The histone deacetylase HDAC11 regulates the expression of interleukin 10 and immune tolerance.

Antigen-presenting cells (APCs) induce T cell activation as well as T cell tolerance. The molecular basis of the regulation of this critical 'decision' is not well understood. Here we show that HDAC11, a member of the HDAC histone deacetylase family with no prior defined physiological function, negatively regulated expression of the gene encoding interleukin 10 (IL-10) in APCs. Overexpression of HDAC11 inhibited IL-10 expression and induced inflammatory APCs that were able to prime naive T cells and restore the responsiveness of tolerant CD4+ T cells. Conversely, disruption of HDAC11 in APCs led to upregulation of expression of the gene encoding IL-10 and impairment of antigen-specific T cell responses. Thus, HDAC11 represents a molecular target that influences immune activation versus immune tolerance, a critical 'decision' with substantial implications in autoimmunity, transplantation and cancer immunotherapy.

T cells lacking HDAC11 have increased effector functions and mediate enhanced alloreactivity in a murine model.

Histone acetylation and the families of enzymes responsible for controlling these epigenetic marks have been implicated in regulating T-cell maturation and phenotype. Here, we demonstrate a previously undefined role of histone deacetylase 11 (HDAC11) in regulating T-cell effector functions. Using EGFP-HDAC11 transgenic reporter mice, we found that HDAC11 expression was lower in effector relative to naive and central memory T-cell populations, and activation of resting T cells resulted in its decreased expression. Experiments using HDAC11 knockout (KO) mice revealed that T cells from these mice displayed enhanced proliferation, proinflammatory cytokine production, and effector molecule expression. In addition, HDAC11KO T cells had increased expression of Eomesodermin (Eomes) and TBX21 (Tbet), transcription factors previously shown to regulate inflammatory cytokine and effector molecule production. Conversely, overexpression of HDAC11 resulted in decreased expression of these genes. Chromatin immunoprecipitation showed the presence of HDAC11 at the Eomes and Tbet gene promoters in resting T cells, where it rapidly disassociated following T-cell activation. In vivo, HDAC11KO T cells were refractory to tolerance induction. HDAC11KO T cells also mediated accelerated onset of acute graft-versus-host disease (GVHD) in a murine model, characterized by increased proliferation of T cells and expression of interferon-γ, tumor necrosis factor, and EOMES. In addition, adoptive transfer of HDAC11KO T cells resulted in significantly reduced tumor burden in a murine B-cell lymphoma model. Taken together, these data demonstrate a previously unknown role of HDAC11 as a negative epigenetic regulator of T-cell effector phenotype and function.

A novel role for histone deacetylase 6 in the regulation of the tolerogenic STAT3/IL-10 pathway in APCs.

APCs are critical in T cell activation and in the induction of T cell tolerance. Epigenetic modifications of specific genes in the APC play a key role in this process, and among them histone deacetylases (HDACs) have emerged as key participants. HDAC6, one of the members of this family of enzymes, has been shown to be involved in regulation of inflammatory and immune responses. In this study, to our knowledge we show for the first time that genetic or pharmacologic disruption of HDAC6 in macrophages and dendritic cells results in diminished production of the immunosuppressive cytokine IL-10 and induction of inflammatory APCs that effectively activate Ag-specific naive T cells and restore the responsiveness of anergic CD4(+) T cells. Mechanistically, we have found that HDAC6 forms a previously unknown molecular complex with STAT3, association that was detected in both the cytoplasmic and nuclear compartments of the APC. By using HDAC6 recombinant mutants we identified the domain comprising amino acids 503-840 as being required for HDAC6 interaction with STAT3. Furthermore, by re-chromatin immunoprecipitation we confirmed that HDAC6 and STAT3 are both recruited to the same DNA sequence within the Il10 gene promoter. Of note, disruption of this complex by knocking down HDAC6 resulted in decreased STAT3 phosphorylation--but no changes in STAT3 acetylation--as well as diminished recruitment of STAT3 to the Il10 gene promoter region. The additional demonstration that a selective HDAC6 inhibitor disrupts this STAT3/IL-10 tolerogenic axis points to HDAC6 as a novel molecular target in APCs to overcome immune tolerance and tips the balance toward T cell immunity.

Histone deacetylase inhibitor LAQ824 augments inflammatory responses in macrophages through transcriptional regulation of IL-10.

