The scaffold nitrogen atoms bearing these substituents in
                    both 1a and 2 occupy similar locations. The
                        para-benzonitrile groups extend into the exit groove, with
                    the positions of the cyano group nitrogen atoms from the two molecules differing
                    by only ~0.4 Å, a value close to the estimated error in
                    crystallo-graphic coordinates. Finally, the phenyl substituents of both
                    compounds take advantage of the aromatic character of the
                        Ca1a2Xa2 residue site, interacting with
                    tryptophan residues 102β and
                        106β. The sulfonamide positions of both compounds
                    exhibit the greatest differences in binding modes. As a consequence, the
                    scaffold nitrogen bearing this group in the tetrahydrobenzodiazepine ring of
                        2 is not oriented in a spatially similar location to the
                    nitrogen bearing the sulfonylated N-methylimidazole of
                        1a, which is sandwiched between the first isoprene of the lipid
                    substrate and the zinc-coordinating N-methylimidazole moiety.
                    By contrast, the thienyl group of 2 is largely solvent-exposed and
                    binds in a manner resembling the a1 residue of the
                        Ca1a2X motif. This ring is primarily stabilized by
                    stacking on the tetrahydrobenzodiazepine ring scaffold itself, as opposed to
                    interacting with the enzyme active site or lipid substrate.</p><p>The crystal structure of the ternary complex of 1a:FPP:
                    rFTase was solved toward the end of this research; the medicinal chemistry
                    research program reported herein was actually driven by piggy-backing on our
                    inhibitors of PfFTase and, to a lesser extent, the GOLD docking
                    result presented in Figure 1. At the end of
                    the SAR discussion, the FTase inhibition data will be evaluated in the context
                    of both this crystal structure and the GOLD docking models.</p><!><p>"Piggy-back" mammalian FTase inhibitors (FTIs) were
                    prepared as described previously,29 with
                    new FTIs furnished by synthetic routes depicted in Schemes 1–3. Inhibitors
                    incorporating the para-benzonitrile moiety were accessed by the
                    route in Scheme 1. Briefly,
                        mono-N-Bocethylenediamine (6) was arylated with
                        para-fluorobenzonitrile at 120 °C for 48 h in DMSO,
                    affording secondary aniline 7.29b
                    Installation of the imidazole was achieved by double deprotonation of 7 with LDA
                    at −78 °C and subsequent chemo-selective alkylation with
                        5-chloromethyl-1-methyl-1H-imidazole 3 HCl,29b affording 829b in a moderate yield. After Boc
                    deprotection, completion of the syntheses of FTase inhibitors 1 was
                    conducted in one of two ways: either sulfonylation of 9,29b followed by alkylation of the resultant
                    sulfonamide, or reductive amination of 9, and then sulfonylation of
                    the resultant secondary amine. Finally, purification of the FTIs was achieved by
                    silica gel flash column chromatography, followed by rpHPLC. Full details can be
                    found in the Experimental Section.</p><!><p>With R1 constrained as methyl and R3 as benzyl, a
                    series of eight R2 sulfonyl groups was surveyed (Table 2), all of which proved potent
                    inhibitors of hFTase. The largest and most basic
                    5-dimethylamino-naphthalene-1-sulfonyl derivative 1h was the least
                    active inhibitor (hFTase IC50 = 160 ± 110 nM) of this series,
                    with the most potent being the pyridine-2-sulfonyl compound 1f
                    (hFTase IC50 =25 ± 20 nM). Additionally, these inhibitors were
                    more selective for hFTase over GGTase-I, with selectivities ranging from around
                    8-fold to 333-fold. Many of these compounds also demonstrated very good whole
                    cell activity, with pyridine-2-sulfonyl derivative 1f proving one
                    of the most effective inhibitors (H-Ras processing IC50 = 0.09
                    ± 0.06 μM). Despite its potency in vitro (hFTase:
                        IC50 = 56 ± 29 nM),
                    1-methyl-1H-imidazole-4-sulfonyl derivative 1a had
                    the poorest H-Ras processing IC50 (1.9 ± 1.2
                        Î¼M). This might be a consequence of the basicity of
                    imidazole, and its protonation could hinder cellular entry. Regardless of this,
                    the potent in vitro activity of 1a against hFTase, coupled with the
                    fact that we already had several inhibitors functionalized with the
                        1-methyl-1H-imidazole-4-sulfonyl group in-house from the
                    related PfFTase project,29 led us to retain this particular sulfonyl group as a benchmark
                    while varying other parts of the molecule. Selectivity of these FTIs for the
                    hFTase isoform over the PfFTase isoform might then later be
                    achieved by taking advantage of the previously mentioned three amino acid
                    differences in the subpocket that is predicted to bind the
                    para-benzonitrile ring.</p><!><p>Thus, constraining R2 as
                    1-methy-1H-imidazole-4-sulfonyl, the R3 group was
                    next investigated with a broad series of derivatives, which included previously
                        reported29 and novel inhibitors
                        (Table 3). Replacement of benzyl in
                    the parent inhibitor 1a with a propargyl group (1aa)
                    led to a reduction in both in vitro activity (IC50 = 720 ± 200
                    nM cf. 56 ± 29 nM) and whole cell activity, as well as in GGTase-I/FTase
                    selectivity. However, the unsubstituted allyl derivative (1ab) was
                    tolerated in vitro (IC50 =54 ± 30 nM), although not in whole
                    cells (H-Ras processing IC50 >10 Î¼M).
                    Substitution of the allyl moiety with a methyl group at position-2
                        (1ac) led to a reduction in hFTase in vitro activity and, at
                    the same time, afforded an improvement in GGTase-I, resulting in a reduced
                    GGTase-I/FTase selectivity of about 5-fold. Again, whole cell activity was poor
                    but this was recovered somewhat by replacement of the methyl in 1ac
                    with a bromine atom, giving an H-Ras processing IC50 of 5.7 ±
                    1.5 Î¼M for inhibitor 1ad. Extension of this
                        sp2-hybridized series to the bulky
                        tert-butylacetamido derivative 1ae was not
                    tolerated, neither in vitro nor in whole cells. Incorporation of nitrogen into
                    the para position of benzyl to give pyridine derivative
                        1af maintained activity in vitro (IC50 = 72 ±
                    20 nM), although it showed little activity in whole cells. Nonetheless,
                        1af exhibited substantially improved selectivity toward hFTase,
                    with a GGTase-I/FTase selectivity of > 139-fold.</p><p>We next examined a focused set of five-membered heteroaromatics
                        1ag–1an. Pyrrole 1ag was
                    around twice as active as the parent inhibitor 1a, disrupting
                    hFTase with an IC50 value of 30 ± 33 nM. Conversely, the
                    series of furans and thiophenes typically proved much poorer inhibitors,
                    although interestingly FTase inhibitory activity was found to be sensitive to
                    heteroatom position. Furan 1ah was more than twice as potent as
                    isomeric 1aj, and thiophene 1ai was more than 34 times
                    as potent as isomeric 1ak and approximately as active as
                        1a. Despite these moderate to poor in vitro data for hFTase
                    (except for 1ai) all members of this series exhibited reasonable
                    H-Ras processing whole cell data and in all cases the IC50 values
                    were more potent than 1a. The remaining heteroaromatics
                        (1al–1an) containing two heteroatoms were
                    poor inhibitors of hFTase.</p><p>Introduction of a methyl group on the benzyl ring of 1a was
                    tolerated in the ortho (1ao)and
                        meta (1ap) positions (in vitro hFTase
                        IC50sof51 ± 26 nM and 54 ± 30 nM, respectively) but
                    not in the para (1aq) position (IC50 =
                    370 ± 210 nM). A similar trend was observed in whole cell activity, with
                        meta-derivative 1ap the most potent (H-Ras
                    processing IC50 =0.3 ± 0.2 μM). The
                    members of this series demonstrated GGTase-I IC50 values between 580
                    and 780 nM, considerably reducing the GGTase-I/hFTase selectivities, relative to
                    that of the parent benzyl inhibitor 1a. The greater potency of
                    meta-substituted benzyl groups over their para counterparts was
                    also observed for the cyano derivatives 1ar and 1as
                    and for the phenyl derivatives 1at and 1au, a finding
                    that was reflected in their corresponding H-Ras processing whole cell
                    activities. Additionally, the same trend was noticed in the in vitro data of
                    GGTase-I inhibition. The poor in vitro inhibition of hFTase by
                        para-phenyl derivative 1au (IC50 =
                    1700 ± 900 nM) was improved by approximately 4-fold by replacement of the
                    terminal phenyl group with the smaller pyrrole heterocycle (1av;
                        IC50 = 390 ± 210 nM), which was also reflected in the
                    whole cell assays (IC50 =5 ± 1 μM
                        (1au) vs 4 ± 1 μM
                        (1av)).</p><p>The small cyclopropylmethyl derivative 1aw was less active
                    than the parent benzyl inhibitor 1a, but the larger
                    cyclohexylmethyl derivative 1ax, a closer match to benzyl, proved a
                    potent inhibitor in vitro (hFTase IC50 =60 ± 10 nM) and was
                    one of our most potent inhibitors in whole cells (H-Ras processing
                        IC50 = 0.1 ± 0.07 μM)). However,
                    inhibitor 1ax was also quite active toward GGTase-I
                        (IC50 = 530 ± 120 nM), giving a much-reduced
                    hFTase/GGTase-I selectivity of about 9-fold. We envisaged probing deeper into
                    the binding pocket that was predicted to bind the R3 group by
                    modifying the cyclohexylmethyl substituent to a 4-piperidinylmethyl group; the
                    piperidine nitrogen provided a handle from which to achieve further
                    functionalization. Accordingly, N-Boc-piperidin-4-ylmethyl
                    derivative 1ay and N-(2-pyrimidinyl)-4-ylmethyl
                    derivative 1az were synthesized. Introduction of the
                        N-Boc group at the 4 position of the cyclohexyl group
                        (1ay) caused more than a 60-fold drop in hFTase inhibition
                    relative to 1ax. Activity was recovered by more than 7-fold through
                    replacement of the bulky tert-butoxycarbonyl group with the
                    planar 2-pyrimidinyl group (1az:IC50 = 510 ± 62
                    nM; cf. 1ay: IC50 = 3700 ± 790 nM) although both
                    piperidinylmethyl derivatives exhibited poor whole cell activities.</p><!><p>At the deepest point of the predominantly hydrophobic pocket of rFTase
                    in which GOLD docking studies consistently predicted the
                    para-benzonitrile moiety (the aniline "R4
                    group") of our inhibitors would bind, there is a hydrophilic domain
                    formed by N165α, Y166α, and
                        H201α. To probe this site further, we replaced the
                    cyano group (X) of the para-benzonitrile moiety with
                    alternative polar groups, such as a carboxylic acid or a carboxamide, in the
                    anticipation that such groups might be able to participate in additional
                    hydrogen-bonding interactions and so furnish more potent hFTase inhibitors. We
                    also investigated nonpolar substituents in the para position,
                    such as Br, Ph and CO2t-Bu, which, provided the
                    aniline group binds as predicted, would be expected to inhibit hFTase less well.
                    Accordingly, para-(tert-butyl ester)
                    derivative 11 was prepared in a similar manner to the synthetic
                    route in Scheme 1 but employing
                        tert-butyl para-fluorobenzoate in place of
                        para-fluorobenzonitrile (see Experimental Section for full
                    details). As shown in Scheme 2, treatment
                    of 11 with TFA furnished the para-carboxylic acid
                        12, which was then converted to
                    para-carboxamide 13 using HBTU and ammonium
                    chloride. para-Phenyl derivative 14 was prepared
                    from the previously reported intermediate
                        1-tert-butoxycarbonylamino-2-[biphenyl-4-yl-(3-methyl-3H-imidazol-4-ylmethyl)amino]-ethane
                    (compound 9a in ref 29b; see
                    Experimental Section for full details). As illustrated in Table 4, all replacements for the cyano group afforded
                    compounds that were poorer inhibitors of hFTase in vitro. Interestingly, FTIs
                    with bulky X groups (11: X = CO2t-Bu;
                        14: X = Ph) exhibited a reversed selectivity for GGTase-I over
                    hFTase, suggesting the aniline-binding domain may be larger in GGTase-I than in
                    hFTase.</p><!><p>In addition to the para-substituent of the aniline
                    affecting inhibitor activity, GOLD docking studies suggested that two of the
                    polar amino acids within the para-benzonitrile-binding pocket
                    might influence binding of the aniline ring itself. Specifically,
                        Y166α could make contact with hydrogen bonding
                    acceptor or donor groups in the ortho and meta
                    positions of the aniline to improve inhibitor binding in mammalian FTase, and
                    likewise, H201α might be exploited with an appropriate
                    group in the meta position. Accordingly, a series of FTase
                    inhibitors was prepared in which the para-benzonitrile portion
                    was modified in an attempt to target these two amino acids; the synthetic route
                    that was pursued is shown in Scheme 3. We
                    selected compound 1az bearing the
                    N-(2-pyrimidinyl)-piper-idin-4-ylmethyl R3 group as
                    the benchmark, anticipating that its moderate activity would better highlight
                    the effects of modifying the para-benzonitrile component.
                    Again, with the only constraint being that the non-methylated
                        τ-nitrogen of the
                    1-methyl-1H-imidazol-5-ylmethyl group should bind the active
                    site Zn2+ ion, several GOLD docking experiments of compound
                        1az were peformed. While the R3 =
                    N-(2-pyrimidinyl)-piperidin-4-ylmethyl and the
                        1-methyl-1H-imidazole-4-sulfonyl groups in 1az
                    bound differently to the corresponding groups in 1a, the
                        R4 = para-benzonitrile group under investigation
                    bound in the same subpocket (tetramethylene side chain of
                        K164α and side chain of
                        Y166α) as the corresponding motif in 1a
                    in four out of the five highest scoring (lowest energy) docking solutions (for
                    example, Figure 4A). Furthermore, in most
                    of those cases, the para-benzonitrile moieties of
                        1a and 1az overlaid excellently, as illustrated in
                        Figure 4B. Thus, we were confident the
                    SAR data that would be acquired by varying the R4 aniline component
                    of 1az would be directly translatable to 1a and
                    related inhibitors.</p><p>As depicted in Scheme 3,
                        mono-N-Boc-ethylenediamine (6) was
                    sulfonylated with 1-methyl-1H-imidazole-4-sulfonyl chloride to
                    give 15 in 95% yield. Chemoselective alkylation of the more acidic
                    and less hindered sulfonamide NH was then readily accomplished by treatment of
                        15 with N-(2-pyrimidinyl)-piperidin-4-ylmethyl
                    bromide in the presence of cesium carbonate in DMF. After TFA-mediated Boc
                    deprotection of 16, arylation of the resultant primary amine was
                    achieved by heating 17 at 120 °C in DMSO with a series of
                    aryl fluorides, giving the secondary anilines 18 in a range of
                    yields from 48 to 97%. Finally, these secondary anilines were then smoothly
                    alkylated in good to excellent yields by deprotonation with NaH in DMF, followed
                    by reaction with 5-chloromethyl-1-methyl-1H-imidazole ·
                        HCl29b to furnish the FTase
                    inhibitors 19a–19h (Table 5).</p><p>Positioning a chlorine (19a) at the ortho
                    position had little effect relative to the parent compound 1az on
                    the inhibition of hFTase. Conversely, replacement of an
                    ortho-CH unit with nitrogen to give pyridine 19c
                    conferred approximately 4-fold improvement in hFTase enzymatic inhibitory
                    activity (19c 110 ± 26 nM vs 1az 510 ±
                    320 nM), while meta-substituted pyridine 19b
                    offered no enhancement in activity, suggesting an important role for a
                    hydrogen-bonding acceptor group in the ortho position. However,
                    incorporation of nitrogens at both ortho positions, as in
                    pyrimidine 19d, caused a reduction in the IC50 value back to
                    approximately the same as that exhibited by the parent compound
                    1az. Inhibitor 19e with a fluorine at one of the
                        meta positions was around twice as potent as
                        1az, while a fluorine at the ortho position
                    led to an especially potent inhibitor (19f) with an hFTase
                        IC50 of 64 ± 8.6 nM, approximately 8-fold as active as
                        1az and twice as potent as the ortho-pyridine
                        19c. Introduction of a second fluorine atom at the other
                        ortho position (inhibitor 19g) caused a
                    reduction in inhibition potency relative to the singly
                    ortho-substituted inhibitor 19f, as was observed
                    with the pyridine derivatives 19c and 19d
                    (pyrimidine), respectively. Replacement of all four ring hydrogens in
                        19h offered no benefit to hFTase inhibition, relative to parent
                        1az. Finally, it is noteworthy that all derivatives based on
                    the parent compound 1az generally exhibited very good to excellent
                    selectivity for inhibition of hFTase over GGTase-I, particularly for
                        ortho-pyridine 19c and
                    ortho-fluoride 19f.</p><!><p>After optimizing the R1,R2,R3,
                        R4, and X groups, we next investigated modifying the
                    ethylenediamine core with a variety of alternative diamino-based scaffolds,
                    whose structures are depicted in Table 6
                    and whose syntheses have been described elsewhere.30 1,3-Diaminopropane-based inhibitor 20a has
                    increased conformational flexibility relative to the corresponding
                    ethylenediamine-based inhibitor 1a. The data in Table 6 suggest that increasing the
                    flexibility in this way caused more than a 7-fold drop in hFTase inhibitory
                    activity. Likewise, reducing the conformational flexibility of the scaffold with
                    the 1,2- and 1,3-diaminocyclopentyl derivatives
                        20b–20e also led to a decrease in hFTase
                    inhibitor potency, with the cis-configurations exhibiting half
                    the potencies of their trans-counterparts. Similar trends were
                    observed in PfFTase.30
                    Of all the different scaffolds examined, the ethylenediamine unit was found to
                    be optimal at delivering the four substituents into the proposed subpockets as
                    well as furnishing the greatest GGTase-I/hFTase selectivity.</p><!><p>Finally, we designed three ethylenediamine-based inhibitors
                    incorporating optimized R1, R2, R3,
                        R4, and X substituents that were identified from our SAR studies,
                    as illustrated by compounds 21a–21c in Table 7. Primary data considered in the
                    selection of moieties that we hoped would furnish optimized inhibitors were in
                    vitro FTase inhibition, whole cell H-Ras processing data, and to a lesser
                    extent, GGTase-I/hFTase selectivity. The R1 group selected was methyl
                        (Table 1), R2 was
                    2-pyridinesulfonyl (Table 2),
                        R3 was cyclohexylmethyl (Table
                        3), R4 was para-benzonitrile (Table 4) substituted at the
                        ortho position (Table
                        5), and the core scaffold incorporated was the simple ethylenediamine
                    unit (Table 6), leading to inhibitors
                        21a, 21b, and 21c in Table 7. As a comparison, we have also
                    included in Table 7 three of our most
                    potent "unoptimized" FTIs: 1a, 1ax, and
                        1f. The trend observed in Table
                        5 upon varying the ortho-CH unit to CF and to N was
                    reproduced with these optimized inhibitors, however none of compounds
                        21a–21c exhibited improved activity over
                    the leads 1a, 1ax, and 1f, possibly due
                    to the adoption of different binding geometries. Much reduced selectivities for
                    hFTase over GGTase-I were also observed. Nonetheless, 21b was
                    equipotent with 1ax in whole cells, whereby both inhibitors
                    disrupted H-Ras processing with IC50 values of about 90 nM.</p><!><p>Figure 5 shows the crystal
                    structure of 1a (yellow; taken from Figure 2A) overlaid with the flexible ligand GOLD docking of
                        1a (green; taken from Figure
                        1B). We correctly predicted that the more basic imidazole
                        1-methyl-1H-imidazol-5-ylmethyl would bind the zinc ion
                    (which was the only constraint implemented in the GOLD docking experiments), and
                    GOLD subsequently correctly placed the benzyl moiety of 1a in very
                    close proximity to the sub-pocket formed by W102β,
                        W106β, and Y361β. However,
                    as Figure 5 illustrates, the
                        para-benzonitrile and the sulfonylimidazole substituents
                    were found to bind in different manners to those predicted. Specifically, the
                        para-benzonitrile moiety was predicted to bind in the
                    subpocket created by K164α,
                        N165α, H201α,
                        Y166α, but was actually found engaged in a weak
                        π–π interaction with
                        Y361β in the product exit groove,
                        F360β, Y93β,
                        L96β, W106β, while the
                    sulfonylimidazole group was predicted to bind the
                        Arg202β, yet was found stacked between the
                    Zn-binding imidazole and the first isoprene of FPP.</p><p>Despite the GOLD prediction of a very different binding mode for the
                        para-benzonitrile moiety, several potent inhibitors (e.g.,
                        19f, Table 5) were
                    serendipitously identified as a consequence of the
                    para-benzonitrile making alternative but still significant
                    contacts. The observed
                        π–π stacking of the
                        para-benzonitrile moiety against the
                        Y361β residue suggests an explanation for the
                    observed in vitro SAR data for the modifications made to the
                        para-benzonitrile ring of 1a (Tables 1 and Table 4) and 1az (Table 5).
