Exploring preferences of different modes of administration of hypomethylating agent treatments among patients with acute myeloid leukemia.

Introduction: About half of patients with Acute Myeloid Leukemia (AML) are not eligible for Standard Induction Chemotherapy (SIC). Hypomethylating Agents (HMAs) intravenously (IV) or subcutaneously (SC) in a clinical setting are typically offered as an alternative. However, injectable HMAs may be burdensome for patients given the frequent hospital visits and side effects. This study explored patient treatment preferences for different modes of administration (MOA) and the relative importance of treatment-related characteristics that influence treatment decisions. Methods: Semi-structured 1:1 interviews were conducted with 21 adult patients with AML in Germany, the United Kingdom, and Spain, who are not eligible for SIC, had experience with HMAs or were scheduled to be treated with HMAs. After discussing their experience of living with AML and its treatments, patients were presented with hypothetical treatment scenarios to explore their preferences, and a ranking exercise to assess the relative importance of treatment characteristics that influence their treatment-decisions for AML. Results: Most patients reported an overall preference for oral administration over parenteral routes (71%), mostly due to convenience. Those preferring IV or SC routes (24%) reasoned with faster speed of action and onsite monitoring. When presented with a hypothetical situation of a patient having to choose between two AML treatments that were identical except for their MOA, the majority preferred the oral route (76%). Regarding treatment characteristics that influence treatment decisions, patients most frequently reported efficacy (86%) and side effects (62%) as important, followed by mode of administration (29%), daily life impacts (24%) and location of treatment (hospital versus home) (14%). However, only efficacy and side effects were rated as number one deciding factors (67% and 19%, respectively). Patients most frequently rated dosing regimen (33%) as least important. Conclusion: The insights gained from this study may help support patients with AML who are receiving HMA treatment instead of SIC. A potential oral HMA with similar efficacy and tolerability profiles to injectable HMAs could influence treatment decisions. Furthermore, an oral HMA treatment might decrease the burden of parenteral therapies and improve patients' overall quality of life. However, the extent of influence MOA has on treatment decisions requires further investigation.

DNA methylome combined with chromosome cluster-oriented analysis provides an early signature for cutaneous melanoma aggressiveness.

Aberrant DNA methylation is a well-known feature of tumours and has been associated with metastatic melanoma. However, since melanoma cells are highly heterogeneous, it has been challenging to use affected genes to predict tumour aggressiveness, metastatic evolution, and patients' outcomes. We hypothesized that common aggressive hypermethylation signatures should emerge early in tumorigenesis and should be shared in aggressive cells, independent of the physiological context under which this trait arises. We compared paired melanoma cell lines with the following properties: (i) each pair comprises one aggressive counterpart and its parental cell line and (ii) the aggressive cell lines were each obtained from different host and their environment (human, rat, and mouse), though starting from the same parent cell line. Next, we developed a multi-step genomic pipeline that combines the DNA methylome profile with a chromosome cluster-oriented analysis. A total of 229 differentially hypermethylated genes was commonly found in the aggressive cell lines. Genome localization analysis revealed hypermethylation peaks and clusters, identifying eight hypermethylated gene promoters for validation in tissues from melanoma patients. Five Cytosine-phosphate-Guanine (CpGs) identified in primary melanoma tissues were transformed into a DNA methylation score that can predict survival (log-rank test, p=0.0008). This strategy is potentially universally applicable to other diseases involving DNA methylation alterations.

Demethylation by low-dose 5-aza-2'-deoxycytidine impairs 3D melanoma invasion partially through miR-199a-3p expression revealing the role of this miR in melanoma.

BACKGROUND: Efficient treatments against metastatic melanoma dissemination are still lacking. Here, we report that low-cytotoxic concentrations of 5-aza-2'-deoxycytidine, a DNA demethylating agent, prevent in vitro 3D invasiveness of metastatic melanoma cells and reduce lung metastasis formation in vivo. RESULTS: We unravelled that this beneficial effect is in part due to MIR-199A2 re-expression by promoter demethylation. Alone, this miR showed an anti-invasive and anti-metastatic effect. Throughout integration of micro-RNA target prediction databases with transcriptomic analysis after 5-aza-2'-deoxycytidine treatments, we found that miR-199a-3p downregulates set of genes significantly involved in invasion/migration processes. In addition, analysis of data from melanoma patients showed a stage- and tissue type-dependent modulation of MIR-199A2 expression by DNA methylation. CONCLUSIONS: Thus, our data suggest that epigenetic- and/or miR-based therapeutic strategies can be relevant to limit metastatic dissemination of melanoma.