APCs are important in the initiation of productive Ag-specific T cell responses and the induction of T cell anergy. The inflammatory status of the APC at the time of encounter with Ag-specific T cells plays a central role in determining such divergent T cell outcomes. A better understanding of the regulation of proinflammatory and anti-inflammatory genes in its natural setting, the chromatin substrate, might provide novel insights to overcome anergic mechanisms mediated by APCs. In this study, we show for the first time, to our knowledge, that treatment of BALB/c murine macrophages with the histone deacetylase inhibitor LAQ824 induces chromatin changes at the level of the IL-10 gene promoter that lead to enhanced recruitment of the transcriptional repressors HDAC11 and PU.1. Such an effect is associated with diminished IL-10 production and induction of inflammatory cells able of priming naive Ag-specific T cells, but more importantly, capable of restoring the responsiveness of anergized Ag-specific CD4(+) T cells.

HDAC11 regulates expression of C/EBPß and immunosuppressive molecules in myeloid-derived suppressor cells.

Myeloid-derived suppressor cells (MDSCs) constitute a heterogeneous population of immature myeloid cells derived from bone marrow and negatively regulate both innate and adaptive immunity in the tumor microenvironment. Previously we have demonstrated that MDSCs lacking histone deacetylase 11 (HDAC11) displayed an increased suppressive activity against CD8+ T-cells. However, the mechanisms of HDAC11 that contribute to the suppressive function of MDSCs remain unclear. Here, we show that arginase activity and NO production is significantly higher in HDAC11 knockout MDSCs when compared with wild-type (WT) controls. In the absence of HDAC11, elevated arginase level and enzymatic activity were observed preferentially in the tumor-infiltrated granulocytic MDSCs, whereas iNOS expression and NO production were increased in the tumor-infiltrated monocytic MDSCs. Of note and for the first time, we demonstrated an association between the elevated expression of immunosuppressive molecules with up-regulation of the transcription factor C/EBPß in MDSCs lacking HDAC11. Interestingly, the highest expression of C/EBPß was observed among CD11b+ Gr-1+ MDSCs isolated from tumor-bearing mice. The additional demonstration that HDAC11 is recruited to the promoter region of C/EBPß in WT MDSCs suggests a novel molecular mechanism by which HDAC11 influence the expression of immunosuppressive molecules in MDSCs through regulation of C/EBPß gene expression.

Selective HDAC6 inhibitors improve anti-PD-1 immune checkpoint blockade therapy by decreasing the anti-inflammatory phenotype of macrophages and down-regulation of immunosuppressive proteins in tumor cells.

Histone deacetylases (HDACs) are involved in diverse cellular regulatory mechanisms including non-canonical functions outside the chromatin environment. Several publications have demonstrated that selective HDAC inhibitors (HDACi) can influence tumor immunogenicity and the functional activity of specific immune cells. In particular, the selective inhibition of HDAC6 has been reported to decrease tumor growth in several malignancies. However, there is still no clarity about the cellular components mediating this effect. In this study, we evaluated the HDAC6i Nexturastat A as a priming agent to facilitate the transition of the tumor microenvironment from "cold" to "hot", and potentially augment immune check-point blockade therapies. This combination modality demonstrated to significantly reduce tumor growth in syngeneic melanoma tumor models. Additionally, we observed a complete neutralization of the up-regulation of PD-L1 and other immunosuppressive pathways induced by the treatment with anti-PD-1 blockade. This combination also showed profound changes in the tumor microenvironment such as enhanced infiltration of immune cells, increased central and effector T cell memory, and a significant reduction of pro-tumorigenic M2 macrophages. The evaluation of individual components of the tumor microenvironment suggested that the in vivo anti-tumor activity of HDAC6i is mediated by its effect on tumor cells and tumor-associated macrophages, and not directly over T cells. Overall, our results indicate that selective HDAC6i could be used as immunological priming agents to sensitize immunologically "cold" tumors and subsequently improve ongoing immune check-point blockade therapies.

Author Correction: Selective HDAC6 inhibitors improve anti-PD-1 immune checkpoint blockade therapy by decreasing the anti-inflammatory phenotype of macrophages and down-regulation of immunosuppressive proteins in tumor cells.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Secretome profiling identifies neuron-derived neurotrophic factor as a tumor-suppressive factor in lung cancer.

