{'fraction_non_alphanumeric': 0.03112497058131325, 'fraction_numerical': 0.022828900917863028, 'mean_word_length': 4.92849668643181, 'pattern_counts': {'":': 0, '<': 4, '<?xml version=': 0, '>': 6, 'https://': 1, 'lorem ipsum': 0, 'www.': 1, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 1, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
Effective low–dose sirolimus regimen for kaposiform haemangioendothelioma with Kasabach–Merritt phenomenon in young infants

Aims Management of kaposiform haemangioendotheliomas (KHE) with Kasabach–Merritt phenomenon is challenging in young infants who are subjected to developmental pharmacokinetic changes. Sirolimus, sometimes combined with corticosteroids, can be used as an effective treatment of KHE. Simultaneously, toxicities such as interstitial pneumonitis related to the use of sirolimus may be fatal. As infants have a very low CYP3‐enzyme expression at birth, which rises during ageing, we hypothesize that a reduced metabolization of sirolimus might lead to high sirolimus serum levels and low dose may be sufficient without the side effects. Methods A case series of 5 infants with kaposiform haemangioendothelioma with Kasabach–Merritt phenomenon was analysed retrospectively. All infants were treated with sirolimus 0.2 mg/m2 every 24 or 48 hours according to their age. Prednisone was added to the therapy for additional effect in 4 patients. Results In all patients, low dose of sirolimus led to therapeutic sirolimus levels (4–6 ng/mL). All infants (aged 4 days–7 months) had a complete haematological response, without serious adverse events. In all patients, the Kasabach–Merritt phenomenon resolved, the coagulation profile normalized and tumour size reduction was seen. Conclusion Low‐dose sirolimus treatment is safe for infants with kaposiform haemangioendothelioma and Kasabach–Merritt phenomenon. It is essential to realize that during the first months of life, metabolism is still developing and enzymes necessary to metabolise drugs like sirolimus still have to mature. To avoid toxic levels, the sirolimus dosage should be based on age and the associated pharmacological developments.