                        π–π Stacking
                    interactions are known to be energetically more favorable between an
                    electron-rich arene, here Y361β, and an electron-poor
                    arene, here the para-benzonitrile of inhibitor 1a.
                    For the series of inhibitors in Table 4,
                    substitution of the para position of the aniline with a
                    strongly electron-withdrawing cyano group led to the most potent inhibitor
                        (1a) while eliminating all substitution from the aniline ring
                    of 1a and with R1 = H essentially abolished activity
                    (compound 2, Table 1).
                    Additionally, the product exit groove where the
                    para-benzonitrile binds is of limited size and mostly
                    hydro-phobic, suggesting that substituents larger than the linear cyano unit,
                    such as the bulky tert-butyl ester in 11 or the
                    phenyl ring in 14, and more polar than the cyano group, such as the
                    acid in 12 and the carboxamide in 13, would be poorly
                    tolerated. This was confirmed experimentally (Table 4). For the series of inhibitors in Table 5, replacement of the ortho (with
                    respect to the aniline nitrogen) CH (1az) with either N
                        (19c)or CF(19f) led to an approximate 5-fold or
                    8-fold improvement in inhibitor activity, respectively. This may be due to the
                    introduced electron-withdrawing groups that render the aniline even more
                    electron-poor, thereby enhancing its interaction with
                        Y361β. On the other hand, replacement of the
                        meta-CH (1az) with N (19b)
                    resulted in no improvement in activity, while substitution of the
                        meta-CH with a CF group (19e) led to only a
                    2-fold increase in inhibitor potency, as opposed to the 8-fold enhancement
                    achieved by ortho-fluoride 19f. The crystal
                    structure of 1a suggests that the incorporation of electronegative
                    groups in the meta position might be poorly tolerated due to a
                    clash with the anionic side chain of D359β, explaining
                    the reduced enhancement in potency of inhibitors 19b and
                        19e (as a means of avoiding this unfavorable interaction,
                    180° rotation about the N–C aniline bond would direct the
                    introduced N or CF group into a hydrophobic domain comprising
                        L96β, W102β and
                        W106β and would therefore also be unfavorable).
                    Replacement of both ortho-CH groups with either two nitrogens
                        (19d) or two CF groups (19g) was not tolerated
                    relative to the mono-ortho-substituted inhibitors
                        (19b and 19e, respectively). This observation may
                    be a consequence of one of the electronegative groups being forced into the
                    hydrophobic L96β, W102β and
                        W106β region in order that the energetically
                    favorable π–π stacking between the aniline
                    ring of the inhibitor and Y361β is maintained.</p><p>Generally, there was little variation in activity among the
                        R2 sulfonyl derivatives shown in Table 2, which may be a consequence of this moiety exhibiting
                    insignificant interactions with the protein itself, being found stacked between
                    the zinc-binding imidazole and FPP. However, there were clear trends in the
                    hFTase inhibition data for the R3 series of sulfonamide analogues
                        (Table 3). The crystal structure of
                        1a illustrates that the R3 phenyl ring can access
                    the a2 residue binding site (L96β,
                        W102β and W106β) of the
                        Ca1a2X substrate. For the most part, small,
                    hydrophobic R3 groups afforded potent hFTase inhibitors, likely due
                    to the R3 group making van der Waals contacts with
                        L96β, W102β and/or
                        W106β. Within a particular series of R3
                    substituents, for example the ortho-, meta-,
                    and para-tolyl derivatives 1ao, 1ap,
                    and 1aq, respectively, ortho- and
                        meta-functionalized compounds were tolerated, whereas
                        para-functionalization was not. This may be a consequence
                    of the para derivative being too large to access the
                        a2 residue binding site. Indeed, the crystal structure of
                        1ay shows that the large
                    N-Bocpiperidin-4-ylmethyl group traverses, rather than binds
                    in, the a2 residue binding site and instead reaches into the X
                    residue binding site.38 We have
                    previously reported on the relative selectivities of our ethylenediamine-based
                    inhibitors for the Plasmodium isoform of FTase over the
                    mammalian isoform.29,30 With this crystal structure data in
                    hand, along with several others,38 it
                    should now be possible to tailor these compounds to become more selective for
                    one enzyme isoform over the other.</p><!><p>In summary, we have developed a potent series of mammalian
                farnesyltransferase inhibitors (FTIs) based on an ethylenediamine scaffold. This
                class of compounds was identified by a "piggy-back" approach on our
                potent antimalarial inhibitors of plasmodium FTase. The simple and cost-effective
                ethylenediamine core allowed facile access to a diverse array of inhibitors, greatly
                facilitating lead inhibitor optimization. We have identified several inhibitors with
                double-digit nano-molar inhibition of hFTase in vitro, the most potent compound
                being 1f (IC50 = 25 nM). In most cases, potent inhibition of
                hFTase in vitro was accompanied by potent whole cell data (inhibition of H-Ras
                processing); for example, inhibitor 1f displayed a whole cell
                    IC50 of 90 nM, one of the most active of the entire series. Moreover,
                in all but two cases inhibitors were selective for hFTase over GGTase-I, with the
                greatest selectivity (333-fold) exhibited by inhibitor 1g. Finally, we
                have solved the crystal structure of one of our ethylenediamine-based FTIs
                    (1a) in the active site of rFTase, and this will assist in the
                future design of more potent and isoform-selective mammalian farnesyltransferase
                inhibitors.</p><!><p>Ligand energy minimization was peformed with the CVFF force field in
                    InsightII on an SGI O2. Flexible ligand docking studies were subsequently
                    performed with GOLD, version 3.0,32 on a
                    Linux PC. Default GOLD parameters were utilized with the following exceptions:
                    (i) maximal ligand flexibility was allowed; (ii) the affinity of nitrogen for
                    zinc ion was increased in the GOLD parameters file to better reflect the ability
                    of an imidazole ring to bind to the active site metal ion. Each ligand was used
                    to seed the genetic algorithm 10 times.</p><!><p>Solvents CH2Cl2,CH3CN, and DMF were
                    dried on an Innovative Techonology SPS-400 dry solvent system. Anhydrous MeOH
                    and DMSO were purchased from Sigma-Aldrich and used directly from their
                    Sure-Seal bottles. Molecular sieves were activated by heating to 300 °C
                    under vacuum overnight. All reactions were performed under an atmosphere of dry
                    nitrogen in oven-dried glassware and were monitored for completeness by
                    thin-layer chromatography (TLC) using silica gel (visualized by UV light, or
                    developed by treatment with KMnO4 stain or Hanessian's stain).
                        1H and 13C NMR spectra were recorded on Bruker AM 400
                    MHz and Bruker AM 500 MHz spectrometers in either CDCl3,
                        MeOH-d4, or DMSO-d6.
                    Chemical shifts (Î´) are reported in parts per million
                    after calibration to residual isotopic solvent. Coupling constants
                        (J) are reported in Hz. Mass spectrometry was performed
                    using electrospray ionization on either a Varian MATCH-5 (HRMS) or Waters
                    Micromass ZQ (LRMS). Before biological testing, all new target molecules
                        (1b, 1c, 1ah–1an,
                        1aw, 11–14, 19a–19h,
                        21a–21c) were subjected to further purification by
                    reversed-phase HPLC (rpHPLC). Analysis and purification by rpHPLC were performed
                    using either a Phenonenex Luna 5 Î¼m C18 (2) 250 mm
                    × 21 mm column run at 15 mL/min (preparative) or a Microsorb-MV 300 A C18
                    250 mm × 4.6 mm column run at 1 mL/min (analytical), using gradient
                    mixtures of (A) water with 0.1% TFA and (B) 10:1 acetonitrile/water with 0.1%
                    TFA. Appropriate product fractions were pooled and lyophilized to dryness,
                    affording the inhibitors as fluffy, white powders as their TFA salts. Inhibitor
                    purity was confirmed by analytical rpHPLC using linear gradients from 100% A to
                    100% B, with changing solvent composition of either (I) 4.5% or (II) 1.5% per
                    min after an initial 2 min of 100% A. For reporting HPLC data, percentage purity
                    is given in parentheses after the retention time for each condition. FTase
                    inhibitors 1a, 1d–1h,
                        1aa–1ag, 1ao–1av,
                        1ax–1az, 3–5, and
                        20a–20e have been previously reported in ref 29b.</p><!><p>The appropriate sulfonyl chloride (1.2 equiv) was added to a solution of
                    the amine (1 equiv) and DIPEA (2 equiv) in anhydrous CH3CN (0.1 M) at
                    0°C. The reaction was warmed to room temperature and stirred for 16 h, at
                    which time the solvent was evaporated. The residue was redissolved in
                        CH2Cl2, washed with 5% NaHCO3, water,
                    brine, dried (Na2SO4), filtered, and concentrated.</p><!><p>To a solution of the sulfonamide (1 equiv) and
                        Cs2CO3 (2 equiv) in DMF (0.1 M) at 0 °C was
                    added the appropriate alkyl bromide or chloride (1.1 equiv). The resulting
                    mixture was stirred at room temperature overnight. The reaction was diluted with
                    water, then extracted into EtOAc (×3). The combined EtOAc extractions
                    were washed with 5% NaHCO3 (×3), brine, dried
                        (Na2SO4), filtered, and concentrated.</p><!><p>To a solution of 9 (1 equiv) in dry methanol (0.2 M) with 4
                    Å molecular sieves was added the appropriate aldehyde (1.1 equiv) and
                    acetic acid (1.3 equiv). The reaction was stirred under nitrogen at room
                    temperature for 30 min. Sodium cyanoborohydride (1.5 equiv) was added, and the
                    resulting suspension was stirred at room temperature overnight. The reaction
                    mixture was decanted, washing thoroughly with further MeOH, and then dry-loaded
                    onto silica gel.</p><!><p>To a stirring solution of the primary amine (1 equiv) in DMSO (0.2 M)
                    were added the aryl fluoride (1.2 equiv) and DIPEA (3 equiv). The reaction
                    mixture was heated to 120 °C for 48 h. After allowing the reaction to
                    cool, H2O was added, and the crude product was extracted with EtOAc
                    (×3). The EtOAc extractions were combined, washed with water (×3),
                    brine, dried (Na2SO4), filtered, and concentrated.</p><!><p>The secondary aniline (1 equiv) was dissolved in DMF (0.07 M), and then
                    the reaction was cooled to 0 °C. After 15 min, NaH (3 equiv) was added in
                    one portion. After a further 15min,
                    5-chloromethyl-1-methyl-1H-imidazole·3 HCl31 (1.1 equiv) was added. The reaction was
                    allowed to stir at 0 °C from 2–3 h, when TLC indicated the
                    reaction was complete or had stalled. Upon quenching the reaction with brine
                    (approximately 1 mL for 1 mmol scale), the reaction was diluted with water and
                    extracted with EtOAc (×3). The EtOAc extractions were combined, and
                    washed with 5% NaHCO3 (×3), brine, dried
                        (Na2SO4), filtered, and concentrated.</p><!><p>A stirring solution of the aryl bromide (1 equiv) in DMF (0.1 M) was
                    degassed, and then Zn(CN)2 (0.6 equiv),
                        Pd(PPh3)4 (0.1 equiv), Zn dust (0.05 equiv), and
                        Zn(OAc)2 (0.05 equiv) were added. The mixture was heated to 120
                    °C for 2 h, after which time the reaction was allowed to cool, diluted
                    with water, and extracted into EtOAc (×3). The EtOAc extractions were
                    combined and washed with 5% NaHCO3 (×3), brine, dried
                        (Na2SO4), filtered, and concentrated.</p><!><p>To a solution of
                        4-[(2-{N-tert-butoxycarbonyl}-aminoethyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-benzonitrile
                    (compound 9c in ref 29b)
                    (8.0 g, 30.6 mmol) in CH2Cl2 (75mL) cooled to 0 °C
                    was added TFA (75mL). The reaction was warmed to room temperature and stirred
                    for 30 min, after which time the solvent was concentrated in
                        vacuo, and then the crude material was repeatedly re-dissolved in
                        CHCl3 and re-evaporated until a constant weight was achieved, to
                    afford the ditrifluoroacetic acid salt of 9 as a light-brown solid
                    (14.8 g, 100%): Î´H (500 MHz,
                        MeOH-d4)3.20(t, J
                        =7.1Hz,2H,CH2CH2NH2), 3.81
                    (t, J = 7.1 Hz, 2H,
                        CH2CH2NH2), 3.87 (s, 3H,
                        CH3 (Im)), 4.82 (s, 2H, CH2Im), 6.96
                    (d, J = 9.0 Hz, 2H, 2 CH(Ar)), 7.19 (s, 1H, CH (Im)), 7.56 (d,
                        J = 9.0 Hz, 2H, 2 CH (Ar)), 8.88 (s, 1H, CH (Im));
                        Î´C (125 MHz,
                        MeOH-d4) 33.9, 37.1, 45.3, 48.2, 100.8, 114.0,
                    118.4, 120.4, 132.3, 134.6, 137.5, 151.0; HRMS (ESI) m/z calcd
                    for [C14H17N5 + H] 256.1562, obsd 256.1559.
