Plasma cell dependence on histone/protein deacetylase 11 reveals a therapeutic target in multiple myeloma.

The clinical utility of histone/protein deacetylase (HDAC) inhibitors in combinatorial regimens with proteasome inhibitors for patients with relapsed and refractory multiple myeloma (MM) is often limited by excessive toxicity due to HDAC inhibitor promiscuity with multiple HDACs. Therefore, more selective inhibition minimizing off-target toxicity may increase the clinical effectiveness of HDAC inhibitors. We demonstrated that plasma cell development and survival are dependent upon HDAC11, suggesting this enzyme is a promising therapeutic target in MM. Mice lacking HDAC11 exhibited markedly decreased plasma cell numbers. Accordingly, in vitro plasma cell differentiation was arrested in B cells lacking functional HDAC11. Mechanistically, we showed that HDAC11 is involved in the deacetylation of IRF4 at lysine103. Further, targeting HDAC11 led to IRF4 hyperacetylation, resulting in impaired IRF4 nuclear localization and target promoter binding. Importantly, transient HDAC11 knockdown or treatment with elevenostat, an HDAC11-selective inhibitor, induced cell death in MM cell lines. Elevenostat produced similar anti-MM activity in vivo, improving survival among mice inoculated with 5TGM1 MM cells. Elevenostat demonstrated nanomolar ex vivo activity in 34 MM patient specimens and synergistic activity when combined with bortezomib. Collectively, our data indicated that HDAC11 regulates an essential pathway in plasma cell biology establishing its potential as an emerging theraputic vulnerability in MM.

Essential role for histone deacetylase 11 (HDAC11) in neutrophil biology.

Epigenetic changes in chromatin structure have been recently associated with the deregulated expression of critical genes in normal and malignant processes. HDAC11, the newest member of the HDAC family of enzymes, functions as a negative regulator of IL-10 expression in APCs, as previously described by our lab. However, at the present time, its role in other hematopoietic cells, specifically in neutrophils, has not been fully explored. In this report, for the first time, we present a novel physiologic role for HDAC11 as a multifaceted regulator of neutrophils. Thus far, we have been able to demonstrate a lineage-restricted overexpression of HDAC11 in neutrophils and committed neutrophil precursors (promyelocytes). Additionally, we show that HDAC11 appears to associate with the transcription machinery, possibly regulating the expression of inflammatory and migratory genes in neutrophils. Given the prevalence of neutrophils in the peripheral circulation and their central role in the first line of defense, our results highlight a unique and novel role for HDAC11. With the consideration of the emergence of new, selective HDAC11 inhibitors, we believe that our findings will have significant implications in a wide range of diseases spanning malignancies, autoimmunity, and inflammation.

An Ex Vivo Platform for the Prediction of Clinical Response in Multiple Myeloma.

Multiple myeloma remains treatable but incurable. Despite a growing armamentarium of effective agents, choice of therapy, especially in relapse, still relies almost exclusively on clinical acumen. We have developed a system, Ex vivo Mathematical Myeloma Advisor (EMMA), consisting of patient-specific mathematical models parameterized by an ex vivo assay that reverse engineers the intensity and heterogeneity of chemosensitivity of primary cells from multiple myeloma patients, allowing us to predict clinical response to up to 31 drugs within 5 days after bone marrow biopsy. From a cohort of 52 multiple myeloma patients, EMMA correctly classified 96% as responders/nonresponders and correctly classified 79% according to International Myeloma Working Group stratification of level of response. We also observed a significant correlation between predicted and actual tumor burden measurements (Pearson r = 0.5658, P < 0.0001). Preliminary estimates indicate that, among the patients enrolled in this study, 60% were treated with at least one ineffective agent from their therapy combination regimen, whereas 30% would have responded better if treated with another available drug or combination. Two in silico clinical trials with experimental agents ricolinostat and venetoclax, in a cohort of 19 multiple myeloma patient samples, yielded consistent results with recent phase I/II trials, suggesting that EMMA is a feasible platform for estimating clinical efficacy of drugs and inclusion criteria screening. This unique platform, specifically designed to predict therapeutic response in multiple myeloma patients within a clinically actionable time frame, has shown high predictive accuracy in patients treated with combinations of different classes of drugs. The accuracy, reproducibility, short turnaround time, and high-throughput potential of this platform demonstrate EMMA's promise as a decision support system for therapeutic management of multiple myeloma. Cancer Res; 77(12); 3336-51. ©2017 AACR.