| INTRODUCTION
Kaposiform haemangioendothelioma (KHE) is a rare locally aggressive vascular tumour that acts as an infiltrative lesion, invading the skin, subcutaneous fat mass, and muscle planes. KHE occurs with Kasabach-Merritt phenomenon (KMP) in 42-71% of cases. [1][2][3][4] The KMP is characterized by a coagulopathy with profound thrombocytopenia, hypofibrinogaenemia and elevation of D-dimers, which can be life-threatening. A form of disseminated intravascular coagulation (DIC) can be seen in the patients having both the propensity of clotting and the high risk of bleeding. The morbidity and mortality rates are up to 37% in patients with KHE due to the KMP which may have an acute onset. 1,3,[5][6][7] Knowledge about this clinical condition is often insufficient, due the fact that the prevalence of KHE is very rare-approximately 0.91/100 000 children. There is not much known about the aetiology of KHE. Regarding genetics, it has been described in 1 patient that there was a somatic balanced translocation between chromosomes 13 and 16. 8 Furthermore, in other patients, activating mutations have been found in GNAQ and its paralogues (such as GNA11 and GNA14). [9][10][11] Immediate recognition of KHE including the KMP is of utmost importance and can be lifesaving. 1 Treatment options include steroids, radiation therapy, vincristine, interferon, propranolol and other chemotherapeutic agents such as cyclophosphamide. In the past decade, more and more cases of KHE with KMP have been treated with the mechanistic target of rapamycin kinase inhibitor, sirolimus. Whereas steroids, vincristine and other chemotherapeutic agents were often toxic and not always successful, sirolimus seems to have a substantial better outcome. [12][13][14][15][16][17][18] Still, toxicity directly related to high plasma levels of sirolimus can lead to severe complications and even fatal outcomes as has been described previously. 12,19 A systematic review of Freixo et al. confirmed the effectiveness of sirolimus in the treatment of KHE with KMP. 20 In 95.5% of the 121 patients the KHE clinically improved and in 93% the coagulopathy normalized. Most frequently used target levels of sirolimus were 10-15 ng/mL (38.4%), 15-20 ng/mL (24.7%) and 5-15 ng/mL (15.1%). Side effects in patients with vascular anomalies included oral mucositis (21.9%), dyslipidaemia (16.5%), leukopenia (12.3%), gastrointestinal symptoms (10.2%) and rash/eczema (8.2%).
In 5.5% of patients with oral sirolimus treatment, complications of infections were reported, such as fatal pneumoniae in 2 children aged 1 and 6 months with KHE. 19 It may be possible that 1 of these patients suffered from this sirolimus-associated interstitial pneumonitis, as signs of infections were lacking and antibiotic therapy failed. It is well known that sirolimus-associated interstitial pneumonitis is a rare serious and sometimes even fatal complication. [21][22][23][24][25] As in most drugs observed, there might be a relation between levels of sirolimus and the occurrence of (severe) adverse events (AEs). 26 For example, there is a correlation of the occurrence of thrombocytopenia, leukopenia and hypertriglyceridaemia and sirolimus levels >15 ng/mL. 27 No toxic values were seen when the steady state concentration was <10 ng/mL. This is also seen in our case series of patients with low flow vascular malformations showing that low-dose sirolimus led to less (serious) AEs compared to intermediate-high dose sirolimus. 28 In some case series of KHE patients described in the literature, a dose of 0.8 mg/m 2 twice daily was prescribed, based on standard dosing for off label treatment of vascular anomalies (such as vascular malformation). This dose regimen led to supratherapeutic levels of >30 ng/mL in patients with KHE, which in turn led to serious AEs or even were fatal. 29 This may be a result of the slow clearance of sirolimus secondary to the maturation of the CYP3A pathway in neonates with an almost absent expression of this enzymes after birth and slowly increases during the first year of life. 30 For this reason, neonates seem to require a lower dose of sirolimus to maintain a serum level that is therapeutic effective without supratherapeutic or even toxic levels. 31 Our case series represents a retrospective evaluation of 5 patients with KHE and KMP who were treated with low-dose sirolimus and prednisone for additional effect. Dosing of sirolimus was based on the hypothetical assumption of the enzymatic maturation of CYP3A4 in time that is needed for the metabolization of sirolimus. Developmental changes of enzymes in infants during first year of life have been described previously. 30 Uniform guidelines how to dose sirolimus in infants with KHE are lacking; however, based on the cases reported here, we postulate a treatment regimen based on the age of the infants that may be useful in clinical practice.

What is already known about this subject
• Kaposiform haemangioendothelioma (KHE) can lead to the life-threatening Kasabach-Merritt phenomenon.
• Sirolimus can be used to treat KHE, although serious toxicities have been described in (very) young infants.
• The CYP3A4 enzyme is nearly absent in neonates and matures in the first months of life, reaching adult levels at 1 year.

What this study adds
• Sirolimus treatment of infants with life-threatening KHE using dosing and titration based on the hypothetical maturation of liver enzymes can be safe and effective.
• Pharmacological insight in treatment of KHE in neonates using sirolimus leading to a safer administration without hampering efficacy of treatment.

| METHODS
A retrospective case review was performed with review of the medical report of patients with kaposiform haemangioendothelioma complicated with KMP, treated with low-dose sirolimus and corticosteroids between January 2019 and June 2021. Informed consent was obtained from all patients' parents reviewing of their child's medical report. The treatment efficacy of sirolimus was rated by the coagulation profile, platelet count, and relevant reduction (clinical or magnetic resonance imaging [MRI]) of the size of the KHE after follow-up >3 months. A complete response was defined as a platelet count >100 Â 10 9 /L, a normalization of the coagulation profile, and a size reduction more than 80% was seen. A partial response was defined as a platelet count of 40-100 Â 10 9 /L, an abnormal/normal coagulation profile, and size reduction between 30 and 80%. There was no response if the platelet count was <40 Â 10 9 /L, the coagulation profile was abnormal, and size reduction was reduced <30% or increased (adjusted criteria based on Wang et al. 12 ). Adverse events were assessed according to the Common Terminology Criteria for Adverse Events version 4.0.