                    Subsequently, the free base form of 9, which is the form used for
                    all subsequent reactions, was furnished by passing the di-TFA salt through a
                    short pad of silica gel (eluent
                    CH2Cl2/MeOH/NH4OH, 92:7:1).</p><!><p>Compound 9 was treated with propane-1-sulfonyl chloride,
                    according to general procedure A, on a 0.15 mmol scale. The crude material was
                    purified by silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1) to give
                        N-{2-[(4-cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}
                    propane-1-sulfonamide as a colorless film (46 mg, 85%):
                        Î´H (500 MHz,
                        MeOH-d4)0.98 (t, J = 7.5 Hz, 3H,
                        CH2CH2CH3), 1.71
                    (sextet, J = 7.5 Hz, 2H,
                        CH2CH2CH3), 2.96 (m,
                    2H, NHCH2CH2N), 3.21–3.26
                    (m, 2H, CH2CH2CH3), 3.61
                    (t, J = 6.5 Hz, 2H,
                        NHCH2CH2N), 3.84 (s, 3H,
                        CH3 (Im)), 4.79 (s, 2H, CH2Im),
                    6.91 (d, J = 9.1 Hz, 2H, 2CH (Ar)), 7.20 (s, 1H, CH (Im)), 7.47
                    (d, J = 9.1 Hz, 2H, 2 CH (Ar)), 8.82 (s, 1H, CH (Im));
                        Î´C (125 MHz,
                        MeOH-d4) 11.7, 16.9, 32.8, 39.7, 44.4, 50.6,
                    53.2, 98.9, 112.6, 117.7, 119.4, 131.5, 133.3, 136.2, 150.4; LRMS (ESI)
                        m/z 384.2
                        [C17H23N5O2S+Na]. Subsequently,
                    the sulfonamide (36 mg, 0.1 mmol) was treated with benzyl bromide according to
                    general procedure B. After the usual workup and flash chromatography over silica
                    gel (eluent CH2Cl2/MeOH/NH4OH, 192:7:1), the
                    title compound 1b was furnished as a glassy film (37 mg, 82%):
                        Î´H (400 MHz,
                        MeOH-d4)1.20(t, J = 7.4 Hz, 3H,
                        CH2CH2CH3), 1.99 (m,
                    2H, CH2CH2CH3), 3.24 (t,
                        J = 7.8 Hz, 2H,
                        N(Bn)CH2CH2N), 3.40 (obscured,
                    4H, NHCH2CH2N,
                        CH2CH2CH3), 3.94 (s,
                    3H, CH3 (Im)), 4.52 (s, 2H, CH2Ph),
                    4.65 (s, 2H, CH2Im), 6.75 (d,
                        J = 8.9 Hz, 2H, 2 CH (Ar)), 7.25 (s, 1H, CH (Im)), 7.55 (m,
                    7H, 5 CH (Ph), 2 CH (Ar)), 9.00 (s, 1H, CH (Im));
                        Î´C (125 MHz,
                        MeOH-d4) 13.7, 18.6, 34.6, 45.4, 46.8, 51.6,
                    53.4, 55.1, 100.3, 114.1, 119.6, 121.2, 129.8, 130.7, 133.1, 135.1, 138.2,
                    138.7, 151.9, 152.9; HRMS (ESI) m/z calcd for
                        [C24H29N5O2S+H] 452.2120, obsd
                    452.2108; HPLC (I) tR = 13.45 min (100%), (II)
                        tR = 19.72 min (100%).</p><!><p>First, amine 9 was reacted with cyclopropanesulfonyl
                    chloride, according to general procedure A, on a 0.15 mmol scale. After workup
                    and flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1),
                        N-{2-[(4-cyanophenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}
                    cyclopropanesulfonamide was yielded as a colorless film (44 mg, 82%):
                        Î´H (500 MHz,
                        MeOH-d4) 0.98 (m, 4H, 2 CH2
                    (cyclopropyl)), 2.51 (m, 1H, CH (cyclopropyl)), 3.33 (t, J =
                    5.8 Hz, 2H, NHCH2CH2N), 3.67 (t,
                        J = 5.8 Hz, 2H,
                        NHCH2CH2N), 3.88 (s, 3H,
                        CH3 (Im)), 4.76 (s, 2H, CH2Im),
                    6.95 (d, J = 8.83 Hz, 2H, 2 CH (Ar)), 7.25 (s, 1H, CH (Im)),
                    7.53 (d, J=8.84 Hz, 2H, 2CH (Ar)), 8.86 (s, 1H, CH (Im));
                        Î´C (125 MHz,
                        MeOH-d4) 4.1, 29.0, 32.8, 39.9, 44.5, 50.5,
                    99.1, 112.7, 117.8, 119.4, 131.6, 133.4, 136.3, 150.4; LRMS (ESI)
                        m/z 382.1
                        [C17H21N5O2S + Na].
                    Subsequently, the sulfonamide (36 mg, 0.1 mmol) was treated with benzyl bromide,
                    according to general procedure B. The crude material was dry-loaded onto silica
                    gel and purified by flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to give 1c as a
                    white foam (40 mg, 89%): Î´H (500 MHz,
                        MeOH-d4) 0.95 (m, 4H, 2 CH2
                    (cyclopropyl)), 2.55 (m, 1H, CH (cyclopropyl)), 3.31 (m, 4H,
                        NCH2CH2N),
                    3.71 (s, 3H, CH3 (Im)), 4.32 (s, 2H,
                        CH2Ph), 4.43 (s, 2H,
                        CH2Im), 6.51 (d, J = 8.89
                    Hz, 2H, 2 CH (Ar)), 7.02 (s, 1H, CH (Im)), 7.26–7.33 (m, 7H, 5 CH (Ph), 2
                    CH (Ar)), 8.76 (s, 1H, CH (Im)); Î´C (125 MHz,
                        MeOH-d4) 5.3, 28.1, 34.3, 45.1, 46.3, 51.1,
                    54.9, 100.3, 113.7, 119.1, 120.8, 129.4, 129.9, 130.3, 132.7, 134.7, 137.8,
                    138.2, 151.5; HRMS (ESI) m/z calcd for
                        [C24H27N5O2S + H] 450.1964, obsd
                    450.1985; HPLC (I) tR = 12.70 min (100%), (II)
                        tR = 18.99 min (100%).</p><!><p>Furan-3-carbaldehyde was reductively aminated with amine 9
                    according to general procedure C, on a 0.15 mmol scale. Purification by flash
                    column chromatography (eluent
                    CH2Cl2/MeOH/NH4OH, 92:7:1) then gave
                        4-[{2-[(furan-3-ylmethyl)-amino]-ethyl}-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-benzonitrile
                    as a film (39 mg, 78%): Î´H (400 MHz,
                        MeOH-d4) 3.24 (obscured t, 2H,
                        NHCH2CH2N), 3.78 (m, 5H,
                        NHCH2CH2N, CH3
                    (Im)), 4.07 (s, 2H, CH2-furan), 4.75 (s, 2H,
                        CH2Im), 6.50 (s, 1H, CH (furan)), 6.88 (d,
                        J =9.0 Hz, 2H, 2 CH (benzonitrile)), 7.11 (s, 1H, CH
                    (furan)), 7.42–7.53 (m, 3H, 2 CH (benzonitrile), CH (Im)), 7.62 (s, 1H,
                    CH (furan)), 8.82 (s, 1H, CH (Im)); Î´C (100
                    MHz, MeOH-d4) 34.5, 43.5, 44.7, 45.9, 47.7, 101.7,
                    111.9, 114.5, 117.3, 119.0, 120.8, 132.9, 135.2, 138.2, 145.1, 146.0, 151.5;
                    LRMS (ESI) m/z 358.2
                    [C19H21N5O + Na]. This secondary amine (34 mg,
                    0.1 mmol) was reacted with 1-methyl-1H-imidazole-4-sulfonyl
                    chloride according to general procedure A to yield the title compound
                        1ah as a white foam (43 mg, 90%):
                        Î´H (500 MHz,
                        MeOH-d4) 3.38 (obscured t, 2H,
                        ((furan-3-ylmethyl)-NHCH2CH2N)),
                    3.56 (t, J = 7.0 Hz, 2H,
                        ((furan-3-ylmethyl)-NHCH2CH2N)),
                    3.81 (s, 3H, CH3 (Im)), 3.89 (s, 3H, CH3 (Im)), 4.19 (s,
                    2H, CH2-furan), 4.70 (s, 2H,
                        CH2Im), 6.57 (s, 1H, CH (furan)), 6.77 (d,
                        J = 8.94 Hz, 2H, 2 CH (benzonitrile)), 7.25 (s, 1H, CH
                    (furan)), 7.46–7.52 (m, 4H, 2 CH (benzonitrile), 2 CH (Im)), 7.78 (s, 1H,
                    CH (furan)), 7.81 (s, 1H, CH (Im)), 8.91 (s, 1H, CH (Im));
                        Î´C (125 MHz,
                        MeOH-d4) 34.3, 34.4, 45.3, 45.5, 45.9, 51.0,
                    100.4, 111.9, 113.8, 119.3, 120.8, 122.0, 126.8, 132.9, 134.8, 137.9, 139.1,
                    141.5, 143.2, 145.3, 151.7; HRMS (ESI) m/z calcd for
                        [C23H25N7O3S + H] 480.1818, obsd
                    480.1809; HPLC (I) tR = 12.62 min (100%).</p><!><p>Thiophene-3-carbaldehyde was reductively aminated with amine
                        9 (38 mg, 0.15 mmol) according to general procedure C to give,
                    after flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1),
                        4-((3-methyl-3H-imidazol-4-ylmethyl)-{2-[(thiophen-3-ylmethyl)-amino]-ethyl}-amino)-benzonitrile
                    as a glassy film (40 mg, 76%): Î´H (400 MHz,
                        MeOH-d4) 3.20 (obscured t, 2H,
                        NHCH2CH2N), 3.70–3.75
                    (m, 5H, NHCH2CH2N, CH3
                    (Im)), 4.16 (s, 2H, CH2-thiophene), 4.69 (s,
                    2H, CH2Im), 6.74 (d, J = 9.0
                    Hz, 2H, 2 CH (benzonitrile)), 7.06 (s, 1H, CH (Im)), 7.09 (d, J
                    = 3.9 Hz, 1H, CH (thiophene)), 7.41–7.48 (m, 4H, 2CH (benoznitrile), 2 CH
                    (thiophene)), 8.77 (s, 1H, CH (Im)); Î´C (100
                    MHz, MeOH-d4) 34.7, 45.0, 46.1, 47.4, 47.9, 102.0,
                    114.7, 119.2, 120.9, 129.0, 129.1, 129.5, 133.1, 133.2, 135.4, 138.4, 151.6;
                    LRMS (ESI) m/z 374.2
                    [C19H21N5S + Na]. This secondary amine (35 mg,
                    0.1 mmol) was then reacted with
                    1-methyl-1H-imidazole-4-sulfonyl chloride according to general
                    procedure A to give, after purification by silica gel flash column
                    chromatography (eluent CH2Cl2/MeOH/NH4OH,
                    192:7:1), compound 1ai as a white foam (46 mg, 93%):
                        Î´H (400 MHz,
                        MeOH-d4)3.29(obscured t, 2H,
                        ((thiophen-3-ylmethyl)NCH2CH2N)),
                    3.39 (t, J =6.9Hz, 2H,
                        ((thiophen-3-ylmethyl)NCH2CH2N)),
                    3.69 (s, 3H, CH3 (Im)), 3.75 (s, 3H, CH3 (Im)), 4.20 (s,
                    2H, 2 CH (CH2-thiophene)), 4.50 (s, 2H,
                        CH2Im), 6.57 (d, J = 9.1
                    Hz, 2H, 2 CH (benoznitrile)), 6.95 (d, J = 3.8 Hz, 1H, CH
                    (thiophene)), 7.09 (s, 1H, CH (thiophene)), 7.22 (s, 1H, CH (Im)), 7.31 (m, 1H,
                    CH (thiophene)), 7.36 (d, J = 9.1 Hz, 2H, 2 CH (benzonitrile)),
                    7.65 (s, 1H, CH (Im)), 7.70 (s, 1H, CH (Im)), 8.79 (s, 1H, CH (Im));
                        Î´C (125 MHz,
                        MeOH-d4) 34.7, 34.8, 45.5, 46.7, 50.3, 51.5,
                    100.7, 114.1, 119.6, 121.2, 126.2, 127.2, 128.3, 129.7, 133.2, 135.2, 138.2,
                    139.0, 139.5, 141.9, 152.0; HRMS (ESI) m/z calcd for
                        [C23H25N7O2S2 + H]
                    496.1589, obsd 496.1576; HPLC (I) tR = 12.64 min
                    (99.0%), (II) tR = 18.84 min (98.2%).</p><!><p>Furan-2-carbaldehyde was reductively aminated with amine 9
                    according to general procedure C, on a 0.15 mmol scale. Purification by silica
                    gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1) afforded
                        4-[{2-[(furan-2-ylmethyl)-amino]-ethyl}-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-benzonitrile
                    as a film (39 mg, 78%): Î´H (500 MHz,
                        MeOH-d4) 3.31 (obscured t, 2H,
                        NHCH2CH2N), 3.84 (t,
                        J = 7.4 Hz, 2H,
                        NHCH2CH2N), 3.87 (s, 3H,
                        CH3 (Im)), 4.33 (s, 2H,
                    CH2-furan), 4.81 (s, 2H,
                        CH2Im), 6.48 (m, 1H, CH (furan)), 6.61 (d,
                        J = 3.19 Hz, 1H, CH (furan)), 6.95 (d, J =
                    8.9 Hz, 2H, 2 CH (benzonitrile)), 7.18, (s, 1H, CH (Im)), 7.54 (d,
                        J = 8.9 Hz, 2H, 2 CH (benzonitrile)), 7.60 (m, 1H, CH
                    (furan)), 8.88 (s, 1H, CH (Im)); Î´C (125 MHz,
                        MeOH-d4) 32.8, 43.0, 43.1, 44.2, 46.0, 100.1,
                    110.7, 112.4, 113.0, 117.4, 119.0, 131.2, 133.5, 136.5, 144.5, 144.8, 149.7;
                    LRMS (ESI) m/z 358.2 [C19H21N5
                    O + Na]. This secondary amine was reacted with
                    1-methyl-1H-imidazole-4-sulfonyl chloride according to general
                    procedure A, on a 0.1 mmol scale. After workup, the crude material was purified
                    by silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to furnish the
                    title compound 1aj as a white foam (43 mg, 90% 47.91818):
                        Î´H (500 MHz,
                        MeOH-d4) 3.34 (t, J = 6.9 Hz,
                    2H, ((furan-3-ylmethyl)NCH2CH2N)),
                    3.46 (t, J = 6.9 Hz, 2H,
                        ((furan-3-ylmethyl)NCH2CH2N)),
                    3.67 (s, 3H, CH3 (Im)), 3.79 (s, 3H, CH3 (Im)), 4.23 (s,
                    2H, CH2-furan), 4.63 (s, 2H,
                        CH2Im), 6.18 (m, 1H, CH (furan)), 6.23 (m,
                    1H, CH (furan)), 6.73 (d, J = 9.1 Hz, 2H, 2 CH (benzonitrile)),
                    7.15 (s, 1H, CH (Im)), 7.29 (s, 1H, CH (furan)), 7.41 (d, J =
                    9.1 Hz, 2H, 2 CH (benzonitrile)), 7.59 (s, 1H, CH (Im)), 7.64 (s, 1H, CH (Im)),
                    8.79 (s, 1H, CH (Im)); Î´C (125 MHz,
                        MeOH-d4) 34.3, 45.4, 46.5, 47.1, 49.6
                    (obscured), 50.8, 100.4, 111.3, 111.7, 113.9, 119.3, 120.9, 126.8, 132.9, 134.8,
                    137.8, 139.1, 141.4, 144.4, 150.9, 151.8; HRMS (ESI) m/z calcd
                    for [C23H25N7O3S + H] 480.1818, obsd
                    480.1833; HPLC (I) tR = 12.51 min (100%).</p><!><p>Thiophene-2-carbaldehyde was reductively aminated with amine 9 (38 mg,
                    0.15 mmol) according to general procedure C, giving, after flash column
                    chromatography (eluent CH2Cl2/MeOH/NH4OH,
                    92:7:1),
                    4-((3-methyl-3H-imidazol-4-ylmethyl)-{2-[(thiophen-2-ylmethyl)-amino]-ethyl}-amino)-benzonitrile
                    as a colorless film (38 mg, 72%): Î´H (500
                    MHz, MeOH-d4)3.37 (obscured t, 2H,
                        NHCH2CH2N), 3.8–3.92 (m,
                    5H, NHCH2-CH2N, CH3 (Im)), 4.54 (s, 2H,
                        CH2-thiophene), 4.86, (s, 2H,
                        CH2Im), 6.87 (m, 1H, CH (thiophene)), 7.00
                    (d, J = 8.8 Hz, 2H, 2 CH (benzonitrile)), 7.13 (m, 1H, CH
                    (thiophene)), 7.23 (s, 1H, CH (Im)), 7.34 (m, 1H, CH (thiophene)), 7.59 (d,
                        J = 8.8 Hz, 2H, 2 CH (benzonitrile)), 8.93 (s, 1H, CH
                    (Im)); Î´C (125 MHz,
                        MeOH-d4) 32.8, 42.9, 44.2, 45.0, 45.9, 100.2,
                    113.0, 117.4, 119.0, 127.3, 128.2, 130.6, 131.2, 131.5, 133.5, 136.6, 149.7;
                    LRMS (ESI) m/z 374.2
                    [C19H21N5S + Na]. This secondary amine (35 mg,
                    0.1 mmol) was reactedwith1-methyl-1H-imidazole-4-sulfonyl chloride according to
                    general procedure A. The usual workup and purification by flash column
                    chromatography (eluent CH2Cl2/MeOH/NH4OH,
                    192:7:1) gave 1ak as a glassy film (44 mg, 89%):
                        Î´H (400 MHz,
                        MeOH-d4)3.31 (t, J = 7.1 Hz,
                    2H, ((thiophen-2-yl)NCH2-CH2N)),
                    3.47 (t, J = 7.1 Hz, 2H,
                        ((thiophen-2-yl)NCH2CH2N)),
                    3.69 (s, 3H, CH3 (Im)), 3.75 (s, 3H, CH3 (Im)), 4.41 (s,
                    2H, CH2-thiophene), 4.55 (s, 2H,