Clinical and preclinical studies show tissue-specific differences in tumorigenesis. Tissue specificity is controlled by differential gene expression. We prioritized genes that encode secreted proteins according to their preferential expression in normal lungs to identify candidates associated with lung cancer. Indeed, most of the lung-enriched genes identified in our analysis have known or suspected roles in lung cancer. We focused on the gene encoding neuron-derived neurotrophic factor (NDNF), which had not yet been associated with lung cancer. We determined that NDNF was preferentially expressed in the normal adult lung and that its expression was decreased in human lung adenocarcinoma and a mouse model of this cancer. Higher expression of NDNF was associated with better clinical outcome of patients with lung adenocarcinoma. Purified NDNF inhibited proliferation of lung cancer cells, whereas silencing NDNF promoted tumor cell growth in culture and in xenograft models. We determined that NDNF is downregulated through DNA hypermethylation near CpG island shores in human lung adenocarcinoma. Furthermore, the lung cancer-related DNA hypermethylation sites corresponded to the methylation sites that occurred in tissues with low NDNF expression. Thus, by analyzing the tissue-specific secretome, we identified a tumor-suppressive factor, NDNF, which is associated with patient outcomes in lung adenocarcinoma.

BCL2 Amplicon Loss and Transcriptional Remodeling Drives ABT-199 Resistance in B Cell Lymphoma Models.

Drug-tolerant "persister" tumor cells underlie emergence of drug-resistant clones and contribute to relapse and disease progression. Here we report that resistance to the BCL-2 targeting drug ABT-199 in models of mantle cell lymphoma and double-hit lymphoma evolves from outgrowth of persister clones displaying loss of 18q21 amplicons that harbor BCL2. Further, persister status is generated via adaptive super-enhancer remodeling that reprograms transcription and offers opportunities for overcoming ABT-199 resistance. Notably, pharmacoproteomic and pharmacogenomic screens revealed that persisters are vulnerable to inhibition of the transcriptional machinery and especially to inhibition of cyclin-dependent kinase 7 (CDK7), which is essential for the transcriptional reprogramming that drives and sustains ABT-199 resistance. Thus, transcription-targeting agents offer new approaches to disable drug resistance in B-cell lymphomas.

Loss of HDAC11 ameliorates clinical symptoms in a multiple sclerosis mouse model.

Multiple sclerosis (MS) is a chronic, immune-mediated, demyelinating disease of the central nervous system (CNS). There is no known cure for MS, and currently available drugs for managing this disease are only effective early on and have many adverse side effects. Results from recent studies suggest that histone deacetylase (HDAC) inhibitors may be useful for the treatment of autoimmune and inflammatory diseases such as MS. However, the underlying mechanisms by which HDACs influence immune-mediated diseases such as MS are unclear. More importantly, the question of which specific HDAC(s) are suitable drug targets for the potential treatment of MS remains unanswered. Here, we investigate the functional role of HDAC11 in experimental autoimmune encephalomyelitis, a mouse model for MS. Our results indicate that the loss of HDAC11 in KO mice significantly reduces clinical severity and demyelination of the spinal cord in the post-acute phase of experimental autoimmune encephalomyelitis. The absence of HDAC11 leads to reduced immune cell infiltration into the CNS and decreased monocytes and myeloid DCs in the chronic progressive phase of the disease. Mechanistically, HDAC11 controls the expression of the pro-inflammatory chemokine C-C motif ligand 2 (CCL2) gene by enabling the binding of PU.1 transcription factor to the CCL2 promoter. Our results reveal a novel pathophysiological function for HDAC11 in CNS demyelinating diseases, and warrant further investigations into the potential use of HDAC11-specific inhibitors for the treatment of chronic progressive MS.

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.

All-trans-retinoic acid eliminates immature myeloid cells from tumor-bearing mice and improves the effect of vaccination.

Tumor-induced immunosuppression is one of the crucial mechanisms of tumor evasion of immune surveillance. It contributes greatly to the failure of cancer vaccines. Immature myeloid cells (ImCs) play an important role in tumor-induced immunosuppression. These cells accumulate in large numbers in tumor-bearing hosts and directly inhibit T-cell functions via various mechanisms. In this study, we tried to eliminate ImCs in an attempt to improve antitumor response. In vivo administration of all-trans-retinoic acid (ATRA) dramatically reduced the presence of ImCs in all tested tumor models. This effect was not because of a direct antitumor effect of ATRA or decreased production of growth factors by tumor cells. Experiments with adoptive transfer demonstrated that ATRA differentiated ImC in vivo into mature dendritic cells, macrophages, and granulocytes. Decreased presence of ImC in tumor-bearing mice noticeably improved CD4- and CD8-mediated tumor-specific immune response. Combination of ATRA with two different types of cancer vaccines in two different tumor models significantly prolonged the antitumor effect of the treatment. These data suggest that elimination of ImC with ATRA may open an opportunity to improve the effect of cancer vaccines.

Extra-lymphatic solid tumor growth is not immunologically ignored and results in early induction of antigen-specific T-cell anergy: dominant role of cross-tolerance to tumor antigens.