Unification of de novo and acquired ibrutinib resistance in mantle cell lymphoma.

The novel Bruton's tyrosine kinase inhibitor ibrutinib has demonstrated high response rates in B-cell lymphomas; however, a growing number of ibrutinib-treated patients relapse with resistance and fulminant progression. Using chemical proteomics and an organotypic cell-based drug screening assay, we determine the functional role of the tumour microenvironment (TME) in ibrutinib activity and acquired ibrutinib resistance. We demonstrate that MCL cells develop ibrutinib resistance through evolutionary processes driven by dynamic feedback between MCL cells and TME, leading to kinome adaptive reprogramming, bypassing the effect of ibrutinib and reciprocal activation of PI3K-AKT-mTOR and integrin-ß1 signalling. Combinatorial disruption of B-cell receptor signalling and PI3K-AKT-mTOR axis leads to release of MCL cells from TME, reversal of drug resistance and enhanced anti-MCL activity in MCL patient samples and patient-derived xenograft models. This study unifies TME-mediated de novo and acquired drug resistance mechanisms and provides a novel combination therapeutic strategy against MCL and other B-cell malignancies.

An Organotypic High Throughput System for Characterization of Drug Sensitivity of Primary Multiple Myeloma Cells.

In this work we describe a novel approach that combines ex vivo drug sensitivity assays and digital image analysis to estimate chemosensitivity and heterogeneity of patient-derived multiple myeloma (MM) cells. This approach consists in seeding primary MM cells freshly extracted from bone marrow aspirates into microfluidic chambers implemented in multi-well plates, each consisting of a reconstruction of the bone marrow microenvironment, including extracellular matrix (collagen or basement membrane matrix) and stroma (patient-derived mesenchymal stem cells) or human-derived endothelial cells (HUVECs). The chambers are drugged with different agents and concentrations, and are imaged sequentially for 96 hr through bright field microscopy, in a motorized microscope equipped with a digital camera. Digital image analysis software detects live and dead cells from presence or absence of membrane motion, and generates curves of change in viability as a function of drug concentration and exposure time. We use a computational model to determine the parameters of chemosensitivity of the tumor population to each drug, as well as the number of sub-populations present as a measure of tumor heterogeneity. These patient-tailored models can then be used to simulate therapeutic regimens and estimate clinical response.

Histone deacetylase 11: A novel epigenetic regulator of myeloid derived suppressor cell expansion and function.

Myeloid-derived suppressor cells (MDSCs), a heterogeneous population of cells capable of suppressing anti-tumor T cell function in the tumor microenvironment, represent an imposing obstacle in the development of cancer immunotherapeutics. Thus, identifying elements essential to the development and perpetuation of these cells will undoubtedly improve our ability to circumvent their suppressive impact. HDAC11 has emerged as a key regulator of IL-10 gene expression in myeloid cells, suggesting that this may represent an important targetable axis through which to dampen MDSC formation. Using a murine transgenic reporter model system where eGFP expression is controlled by the HDAC11 promoter (Tg-HDAC11-eGFP), we provide evidence that HDAC11 appears to function as a negative regulator of MDSC expansion/function in vivo. MDSCs isolated from EL4 tumor-bearing Tg-HDAC11-eGFP display high expression of eGFP, indicative of HDAC11 transcriptional activation at steady state. In striking contrast, immature myeloid cells in tumor-bearing mice display a diminished eGFP expression, implying that the transition of IMC to MDSC's require a decrease in the expression of HDAC11, where we postulate that it acts as a gate-keeper of myeloid differentiation. Indeed, tumor-bearing HDAC11-knockout mice (HDAC11-KO) demonstrate a more suppressive MDSC population as compared to wild-type (WT) tumor-bearing control. Notably, the HDAC11-KO tumor-bearing mice exhibit enhanced tumor growth kinetics when compare to the WT control mice. Thus, through a better understanding of this previously unknown role of HDAC11 in MDSC expansion and function, rational development of targeted epigenetic modifiers may allow us to thwart a powerful barrier to efficacious immunotherapies.