                        CH2Im), 6.61, (d, J = 9.0
                    Hz, 2H, 2 CH (benzonitrile)), 6.85 (m, 1H, CH (thiophene)), 6.89 (d,
                        J = 2.70 Hz, 1H), 7.11 (s, 1H, CH (Im)), 7.29 (m, 1H, CH
                    (thiophene)), 7.37 (d, J = 9.0 Hz, 2H, 2 CH (benzonitrile)),
                    7.67 (s, 1H, CH (Im)), 7.69 (s, 1H, CH (Im)), 8.78 (s, 1H, CH (Im));
                        Î´C (100 MHz,
                        MeOH-d4) 34.7, 34.8, 45.7, 46.6, 50.3
                    (obscured), 51.0, 100.9, 114.2, 119.6, 121.2, 126.5, 127.2, 128.5, 130.0, 133.2,
                    135.2, 138.2, 139.5, 140.7, 141.9, 152.0; HRMS (ESI) m/z calcd
                    for [C23H25N7O2S2 + H]
                    496.1589, obsd 496.1585; HPLC (I) tR = 12.90 min
                    (100%).</p><!><p>Amine 9 was reacted with
                    1-methyl-1H-imidazole-4-sulfonyl chloride according to general
                    procedure A, on a 0.15 mmol scale. Flash column chromatography of the crude
                    material over silica gel (eluent CH2Cl2/MeOH/
                        NH4OH, 92:7:1) afforded
                    1-methyl-1H-imidazole-4-sulfonic acid
                        {2-[(4-cyano-phenyl)-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-ethyl}-amide
                    as a white powder (55 mg, 92%): Î´H (400 MHz,
                        MeOH-d4)3.19(t, J = 6.3 Hz, 2H,
                        NHCH2CH2N), 3.64 (t,
                        J = 6.3 Hz, 2H,
                        NHCH2CH2N), 3.72 (s, 3H,
                        CH3 (Im)), 3.88 (s, 3H, CH3 (Im)), 4.81 (s, 2H,
                        CH2Im), 6.89 (d, J = 9.0
                    Hz, 2H, 2 CH (benzonitrile)), 7.22 (s, 1H, CH (Im)), 7.50 (d, J
                    = 9.0 Hz, 2H, 2 CH (benzonitrile)), 7.62 (s, 1H, CH (Im)), 7.71 (s, 1H, CH
                    (Im)), 8.87 (s, 1H, CH (Im)); Î´C (100 MHz,
                        MeOH-d4) 33.8, 33.9, 40.7, 45.4, 50.9, 99.5,
                    113.5, 118.5, 120.5, 125.3, 132.3, 134.2, 137.2, 139.9, 140.7, 151.2; HRMS (ESI)
                        m/z calcd for
                        [C18H21N7O2S + H] 400.1556, obsd
                    400.1545. This secondary sulfonamide was then alkylated with
                    4-chloromethyl-3,5-dimethyl-isoxazole according to general procedure B, on a 0.1
                    mmol scale. After workup, purification was accomplished by silica gel flash
                    column chromatography (eluent
                    CH2Cl2/MeOH/NH4OH, 192:7:1) to furnish the
                    title compound 1al as a white foam (40 mg, 79%):
                        Î´H (500 MHz,
                        MeOH-d4)2.15 (s, 3H, CH3
                    (isoxazole)), 2.20 (s, 3H, CH3 (isoxazole)), 3.23 (t,
                        J = 6.7 Hz, 2H,
                        ((isoxazol-4-ylmethyl)NCH2-CH2N)),
                    3.53 (t, J = 6.7 Hz, 2H,
                        ((isoxazol-4-ylmethyl)NCH2-CH2N)), 3.77 (s, 3H,
                        CH3 (Im)), 3.82 (s, 3H, CH3 (Im)), 4.02 (s, 2H,
                        CH2-isoxazole), 4.56 (s, 2H,
                        CH2Im), 6.66 (d, J = 8.7
                    Hz, 2H, 2 CH (benzonitrile)), 7.17 (s, 1H, CH (Im)), 7.45 (d, J
                    =8.7 Hz, 2H, 2 CH (benzonitrile)), 7.74 (s, 1H, CH (Im)), 7.78 (s, 1H, CH (Im)),
                    8.85 (s, 1H, CH (Im)); Î´C (125 MHz,
                        MeOH-d4) 10.2, 10.9, 34.3, 34.4, 44.4, 45.0,
                    47.0, 51.2, 100.6, 110.7, 113.7, 119.3, 120.8, 127.1, 132.6, 134.9, 137.9,
                    138.2, 141.7, 151.4, 161.6, 169.6; HRMS (ESI) m/z calcd for
                        [C24H28N8O3S H] 509.2083, obsd
                    509.2072; HPLC (I) tR = 12.27 min (100%), (II)
                        tR = 17.76 min (100%).</p><!><p>2,4-Dimethyl-thiazole-5-carbaldehyde was reductively aminated with amine
                        9 (38 mg, 0.15 mmol) according to general procedure C to give,
                    after flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1),
                        4-[{2-[(2,4-dimethyl-thiazol-5-ylmethyl)-amino]-ethyl}-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-benzonitrile
                    as a glassy film (42 mg, 74%): Î´H (500 MHz,
                        MeOH-d4)2.34(s,3H,CH3 (thiazole)),
                    2.60 (s, 3H, CH3 (thiazole)), 3.32 (t, J = 7.06 Hz,
                    2H, NHCH2CH2N), 3.83 (m, 5H,
                        NHCH2CH2N, CH3
                    (Im)), 4.39 (s, 2H, CH2-thiazole), 4.78 (s,
                    2H, CH2Im), 6.92 (d, J = 9.0
                    Hz, 2H, 2 CH (benzonitrile)), 7.14 (s, 1H, CH (Im)), 7.51 (d, J
                    = 9.0 Hz, 2H, 2 CH (benzonitrile)), 8.85 (s, 1H, CH (Im)); LRMS (ESI)
                        m/z 403.2 [C20H24N6S +
                    Na]. The secondary amine (38 mg, 0.1 mmol) was then treated with
                        1-methyl-1H-imidazole-4-sulfonyl chloride according to
                    general procedure A. Workup and purification (flash column chromatography
                    (eluent CH2Cl2/MeOH/NH4OH, 192:7:1)) as usual
                    yielded 1am as a white foam (47 mg, 90%):
                        Î´H (500 MHz,
                        MeOH-d4)2.21 (s, 3H, CH3 (thiazole)),
                    2.55 (s, 3H, CH3 (thiazole)), 3.39 (t, J = 6.5 Hz,
                    2H,
                    ((thiazol-5-ylmethyl)-NCH2CH2N)),
                    3.64 (t, J = 6.5 Hz, 2H,
                        ((thiazol-5-ylmethyl)-NCH2CH2N)),
                    3.77 (s, 3H, CH3 (Im)), 3.86 (s, 3H,
                        CH3 (Im)), 4.38 (s, 2H,
                        CH2-thiazole), 4.73 (s, 2H,
                        CH2Im), 6.71 (d, J =9.1
                    Hz, 2H, 2 CH (benzonitrile)), 7.20 (s, 1H, CH (Im)), 7.46 (d, J
                    = 9.1 Hz, 2H, 2 CH (benzonitrile)), 7.76 (s, 1H, CH (Im)), 7.79 (s, 1H, CH
                    (Im)), 8.87 (s, 1H, CH (Im)); Î´C (125 MHz,
                        MeOH-d4) 14.8, 18.9, 34.7, 34.8, 45.9, 47.3,
                    47.6, 51.3, 100.8, 114.0, 119.6, 121.1, 127.5, 128.6, 133.2, 135.2, 138.3,
                    138.8, 142.0, 142.1, 150.9, 151.8; HRMS (ESI) m/z calcd for
                        [C24H28N8O2S2 + H]
                    525.1855, obsd 525.1828; HPLC (I) tR = 12.14 min (96.61%), (II)
                        tR = 17.00 min (96.03%).</p><!><p>Trimethyl-1H-pyrazole-4-carbaldehyde was reductively
                    aminated with amine 9, according to general procedure C, on a 0.15 mmol scale.
                    The crude material was chromatographed over silica gel (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1) to give
                        4-((3-methyl-3H-imidazol-4-ylmethyl)-{2-[(1,3,5-trimethyl-1H-pyrazol-4-ylmethyl)-amino]-ethyl}-amino)-benzonitrile
                    as a film (46 mg, 81%): Î´H (400 MHz,
                        MeOH-d4)2.28(s, 3H, CH3 (pyrazole)),
                    2.34 (s, 3H, CH3 (pyrazole)), 3.39 (t, J = 7.5 Hz,
                    2H, NHCH2CH2N), 3.75 (s, 3H,
                        N–CH3 (pyrazole)), 3.88–3.95 (m, 5H,
                        NHCH2CH2N, CH3
                    (Im)), 4.15 (s, 2H, CH2-pyrazole), 4.88 (s,
                    2H, CH2Im), 7.01 (d, J = 8.9
                    Hz, 2H, 2 CH (benzonitrile)), 7.22 (s, 1H, CH (Im)), 7.59 (d, J
                    = 8.9 Hz, 2H, 2 CH (benzonitrile)), 8.94 (s, 1H, CH (Im));
                        Î´C (125 MHz,
                        MeOH-d4) 9.8, 11.4, 34.3, 36.1, 42.3, 44.7,
                    45.6, 47.5, 99.9, 108.3, 114.3, 118.8, 120.6, 132.8, 135.0, 138.0, 142.6, 148.5,
                    151.3; LRMS (ESI) m/z 400.2
                        [C21H27N7 + Na]. This secondary amine (38
                    mg, 0.1 mmol) was then reacted with
                    1-methyl-1H-imidazole-4-sulfonyl chloride according to general
                    procedure A. Workup then the usual purification (silica gel flash column
                    chromatography (eluent CH2Cl2/MeOH/NH4OH,
                    192:7:1)) furnished the title compound 1an as a white foam (44 mg,
                    84%): Î´H (500 MHz,
                        MeOH-d4)2.00 (s, 3H, CH3 (pyrazole)),
                    2.06 (s, 3H, CH3 (pyrazole)), 3.15 (t,
                        J = 6.5 Hz, 2H,
                        ((pyrazole-4-ylmethyl)-NCH2CH2N)),
                    3.44 (t, J = 6.5 Hz, 2H,
                        ((pyrazole-4-ylmethyl)-NCH2CH2N)),
                    3.51 (s, 3H, N-CH3 (pyrazole)), 3.73 (s, 3H, CH3 (Im)),
                    3.78 (s, 3H, CH3 (Im)), 3.92 (s, 2H,
                        CH2-pyrazole), 4.48 (s, 2H,
                        CH2Im), 6.50 (d, J = 8.9
                    Hz, 2H, 2 CH (benzonitrile)), 7.11 (s, 1H, CH (Im)), 7.39 (d, J
                    = 8.9 Hz, 2H, 2 CH (benzonitrile)), 7.72 (s, 1H, CH (Im)), 7.75 (s, 1H, CH
                    (Im)), 8.80 (s, 1H, CH (Im)); Î´C (125 MHz,
                        MeOH-d4)9.5, 11.7, 34.3, 34.4, 36.0, 45.1, 45.4,
                    46.5, 51.2, 100.2, 111.8, 113.4, 119.2, 120.8, 127.1, 132.7, 134.8, 137.9,
                    138.1, 141.2, 141.7, 147.9, 151.4; HRMS (ESI) m/z calcd for
                        [C25H31-N9O2S + H] 522.2400,
                    obsd 522.2377; HPLC (I) tR = 12.49 min (95.83%),
                    (II) tR = 17.78 min (96.33%).</p><!><p>First, cyclopropanecarbaldehyde was reductively aminated with amine
                        9 (38 mg, 0.15 mmol), according to general procedure C. Flash
                    column chromatography (eluent
                    CH2Cl2/MeOH/NH4OH, 92:7:1) purified the
                    material to give
                        4-[[2-(cyclopropylmethyl-amino)-ethyl]-(3-methyl-3H-imidazol-4-ylmethyl)-amino]-benzonitrile
                    as a colorless film (39 mg, 84%): Î´H (400
                    MHz, CDCl3)0.07 (m, 2H, 2 CH (cyclopropyl)), 0.46 (m, 2H, 2 CH
                    (cyclopropyl)), 0.78 (m, 1H,
                        CHCH2NHCH2CH2N),
                    2.41 (m, 2H, CH2-cyclopropyl), 2.75 (s, 2H,
                        NHCH2CH2N), 3.50 (m, 2H,
                        NHCH2CH2N), 3.55 (s, 3H,
                        CH3 (Im)), 4.88 (s, 2H, CH2Im),
                    6.68 (d, J =8.8 Hz, 2H, 2 CH (benzonitrile)), 6.86 (s, 1H, CH
                    (Im)), 7.26 (s, 1H, CH (Im)), 7.46 (d, J = 8.8 Hz, 2H, 2 CH
                    (benzonitrile); LRMS (ESI) m/z 332.2
                        [C18H23N5 + Na]. This secondary amine was
                    then reacted with 1-methyl-1H-imidazole-4-sulfonyl chloride
                    according to general procedure A, on a 0.1 mmol scale. After workup, the crude
                    material was purified by silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to furnish
                        1aw as a white powder (39 mg, 86%):
                        Î´H (500 MHz,
                        MeOH-d4) 0.14 (m, 2H, 2 CH (cyclopropyl)), 0.43
                    (m, 2H, 2 CH (cyclopropyl)), 0.84 (m, 1H,
                        CHCH2NCH2-CH2N),
                    2.50 (d, J =6.9,2H,
                    CH2-cyclopropyl), 2.99 (t, J
                    = 6.9 Hz, 2H,
                        ((cyclopropylmethyl)NCH2CH2N)),
                    3.73 (s, 3H, CH3 (Im)), 3.81 (t, J = 6.9 Hz, 2H,
                        ((cyclopropylmethyl)-NCH2CH2N)),
                    3.88 (s, 3H, CH3 (Im)), 4.85 (s, 2H,
                        CH2Im), 6.97 (d, J = 9.1
                    Hz, 2H, 2 CH (benzonitrile)), 7.28 (s, 1H, CH (Im)), 7.62 (d, J
                    = 9.1 Hz, 2H, 2 CH (benzonitrile)), 7.67 (s, 1H, CH (Im)), 7.71 (s, 1H, CH
                    (Im)), 8.87 (s, 1H, CH (Im)); Î´C (125 MHz,
                        MeOH-d4) 4.6, 11.0, 34.3, 34.4, 45.8, 46.8,
                    51.6, 55.9, 100.4, 114.1, 119.3, 120.9, 126.5, 133.1, 134.9, 137.9, 139.5,
                    141.4, 151.9; HRMS (ESI) m/z calcd for
                        [C22H27N7-O2S + H] 454.2025,
                    obsd 454.2009; HPLC (I) tR = 12.55 min (99.05%),
                    (II) tR = 18.46 min (98.47%).</p><!><p>tert-Butyl p-fluorobenzoate (750 mg,
                    3.93 mmol, 1 equiv) was dissolved in ethylenediamine
                    (9;4mL,59.8mmol,15.2equiv)and heated to 120 °C for 24 h. All
                    solvent was evaporated, and then the residue was dry-loaded onto silica gel and
                    purified by flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1) to furnish
                        tert-butyl 4-(2-amino-ethylamino)-benzoate as a white
                    powder (891 mg, 96%): Î´H (400 MHz,
                        CD2Cl2)1.59 (s, 9H,
                        CO2-(CH3)3), 2.98 (t, J =
                    5.8 Hz, 2H,
                    H2NCH2CH2NH), 3.23 (q,
                        J = 5.8 Hz, 2H,
                        H2NCH2CH2NH), 4.67
                    (br, 1H, H2NCH2CH2NH),
                    6.62 (d, J = 8.6 Hz, 2H, 2 CH (Ar)), 7.81 (d,
                        J = 8.6 Hz, 2H, 2 CH (Ar));
                        Î´C (125 MHz,
                    CD2Cl2) 28.9, 41.7, 46.6, 80.3, 112.1, 120.9, 131.9,
                    152.9, 166.7; HRMS (EI+) m/z calcd for
                        [C13H20N2O2] 236.1525, obsd
                    236.1527. tert-Butyl 4-(2-amino-ethylamino)-benzoate (700 mg,
                    3.03 mmol, 1 equiv) was then sulfonylated with
                    1-methyl-1H-imidazole-4-sulfonyl chloride according to general
                    procedure A. The crude material was dry-loaded onto silica gel,
                    andthenpurifiedbyflash column chromatography (eluent
                        CH2-Cl2/MeOH/NH4OH, 192:7:1) to give
                        tert-butyl
                    4-[2-(1-methyl-1H-imidazole-4-sulfonylamino)-ethylamino]-benzoate
                    as a white powder (1.04 g, 90%): Î´H (500 MHz,
                        CDCl3) 1.56 (s, 9H, CO2-(CH3)3),
                    3.17 (m, 2H,
                    SO2NHCH2CH2NH), 3.28 (m,
                    2H, SO2NHCH2CH2NH), 3.67
                    (s, 3H, CH3 (Im)), 6.51 (d, J = 8.5 Hz, 2H, 2 CH
                    (Ar)), 7.46 (s, 1H, CH (Im)), 7.51 (s, 1H, CH (Im)), 7.75 (d, J
                    = 8.5 Hz, 2H, 2 CH (Ar)); Î´C (125 MHz,
                        MeOH-d4) 28.7, 34.6, 42.4, 43.1, 80.8, 111.9,
                    120.5, 125.2, 131.8, 140.1, 140.3, 152.4, 167.4; HRMS (ESI+)
                        m/z calcd for
                        [C17H24N4O4S + H] 381.1597, obsd
                    381.1573. tert-Butyl
                    4-[2-(1-methyl-1H-imidazole-4-sulfonylamino)-ethylamino]-benzoate
                    (610 mg, 1.63 mmol, 1 equiv) was then chemoselectively benzylated on the
                    sulfonamide NH with benzyl bromide according to general procedure B. The crude
                    material was then flash chromatographed over silica gel to furnish
                        tert-butyl
                    4-{2-[benzyl-(1-methyl-1H-imidazole-4-sulfonyl)-amino]-ethylamino}-benzoate
                    as a white solid (703 mg, 92%): Î´H (400 MHz,
                        CDCl3)1.52(s,9H,CO2(CH3)3), 3.13
                    (t, J = 6.2 Hz, 2H,
                        (Bn)NCH2CH2NH), 3.43 (t,
                        J = 6.2 Hz, 2H,
                        (Bn)NCH2CH2NH), 3.68 (s, 3H,
                        CH3 (Im)), 4.32 (s, 2H, CH2Ph),
                    6.30 (d, J = 8.8 Hz, 2H, 2 CH (Ar)), 7.23–7.30 (m, 5H, 5