A better understanding of how solid malignancies arise in an immunocompetent host, avoid immune recognition, and ultimately progress to widely disseminated cancer is essential to effectively harness the immune system against solid tumors. Because of their extra-lymphatic localization, it has been proposed that solid malignancies are just ignored by the immune system, thereby allowing their uncontrolled growth and dissemination. Alternatively, as most of the solid tumors are unable to express costimulatory molecules, the "signal one without signal two" model of tolerance induction has been frequently evoked to account for the failure of the immune system to reject antigenic tumors in vivo. In this study, we showed, however, that the extra-lymphatic growth of solid tumors is not immunologically ignored by the lymphoid compartment, resulting instead in the early induction of antigen-specific CD4(+) T-cell tolerance. Furthermore, analysis of parent-into-F1 bone marrow (BM) chimeras demonstrates that presentation of tumor antigens by BM-derived antigen-presenting cells represents the dominant mechanism in solid tumor-induced CD4(+) T-cell tolerance. Our findings of early development of antigen-specific T-cell unresponsiveness mediated by BM-derived antigen-presenting cells, not only provides a plausible explanation for the failure of the immune system to reject antigenic solid tumors in vivo, but more importantly, they have identified a barrier that, if appropriately manipulated, may lead to approaches to effectively harness the immune system against solid malignancies.

Effective combination of chemotherapy and dendritic cell administration for the treatment of advanced-stage experimental breast cancer.

PURPOSE: The goal of this study was to investigate the utility of a new approach to the treatment of advanced stage breast cancer, a combination of chemotherapy and dendritic cell (DC) administration. EXPERIMENTAL DESIGN: Mice bearing mammary adenocarcinoma expressing a model tumor antigen, influenza virus HA (DA3-HA), and parental tumor (DA3) were treated with different doses of paclitaxel with or without DCs. Paclitaxel was injected three times weekly, DCs were injected either i.v. or into tumor site (t.s.) 36 h after each injection of paclitaxel. Apoptosis was measured using Annexin V binding or terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) assays. CD8-mediated response of T cells to HA-derived peptide epitope was measured in an enzyme-linked immunospot (ELISPOT) assay. CD4-mediated response of T cells to HA-derived peptide was measured in proliferation assay. Nonspecific T-cell proliferation was measured in response to ConA and immobilized anti-CD3 and anti-CD28 antibodies. RESULTS: We have selected the dose of paclitaxel that induced a substantial level of apoptosis and moderate inhibition of T-cell function. Combined treatment resulted in the induction of HA-specific CD8-mediated response in all nine of the tested mice, and CD4-mediated responses in four of six treated mice. These effects were observed only if DCs were injected into tumor site, but not when injected i.v. No specific responses were found in mice treated with either chemotherapy or DCs alone. Injection of dexamethasone together with paclitaxel did not affect the induction of immune responses. Significant antitumor effect of combined treatment was observed in DA3-HA tumor-bearing mice as well as in mice bearing parental DA3 tumor. CONCLUSION: The combination of DC administration with repeated cycles of chemotherapy and dexamethasone (conditions similar to real clinical practice) resulted in the induction of antitumor response despite the immunosuppression induced by such treatment.

Combination therapy with AG-490 and interleukin 12 achieves greater antitumor effects than either agent alone.

Constitutive activation of Janus kinases (JAKs) and signal transducers and activators of transcription (STAT) occurs at very high frequency in various hematopoietic malignancies and solid tumors. It has been demonstrated that the tyrosine kinase inhibitor, AG-490, selectively blocks JAK activity and completely eliminates leukemia cells in a severe combined immunodeficient (SCID) mouse model. Because many cytokines, including interleukin (IL)-12, have been shown to signal through JAK/STAT pathways, AG-490 may inhibit cytokine-based cancer therapy. In this study, we evaluated the effects of AG-490 on IL-12 functional signaling and IL-12-mediated antitumor response in vivo. Previous studies have established the critical roles of macrophages and IFN-gamma in mediating IL-12-induced antitumor effects. Our results show that in vivo administration of AG-490 causes tumor cell apoptosis but does not inhibit IL-12-mediated macrophage activation and IFN-gamma production by lymphocytes. Furthermore, our data indicate that combined therapy with AG-490 and IL-12-induces greater antitumor effects than either agent alone in a murine myeloma tumor model. These results suggest that JAK/STAT inhibitors deserve further investigation for use with IL-12 therapy in treating human cancers with elevated JAK/STAT activity.

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.

Immune tolerance in breast cancer.