Integrated Pharmacokinetic/Pharmacodynamic Model of a Bispecific CD3xCD123 DART Molecule in Nonhuman Primates: Evaluation of Activity and Impact of Immunogenicity.

Purpose: Flotetuzumab (MGD006 or S80880) is a bispecific molecule that recognizes CD3 and CD123 membrane proteins, redirecting T cells to kill CD123-expressing cells for the treatment of acute myeloid leukemia. In this study, we developed a mathematical model to characterize MGD006 exposure-response relationships and to assess the impact of its immunogenicity in cynomolgus monkeys.Experimental Design: Thirty-two animals received multiple escalating doses (100-300-600-1,000 ng/kg/day) via intravenous infusion continuously 4 days a week. The model reflects sequential binding of MGD006 to CD3 and CD123 receptors. Formation of the MGD006/CD3 complex was connected to total T cells undergoing trafficking, whereas the formation of the trimolecular complex results in T-cell activation and clonal expansion. Activated T cells were used to drive the peripheral depletion of CD123-positive cells. Anti-drug antibody development was linked to MGD006 disposition as an elimination pathway. Model validation was tested by predicting the activity of MGD006 in eight monkeys receiving continuous 7-day infusions.Results: MGD006 disposition and total T-cell and CD123-positive cell profiles were well characterized. Anti-drug antibody development led to the suppression of T-cell trafficking but did not systematically abolish CD123-positive cell depletion. Target cell depletion could persist after drug elimination owing to the self-proliferation of activated T cells generated during the first cycles. The model was externally validated with the 7-day infusion dosing schedule.Conclusions: A translational model was developed for MGD006 that features T-cell activation and expansion as a key driver of pharmacologic activity and provides a mechanistic quantitative platform to inform dosing strategies in ongoing clinical studies. Clin Cancer Res; 24(11); 2631-41. ©2018 AACR.

First-in-human phase I study of oral S49076, a unique MET/AXL/FGFR inhibitor, in advanced solid tumours.

BACKGROUND AND OBJECTIVES: S49076 is a novel ATP-competitive tyrosine kinase inhibitor of MET, AXL and FGFR with a unique selectivity profile. A phase I open-label study was undertaken to establish the tolerability profile and determine the recommended dose (RD) and administration schedule. MATERIALS AND METHODS: Patients with advanced solid tumours received S49076 orally once-daily (qd) or twice-daily (bid) in continuous 21-day cycles at escalating doses guided by a 3 + 3 design and followed by an expansion phase at the RD. Pharmacokinetic (PK) parameters were assessed and pharmacodynamic end-points were evaluated in pre- and post-treatment tumour biopsies. Preliminary anti-tumour activity was evaluated as per the Response Evaluation Criteria In Solid Tumours 1.1 criteria. RESULTS: A total of 103 patients were treated: 79 in the dose-escalation and 24 in the expansion. Doses from 15 to 900 mg were evaluated. Dose-limiting toxicities were reported in 9 patients and occurred at 30, 760 and 900 mg in the qd arm and at 180, 225 and 285 mg in the bid arm. The RD was defined at 600 mg qd. Adverse events (AEs) occurred with similar frequency in both regimens at an equivalent total daily dose. Overall, 83 patients (81.4%) had drug-related AEs, the majority (93%) of which were grade I-II (National Cancer Institute Common Terminology Criteria for Adverse Events version 4.0) and only 3% led to drug discontinuation. Intratumoural PK analysis at the RD suggested hitting of MET, AXL and FGFR. CONCLUSION: S49076 demonstrated a tolerable safety profile with limited single-agent activity. PK/pharmacodynamic readouts of S49076 are encouraging for further investigation of S49076 in combination therapies. TRIAL REGISTRATION NUMBER: ISRCTN00759419.