| Treatment regime
The used local protocol was based on the estimated expression of CYP3A4 in the liver depending on the age of the infant and closely therapeutic drug monitoring (TDM). 30,32 This protocol is described below. We used low target trough levels of sirolimus under 6 ng/mL, as even with a very low dose of sirolimus 0.2 mg/m 2 every 48 hours sirolimus levels above 10 ng/mL were seen in the literature.

| Sirolimus TDM and titration
We performed, when the child was older than 1 week, a TDM at 24 hours after the second gift to prevent supratherapeutic levels at start of sirolimus. Thereafter, weekly TDM was performed to achieve the steady state concentration and maintain the target trough levels.
In case sirolimus level was <2 ng/mL at age >4 weeks, dose frequency was increased to 1 dose per 24 hours. If sirolimus levels were above 6/ng/mL the dosing interval was extended to 1 dose per 72 or 96 hours.
We advise to use the propylene glycol-free liquid soluble capsules in children younger than 6 months, since the oral solution of sirolimus contains propylene glycol, which can be associated with severe neurotoxicities in neonates. [33][34][35][36][37] Addition of prednisone 1 mg/kg/d in 2 doses to sirolimus for supplementary effect in patients younger than 3 months is recommended. For pneumocystis prophylaxis we started co-trimoxazole according to guidelines.

| Nomenclature of targets and ligands
Key protein targets and ligands in this article are hyperlinked to corresponding entries in https://www.guidetopharmacology.org, and are permanently archived in the Concise Guide to PHARMACOLOGY 2019/2020. 68

| RESULTS
In total, 5 patients were treated in the acute phase at the Radboud University Medical centre and 1 patient continued treatment in a later phase at the University Medical Centre Groningen. All patients were diagnosed with a kaposiform haemangioendothelioma with KMP at age between 4 days and 6 months. Patient characteristics are summarized in Table 1  and clinical size reduction-was seen. The follow-up of 1 patient (patient 5) was 4.13 months at the time of this report and D-dimers highly decreased; however, they were not normalized yet, resulting in a partial response (Table 3).
Overall clinical response, consisting of relevant decrease of the size or discoloration of the vascular tumour, was seen at median of 14 days (IQR 2.50-29.50).
Two patients did not experience any AEs, while 1 patient developed multiple mild-moderate AEs. In total, 19 possible or probable related to sirolimus AEs occurred (   Figure 3 shows the significant effect of low-dose sirolimus without any sirolimus related side effects.

| Patient 4
A term male infant was born with a tumour of his scrotum, right buttock and lower extremity (Figure 3). The MRI ( Figure A1)  after cessation of his intravenously antibiotics co-trimoxazole was restarted. This patient experienced elevated liver enzymes and alkalic phosphatase, which may be possibly caused by sirolimus or by total parenteral feeding, which may also lead to these elevations. At age