                    CH (Ph)), 7.42 (s, 1H, CH (Im)), 7.49 (s, 1H, CH (Im)), 7.70 (d,
                        J = 8.8 Hz, 2H, 2 CH (Ar));
                        Î´C (500 MHz, CDCl3) 28.1,
                    33.9, 41.6, 46.6, 52.6, 79.6, 111.1, 119.9, 124.4, 127.9, 128.3, 128.6, 131.1,
                    136.0, 139.0, 139.8, 151.1, 166.1; HRMS (ESI ) m/z calcd for
                        [C24H30N4O4S + H] 471.2066, obsd
                    471.2053. tert-Butyl
                    4-{2-[benzyl-(1-methyl-1H-imidazole-4-sulfonyl)-amino]-ethyl-amino}-benzoate
                    (220 mg, 0.473 mmol, 1 equiv) was alkylated on the aniline NH with
                        5-chloromethyl-1-methyl-1H-imidazole 3 HCl31 according to general procedure E. The
                    crude material was flash chromatographed over silica gel (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to afford the
                    title compound 11 as a colorless foam (200 mg, 76%):
                        Î´H (500 MHz, CDCl3) 1.53 (s,
                    9H, C(CH3)3), 3.09–3.15 (m, 2H,
                        CH2CH2NSO2),
                    3.29–3.34 (m, 2H,
                        CH2CH2-NSO2), 3.43
                    (s, 3H, CH3), 3.73 (s, 3H, CH3), 4.26 (s, 2H,
                        CH2Ph), 4.30 (s, 2H,
                        CH2Im), 6.35 (d, J = 9.0
                    Hz, 2H, 2 CH (Ar)), 6.69 (s, 1H, CH (Im)), 7.31–7.37 (m, 5H, 5 CH (Ph)),
                    7.39 (s, 1H, CH (Im)), 7.42 (s, 1H, CH (Im)), 7.46 (s, 1H, CH (Im)), 7.68 (d,
                        J = 9.0 Hz, 2H, 2 CH (Ar));
                        Î´C (125 MHz, CDCl3) 28.2,
                    31.6, 33.9, 44.1, 45.0, 49.7, 54.3, 79.8, 111.0, 120.0, 124.4, 127.5, 128.1,
                    128.6, 128.7, 128.9, 131.1, 136.2, 138.5, 139.0, 139.1, 150.3, 165.8; HRMS
                    (ESI+) m/z calcd for
                        [C29H36N6O4S + H] 565.2597, obsd
                    565.2576; HPLC (I) tR = 12.53 min (100%),(II)
                        tR = 18.77 min (100%).</p><!><p>TFA (1.5 mL) was added to a solution of 11 (150 mg, 0.268
                    mmol, 1 equiv) in CH2Cl2 (1.5 mL), and the resulting
                    mixture was stirred for 3 h at room temperature, after which time all solvents
                    were evaporated. Residual TFA was removed by repeated coevaporation with a 2:1
                    mixture of CHCl3/MeOH, then the residue was purified by silica gel
                    flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 25:7:1) to afford the
                    title compound as a sticky foam (131 mg, 96%):
                        Î´H (500 MHz,
                        MeOH-d4)3.34–3.39 (m, 2H,
                        CH2CH2NSO2),
                    3.42–3.46 (m, 2H,
                        CH2CH2NSO2), 3.81
                    (s, 3H, CH3), 3.84 (s, 3H, CH3), 4.30 (s, 2H,
                        CH2Ph), 4.55 (s, 2H, CH2Im),
                    6.53 (d, J = 9.0 Hz, 2H, 2 CH (Ar)), 7.15 (s, 1H, CH (Im)),
                    7.34–7.38 (m, 5H, 5 CH (Ph)), 7.76–7.79 (m, 3H, 2 CH (Ar), CH
                    (Im)), 7.82 (s, 1H, CH (Im)), 8.87 (s, 1H, CH (Im));
                        Î´C (125 MHz,
                        MeOH-d4) 34.3, 34.4, 45.0, 46.5, 51.2, 55.2,
                    112.8, 119.3, 120.3, 126.8, 129.3, 129.9 (2 CH), 132.7, 133.1, 137.7, 137.9,
                    139.1, 141.5, 152.0, 170.1; HRMS (ESI+) m/z calcd for
                        [C25H28N6O4S + H] 509.1971, obsd
                    509.1953; HPLC (I) tR = 11.24 min (100%), (II)
                        tR = 15.21 min (100%).</p><!><p>To a solution of acid 12 (50 mg, 0.0984 mmol, 1 equiv),
                        NH4Cl (10.5 mg, 0.197 mmol, 2 equiv), and DIPEA (34
                        Î¼L) in DMF (1 mL) was added HBTU (48.5 mg, 0.128
                    mmol, 1.3 equiv). The reaction mixture was stirred for 1 h, after which time TLC
                    indicated the reaction was complete. Water (25 mL) was added, and then the crude
                    product was extracted into EtOAc (5 × 5mL). The EtOAc extractions were
                    combined, washed with water (3 × 5 mL), brine (5 mL), dried
                        (Na2SO4), filtered, and concentrated. The residue was
                    dry-loaded onto silica gel, then flash chromatographed (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1) to furnish
                    primary amide 13 as a white powder (44 mg, 89%):
                        Î´H (500 MHz, CDCl3) 3.15 (m,
                    2H, (Bn)NCH2CH2N), 3.31 (m, 2H,
                        (Bn)NCH2CH2N), 3.41 (s, 3H,
                        CH3 (Im)), 3.71 (s, 3H, CH3 (Im)), 4.23 (s, 2H,
                        CH2Ph), 4.29 (s, 2H,
                        CH2Im), 6.02 (br, 2H, NH2),
                    6.42 (d, J = 9.0 Hz, 2H, 2 CH (Ar)), 6.66 (s, 1H, CH (Im)),
                    7.29–7.34 (m, 6H, 5 CH (Ph), CH (Im)), 7.41 (s, 1H, CH (Im)), 7.45 (s,
                    1H, CH (Im)), 7.56 (d, J = 9.0 Hz, 2H, 2 CH (Ar));
                        Î´C (125 MHz, CDCl3) 31.3,
                    33.7, 43.9, 44.8, 49.4, 50.2, 54.0, 111.1, 121.0, 124.2, 127.2, 127.9, 128.4,
                    128.6, 128.9, 136.0, 138.4, 138.8, 138.9, 149.8, 168.9; HRMS (ESI+)
                        m/z calcd for
                        [C25H29N7O3S + H] 508.2137, obsd
                    508.2131; HPLC (I) tR = 11.72 min (100%), (II)
                        tR = 16.29 min (100%).</p><!><p>Previously reported
                        1-tert-butoxycarbonylamino-{2-[biphenyl-4-yl-(3-methyl-3H-imidazol-4-ylmethyl)-amino]}-ethane
                    (compound 9a in ref 31) (30
                    mg, 0.0739 mmol) was deprotected by stirring in a 1:1 mixture of
                        TFA-CH2Cl2 (1 mL) for 30 min. After removal of all
                    solvent in vacuo, the residue (23 mg, 0.0739 mmol) was sulfonylated with
                        1-methyl-1H-imidazole-4-sulfonyl chloride according to
                    general procedure A, but with 5 equiv of DIPEA. After the usual workup, the
                    residue was purified by silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1) to afford
                        1-methyl-1H-imidazole-4-sulfonic acid
                    benzyl-[2-(biphenyl-4-ylamino)-ethyl]-amide as a colorless film (30 mg, 91%)
                    Next, the secondary aniline of 1-methyl-1H-imidazole-4-sulfonic
                    acid benzyl-[2-(biphenyl-4-ylamino)-ethyl]-amide (30 mg, 0.067 mmol, 1 equiv)
                    was alkylated with the HCl salt of
                        5-chloromethyl-1-methyl-1H-imidazole31 according to general procedure E. After the usual
                    workup, the crude material was dry-loaded onto silica gel and purified by flash
                    column chromatography (eluent
                    CH2Cl2/MeOH/NH4OH, 92:7:1) to give
                        14 as a glassy film (24 mg, 65%):
                        Î´H (500 MHz, CDCl3) 3.16 (m,
                    2H, (Bn)NCH2-CH2N), 3.33 (m, 2H,
                        (Bn)NCH2CH2N), 3.44 (s, 3H,
                        CH3 (Im)), 3.75 (s, 3H, CH3 (Im)), 4.27 (s, 2H,
                        CH2Ph), 4.38 (s, 2H,
                        CH2Im), 6.56 (d, J = 9.0
                    Hz, 2H, 2 CH (Ar)), 6.79 (s, 1H, CH (Im)), 7.34 – 7.51 (m, 15 H, 10 CH
                    (Ph), 3 CH (Im), 2 CH (Ar)); Î´C (125 MHz,
                        CDCl3) 31.9, 34.2, 44.9, 45.4, 50.0, 54.5, 113.5, 124.6, 126.4,
                    126.5, 128.1, 128.3 (2), 128.9, 129.0, 129.2, 129.4, 130.5, 136.8, 138.8, 139.3,
                    139.9, 141.1, 147.2; HRMS (ESI+) m/z calcd for
                        [C30H32N6-O2S + H] 541.2380,
                    obsd 541.2386; HPLC (I) tR = 13.01 min (100%), (II)
                        tR = 20.24 min (100%).</p><!><p>Mono-N-Boc-ethylenediamine (6; 5 g, 31.2
                    mmol, 1 equiv) was sulfonylated with
                    1-methyl-1H-imidazole-4-sulfonyl chloride (6.76 g, 37.4 mmol,
                    1.2 equiv), according to general procedure A. After 16 h, the solvent was
                    reduced and then the reaction mixture was dry-loaded onto silica gel and
                    purified by flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 1092:7:1) to give the
                    title compound as a white powder (9 g, 95%):
                        Î´H (500 MHz,
                    CD2Cl2) 1.37 (s, 9H, C(CH3)3), 2.99
                    (t, J = 5.8 Hz, 2H,
                        CH2NHSO2), 3.11 (q,
                        J = 5.8 Hz, 2H,
                    CH2NHBoc), 3.71 (s, 3H, CH3), 5.53
                    (s, 1H, NHSO2), 5.68 (s, 1H, NHBoc), 7.48 (s,
                    1H, Im), 7.56 (s, 1H, Im); Î´C (125 MHz,
                        CD2Cl2) 27.6, 33.5, 39.8, 42.6, 78.9, 124.0, 127.2,
                    139.2, 156.1; HRMS (ESI+) m/z calcd for
                        [C11H20N4O4S + H] 305.1284, obsd
                    305.1294.</p><!><p>N-(2-tert-Butoxycarbonylaminoethyl)
                        1-methyl-1H-imidazole-4-sulfonamide (5.94 g, 19.5 mmol, 1
                    equiv) was chemoselectively alkylated on the sulfonamide NH with
                        N-(2-pyrimidinyl)-piperidin-4-ylmethyl bromide (6 g, 23.4
                    mmol, 1.2 equiv), according to general procedure B, but the
                    reaction was stirred for 4 d at room temperature.
                    [N-(2-Pyrimidinyl)-piperidin-4-ylmethyl bromide was made from
                    the reaction of N-(2-pyrimidinyl)-piperidin-4-ylmethyl alcohol
                    (5 g, 25.9 mmol, 1 equiv) with PPh3Br2 (14.2 g, 33.7 mmol,
                    1.3 equiv) in CH2Cl2 (130 mL) at 0°C to room
                    temperature over 6 h. At this point, the reaction was complete. Water (100 mL)
                    was added to quench, and then the CH2Cl2 layer was diluted
                    further (500 mL total volume). The organic layer was collected, and the aqueous
                    layer was extracted twice further (2 × 50 mL). The organic layers were
                    collected, dried (Na2SO4), filtered, and concentrated. The
                    residue was redissolved in CH2Cl2, dry-loaded onto silica
                    gel, then purified by flash column chromatography (eluent Hex/EtOAc, 7:1) to
                    furnish N-(2-pyrimidinyl)-piperidin-4-ylmethyl bromide as a
                    white powder (6.04 g, 91%).] After workup, the crude material was dry-loaded
                    onto silica gel and purified by silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1), affording the
                    title compound as a white solid (8.81 g, 94%):
                        Î´H (500 MHz, CDCl3)1.09(qd,
                        J = 12.3, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)), 1.36 (s,
                    9H, C(CH3)3), 1.69–1.78 (m, 2H, 2 CH
                    (piperidinylmethyl)), 1.84–1.92 (m, 1H, CH (piperidinylmethyl)),
                    2.77–2.83 (m, 2H, 2 CH (piperidinylmethyl)), 2.97 (br d,
                        J = 8.0 Hz, 2H, 2 CH (piperidinylmethyl)), 3.25–3.34
                    (m, 4H,
                        CH2CH2NHBoc),
                    3.67 (s, 3H, CH3), 4.64–4.69 (m, 2H, 2 CH
                    (piperidinylmethyl)), 5.53 (m, 1H, NHBoc), 6.34 (t, J = 4.9 Hz,
                    1H, CH (pyrimidine)), 7.36 (s, 1H, CH (Im)), 7.40 (s, 1H, CH (Im)), 8.20 (t,
                        J = 4.9 Hz, 2H, 2 CH (pyrimidine));
                        Î´C (125 MHz, CDCl3) 28.3,
                    29.4, 34.8, 35.4, 39.6, 43.4, 49.1, 55.2, 79.0, 109.2, 124.2, 138.9, 140.6,
                    155.9, 157.5, 161.4; HRMS (ESI+) m/z calcd for
                        [C21H33N7O4S + H] 480.2393, obsd
                    480.2409.</p><!><p>To a cooled (0 °C), stirring solution of 16 (8.67 g,
                    18.1 mmol) in CH2Cl2 (47.5 mL) was added
                        triisopropylsilane (TIPS; 2.5 mL), H2O (2.5 mL)
                    and TFA (47.5 mL). After 5 min, the reaction was removed from the ice bath and
                    allowed to stir for 1 h at room temperature, and then all solvents were
                    evaporated. The residue was redissolved in CH2Cl2 and the
                    minimal required volume of MeOH, then dry-loaded onto silica gel and purified by
                    flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1) to give
                        17 as a white, sticky foam (7.2 g, 100%):
                        Î´H (500 MHz, CDCl3)1.10 (qd,
                        J = 12.2, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.73–1.78 (m, 2H, 2 CH (piperidinylmethyl)), 1.87–1.95 (m, 1H, CH
                    (piperidinyl)), 2.77–2.83 (m, 2H, 2 CH (piperidinylmethyl)), 2.92 (t,
                        J = 6.5 Hz, 2H,
                        CH2NH2) 3.00 (br d,
                        J = 7.5 Hz, 2H, 2 CH (piperidinylmethyl)), 3.32 (t,
                        J = 6.4 Hz, 2H,
                        CH2CH2NH2),
                    3.46–3.51 (br s, NH2), 3.69 (s, 3H, CH3 (Im)),
                    4.66–4.72 (m, 2 H, 2 CH (piperidinylmethyl)), 6.38 (t, J
                    = 4.7 Hz, 1H, CH (pyrimidine)), 7.40 (s, 1H, CH (Im)), 7.47 (s, 1H, CH (Im)),
                    8.22 (t, J = 4.7 Hz, 2H, 2 CH (pyrimidine));
                        Î´C (125 MHz, CDCl3) 29.5,
                    33.8, 35.5, 40.1, 43.4, 52.5, 54.9, 109.2, 124.2, 139.0, 139.4, 158.5, 161.5;
                    HRMS (ESI+) m/z calcd for
                        [C16H25N7O2S + H] 380.1869, obsd
                    380.1886.</p><!><p>Compound 17 was reacted with 2-chloro-4-fluorobenzonitrile
                    on a 0.338 mmol scale, according to general procedure D. After the usual workup,
                    the crude material was purified by silica gel flash column chromatography
                    (eluent CH2Cl2/MeOH/NH4OH, 192:7:1) to give the
                    title compound as a colorless, viscous oil (158 mg, 91%):
                        Î´H (500 MHz, CDCl3)1.08 (qd,
                        J = 12.3, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.68–1.74 (m, 2H, 2 CH (piperidinylmethyl)), 1.77–1.86 (m, 1H, CH
                    (piperidinylmethyl)), 2.72 (td, J = 12.5, 2.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.00 (br d, J = 7.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.48–3.55 (m, 4H,
                    CH2CH2NHAr), 3.71 (m, 3H, CH3 (Im)),
                    4.63–4.69 (m, 2 H, 2 CH (piperidinylmethyl)), 5.77 (t, J
                    = 5.3 Hz, 1H, CH2NHAr), 6.40 (t,
                        J = 4.7 Hz, 1H, CH (pyrimidine)), 6.66 (d,
                        J = 8.5 Hz, 1H, CH (Ar)), 7.37 (dd, J =
                    8.5, 2.0 Hz, 1H, CH (Ar)), 7.43 (s, 1H, CH (Im)), 7.45 (d, J =
                    2.0 Hz, 1H, CH (Ar)), 7.46 (s, 1H, CH (Im)), 8.22 (t, J = 4.7
                    Hz, 2H, 2 CH (pyrimidine)); Î´C (125 MHz,
                        CDCl3) 29.6, 33.9, 36.0, 42.2, 43.4, 48.7, 56.1, 98.7, 109.4,
                    110.0, 118.6, 119.0, 124.4, 132.4, 132.5, 139.1, 139.5, 147.0, 157.5, 161.4;
                    HRMS (ESI+) m/z calcd for
                        [C23H27N8O2SCl + H] 515.1744,
                    obsd 515.1754.</p><!><p>Compound 17 was reacted with 2-cyano-5-fluoropyridine on a
                    0.791 mmol scale, according to general procedure D. After the usual workup, the
