Venetoclax-based therapy can induce responses in approximately 70% of older previously untreated patients with acute myeloid leukemia (AML). However, upfront resistance as well as relapse following initial response demonstrates the need for a deeper understanding of resistance mechanisms. In the present study, we report that responses to venetoclax + azacitidine in patients with AML correlate closely with developmental stage, where phenotypically primitive AML is sensitive, but monocytic AML is more resistant. Mechanistically, resistant monocytic AML has a distinct transcriptomic profi le, loses expression of venetoclax target BCL2, and relies on MCL1 to mediate oxidative phosphorylation and survival. This differential sensitivity drives a selective process in patients which favors the outgrowth of monocytic subpopulations at relapse. Based on these fi ndings, we conclude that resistance to venetoclax + azacitidine can arise due to biological properties intrinsic to monocytic differentiation. We propose that optimal AML therapies should be designed so as to independently target AML subclones that may arise at differing stages of pathogenesis. SIGNIFICANCE: Identifying characteristics of patients who respond poorly to venetoclax-based therapy and devising alternative therapeutic strategies for such patients are important topics in AML. We show that venetoclax resistance can arise due to intrinsic molecular/metabolic properties of monocytic AML cells and that such properties can potentially be targeted with alternative strategies.

: Suppression of apoptosis by expression of antiapoptotic BCL2 family members is a hallmark of acute myeloblastic leukemia (AML). Induced myeloid leukemia cell differentiation protein (MCL1), an antiapoptotic BCL2 family member, is commonly upregulated in AML cells and is often a primary mode of resistance to treatment with the BCL2 inhibitor venetoclax. Here, we describe VU661013, a novel, potent, selective MCL1 inhibitor that destabilizes BIM/MCL1 association, leads to apoptosis in AML, and is active in venetoclax-resistant cells and patient-derived xenografts. In addition, VU661013 was safely combined with venetoclax for synergy in murine models of AML. Importantly, BH3 profiling of patient samples and drug-sensitivity testing accurately predicted cellular responses to selective inhibitors of MCL1 or BCL2 and showed benefit of the combination. Taken together, these data suggest a strategy of rationally using BCL2 and MCL1 inhibitors in sequence or in combination in AML clinical trials. SIGNIFICANCE: Targeting antiapoptotic proteins in AML is a key therapeutic strategy, and MCL1 is a critical antiapoptotic oncoprotein. Armed with novel MCL1 inhibitors and the potent BCL2 inhibitor venetoclax, it may be possible to selectively induce apoptosis by combining or thoughtfully sequencing these inhibitors based on a rational evaluation of AML...

A20 is a NF-kappaB-dependent gene that has dual anti-inflammatory and antiapoptotic functions in endothelial cells (EC). The function of A20 in smooth muscle cells (SMC) is unknown. We demonstrate that A20 is induced in SMC in response to inflammatory stimuli and serves an anti-inflammatory function via blockade of NF-kappaB and NF-kappaB-dependent proteins ICAM-1 and MCP-1. A20 inhibits SMC proliferation via increased expression of cyclin-dependent kinase inhibitors p21waf1 and p27kip1. Surprisingly, A20 sensitizes SMC to cytokine- and Fas-mediated apoptosis through a novel NO-dependent mechanism. In vivo, adenoviral delivery of A20 to medial rat carotid artery SMC after balloon angioplasty prevents neointimal hyperplasia by blocking SMC proliferation and accelerating re-endothelialization, without causing apoptosis. However, expression of A20 in established neointimal lesions leads to their regression through increased apoptosis. This is the first demonstration that A20 exerts two levels of control of vascular remodeling and healing. A20 prevents neointimal hyperplasia through combined anti-inflammatory and antiproliferative functions in medial SMC. If SMC evade this first barrier and neointima is formed, A20 has a therapeutic potential by uniquely sensitizing neointimal SMC to apoptosis. A20-based therapies hold promise for the prevention and treatment of neointimal disease.

Key Points• Rapid growth and rising platelet counts result in a significant expansion of platelet mass during neonatal life.• The rise in platelet counts is mediated by a prolongation in the neonatal platelet lifespan.The fetal/neonatal hematopoietic system must generate enough blood cells to meet the demands of rapid growth. This unique challenge might underlie the high incidence of thrombocytopenia among preterm neonates. In this study, neonatal platelet production and turnover were investigated in newborn mice. Based on a combination of blood volume expansion and increasing platelet counts, the platelet mass increased sevenfold during the first 2 weeks of murine life, a time during which thrombopoiesis shifted from liver to bone marrow. Studies applying in vivo biotinylation and mathematical modeling showed that newborn and adult mice had similar platelet production rates, but neonatal platelets survived 1 day longer in circulation. This prolonged lifespan fully accounted for the rise in platelet counts observed during the second week of murine postnatal life. A study of pro-apoptotic and anti-apoptotic Bcl-2 family proteins showed that neonatal platelets had higher levels of the anti-apoptotic protein Bcl-2 and were more resistant to apoptosis induced by the Bcl-2/Bcl-xL inhibitor ABT-737 than adult platelets. However, genetic ablation or pharmacologic inhibition of Bcl-2 alone did not shorten neonatal platelet survival or reduce platelet counts in newborn mice, indicating the existence of redundant or alternative mechanisms mediating the prolonged lifespan of neonatal platelets. (Blood. 2014;123(22):3381-3389) IntroductionThrombocytopenia affects 18% to 35% of infants admitted to the Neonatal Intensive Care Unit (NICU).1,2 The incidence of thrombocytopenia is inversely proportional to the gestational age, reaching 70% among the most preterm of infants.3 The reasons underlying the predisposition of neonates to develop thrombocytopenia are unclear, but are likely related to the unique characteristics of fetal/neonatal megakaryocytes (MKs), which are regulated by a developmental stage-specific interplay of pathways, transcription factors, and microRNAs. [4][5][6] Although the molecular mechanisms accounting for the differences between fetal/neonatal and adult MKs are becoming increasingly clear, 7 our understanding of thrombopoiesis and platelet homeostasis during fetal/neonatal life has lagged behind.Platelets have a finite lifespan in the circulation, normally 7 to 10 days in humans and 3 to 5 days in mice. Studies have shown this lifespan to be underpinned by a Bcl-2 family-mediated program that initiates the apoptotic death and subsequent clearance of aged platelets from the bloodstream. 8,9 The Bcl-2 family includes the prosurvival proteins Bcl-2, Bcl-x L and Mcl-1, the pro-apoptotic members Bak and Bax, and the BH3-only proteins. The latter promote apoptosis in nucleated cells by directly activating Bak and Bax, and/or by inactivating the prosurvival family members. 10,11 In platelets, the critical p...