Dynamic BH3 Profiling Predicts for Clinical Response to Lenalidomide Plus Chemotherapy in Relapsed Acute Myeloid Leukemia
Authors: Garcia Jacqueline S , Bhatt Shruti , Fell Geoffrey , Blonquist Traci M. , Taw Jack , Iberri David , Letai Anthony , Medeiros Bruno C. 
Published: October 2019

Background: Early prediction of response to novel chemotherapy combinations for patients diagnosed with relapsed or refractory acute myeloid leukemia (R/R AML) may have clinical utility. We conducted a phase 1 clinical trial (NCT01904643) of lenalidomide (LEN) given prior to MEC (Mitoxantrone + Etoposide + Cytarabine) salvage chemotherapy (LEN+MEC) for patients with R/R AML. Although the clinical trial was terminated due to administrative reasons, we explored the utility of FACS-based BH3 and dynamic BH3 profiling (DBP) assays to predict for response. Standard BH3 profiling is a functional assay that uses synthetic peptides derived from the BH3 domains of pro-apoptotic BCL-2 family members to measure mitochondrial "priming" (a cell's readiness for apoptosis) (Ni Chonghaile, Science, 2011). DBP measures whether short term incubation with drug enhances priming (Montero J, Cell, 2015).

Methods: Primary objective was to assess the safety/toxicity profile (DLTs and MTD) of LEN+MEC. LEN was administered 5-7 days prior to MEC and dose was escalated using an mTPI design with a target toxicity level of 30%. Dose levels (DL) included LEN given at doses of 15mgx5d (DL1), 15mgx7d (DL2), 25 mgx5d (DL3), 25mgx7d (DL4), 50mgx5d (DL5), and 50 mgx7d (DL6). Dose-escalation decisions were made based on DLTs that occurred during induction chemotherapy. DLT was defined as persistent cytopenias in absence of disease, gr 3 or higher non-hematologic toxicity not due to underlying disease or intolerable gr 2 toxicities attributable to LEN. Blood/bone marrow samples were collected at pre-treatment, post LEN only, and post LEN+MEC therapy. Clinical response was defined per ELN guidelines. For correlative studies, standard BH3 and DBP assays were used to measure overall mitochondrial priming and individual anti-apoptotic protein dependencies. DBP was performed after overnight (~16h) treatment with LEN at various concentrations on pretreatment leukemic blasts. Delta priming, utilized as a readout, represents the change in mitochondrial outer membrane depolarization after LEN treatment as measured by cytochrome C release following exposure to BH3 peptides. Data analysis was performed blinded to clinical response. Receiver operating characteristic (ROC) curve analysis was used to determine the optimal cut-off level for respective clinical end points.

Results: 17 total patients with R/R AML received LEN+MEC. Median age was 55 years (range: 22-72) with 12% Fav, 47% Int and 41% Adv risk by ELN risk stratification. 60% were heavily pre-treated. DLTs were observed in DL2 (n=1, gr 4 sepsis due to bacteremia), DL3 (n=1, gr 5 septic shock), and DL5 (n=1, gr3 bacteremia). MTD was not yet reached due to early study termination (although patients were treated at all 6 dose levels). No GVHD was observed among post-transplant patients. Composite remission rate among the 17 patients (CR + PR) was 35% (5 CR and 1 PR). Of the 5 patients who achieved CR, 3 of them eventually progressed. Median PFS and OS for the entire cohort was 2.30 (90% CI 1.84-3.47) and 4.60 (90% CI 2.84-22.10) months, respectively.

To assess whether mitochondrial priming might correlate with clinical response, we first performed standard BH3 profiling with a series of peptides on 16 pretreatment bone marrow samples. Mitochondrial cytochrome c release in response to BIM-BH3 correlated weakly with clinical response. DBP on pretreatment myeloblasts in response to BIM (activator) or PUMA (sensitizer) peptides were able to stratify responders based on an increase in mitochondrial priming caused by ex vivo LEN treatment (P<0.01 and P<0.0005, respectively). Using ROC curve analysis, it was determined that BIM and PUMA peptides served as binary predictors of achievement of CR, with AUC 0.95 and 1.0, respectively.

Conclusions: LEN+MEC was tolerable and had modest efficacy in this small study. Correlative studies showed that leukemic blasts briefly treated ex vivo with LEN which then underwent DBP were predictive of in vivo response among a heterogeneous patient population to LEN+MEC treatment. This study is an example of how a powerful biomarker can potentially identify those who are more likely to respond to novel therapies to avoid undesirable toxicity in those who would otherwise have less chance to benefit. Further validation is required to confirm the utility of DBP as a predictive biomarker to targeted agents given with chemotherapy.

Disclosures

Garcia: Celgene: Consultancy. Letai:AbbVie: Consultancy, Other: Lab research report; Vivid Biosciences: Equity Ownership; Flash Therapeutics: Equity Ownership; AstraZeneca: Consultancy, Other: Lab research report; Novartis: Consultancy, Other: Lab research report. Medeiros:Genentech: Employment; Celgene: Consultancy, Research Funding.