                    crude material was purified by silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to give the
                    title compound as a colorless, viscous oil (320 mg, 84%):
                        Î´H (500 MHz, CDCl3) 0.95 (qd,
                        J = 12.3, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.63–1.71 (m, 2H, 2 CH (piperidinylmethyl)), 1.74–1.83 (m, 1H, CH
                    (piperidinylmethyl)), 2.72 (td, J = 12.5, 2.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 2.93 (br d, J = 7.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.23–3.27 (m, 2H,
                    CH2CH2NHAr), 3.34–3.36 (m, 2H,
                        CH2CH2NHAr), 3.65 (m, 3H,
                        CH3 (Im)), 4.55–4.59 (m, 2 H, 2 CH (piperidinylmethyl)),
                    6.54 (m, 1H, CH2NHAr), 6.96 (t,
                        J = 4.9 Hz, 1H, CH (pyrimidine)), 7.04 (m, 1H, CH (Ar)),
                    7.62 (s, 1H, CH (Im)), 7.77–7.78 (m, 2H, CH (Im), CH (Ar)), 8.02 (m, 1H,
                    CH (Ar)), 8.28 (t, J = 4.9 Hz, 2H, 2 CH (pyrimidine));
                        Î´C (125 MHz,
                        DMSO-d6) 28.96, 33.34, 34.88, 40.33, 41.20,
                    43.03, 47.87, 54.82, 109.46, 117.39, 118.92, 124.99, 129.69, 136.90, 137.56,
                    139.75, 146.98, 157.72, 161.02; HRMS (ESI+) m/z calcd for
                        [C22H27N9O2S + H] 482.2087, obsd
                    482.2086.</p><!><p>Compound 17 was reacted with 5-bromo-2-fluoropyridine on a
                    0.512 mmol scale, according to general procedure D. After the usual workup, the
                    crude material was purified by silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to give the
                    title compound as a colorless, viscous oil (200 mg, 73%):
                        Î´H (500 MHz, CDCl3) 1.11 (qd,
                        J = 12.3, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.72–1.78 (m, 2H, 2 CH (piperidinylmethyl)), 1.87–1.97 (m, 1H, CH
                    (piperidinylmethyl)), 2.78 (td, J = 12.8, 2.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.03 (br d, J = 8.0 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.49–3.57 (m, 4H,
                        CH2CH2-NHAr),
                    3.72 (m, 3H, CH3 (Im)), 4.67–4.73 (m, 2 H, 2 CH
                    (piperidinylmethyl)), 5.62 (t, J = 5.3 Hz, 1H,
                        CH2NHAr), 6.34 (dd, J =
                    9.0 Hz, 1H, CH (Ar)), 6.41 (t, J = 4.9 Hz, 1H, CH
                    (pyrimidine)), 7.40 (dd, J = 9.0, 2.5 Hz, 1H, CH (Ar)), 7.42
                    (s, 1H, CH (Im)), 7.47 (s, 1H, CH (Im)), 8.05 (d, J = 2.5 Hz,
                    1H, CH (Ar)), 8.25 (t, J = 4.9 Hz, 2H, 2 CH (pyrimidine));
                        Î´C (125 MHz, CDCl3) 29.7,
                    33.9, 35.6, 40.5, 43.6, 48.5, 55.0, 106.8, 109.3, 109.7, 124.4, 138.9, 139.3,
                    140.0, 148.4, 156.9, 157.6, 161.5; HRMS (ESI+) m/z calcd for
                        [C21H27N8O2SBr + H] 535.1239,
                    obsd 535.1259.</p><!><p>Compound 17 was reacted with 5-bromo-2-fluoropyrimidine on
                    a 0.528 mmol scale, according to general procedure D. After the usual workup,
                    the crude material was purified by silica gel flash column chromatography
                    (eluent CH2Cl2/MeOH/NH4OH, 192:7:1) to give the
                    title compound as a white foam (255mg, 90%):
                        Î´H (500 MHz, CDCl3) 1.08 (qd,
                        J = 12.3, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.70–1.77 (m, 2H, 2 CH (piperidinylmethyl)), 1.83–1.92 (m, 1H, CH
                    (piperidinylmethyl)), 2.78 (td, J = 12.8, 2.5Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.03 (br d, J = 8.0 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.43 (t, J = 6.3 Hz, 2H
                        ,CH2CH2NHAr), 3.57 (t,
                        J = 6.3 Hz, 2H,
                        CH2CH2NHAr), 3.68 (m, 3H, CH3
                    (Im)), 4.60–4.66 (m, 2 H, 2 CH (piperidinylmethyl)), 6.39 (t,
                        J = 4.6 Hz, 1H, CH (pyrimidine)), 7.40 (s, 1H, CH (Im)),
                    7.47 (s, 1H, CH (Im)), 8.19 (s, 2H, 2CH(Ar)), 8.21 (t, J = 4.6
                    Hz, 2H, 2CH (pyrimidine)); Î´C (125 MHz,
                        CDCl3) 29.5, 33.8, 35.5, 40.4, 43.6, 48.3, 55.0, 106.1, 109.3,
                    124.3, 139.1, 139.6, 157.6, 158.1, 160.3, 161.3; HRMS (ESI+)
                        m/z calcd for
                        [C20H26N9O2SBr + H] 536.1192,
                    obsd 536.1213.</p><!><p>Compound 17 was reacted with 2,4-difluorobenzonitrile on a
                    0.417 mmol scale, according to general procedure D, to give a separable mixture
                    of the two expected isomers. After the usual workup, the crude material was
                    purified by prepTLC (eluent CH2Cl2/MeOH/NH4OH,
                    92:7:1) to give the title compound as a white foam (99 mg, 48%):
                        Î´H (500 MHz, CDCl3) 1.15 (qd,
                        J = 12.5, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.73–1.79 (m, 2H, 2 CH (piperidinylmethyl)), 1.87–1.95 (m, 1H, CH
                    (piperidinylmethyl)), 2.80 (td, J = 12.8, 2.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.03 (br d, J = 7.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.36 (q, J = 5.5 Hz, 2H,
                        CH2CH2NHAr), 3.65 (t, J = 5.8 Hz, 2H,
                        CH2CH2NHAr), 3.77 (m, 3H,
                        CH3 (Im)), 4.70–4.75 (m, 2 H, 2 CH (piperidinylmethyl)),
                    6.10–6.13 (m, 1H, CH2NHAr), 6.30 (dd,
                        J = 12.0, 2.0 Hz, 1H, CH (Ar)), 6.39 (dd,
                        J = 8.8, 2.0 Hz, 1H, CH (Ar)), 6.44 (t, J
                    = 4.9 Hz, 1H, CH (pyrimidine)), 7.32 (dd, J = 8.8, 7.3 Hz, 1H,
                    CH (Ar)), 7.47 (s, 1H, CH (Im)), 7.52 (s, 1H, CH (Im)), 8.27 (t,
                        J = 4.9 Hz, 2H, CH (pyrimidine));
                        Î´C (125 MHz, CDCl3) 29.5,
                    34.0, 35.5, 41.5, 43.5, 47.7, 54.4, 87.0 (d, JCF =
                    15.5 Hz), 98.1 (d, JCF = 22.8 Hz), 108.8 (br),
                    109.4, 115.7, 124.6, 133.9 (d, JCF = 2.75 Hz),
                    139.0, 139.6, 153.4 (d, JCF = 10.9 Hz), 157.6,
                    161.5, 165.0 (d, JCF = 252 Hz); HRMS (ESI+)
                        m/z calcd for
                        [C23H27N8O2FS + H] 499.2040,
                    obsd 499.2057.</p><!><p>Compound 17 was reacted with 3,4-difluorobenzonitrile on a
                    0.343 mmol scale, according to general procedure D. After the usual workup, the
                    crude material was purified by silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to give the
                    title compound as a white foam (138 mg, 82%):
                        Î´H (500 MHz, CDCl3)1.13 (qd,
                        J = 12.3, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.73–1.78 (m, 2H, 2 CH (piperidinylmethyl)), 1.83–1.91 (m, 1H, CH
                    (piperidinylmethyl)), 2.78 (td, J = 12.8, 2.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.04 (br d, J = 7.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.51 (q, J = 5.7 Hz, 2H,
                        CH2CH2NHAr), 3.59 (t,
                        J = 6.0 Hz, 2H,
                        CH2CH2NHAr), 3.75 (m, 3H,
                        CH3 (Im)), 4.69–4.75 (m, 2 H, 2 CH (piperidinylmethyl)),
                    5.74–5.78 (m, 1H, CH2NHAr), 6.43 (t,
                        J = 4.7 Hz, 1H, CH (pyrimidine)), 6.68 (t,
                        J = 8.5 Hz, 1H, CH (Ar)), 7.19 (dd, J =
                    11.5, 1.5 Hz, 1H, CH (Ar)), 7.31 (br dd, J = 8.5, 1.5 Hz, 1H,
                    CH (Ar)), 7.44 (s, 1H, CH (Im)), 7.49 (s, 1H, CH (Im)), 8.27 (t,
                        J = 4.7 Hz, 2H, 2 CH (pyrimidine));
                        Î´C (125 MHz, CDCl3) 29.5,
                    33.9, 35.7, 41.6, 43.4, 48.3, 55.1, 97.4 (d, JCF =
                    9.13 Hz), 109.4, 110.8 (d, JCF = 4.5 Hz), 117.5 (d,
                        JCF = 21.9 Hz), 119.2 (d,
                        JCF = 2.75 Hz), 124.3, 130.1 (d,
                        JCF = 2.75 Hz), 139.1, 139.6, 140.5 (d,
                        JCF = 11.9 Hz), 149.8 (d,
                        JCF = 241 Hz), 157.6, 161.4; HRMS (ESI+)
                        m/z calcd for
                        [C23H27N8O2FS + H] 499.2040,
                    obsd 499.2055.</p><!><p>Compound 17 was reacted with 3,4,5-trifluorobenzonitrile on
                    a 0.359 mmol scale, according to general procedure D. After the usual workup,
                    the crude material was purified by silica gel flash column chromatography
                    (eluent CH2Cl2/MeOH/NH4OH, 192:7:1) to give the
                    title compound as a white foam (179 mg, 97%):
                        Î´H (500 MHz, CDCl3) 1.09 (qd,
                        J = 12.3, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.68–1.75 (m, 2H, 2 CH (piperidinylmethyl)), 1.81–1.90 (m, 1H, CH
                    (piperidinylmethyl)), 2.75 (td, J = 12.5, 2.5 Hz, 2H, 2
                    CH(piperidinylmethyl)), 2.98 (br d, J = 7.0 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.51 (t, J = 5.8 Hz, 2H,
                        CH2CH2NHAr), 3.72 (m, 5H,
                        CH2CH2-NHAr, CH3
                    (Im)), 4.65–4.73 (m, 2H, 2 CH (piperidinylmethyl)), 5.67 (m, 1H,
                        CH2NHAr), 6.40 (t, J = 4.9 Hz, 1H, CH
                    (pyrimidine)), 7.05 (dd, J = 7.5, 2.5 Hz, 2H, CH (Ar)), 7.43
                    (s, 1H, CH (Im)), 7.47 (s, 1H, CH (Im)), 8.23 (t, J = 4.9 Hz,
                    2H, 2CH (pyrimidine)); Î´C (125 MHz,
                        CDCl3) 29.6, 33.9, 35.7, 43.5, 43.7, 50.2, 55.4, 96.7 (t,
                        JCF = 11.4 Hz), 109.3, 115.8 (dd,
                        JCF = 18.3, 8.13 Hz), 117.9 (t,
                        JCF = 3.19 Hz), 124.4, 130.8 (t,
                        JCF = 12.3 Hz), 139.0, 139.7, 150.8 (dd,
                        JCF = 241, 10.0 Hz), 157.5, 161.4; HRMS (ESI+)
                        m/z calcd for
                        [C23H26N8O2F2S + H]
                    517.1946, obsd 517.1953.</p><!><p>Compound 17 was reacted with 2-chloro-4-fluorobenoznitrile
                    on a 0.338 mmol scale, according to general procedure D, although the reaction
                    was complete within 12 h at room temperature. After the usual workup, the crude
                    material was purified by silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to give the
                    title compound as a white foam (158 mg, 85%):
                        Î´H (500 MHz, CDCl3) 1.13 (qd,
                        J = 12.2, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.72–1.79 (m, 2H, 2 CH (piperidinylmethyl)), 1.88–1.97 (m, 1H, CH
                    (piperidinylmethyl)), 2.81 (td, J = 12.8, 2.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 2.97 (br d, J = 7.0 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.63 (t, J = 5.0 Hz, 2H,
                        CH2CH2NHAr), 3.77 (s, 3H,
                        CH3 (Im)), 3.78–3.84 (m, 2H,
                        CH2-CH2NHAr), 4.71–4.77
                    (m, 2H, 2CH (piperidinylmethyl)), 6.44 (t, J = 4.8 Hz, 1H, CH
                    (pyrimidine)), 6.81–6.86 (m, 1H,
                    CH2NHAr), 7.47 (s, 1H, CH (Im)), 7.51 (s, 1H,
                    CH (Im)), 8.27 (t, J = 4.8 Hz, 2H, CH (pyrimidine));
                        Î´C (125 MHz, CDCl3) 29.7,
                    34.1, 35.9, 43.4, 43.5, 50.2, 55.0, 78.4 (t, JCF =
                    18.3 Hz), 109.0 (m), 109.5, 124.7, 133.5 (m), 135.9 (dm), 139.0, 140.1, 148.1
                    (dm), 157.7, 161.6; HRMS (ESI+) m/z calcd for
                        [C23H24N8O2F4S + H]
                    553.1757, obsd 553.1774.</p><!><p>As per general procedure E with 18a on a 0.068 mmol scale.
                    After the usual workup, the crude material was purified by silica gel flash
                    column chromatography (eluent CH2Cl2/
                    MeOH/NH4OH, 192:7:1) to afford the title compound as a white foam (37
                    mg, 90%): Î´H (500 MHz, CDCl3) 1.04
                    (qd, J = 12.2, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.63–1.68 (m, 2H, 2 CH (piperidinylmethyl)), 1.70–1.76 (m, 1H, CH
                    (piperidinylmethyl)), 2.76 (td, J = 12.5, 2.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 2.94 (br d, J = 7.0 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.31–3.36 (m, 2H,
                        SO2NCH2CH2N),
                    3.39–3.44 (m, 2H,
                        SO2-NCH2CH2N), 3.51
                    (s, 3H, CH3 (Im)), 3.73 (m, 3H, CH3 (Im)), 4.41 (s, 2H,
                        CH2Im), 4.65–4.71 (m, 2 H, 2 CH
                    (piperidinylmethyl)), 6.42 (t, J = 4.9 Hz, 1H, CH
                    (pyrimidine)), 6.97 (s, 1H, CH (Im)), 7.23 (d, J = 8.5 Hz, 1H,
                    CH (Ar)), 7.36 (s, 1H, CH (Im)), 7.41 (s, 1H, CH (Im)), 7.43 (s, 1H, CH (Im)),
                    7.48 (dd, J = 8.5, 2.0 Hz, 1H, CH (Ar)), 7.63 (d,
                        J = 2.0 Hz, 1H, CH (Ar)), 8.26 (t, J = 4.9
                    Hz, 2H, 2 CH (pyrimidine)); Î´C (125 MHz,
                        CDCl3) 29.6, 31.7, 33.9, 36.0, 43.5, 45.7, 47.1, 50.7, 55.6,
                    107.1, 109.4, 117.6, 123.7, 124.1, 126.6, 129.0, 129.9, 131.4, 134.3, 138.9,
                    139.0, 139.7, 151.0, 157.6, 161.5; HRMS (ESI+) m/z calcd for
                        [C28H33-N10O2SCl + H] 609.2275,
                    obsd 609.2296; HPLC (I) tR = 12.90 min (99.13%),
                    (II) tR = 19.65 min (98.61%).</p><!><p>As per general procedure E with 18b on a 0.415 mmol scale. After the
                    usual workup, the crude material was purified by silica gel flash column
                    chromatography (eluent CH2Cl2/MeOH/NH4OH,
                    192:7:1) to afford the title compound as a white foam (186 mg, 78%):
                        Î´H (500 MHz,
                        DMSO-d6) 1.05–1.13 (m, 4H, 4 CH
                    (piperidinylmethyl)), 1.69–1.74 (m, 1H, CH (piperidinylmethyl)), 2.79
                    (td, J = 12.5, 2.5 Hz, 2H, 2 CH (piperidinylmethyl)), 2.95 (br
                    d, J = 7.0 Hz, 2H, 2 CH(piperidinylmethyl)), 3.28 (s, 3H,
                        CH3 (Im)), 3.34–3.39 (m, 2H,
                        SO2NCH2CH2N),
                    3.41–3.42 (m, 2H,
                        SO2NCH2CH2N), 3.68
                    (s, 3H, CH3 (Im)), 4.38 (s, 2H,
                    CH2Im), 4.60–4.64 (m, 2 H, 2 CH
                    (piperidinylmethyl)), 6.41 (t, J = 4.8 Hz, 1H, CH
                    (pyrimidine)), 6.86 (s, 1H, CH (Ar)), 7.02 (s, 1H, CH (Im)), 7.20 (m, 1H, CH
                    (Ar)), 7.41–7.50 (m, 2H, CH (Im), CH (Ar)), 7.65 (s, 1H, CH (Im)), 7.80
                    (s, 1H, CH (Im)), 8.26 (t, J = 4.9 Hz, 2H, 2 CH (pyrimidine));
                        Î´C (125 MHz,
                        DMSO-d6) 29.5, 32.0, 32.8, 34.0, 35.9, 43.5,
                    44.2, 48.9, 49.2, 49.4, 49.7, 53.2, 56.2, 109.5, 118.1, 120.8, 122.3, 124.6,
                    129.4, 135.8, 139.3, 145.5, 157.7, 161.4; HRMS (ESI+) m/z calcd
                    for [C27H33N11O2S + H] 576.2618,
                    obsd 576.2617.</p><!><p>As per general procedure E with 18c on a 0.178 mmol scale.
