Javier Pinilla

Abstract 4055: Enhancing anti-PD-1 immune blockade in melanoma through selective inhibition of histone deacetylase 6

Histone deacetylases (HDACs), were originally described in a limited context as histone modifiers. New evidence has demonstrated that HDACs are also involved in a diverse range of cellular processes that are not restricted to the chromatin environment, such as the regulation of the cell cycle/apoptosis and, more recently, a modulator of immune response. However, much remains unknown about the mechanism of action of HDACs and their roles in the immune-biology of cancer. The non-specific nature of pan-HDAC inhibitors results in a narrow therapeutic window of use, limiting the dose and duration due to toxicity. Our group has focused in one specific HDAC, HDAC6, and shown that both the genetic abrogation and pharmacological inhibition of this HDAC modulates the expression of a variety of immune-regulatory proteins in the tumor microenvironment, including PD-L1, PD-L2, MHC class I, B7-H4 and TRAIL-R1. In particular, we have previously demonstrated that both pharmacological inhibition and/or genetic abrogation of HDAC6 plays a critical role in the immune check point blockade by down-regulating the expression of PD-L1 and other check-point modulators such as PD-L2, B7-H4, etc. Moreover, we have also observed that in vivo inhibition of HDAC6 reduces tumor growth in B16 and SM1 murine melanoma models within syngeneic immunocompetent hosts. Additionally, we have found that the combination of low doses of the HDAC6i Nexturastat A and checkpoint immune blockade, including anti-PD-1 and anti-CTLA4, results in an important improvement in anti-tumor immune responses as evidenced by the reduction of tumor growth when compared to treatment with individual stand-alone agents. In these studies we also evidenced an increased production of IFNγ and IL-2 in the stand-alone check-point inhibitor treatments, which leads to an upregulation of PD-L1 and PD-L2. Similar levels of IFNγ and IL-2 were found in the combination groups. However, the expression level of PD-L1 and PD-L2 were comparable to the non-treated group. Taking all together, we have found that HDAC6i could be used as a potential adjuvant in ongoing therapeutic options involving immune check-point blockade. Citation Format: Tessa Knox, Eva Sahakian, Jayakumar Nair, Jennifer Kim, Debarati Banik, Melissa Hadley, John Powers, Fengdong Cheng, Sida Shen, Javier Pinilla, Jeffrey Weber, Alan Kozikowski, Eduardo Sotomayor, Alejandro Villagra. Enhancing anti-PD-1 immune blockade in melanoma through selective inhibition of histone deacetylase 6 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4055. doi:10.1158/1538-7445.AM2017-4055

Abstract 2331: HDAC6, new role as master regulator of PD-L1 and immune-related pathways

Histone deacetylases (HDACs), originally described as histone modifiers, have more recently been demonstrated to modify a variety of other proteins involved in diverse cellular processes unrelated to the chromatin environment, including the modulation of proteins related to cell cycle/apoptosis and immune regulation. In contrast to the well-documented effects of HDAC inhibitors (HDACi) in the control of cell cycle and apoptosis, their role in immunobiology is still not completely understood, and the reported immunological outcomes when using HDACi are heterogeneous. Our group recently reported that the pharmacological or genetic abrogation of a single HDAC, HDAC6, modulates the expression of immuno-regulatory proteins, including PD-L1, PD-L2, MHC class I, B7-H4 and TRAIL-R1. We primarily focused in PD-L1, which is an important negative regulator of T-cell function and often over-expressed in cancer cells. In a mechanistic point of view, we have found that the pharmacological inhibition and genetic abrogation of HDAC6 inactivates the STAT3 pathway, impairs the nuclear translocation and the recruitment of STAT3 to the PD-L1 promoter and subsequently down-regulates the expression of PD-L1. Moreover, the in vivo abrogation of HDAC6 reduces tumor growth in melanoma models, effect that is enhanced in the presence of the immune check-point blocking antibodies anti-PD-1 and anti-CTLA4. These results provide a key pre-clinical rationale and justification to further study isotype selective HDAC6 inhibitors as potential immunomodulatory agents in cancer. Citation Format: Tessa Knox, Maritza Lienlaf, Patricio Perez, Mibel Pabon, Calvin Lee, Fengdong Cheng, Eva Sahakian, John Powers, Susan Deng, Smalley Keiran, Alan Kozikowski, Javier Pinilla, Amod Sarnaik, Ed Seto, Jeffrey Weber, Eduardo Sotomayor, Alejandro Villagra. HDAC6, new role as master regulator of PD-L1 and immune-related pathways. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2331.

Abstract 4260: Histone deacetylase 11 (HDAC11) as a novel therapeutic target in the regulation of myeloid-derived suppressor cell (MDSC)

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL The physiological role of this HDAC11, the newest member of the HDAC family was mainly unknown until the discovery by our group that HDAC11 regulates IL-10 gene expression in myeloid cells in-vitro. To better elucidate the role of HDAC11 in these cells, we have utilized HDAC11 promoter-driven eGFP reporter transgenic mice (TgHDAC11-eGFP) which allow us to “visualize” dynamic changes in HDAC11 gene expression /transcriptional activity in immune cells in vivo. Immature myeloid cells (IMCs) differentiate into macrophages, dendritic cells, and neutrophils and are also considered to be precursors of MDSCs in tumor-bearing hosts. Here, we show for the first time that HDAC11 plays an important role in this process. First, IMCs from the bone marrow and spleen of TgHDAC11-eGFP mice display high expression of eGFP indicative of HDAC11 transcriptional activation in these cells in the steady state. Subcutaneous injection of EL4 tumor cells into these mice resulted in expansion of MDSCs (identified by the expression of CD11b+/GR1+ [Ly6G and Ly6C] with variable expression of CD49d and CD115) in their lymphoid organs which was similar in extent to the expansion observed in tumor-bearing wild type (WT) mice. Of note, flow cytometric analysis revealed that expression of eGFP was significantly decreased in the myeloid compartment of tumor bearing TgHDAC11-eGFP mice, with greatest decrease in the Ly6Chigh population suggesting that the transition of IMC into MDSCs might require a decrease in HDAC11 expression. Reminiscent of our findings in the eGFP mice, studies in nontransgenic mice also demonstrated that tumor derived MDSCs display less HDAC11 mRNA expression compared to splenic MDSCs, with lowest expression of HDAC11 in the Ly6Chigh MDSCs. Additional support for the regulatory role of HDAC11 in MDSC expansion/function has been recently provided by our preliminary functional studies in TgHDAC11-eGFP mice which demonstrates that isolated GR1+/eGFP- cells demonstrate more suppressive phenotype than that of GR1+/eGFP+ cells, in the presence of tumor challenge. Taken together, these results suggest that HDAC11 plays a regulatory role in the expansion and function of MDSCs in vivo. A better understanding of this previously unknown role of HDAC11 in MDSC biology might lead to targeted epigenetic therapies to influence the suppressive abilities of these cells and augment the efficacy of immunotherapeutic approaches against malignancies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4260. doi:1538-7445.AM2012-4260