In spite of the demonstrated coexistence of tumor-associated rejection antigens expressed by breast tumors along with T cells that are capable of recognize them, breast cancers arises in immunocompetent hosts, outmaneuver immune recognition and ultimately progress to widely disseminated disease. In recent years, several explanations have been proposed to account for the inability of the immune system to recognize and reject antigenic breast tumors. Among them, the immune tolerance mechanisms have gained particular attention. Here, we discuss the increasing evidence pointing to tolerance towards tumor antigens as an important explanation for the failure of the immune system to reject breast tumors. A better understanding of the cellular and molecular mechanisms involved in breast tumor-induced antigen-specific T-cell tolerance may lead to approaches to effectively harness the immune system against this malignancy.

Divergent roles of histone deacetylase 6 (HDAC6) and histone deacetylase 11 (HDAC11) on the transcriptional regulation of IL10 in antigen presenting cells.

The anti-inflammatory cytokine IL-10 is a key modulator of immune responses. A better understanding of the regulation of this cytokine offers the possibility of tipping the balance of the immune response toward either tolerance, or enhanced immune responses. Histone deacetylases (HDACs) have been widely described as negative regulators of transcriptional regulation, and in this context, the primarily nuclear protein HDAC11 was shown to repress il-10 gene transcriptional activity in antigen-presenting cells (APCs). Here we report that another HDAC, HDAC6, primarily a cytoplasmic protein, associates with HDAC11 and modulates the expression of IL-10 as a transcriptional activator. To our knowledge, this is the first demonstration of two different HDACs being recruited to the same gene promoter to dictate divergent transcriptional responses. This dynamic interaction results in dynamic changes in the expression of IL-10 and might help to explain the intrinsic plasticity of the APC to determine T-cell activation versus T-cell tolerance.

The antimelanoma activity of the histone deacetylase inhibitor panobinostat (LBH589) is mediated by direct tumor cytotoxicity and increased tumor immunogenicity.

Melanoma is the deadliest skin cancer, and its incidence has been increasing faster than any other cancer. Although immunogenic, melanoma is not effectively cleared by host immunity. In this study, we investigate the therapeutic, antimelanoma potential of the histone deacetylase inhibitor (HDACi) panobinostat (LBH589) by assessing both its cytotoxic effects on melanoma cells as well as enhancement of immune recognition of melanoma. Utilizing murine and human melanoma cell lines, we analyzed the effects of LBH589 on proliferation and survival. In addition, we analyzed the expression of several immunologically relevant surface markers and melanoma differentiation antigens, and the ability of LBH589-treated melanoma to activate antigen-specific T cells. Finally, we assessed the in-vivo effects of LBH589 in a mouse melanoma model. Low nanomolar concentrations of LBH589 inhibit the growth of all melanoma cell lines tested, but not normal melanocytes. This inhibition is characterized by increased apoptosis as well as a G1 cell cycle arrest. In addition, LBH589 augments the expression of major histocompatibility complex and costimulatory molecules on melanoma cells leading to an increased ability to activate antigen-specific T cells. Treatment also increases expression of melanoma differentiation antigens. In vivo, LBH589 treatment of melanoma-bearing mice results in a significant increase in survival. However, in immunodeficient mice, the therapeutic effect of LBH589 is lost. Taken together, LBH589 exerts a dual effect upon melanoma cells by affecting not only growth/survival but also by increasing melanoma immunogenicity. These effects provide the framework for future evaluation of this HDAC inhibitor in melanoma treatment.

A microenvironment-mediated c-Myc/miR-548m/HDAC6 amplification loop in non-Hodgkin B cell lymphomas.

A dynamic interaction occurs between the lymphoma cell and its microenvironment, with each profoundly influencing the behavior of the other. Here, using a clonogenic coculture growth system and a xenograft mouse model, we demonstrated that adhesion of mantle cell lymphoma (MCL) and other non-Hodgkin lymphoma cells to lymphoma stromal cells confers drug resistance, clonogenicity, and induction of histone deacetylase 6 (HDAC6). Furthermore, stroma triggered a c-Myc/miR-548m feed-forward loop, linking sustained c-Myc activation, miR-548m downregulation, and subsequent HDAC6 upregulation and stroma-mediated cell survival and lymphoma progression in lymphoma cell lines, primary MCL and other B cell lymphoma cell lines. Treatment with an HDAC6-selective inhibitor alone or in synergy with a c-Myc inhibitor enhanced cell death, abolished cell adhesion­mediated drug resistance, and suppressed clonogenicity and lymphoma growth ex vivo and in vivo. Together, these data suggest that the lymphoma-stroma interaction in the lymphoma microenvironment directly impacts the biology of lymphoma through genetic and epigenetic regulation, with HDAC6 and c-Myc as potential therapeutic targets.