                    After the usual workup, the crude material was purified by silica gel flash
                    column chromatography (eluent
                    CH2Cl2/MeOH/NH4OH, 192:7:1) to furnish
                        [N-{2-[(5-bromopyridin-2-yl)-(3-methyl-3H-imidazol-4-ylmethyl)amino]ethyl},N-{N-(2-pyrimidinyl)-piperidin-4-ylmethyl}]
                        1-methyl-1H-imidazole-4-sulfonamide as a white foam (107
                    mg, 96%): Î´H (500 MHz, CDCl3) 1.12
                    (qd, J = 12.5, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.72–1.80 (m, 3H, 3 CH (piperidinylmethyl)), 2.84 (td, J
                    = 13.0, 2.5 Hz, 2H, 2 CH (piperidinylmethyl)), 2.98 (br d, J =
                    7.0 Hz, 2H, 2 CH (piperidinylmethyl)), 3.14–3.18 (m, 2H,
                        NSO2CH2CH2N), 3.55
                    (s, 3H, CH3 (Im)), 3.62–3.67 (m, 2H,
                        NSO2CH2CH2N), 3.73
                    (m, 3H, CH3 (Im)), 4.69–4.78 (m, 2H, 2 CH
                    (piperidinylmethyl)), 4.80 (s, 2H, CH2Im),
                    6.43 (t, J = 4.9 Hz, 1H, CH (pyrimidine)), 6.69 (d,
                        J = 9.5 Hz, 1H, CH (Ar)), 6.96 (s, 1H, CH (Im)), 7.38 (s,
                    1H, CH (Im)), 7.43 (s, 1H, CH (Im)), 7.46 (s, 1H, CH (Im)), 7.55 (dd,
                        J = 9.0, 2.5 Hz, 1H, CH (Ar)), 8.15 (d, J
                    = 2.5 Hz, 1H, CH (Ar)), 8.27 (t, J = 4.9 Hz, 2H, 2 CH
                    (pyrimidine)); Î´C (125 MHz, CDCl3)
                    29.6, 31.8, 33.9, 35.6, 41.0, 43.5, 46.2, 47.0, 55.9, 107.1, 107.6, 109.3,
                    124.2, 128.2, 129.1, 138.8, 138.9, 139.3, 140.1, 148.2, 155.9, 157.6, 161.5;
                    HRMS (ESI+) m/z calcd for
                        [C26H33N10O2SBr + H] 629.1770,
                    obsd 629.1780; HPLC (I) tR = 12.78 min (100%), +
                    (II) tR = 18.88 min (100%). The aryl bromide was
                    then converted to the corresponding aryl nitrile on a 0.115 mmol scale,
                    according to general procedure F. After workup, the crude material was
                    dry-loaded onto silica gel, and purified by flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to give
                        19c as a white foam (51 mg, 77%):
                        Î´H (500 MHz, CDCl3) 1.11 (qd,
                        J = 12.5, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.67–1.77 (m, 3H, 3 CH (piperidinylmethyl)), 2.78–2.87 (m, 2H, 2
                    CH (piperidinylmethyl)), 2.93 (br d, J = 7.0 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.15–3.22 (m, 2H,
                        SO2NCH2CH2N), 3.54
                    (s, 3H, CH3 (Im)), 3.69–3.78 (m, 5H,
                        SO2NCH2CH2N,
                        CH3 (Im)), 4.68–4.76 (m, 2H, 2 CH (piperidinylmethyl)),
                    4.91 (s, 2H, CH2Im), 6.43 (t,
                        J = 4.9 Hz, 1H, CH (pyrimidine)), 6.85 (d,
                        J = 9.5 Hz, 1H, CH (Ar)), 6.97 (s, 1H, CH (Im)), 7.39 (s,
                    1H, CH (Im)), 7.42 (s, 1H, CH (Im)), 7.43 (s, 1H, CH (Im)), 7.66 (dd,
                        J = 9.0, 2.5 Hz, 1H, CH (Ar)), 8.27 (t, J
                    = 4.9 Hz, 2H, 2 CH (pyrimidine)), 8.41 (d, J = 2.5 Hz, 1H, CH
                    (Ar)); Î´C (125 MHz, CDCl3) 29.7,
                    31.9, 34.0, 35.7, 41.0 (br), 43.6, 46.3, 47.1, 56.0, 97.1, 106.1, 109.4, 118.3,
                    124.3, 127.3, 129.7, 139.0, 139.2, 139.4, 140.3, 152.4, 157.6, 158.5, 161.6;
                    HRMS (ESI+) m/z calcd for
                        [C27H33N11O2S + H] 576.2618,
                    obsd 576.2628; HPLC (I) tR = 12.43 min (100%), (II)
                        tR = 17.87 min (100%).</p><!><p>As per general procedure E with 18d on a 0.187 mmol scale
                    to afford
                        [N-{2-[(5-bromopyrimidin-2-yl)-(3-methyl-3H-imidazol-4-ylmethyl)amino]ethyl},N-{N-(2-pyrimidinyl)-piperidin-4-ylmethyl}]
                        1-methyl-1H-imidazole-4-sulfonamide as a white foam (109
                    mg, 93%): Î´H (500 MHz, CDCl3) 1.17
                    (qd, J = 12.2, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.78–1.84 (m, 2H, 2 CH (piperidinylmethyl)), 1.92–1.99 (m, 1H, CH
                    (piperidinylmethyl)), 2.82 (td, J = 12.8, 2.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.10 (br d, J = 7.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.19–3.23 (m, 2H,
                        SO2NCH2-CH2N), 3.58
                    (s, 3H, CH3 (Im)), 3.71–3.75 (m, 5H,
                        SO2NCH2-CH2N,
                        CH3 (Im)), 4.71–4.77 (m, 2 H, 2 CH (piperidinylmethyl)),
                    4.89 (s, 2H, CH2Im), 6.43 (t,
                        J = 4.9 Hz, 1H, CH (pyrimidine)), 7.05 (s, 1H, CH (Im)),
                    7.42 (s, 1H, CH (Im)), 7.43 (s, 1H, CH (Im)), 7.46 (s, 1H, CH (Im)), 8.28 (t,
                        J = 4.9 Hz, 2H, 2 CH (pyrimidine)) 8.30 (s, 2H, 2 CH (Ar));
                        Î´C (125 MHz, CDCl3) 29.7,
                    31.9, 33.9, 35.6, 40.7, 43.7, 46.0, 46.2, 55.8, 106.4, 109.3, 124.4, 128.1,
                    129.5, 138.5, 138.9, 139.3, 157.6, 158.0, 159.3, 161.6; HRMS (ESI )
                        m/z calcd for
                        [C25H32N11O2SBr + H] 630.1723,
                    obsd 630.1748; +HPLC (I) tR = 13.05 min (100%), (II)
                        tR = 19.52 min (100%). The aryl bromide was then
                    converted to the corresponding aryl nitrile on a 0.127 mmol scale, according to
                    general procedure F. After workup, the crude material was dry-loaded onto silica
                    gel and purified by flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to give
                        19d as a white foam (50 mg, 69%):
                        Î´H (500 MHz, CDCl3) 1.16 (qd,
                        J = 12.2, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.75–1.82 (m, 2H, 2 CH (piperidinylmethyl)), 1.86–1.94 (m, 1H, CH
                    (piperidinylmethyl)), 2.80 (td, J = 12.8, 2.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.07 (br d, J = 7.5 Hz, 2H, 2CH
                    (piperidinylmethyl)), 3.26–3.30 (m, 2H,
                        SO2NCH2CH2N), 3.58
                    (s, 3H, CH3 (Im)), 3.74 (s, 3H, CH3 (Im)),
                    3.83–3.87 (m, 2H, SO2NCH2-CH2N),
                    4.70–4.76 (m, 2 H, 2 CH (piperidinylmethyl)), 4.99 (s, 2H,
                        CH2Im), 6.44 (t, J = 4.8
                    Hz, 1H, pyrimidine), 7.07 (s, 1H, CH (Im)), 7.39–7.44, (s, 3H, 3 CH
                    (Im)), 8.28 (t, J = 4.9 Hz, 2H, 2 CH (pyrimidine)), 8.54 (br s,
                    2H, 2 CH (Ar)); Î´C (125 MHz,
                    CDCl3) 29.7, 32.0, 34.0, 35.8, 40.9, 43.7, 46.3, 46.5, 55.9, 96.8,
                    109.5, 116.2, 124.5, 127.2, 129.8, 138.8, 139.0, 139.3, 157.7, 160.6, 161.0,
                    161.6; HRMS (ESI+) m/z calcd for
                        [C26H32N12O2S + H] 577.2570,
                    obsd 577.2569; HPLC (I) tR = 12.45 min (100%), (II)
                        tR = 17.94 min (100%).</p><!><p>As per general procedure E with 18e on a 0.0683 mmol scale.
                    After the usual workup, the crude material was purified by silica gel flash
                    column chromatography (eluent
                    CH2Cl2/MeOH/NH4OH, 192:7:1) to furnish
                        19e as a white foam (36 mg, 89%):
                        Î´H (500 MHz, CDCl3) 1.10 (qd,
                        J = 12.2, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.60–1.67 (m, 1H, CH (piperidinylmethyl)), 1.69–1.75 (m, 2H, 2 CH
                    (piperidinylmethyl)), 2.80 (td, J = 12.8, 2.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 2.95 (br d, J = 7.5 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.20–3.24 (m, 2H,
                        SO2NCH2CH2N), 3.56
                    (s, 3H, CH3 (Im)), 3.62–3.66 (m, 2H,
                        SO2NCH2CH2N),
                    3.72 (m, 3H, CH3 (Im)), 4.52 (s, 2H,
                        CH2Im), 4.67–4.73 (m, 2H, 2 CH
                    (piperidinylmethyl)), 6.43 (t, J = 4.7 Hz, 1H, CH
                    (pyrimidine)), 6.60 (dd, J = 13.0, 2.5 Hz, 1H, CH (Ar)), 6.67
                    (dd, J = 9.0, 2.5 Hz, 1H, CH (Ar)), 6.87 (s, 1H, CH (Im)),
                    7.37–7.40 (m, 2H, CH (Im), CH (Ar)), 7.43 (s, 1H, CH (Im)), 7.48 (s, 1H,
                    CH (Im)), 8.26 (t, J = 4.7 Hz, 2H, 2 CH (pyrimidine));
                        Î´C (125 MHz, CDCl3) 29.5,
                    31.7, 34.0, 35.9, 43.5, 44.5, 46.5, 49.3, 56.3, 88.1 (d,
                        JCF = 15.5 Hz), 99.3 (d,
                        JCF = 24.6 Hz), 108.4 (d,
                        JCF = 1.9Hz), 109.5, 115.2, 124.4, 126.3, 129.1,
                    134.3 (d, JCF = 2.75 Hz), 139.0, 139.1, 139.2, 152.7
                    (d, JCF = 10.9 Hz), 157.6, 161.5, 164.9 (d,
                        JCF = 252 Hz); HRMS (ESI+) m/z
                    calcd for [C28H33N10O2SF + H]
                    593.2571, obsd 593.2588; HPLC (I) tR = 12.54 min
                    (98.51%), (II) tR = 18.51 min (98.09%).</p><!><p>As per general procedure E with 18f on a 0.060 mmol scale.
                    After the usual workup, the crude material was purified by silica gel flash
                    column chromatography (eluent
                    CH2Cl2/MeOH/NH4OH, 192:7:1) to afford
                        19f as a white foam (33 mg, 93%):
                        Î´H (500 MHz, CDCl3) 1.09 (qd,
                        J = 12.3, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.70–1.78 (m, 3H, 3 CH (piperidinylmethyl)), 2.76–2.83 (m, 2H, 2
                    CH (piperidinylmethyl)), 2.95 (br d, J = 7.0 Hz, 2H, 2 CH
                    (piperidinylmethyl)), 3.31 (br t, J =7.5Hz,2H,
                        SO2NCH2CH2N), 3.53
                    (br t, J =7.5Hz, 2H,
                        SO2NCH2CH2N), 3.58
                    (m, 3H, CH3 (Im)), 3.73 (m, 3H, CH3 (Im)), 4.50 (s, 2H,
                        CH2Im), 4.67–4.73 (m, 2H, 2 CH
                    (piperidinylmethyl)), 6.43 (t, J = 4.7 Hz, 1H, CH
                    (pyrimidine)), 6.93 (s, 1H, CH (Im)), 7.06 (t, J = 8.8 Hz, 1H,
                    CH (Ar)), 7.28 (dd, J = 13.0, 1.8 Hz, 1H, CH (Ar)), 7.31 (br
                    dd, J = 8.5, 1.8 Hz, 1H, CH (Ar)), 7.37 (s, 1H, CH (Im)), 7.41
                    (s, 1H, CH (Im)), 7.48 (s, 1H, CH (Im)), 8.26 (t, J = 4.7 Hz,
                    2H, 2 CH (pyrimidine)); Î´C (125 MHz,
                        CDCl3) 29.6, 31.8, 34.0, 35.9, 43.5, 45.9, 47.5, 50.5, 55.8,
                    103.0 (d, JCF = 10 Hz), 109.4, 118.2 (d,
                        JCF = 1.8 Hz), 119.5 (d,
                        JCF = 4.6 Hz), 120.2 (d,
                        JCF = 25.5 Hz), 124.2, 126.9, 129.3, 129.5 (d,
                        JCF = 2.8 Hz), 138.9, 139.0, 139.6, 141.5 (d,
                        JCF = 8.1 Hz), 152.9 (d,
                        JCF =244 Hz), 157.6, 161.5; HRMS (ESI+)
                        m/z calcd for
                        [C28H33N10O2-FS + H] 593.2571,
                    obsd 593.2568; HPLC (I) tR = 12.75 min (100%), (II),
                        tR = 19.09 (99.22%).</p><!><p>As per general procedure E with 18g on a 0.068 mmol scale.