| DISCUSSION
This case series of infants with KHE with life-threatening KMP shows that treatment with sirolimus using dosing and titration based on the hypothetical maturation of liver enzymes can be safe and effective.
Sirolimus is mainly metabolized by the CYP3A4 enzyme. A significant smaller part is metabolized by the CYP3A5 and/or by the CYP2C8 enzyme. 38 Based on age, body weight and estimated expression of the CYP3A enzymes, treatment was initiated. Intensive therapeutic drug monitoring was used to adjust sirolimus dose either by increasing or decreasing the interval (see methods). All patients in our case series had a quick positive response using low-dose sirolimus corresponding to findings in literature. 13,20,39 Table 4 shows that only 17 articles   To prevent toxic blood levels at initiation of sirolimus therapy it is important that the initial dose is as low as possible. A high initial sirolimus blood level is seen in several studies using high initial doses (0.14-0.8 mg/m 2 /d), leading to high supratherapeutic or even toxic sirolimus target levels >20 ng/mL. 12,45 Another retrospective study described a Chinese boy aged 6 months. He suffered of a multifocal KHE of his right shoulder, chest, lung, right axilla and was treated with an initial sirolimus dose of 1.6 mg/m 2 /24 h in 2 doses. 12 Despite his positive response to sirolimus, he died of complications probably due to sirolimus toxicities.
Interestingly, this was the only patient described in this study who had initial sirolimus range above the normal target range (4-20 ng/ mL), namely 30 ng/mL.
It is possible that there is an inter-racial variability of the activity of the CYP3A4 and CYP3A5 enzyme, leading to higher sirolimus levels, which might result in more severe adverse events. It has been described that in Asian patients the genetic variants of CYP3A4/A5 (CYP3A4*1G, CYP3A5*1/*1 and *1/*3) are frequently found and associated with the need of higher dose of sirolimus to reach the therapeutic levels, [46][47][48] in contrast to patients with CYP3A5 *3/*3 genotypes frequently found in Caucasian patients, who may are more at risk of high sirolimus level. 49 In our case series, patient 5 needed a dose elevation in a short time. Since the patient has an Italic/Asian ethnicity it is possible that he had a functional variant from the CYP3A5 gene, which may lead to a higher quantity of this CYP3A5 enzymes. 50,51 However, there are contradictive reports, suggesting that there is no difference between T A B L E 3 Overview of efficacy and safety per patient treated with low-dose sirolimus and corticosteroids Asian and Caucasian races. 49,52,53 Further research is necessary for investigation of the interracial differences and their pharmacogenetic role of the CYP3 enzymes in infants. In general, we have to realize that each patient is different with perhaps different genetic backgrounds. Therefore, we can only recommend a low starting dose and that sirolimus dosage has to be adapted based on TDM.
In addition to the mentioned PK aspects, frequent TDM is needed when prednisone is added to sirolimus treatment. Since corticosteroids are competitive inhibitors of CYP3A4, they may induce the CYP3A4 enzyme. 54,55 This might affect sirolimus PK leading to lower sirolimus levels. 56 By contrast, sirolimus may affect the PK of prednisone, leading to a decrease of maximum plasma concentration found in kidney transplant recipients. No significant effect was seen in doses sirolimus of ≤5 mg/m 2 /d. 57 More research is necessary to analyse the exact interaction of sirolimus and corticosteroids. In summary, our case series showed that even with very low target trough levels of 2-6 ng/mL sirolimus is still effective in patients with KHE and KMP. Low-dose sirolimus may decrease the need for platelet transfusions and can reduce the risk of morbidity and mortality, accompanied with a lower incidence of AEs. The overall response of our case series is comparable to other used target levels, while no grade III AEs were seen in contrast to the patients described previously.

Management of KHE with KMP is difficult and challenging in infants
and especially in neonates, as developmental changes in the metabolization, pharmacogenetics and clearance of drugs play an important role. Treatment dosing and titration can be based on the developing metabolism in infants and other processes such as absorption, distribution and clearance. By frequent TDM, supratherapeutic levels of sirolimus can be avoided whereas the desired effect of treatment is reached. Our case series offers a proposal for treatment dose regimen covering the neonatal period and young infancy.

ACKNOWLEDGEMENT
No funding was secured for this study.

ETHICS APPROVAL
All caregivers of the patients included in this case series gave their informed consent for describing and images of their children.

COMPETING INTERESTS
All authors have no conflicts of interest to disclose.

AUTHOR CONTRIBUTIONS
Dr Harbers reviewed the cases, drafted the initial manuscript, All authors approved the final manuscript as submitted and agree to be accountable for all aspects of the work.

DATA AVAILABILITY STATEMENT
The data that support the findings of this study are available from the corresponding author upon reasonable request.