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Targeting p85β nuclear translocation for the tumors with PIK3CA helical domain mutations

helical domain mutations promote tumorigenesis through two independent pathways: (1) p110 a helical domain mutant protein directly interacts with IRS1 to activate the canonical AKT pathway; and (2) p85 b disassociates from p110 a helical-domain-mutated PI3K complex and translocates into the nucleus to stabilize EZH1/2 and enhance H3K27 trimethylation. Simultaneously targeting PI3K-AKT signaling and the nuclear p85 b -EZH1/2 pathways could be an effective therapy strategy for PIK3CA helical domain mutant cancers.


PIK3CA;
Helical domain mutation; p85b; EZH2; USP7 Phosphatidylinositol 3-kinases (PI3Ks) play key roles in tumorigenesis. PIK3CA, which encodes PI3K complex catalytic subunit p110a, is one of the most frequently mutated oncogenes in human cancers. 1 So, targeting p110a holds great promise for cancer therapy. Alpelisib, a small molecule inhibitor specifically targeting PIK3CA/p110a, has been approved by FDA to treat HR-positive and HER2-negative breast cancer patients harboring PIK3CA gene mutations. 2 Most PIK3CA/p110a mutations occur at two hot spot regions: an acidic cluster (E542, E545, and Q546) in the helical domain and a histidine residue (H1047) in the kinase domain. 3 Although all these hot-spot mutations activate the PI3-kinase activity, p110a helical domain mutations and kinase domain mutations promote tumorigenesis through different molecular mechanisms. 4 Moreover, PIK3CA helical domain mutant tumors were less responsive to alpelisib treatment compared with PIK3CA H1047R mutant tumors in early clinical trials, 5 while the mechanism has not been clearly clarified. Therefore, it is important to investigate the oncogenic mechanism and develop a more effective therapeutic strategy for tumors with PIK3CA helical domain mutations (Fig. 1).
The regulatory subunits of PI3K complex p85s normally stabilize p110 subunits and inhibit their enzymatic activity. Our previous studies have demonstrated that the p110a proteins with helical domain mutations can directly interact with IRS1 to activate downstream AKT signaling. 3 This process is independent of p85s. 3 Whether p85s play important role in tumors with PIK3CA helical domain mutations remains unknown. Recently, we reported that p85b translocated into the nucleus and epigenetically regulated genes expression via stabilizing EZH1/2 proteins in PIK3CA helical mutant tumors. A combination of EZH inhibitors and the p110a inhibitor alpelisib induced the repression of tumors with PIK3CA helical domain mutations. 4 This study indicates that simultaneously targeting EZHs and p110a could be a potential effective therapeutic strategy for PIK3CA helical domain mutant tumors.
We identified a nuclear localization signal (NLS) at amino acids 474 to 484 of p85b which mediated its nuclear translocation in cancer cells with a PIK3CA helical domain mutation. 4 p85b could disassociate from p110a helical-domain-mutated PI3K complex. p110-free p85b exposes NLS which is normally embedded in the interface of p85b/p110 complex to trigger the nuclear translocation. Furthermore, our unpublished data indicates that the disassociation of p85b/p110a rely on p85b tyrosine phosphorylation. These data suggest that p110-free p85s could actively translocate into the nucleus under certain physiological conditions.
Our study reveals the oncogenic function of nuclearlocalized p85b in cancer development. Several studies showed p85b promoted the tumorigenesis through regulating PI3K activity and downstream AKT pathway. However, it is unknown whether p110-free p85s play important role in the tumorigenesis. In our study, we provided compelling evidence to show nuclear but not cytoplasmic p85b functions as an oncogene in tumors with PIK3CA helical domain mutations, which is independent of the canonical AKT signaling Figure 1 The oncogenic mechanism in tumors with PIK3CA helical domain mutations. PIK3CA helical domain mutations promote tumorigenesis through two independent pathways: (1) p110a helical domain mutant protein directly interacts with IRS1 to activate the canonical AKT pathway; and (2) p85b disassociates from p110a helical-domain-mutated PI3K complex and translocates into the nucleus to stabilize EZH1/2 and enhance H3K27 trimethylation. Simultaneously targeting PI3K-AKT signaling and the nuclear p85b-EZH1/2 pathways could be an effective therapy strategy for PIK3CA helical domain mutant cancers.
pathway. In the patients with PIK3CA helical domain mutations, high PIK3R2/p85b levels were significantly correlated with poor overall survival. In cancer cells with PIK3CA helical domain mutations, depleting p85b or blocking p85b nuclear translocation could reduce cell proliferation, colony formation, and xenograft tumor growth. On the contrary, p85b didn't show oncogenic function in cancer cells with PIK3CA kinase domain mutations or wild-type PIK3CA, which barely had nuclear-localized p85b. Together, we speculate p85b might perform oncogenic role through different signaling pathways in different cancers.
Our study demonstrates that the nuclear p85b stabilizes EZH1/2 and enhances H3K27 trimethylation in cancer cells with PIK3CA helical domain mutations. 4 Nuclear p85b recruits deubiquitinase USP7 to histone methyltransferase EZH1/2 protein to prevent them from ubiquitin-mediated protein degradation, therefore enhancing the trimethylation of histone H3K27. As repressive histone mark, H3K27me3 was enriched at specific genome regions, especially the promoter region of downstream target genes such as tumor suppressor DLG2, thereby promoting the growth of PIK3CA helical domain mutant tumors. Thus, nuclearlocalized p85b could directly involve in chromatin remodeling processes to serve as a transcriptional modulator in tumorigenesis.
Our study has important therapeutic implications. PIK3CA helical domain mutations promote tumorigenesis through both the canonical PI3K-AKT pathway and nuclear p85b-USP7-EZH1/2 pathways. It could partially explain that the patients with PIK3CA helical domain mutations are less responsive to alpelisib treatment. Therefore, simultaneously targeting EZHs and p110a could be a potentially effective therapeutic strategy for the patients with PIK3CA helical domain mutations. We observed that combination of alpelisib and EZH inhibitors inhibited the growth synergistically of CRC cell-derived xenograft (CDX) and patient-derived xenograft (PDX) tumors with a PIK3CA helical domain mutation, but only had additive effect on CRCs with either wildtype PIK3CA or PIK3CA H1047R mutation. As both EZH2 inhibitor tazemetostat and alpelisib are FDA-approved drugs for cancer, the evaluation of combinational effect of alpelisib and tazemetostat on the patients with PIK3CA helical domain mutations could be achieved in clinical trials soon. Currently, we have only tested the drug combination in CRC models. It is worth investigating whether this drug combination could be effective in other types of cancer patients harboring PIK3CA helical domain mutations.
In summary, our studies provide both conceptual advances to the field and therapeutic implications. Firstly, it sheds new light on the nuclear translocation and function of p85b. Secondly, nuclear-localized p85b is identified as an epigenetic regulator. Thirdly, a combination of an EZH inhibitor and a p110a inhibitor would be an effective approach to treat PIK3CA helical domain mutant cancers.