                    After the usual workup, the crude material was purified by silica gel flash
                    column chromatography (eluent
                    CH2Cl2/MeOH/NH4OH, 192:7:1) to yield
                        19g as a white foam (40 mg, 97%):
                        Î´H (500 MHz, CDCl3) 1.06 (qd,
                        J = 12.3, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.67–1.77 (m, 3H, 3 CH (piperidinylmethyl)), 2.78 (td, J
                    = 12.8, 2.5 Hz, 2H, 2 CH (piperidinylmethyl)), 2.96 (br d, J =
                    6.5 Hz, 2H, 2 CH (piperidinylmethyl)), 3.26–3.30 (m, 2H,
                        SO2NCH2CH2N),
                    3.39–3.43 (m, 2H,
                        SO2NCH2CH2N), 3.57
                    (s, 3H, CH3 (Im)), 3.73 (s, 3H, CH3 (Im)), 4.38 (s, 2H,
                        CH2Im), 4.66–4.73 (m, 2H, 2 CH
                    (piperidinylmethyl)), 6.42 (t, J = 4.9 Hz, 1H, CH
                    (pyrimidine)), 6.92 (s, 1H, CH (Im)), 7.12–7.17 (m, 2H, 2 CH (Ar)), 7.35
                    (s, 1H, CH (Im)), 7.40 (s, 1H, CH (Im)), 7.42 (s, 1H, CH (Im)), 8.26 (t,
                        J = 4.9 Hz, 2H, 2 CH (pyrimidine));
                        Î´C (125 MHz, CDCl3) 29.6,
                    31.4, 33.9, 36.0, 43.5, 46.3 (t, JCF = 4.5 Hz),
                    47.8, 51.9 (t, JCF = 2.6 Hz), 55.6, 106.9 (t,
                        JCF = 11.9 Hz), 109.4, 116.6 (m), 116.6 (t,
                        JCF = 2.8 Hz), 124.1, 126.9, 129.9, 131.1 (t,
                        J = 13.3 Hz), 138.9, 139.2, 139.7, 157.6, 158.3 (dd,
                        JCF = 249, 8.3 Hz), 161.5; HRMS (ESI+)
                        m/z calcd for
                        [C28H32N10O2F2S + H]
                    611.2477, obsd 611.2494; + HPLC (I) tR = 12.93 min
                    (100%), (II) tR = 19.59 min (100%).</p><!><p>As per general procedure E with 18h on a
                    0.0688 mmol scale. After the usual workup, the crude material was purified by
                    silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1) to afford
                        19h as a white foam (28 mg, 63%):
                        Î´H (500 MHz, CDCl3) 1.08 (qd,
                        J = 12.2, 4.0 Hz, 2H, 2 CH (piperidinylmethyl)),
                    1.69–1.81 (m, 3H, 3 CH (piperidinylmethyl)), 2.80 (td, J
                    = 12.8, 2.5 Hz, 2H, 2 CH (piperidinylmethyl)), 2.96 (br d, J =
                    7.5 Hz, 2H, 2 CH (piperidinylmethyl)), 3.34–3.40 (m, 2H,
                        SO2NCH2CH2N),
                    3.51–3.56 (m, 2H,
                        SO2NCH2CH2N), 3.57
                    (s, 3H, CH3 (Im)), 3.74 (s, 3H, CH3 (Im)), 4.53 (s, 2H,
                        CH2Im), 4.68–4.74 (m, 2 H, 2 CH
                    (piperidinylmethyl)), 6.43 (t, J = 4.7 Hz, 1H, CH
                    (pyrimidine)), 6.98 (s, 1H, CH (Im)), 7.38 (s, 1H, CH (Im)), 7.41–7.46
                    (m, 2H, 2 CH (Im)), 8.27 (t, J = 4.7 Hz, 2H, CH (pyrimidine));
                        Î´C (125 MHz, CDCl3) 29.6,
                    31.5, 33.9, 36.0, 43.5, 46.2, 47.9, 51.9 (m), 55.7, 86.9 (t,
                        JCF = 17.4 Hz), 107.8 (m), 109.5, 124.2, 126.2,
                    130.2, 133.9 (m), 139.0, 139.4, 139.6, 142.1 (dm), 147.8 (dm), 157.6, 161.5;
                    HRMS (ESI+) m/z calcd for
                        [C28H30N10O2F4S + H]
                    647.2288, obsd 647.2314; HPLC (I) tR = 12.74 min
                    (98.92%), (II) tR = 19.07 min (98.14%).</p><!><p>Mono-N-Boc-ethylenediamine (6; 860 mg,
                    5.37 mmol, 1 equiv) was reacted with pyridine-2-sulfonyl chloride according to
                    general procedure A. After workup, the crude material was dry-loaded onto silica
                    gel and purified by flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1) to afford
                        tert-butyl 2-(pyridine-2-sulfonamido)ethylcarbamate as a
                    white powder (1.26 g, 80%): Î´H (400 MHz,
                        DMSO-d6) 0.89 (s, 9H,
                        C(CH3)3), 2.04 (m, 2H,
                        SO2NHCH2CH2NHBoc),
                    2.86 (m, 2H,
                    SO2NHCH2CH2NHBoc), 6.30
                    (br, 1H, NHSO2), 7.21 (m, 1H, CH (Py)), 7.414
                    (m, 1H, NHBoc), 7.47 (m, 1H, CH (Py)), 7.63, (m, 1H, CH
                    (Py)), 8.27 (m, 1H, CH (Py)); Î´C (125 MHz,
                        DMSO-d6) 28.0, 39.9, 42.5, 77.6, 121.4, 126.8,
                    138.5, 149.8, 155.3, 157.7; HRMS (ESI+) m/z
                    calcd for [C12H19N3O4S + H]
                    302.1175, obsd 302.1174. The sulfonamide NH of tert-butyl
                    2-(pyridine-2-sulfonamido)ethylcarbamate (1.1 g, 3.65 mmol, 1 equiv) was then
                    chemoselectively alkylated with cyclohexylmethyl bromide (840 mg, 4.75 mmol, 1.3
                    equiv) according to general procedure B, but the reaction was
                    stirred for 4 days at room temperature. After workup, the crude material was
                    dry-loaded onto silica gel and purified by flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 92:7:1) to furnish
                        tert-butyl
                    2-(N-(cyclohexylmethyl)pyridine-2-sulfonamido)ethylcarbamate in
                    quantitative yield (1.53 g) as a white solid:
                        Î´H (500 MHz,
                        DMSO-d6) 0.77–0.83 (m, 2H, 2 CH
                    (cyclohexylmethyl)), 1.09–1.19 (m, 3H, 3 CH (cyclohexylmethyl)), 1.36 (s,
                    9H, C(CH3)3), 1.54–1.65 (m, 6H, 6 CH
                    (cyclohexylmethyl)), 3.03–3.07 (m, 4H,
                        SO2NCH2CH2-NHBoc,
                        NCH2CH (cyclohexylmethyl)), 3.24 (t,
                        J = 7.0 Hz, 2H,
                        SO2NCH2CH2NHBoc),
                    6.79 (br, 1H, NHBoc)), 7.67 (m, 1H, CH (Py)), 7.92 (m,
                    1H, CH (Py)), 8.09 (m, 1H, CH (Py)), 8.72 (m, 1H, CH (Py));
                        Î´C (125 MHz,
                        DMSO-d6) 25.1, 25.8, 28.0, 29.9, 35.6, 39.9,
                    48.4, 55.4, 77.6, 122.1, 127.0, 138.6, 150.0, 155.3, 157.1; HRMS (ESI+)
                        m/z calcd for
                        [C19H31N3O4S + H] 398.2114, obsd
                    398.2113. tert-Butyl
                    2-(N-(cyclohexylmethyl)pyridine-2-sulfonamido)ethyl-carbamate
                    (1.28 g, 3.22 mmol, 1 equiv) was dissolved in propan-2-ol (15 mL). Upon complete
                    dissolution of the solid, 4 M HCl (15 mL) was added, and the reaction was
                    stirred for one hour. All solvents were then evaporated. The residue was
                    redissolved in CH2Cl2 (250 mL) and carefully washed with
                    saturated NaHCO3 (25 mL × 3), dried
                        (Na2SO4), filtered, and concentrated to afford
                        N-(2-aminoethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide
                        (21) as a viscous oil (945 mg, 97%):
                        Î´H (500 MHz,
                        DMSO-d6) 0.78–0.84 (m, 2H, 2 CH
                    (cyclohexylmethyl)), 1.09–1.17 (m, 3H, 3 CH (cyclohexylmethyl)),
                    1.57–1.68 (m, 6H, 6 CH (cyclohexylmethyl)), 2.75 (t, J =
                    7.0 Hz, 2H, NCH2CH (cyclohexylmethyl)), 3.04
                    (t, J = 7.4 Hz, 2H,
                        SO2NCH2CH2NH2),
                    3.34 (t, J = 7.4 Hz, 2H,
                        SO2NCH2CH2NH2),
                    5.15 (br, 2H, NH2), 7.68 (m, 1H, CH (Py)), 7.93 (m, 1H, CH (Py)),
                    8.10 (m, 1H, CH (Py)), 8.73 (m, 1H, CH (Py));
                        Î´C (125 MHz, DMSO) 25.1, 25.8, 29.9,
                    35.7, 49.4, 55.5, 66.2, 122.2, 127.1, 138.7, 150.0, 157.1; HRMS (ESI+)
                        m/z calcd for
                        [C14H23N3O2S + H] 298.1589, obsd
                    298.1589.</p><!><p>The primary amine of
                        N-(2-aminoethyl)-N-(cyclohexylmethyl)-pyridine-2-sulfonamide
                        (21) was arylated with p-fluorobenzonitrile on
                    a 0.336 mmol scale, according to general procedure D. After workup
                    and purification by silica gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1),
                        N-(2-(4-cyanophenylamino)ethyl)-N-(cyclohexylmethyl)-pyridine-2-sulfonamide
                    was obtained as a colorless gum (119 mg, 89%):
                        Î´H (500 MHz,
                        DMSO-d6)0.77–0.83 (m, 2H, 2 CH
                    (cyclohexylmethyl)), 1.09 (br, 3H, 3 CH (cyclohexyl)), 1.48 1.63 (m, 6H, 6 CH
                    (cyclohexylmethyl)), 3.06 (d, J = 6.4 Hz, 2H,
                        NCH2CH (cyclohexylmethyl)),
                    3.28–3.38 (m, 4H,
                        SO2NCH2CH2NHAr),
                    6.61 (d, J = 8.5 Hz, 2H, 2 CH (Ar)), 6.70 (t,
                        J = 6.4 Hz, 1H, NH), 7.45 (d, J = 8.5 Hz,
                    2H, 2 CH (Ar)), 7.66 (m, 1H, CH (Py)), 7.94 (m, 1H, CH (Py)), 8.07 (m, 1H, CH
                    (Py)), 8.73 (m, 1H, CH (Py)); Î´C (125 MHz,
                    DMSO) 25.1, 25.8, 29.9, 35.8, 41.5, 47.7, 55.6, 95.9, 111.6, 120.3, 122.2,
                    127.1, 133.3, 138.6, 150.1, 151.6, 157.0; HRMS (ESI+)
                        m/z calcd for
                        [C21H26N4O2S + H] 399.1855, obsd
                    399.1857.
                    N-(2-(4-Cyanophenylamino)ethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide
                    was then alkylated with 5-chloromethyl-1H-imidazole·3
                    HCl according to general procedure E on a 0.117 mmol scale. After workup and
                    chromatography over silica gel (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1), the title
                    compound
                        N-(2-((4-cyanophenyl)((1-methyl-1H-imidazol-5-yl)methyl)amino)ethyl)-N-(cyclohexylmethyl)-pyridine-2-sulfonamide
                        (21a) was furnished as a glassy film (46 mg, 80%):
                        Î´H (500 MHz,
                    CDCl3)0.84–0.78 (m, 2H, 2 CH (cyclohexylmethyl)),
                    1.06–1.14 (m, 3H, 3 CH (cyclohexylmethyl)), 1.35–1.27 (m, 1H, CH
                    (cyclohexylmethyl)), 1.66–1.59 (m, 5H, 5 CH (cyclohexylmethyl)), 2.95 (d,
                        J = 7.0 Hz, 2H, NCH2CH
                    (cyclohexyl)), 3.36 (t, J = 7.9 Hz, 2H,
                        SO2NCH2CH2N), 3.61
                    (s, 3H, CH3 (Im)), 3.65 (t, J = 7.9 Hz, 2H,
                        SO2NCH2CH2N), 4.55
                    (s, 2H, CH2Im), 6.85 (d, J =
                    9.0 Hz, 2H, 2 CH (Ar)), 6.88 (s, 1H, CH (Im)), 7.47–7.44 (m, 3H, 2 CH
                    (Ar), CH (Im)), 7.71 (s, 1H, CH (Py)), 7.91–7.86 (m, 2H, 2 CH (Py)), 8.61
                    (m, 1H, CH (Py)); Î´C (125 MHz,
                        CDCl3) 25.6, 26.2, 30.6, 32.0, 36.9, 44.3, 46.8, 49.4, 57.1,
                    99.5, 112.3, 119.9, 122.5, 126.6, 127.2, 127.7, 133.8, 137.9, 138.7, 149.9,
                    150.4, 157.5; HRMS (ESI+) m/z calcd for
                        [C26H32N6O2S + H] 493.2386, obsd
                    493.2364.</p><!><p>The primary amine of
                        N-(2-aminoethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide
                        (21) was arylated with 3,4-difluorobenzonitrile on a 0.337 mmol
                    scale, according to general procedure D. After workup and purification by silica
                    gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1),
                        N-(2-(4-cyano-2-fluorophenylamino)ethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide
                    was obtained as a white foam (111 mg, 79%):
                        Î´H (500 MHz,
                        DMSO-d6) 0.83–0.77 (m, 2H, 2 CH
                    (cyclohexylmethyl)), 1.09 (br, 3H, 3 CH (cyclohexylmethyl)), 1.62–1.51
                    (m, 6H, 6 CH (cyclohexylmethyl)), 3.06 (d, J = 7.5 Hz, 2H,
                        NCH2CH (cyclohexyl)), 3.42–3.34 (m, 4H,
                        SO2NCH2CH2N),
                    6.54 (br, 1H, NHAr), 6.80 (t, J = 8.5
                    Hz, 1H, CH (Ar)), 7.45 (dd, J = 11.5, 1.5 Hz, 1H, CH (Ar)),
                    7.53 (dd, J = 8.5, 1.5 Hz, 1H, CH (Ar)), 7.66 (m, 1H, CH (Py)),
                    7.93 (m, 1H, CH (Py)), 8.07 (m, 1H, CH (Py)), 8.71 (m, 1H, CH (Py));
                        Î´C (125 MHz,
                        DMSO-d6) 25.1, 25.9, 30.0, 35.7, 37.1, 41.3,
                    47.5, 55.5, 61.8, 94.4, 122.2, 127.1, 130.4, 138.7, 140.7, 142.5, 150.1, 156.9,
                    179.7; HRMS (ESI+) m/z calcd for
                        [C21H25FN4O2S + H] 417.1760,
                    obsd 417.1749.
                        N-(2-(4-Cyano-2-fluorophenylamino)ethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide
                    was then alkylated with 5-chloromethyl-1H-imidazole·HCl
                    according to general procedure E on a 0.216 mmol scale. After workup and
                    chromatography over silica gel (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1), the title
                    compound
                        N-(2-((4-cyano-2-fluorophenyl)((1-methyl-1H-imidazol-5-yl)methyl)amino)ethyl)-N-(cyclohexylmethyl)-pyridine-2-sulfonamide
                    was afforded as a white foam (90 mg, 81%): Î´H
                    (500 MHz, CDCl3) 0.68–0.75 (m, 2H, 2 CH (cyclohexylmethyl)),
                    1.03 (br, 3H, 3 CH (cyclohexylmethyl)), 1.23–1.32 (m, 1H, CH
                    (cyclohexylmethyl)), 1.50–1.60 (m, 5H, 5 CH (cyclohexylmethyl)), 2.80 (s,
                    2H, NCH2CH (cyclohexylmethyl)), 3.34–3.47 (m, 4H,
                        SO2NCH2CH2N),
                    3.54 (s, 3H, CH3 (Im)), 4.46 (s, 2H,
                        CH2Im), 6.61 (br, 1H, CH (Ar)), 6.95 (s, 1H, CH (Im)), 7.11 (m,
                    1H, CH (Ar)), 7.26 (br, 1H, CH (Ar)), 7.53 (m, 2H, CH (Py), CH (Im)) 7.65 (br,
                    1H, CH (Py)), 8.26 (m, 1H, CH (Py)), 8.69 (m, 1H, CH (Py));
                        Î´C (125 MHz, CDCl3) 25.5,
                    26.1, 30.5, 31.3, 31.8, 36.6, 45.7, 47.3, 50.4, 56.4, 102.9, 118.1, 119.5,
                    120.2, 122.4, 126.5, 128.7, 129.3, 138.8, 141.3, 149.8, 153.9, 157.6, 162.4;
                    HRMS (ESI+) m/z calcd for
                        [C26H31FN6O2S + H] 511.2291,
                    obsd 511.2265; + HPLC (I) tR = 18.69 min (99.3%),
                    (II) tR = 36.03 min (99.7%).</p><!><p>The primary amine of
                        N-(2-aminoethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide
                        (21) was arylated with 5-cyano-2-fluoropyridine on a 0.350 mmol
                    scale, according to general procedure D. After workup and purification by silica
                    gel flash column chromatography (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1),
                        N-(2-(5-cyanopyridin-2-ylamino)ethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide
                    was obtained as a colorless gum (115 mg, 82%):
                        Î´H (500 MHz,
                        DMSO-d6) 0.78–0.82 (m, 2H, 2 CH
                    (cyclohexylmethyl)), 1.09 (br, 3H, 3 CH (cyclohexylmethyl)), 1.57–1.63
                    (m, 6H, 6 CH (cyclohexylmethyl)), 3.06 (d, J = 7.5 Hz, 2H,
                        NCH2CH (cyclohexylmethyl)),
                    3.37–3.47 (m, 4H,
                        SO2-NCH2CH2N),
                    6.50 (t, J = 8.5 Hz, NH), 6.87 (d, J = 9.0 Hz,
                    1H, CH (Ar)), 7.61–7.66 (m, 3H, 2 CH (Ar), CH (Py)), 7.92 (d,
                        J = 7.5 Hz, 1H, CH (Py)), 8.06 (m, 1H, CH (Py)), 8.36 (m,
                    1H, CH (Ar)), 8.71 (m, 1H, CH (Py)); Î´C (125
                    MHz, DMSO-d6) 25.1, 25.8, 30.0, 35.6, 47.7, 54.7,
                    55.5, 69.3, 94.7, 108.7, 118.7, 122.1, 126.9, 138.5, 150.0, 152.8, 157.0, 159.6;
                    HRMS (ESI+) m/z calcd for
                        [C20H25N5O2S + H] 400.1807, obsd
                    400.1795.
                        N-(2-(5-Cyanopyridin-2-ylamino)ethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide
                    was then alkylated with 5-chloromethyl-1H-imidazole·HCl
                    according to general procedure E on a 0.240 mmol scale. After workup and
                    chromatography over silica gel (eluent
                        CH2Cl2/MeOH/NH4OH, 192:7:1), the title
                    compound
                        N-(2-((5-cyanopyridin-2-yl)((1-methyl-1H-imidazol-5-yl)methyl)-amino)ethyl)-N-(cyclohexylmethyl)pyridine-2-sulfonamide
                    was afforded as a white foam (110 mg, 93%):
                        Î´H (500 MHz, CDCl3)
                    0.75–0.83 (m, 2H, 2 CH (cyclohexylmethyl)), 1.07–1.16 (m, 3H, 3 CH
                    (cyclohexylmethyl)), 1.33–1.43 (m, 1H, CH (cyclohexylmethyl)),
                    1.59–1.65 (m, 5H, 5 CH (cyclohexylmethyl)), 2.94 (d, J =
                    7.0 Hz, 2H, NCH2CH (cyclohexylmethyl)), 3.28
                    (t, J = 8.1 Hz, 2H,
                        SO2NCH2CH2N), 3.55
                    (s, 3H, CH3 (Im)), 3.70 (t, J = 8.1 Hz, 2H,
                        SO2NCH2CH2N),
                    4.89 (s, 2H, CH2Im), 6.81 (d,
                        J = 9.0 Hz, 1H, CH (Ar)), 6.97 (s, 1H, CH (Im)), 7.45 (m,
                    1H, CH (Im)), 7.57 (s, 1H, CH (Ar)), 7.64 (m, 1H, CH (Ar)), 7.89–7.85 (m,
                    2H, 2 CH (Py)), 8.39–8.38 (m, 1H, CH (Ar)), 8.60 (m, 1H, CH (Py));
                        Î´C (125 MHz, CDCl3) 25.6,
                    26.2, 30.5, 32.0, 36.6, 40.9, 47.2, 56.8, 97.1, 105.9, 118.2, 122.5, 126.6,
                    127.4, 128.8, 137.9, 138.9, 140.2, 149.9, 152.3, 157.5, 158.4, 162.4; HRMS (ESI
                    ) m/z calcd for
                        [C25H31N7O2S + H] 494.2338, obsd
                    494.2313; HPLC + (I) tR = 18.15 min (99.5%), (II)
                        tR = 34.66 min (97.4%).</p><!><p>In vitro inhibition assays for FTase and GGTase-I were conducted by
                    measuring the incorporation of [3H]FPP and [3H]GGPP into
                    recombinant H-Ras-CVLS and H-Ras-CVLL, respectively, as previously
                        described.36 The in vivo inhibition
                    of farnesylation and geranylgeranylation was determined based on the level of
                    inhibition by synthetic compounds of H-Ras and Rap1A processing,
                        respectively.37 Briefly, oncogenic
                    H-Ras-transformed NIH3T3 cells were treated with various concentrations of
                    inhibitors, and the cell lysates were isolated and proteins separated on a 12.5%
                    SDS-PAGE gel. The separated proteins were transferred to nitrocellulose and
                    immunoblotted using an anti-Ras antibody (Y13–258) or an anti-Rap1A
                    antibody (SantaCruz Biotechnology, Santa Cruz, CA), respectively. Antibody
                    reactions were visualized using either peroxidase-conjugated goat antirat IgG or
                    goat antirabbit IgG (Jackson ImmunoResearch, West Grove, PA) and an enhanced
                    chemiluminescence detection system.</p>