Direct and quantitative evaluation of the human CYP3A4 contribution (fm) to drug clearance using the in vitro SILENSOMES model.

1. Among the different in vitro studies recommended by the regulatory agencies, no gold-standard model can easily and directly measure the quantitative CYP450 contributions to drug biotransformation. In this article, we propose an original strategy, called SilensomesTM, to produce human liver microsomes silenced for one specific CYP450, thanks to specific mechanism-based inhibitors (MBI). 2. Using azamulin as a specific CYP3A4 MBI, we demonstrated the proof of concept that CYP3A4 can be totally, specifically (even against 3A5) and permanently (at least for six years) inhibited by our process. Thus, comparing clearance in control and CYP3A4-SilensomesTM, CYP3A4 contributions were determined for 11 CYP3A4 substrates which correlated with known in vivo contributions and revealed accuracy with less than 10% error. In comparison, contributions determined using recombinant human CYP450 (rhCYP450s) were less accurate (more than 10% error for 30% of the tested CYP3A4 substrates). 3. This easy and ready-to-use in vitro method combines the advantages of existing models (specificity of rhCYP450s and representativeness of HLM) without their drawbacks. The same strategy could be used to silence other major CYP450s one-by-one to provide a complete direct CYP450 quantitative phenotyping kit.

The c-Jun/RHOB/AKT pathway confers resistance of BRAF-mutant melanoma cells to MAPK inhibitors.

The response of BRAF-mutant melanoma patients to BRAF inhibitors is dramatically impaired by secondary resistances and rapid relapse. So far, the molecular mechanisms driving these resistances are not completely understood. Here, we show that, in BRAF-mutant melanoma cells, inhibition of BRAF or its target MEK induces RHOB expression by a mechanism that depends on the transcription factor c-Jun. In those cells, RHOB deficiency causes hypersensitivity to BRAF and MEK inhibitors-induced apoptosis. Supporting these results, loss of RHOB expression in metastatic melanoma tissues is associated with an increased progression-free survival of BRAF-mutant patients treated with vemurafenib. Following BRAF inhibition, RHOB activates AKT whose inhibition causes hypersensitivity of BRAF-mutant melanoma cells to BRAF inhibitors. In mice, AKT inhibition synergizes with vemurafenib to block tumor growth of BRAF-mutant metastatic melanoma. Our findings reveal that BRAF inhibition activates a c-Jun/RHOB/AKT pathway that promotes tumor cell survival and further support a role of this pathway in the resistance of melanoma to vemurafenib. Our data also highlight the importance of using RHOB tumor levels as a biomarker to predict vemurafenib patient's response and to select those that would benefit of the combination with AKT inhibitors.

Establishment and mitotic characterization of new Drosophila acentriolar cell lines from DSas-4 mutant.

In animal cells the centrosome is commonly viewed as the main cellular structure driving microtubule (MT) assembly into the mitotic spindle apparatus. However, additional pathways, such as those mediated by chromatin and augmin, are involved in the establishment of functional spindles. The molecular mechanisms involved in these pathways remain poorly understood, mostly due to limitations inherent to current experimental systems available. To overcome these limitations we have developed six new Drosophila cell lines derived from Drosophila homozygous mutants for DSas-4, a protein essential for centriole biogenesis. These cells lack detectable centrosomal structures, astral MT, with dispersed pericentriolar proteins D-PLP, Centrosomin and γ-tubulin. They show poorly focused spindle poles that reach the plasma membrane. Despite being compromised for functional centrosome, these cells could successfully undergo mitosis. Live-cell imaging analysis of acentriolar spindle assembly revealed that nascent MTs are nucleated from multiple points in the vicinity of chromosomes. These nascent MTs then grow away from kinetochores allowing the expansion of fibers that will be part of the future acentriolar spindle. MT repolymerization assays illustrate that acentriolar spindle assembly occurs "inside-out" from the chromosomes. Colchicine-mediated depolymerization of MTs further revealed the presence of a functional Spindle Assembly Checkpoint (SAC) in the acentriolar cells. Finally, pilot RNAi experiments open the potential use of these cell lines for the molecular dissection of anastral pathways in spindle and centrosome assembly.