Combination Of Romidepsin Research Articles

Compensatory combination of romidepsin with gemcitabine and cisplatin to effectively and safely control urothelial carcinoma

BackgroundHuman urothelial carcinoma (UC) has a high tendency to recur and progress to life-threatening advanced diseases. Advanced therapeutic regimens are needed to control UC development and recurrence.MethodsWe pursued in vitro and in vivo studies to understand the ability of a triple combination of gemcitabine, romidepsin, and cisplatin (Gem+Rom+Cis) to modulate signalling pathways, cell death, drug resistance, and tumour development.ResultsOur studies verified the ability of Gem+Rom+Cis to synergistically induce apoptotic cell death and reduce drug resistance in various UC cells. The ERK pathway and reactive oxygen species (ROS) played essential roles in mediating Gem+Rom+Cis-induced caspase activation, DNA oxidation and damage, glutathione reduction, and unfolded protein response. Gem+Rom+Cis preferentially induced death and reduced drug resistance in oncogenic H-Ras-expressing UC vs. counterpart cells that was associated with transcriptomic profiles related to ROS, cell death, and drug resistance. Our studies also verified the efficacy and safety of the Gem plus Rom+Cis regimen in controlling UC cell-derived xenograft tumour development and resistance.ConclusionsMore than 80% of UCs are associated with aberrant Ras-ERK pathway. Thus the compensatory combination of Rom with Gem and Cis should be seriously considered as an advanced regimen for treating advanced UCs, especially Ras-ERK-activated UCs.

A Phase I/II Study to Examine the Safety and Efficacy of Pembrolizumab 200 Mg Fixed Dose Administered Every 3 Weeks (Q3W) in Combination with Romidepsin in Relapsed or Refractory Peripheral T-Cell Lymphoma (PTCL)

A Phase I/II Study to Examine the Safety and Efficacy of Pembrolizumab 200 Mg Fixed Dose Administered Every 3 Weeks (Q3W) in Combination with Romidepsin in Relapsed or Refractory Peripheral T-Cell Lymphoma (PTCL)

Romidepsin in Combination with Gemcitabine, Oxaliplatin, and Dexamethasone Shows Durable Responses in Aggressive Lymphomas

Romidepsin in Combination with Gemcitabine, Oxaliplatin, and Dexamethasone Shows Durable Responses in Aggressive Lymphomas

Strategy for Overcoming Crebbp and EP300 Mutations in Lymphoma: Development of First-in-Class HAT Activators

Epigenetic dysregulation is a hallmark of germinal center derived B cell lymphomas. In particular, the monoallelic mutations in histone acetyltransferases (HATs), p300 and CBP are present in approximately 40% of DLBCL and FL patients. The haploinsufficiency of CBP leads to loss of MHC which contributes to both immune escape and lymphomagenesis. The intact wild-type allele offers an opportunity to leverage the normal enzyme to overcome the pathogenic impact of the mutated allele. We reported our lead compound YF2 out of a library (n=70) of HAT activators led to significant cytotoxic effect in lymphoma cell lines compared to its parent compound CTPB, which is not cell membrane permeable. YF2 induced p300-mediated histone acetylation as well as p53 acetylation in cell-free acetylation assay. YF2 also induced the acetylation of H3K14 and H3K27 at the cellular level. Since CREBBP mutations are more frequent than EP300 and are mutually exclusive in patients, it is of high insterest to determine if YF2 affects CBP-mediated acetylation. We measured CBP mediated histone acetylation in a cell free assay by combining the catalytic core of CBP, histone H3 and acetyl-CoA. YF2 increased the level of CBP-mediated Ac-H3K27 (EC50=175.4nM) and Ac-H3K18 (EC50=1330 nM). Studies have shown that in mice lacking Crebbp there is reduced MHC II surface expression on GC cells impairing antigen presentation. To determine if YF2 affects the expression of MHC class I, II and related genes in B cell lymphoma cells, qPCR based gene expression profiling was performed pre and post YF2 exposure. YF2 significantly induces mRNA expression level of HLA-DMA (2.9 fold), PSMB10 (2 fold), and reduced the expression of HLA-G (10 fold). Encode ChIP-seq data confirms enrichement of CBP and p300 in the promotor regions of HLA-DMA and PSMB10 in germinal center B cells, indicating a regulatory effect of YF2 on these genes via HAT modulation. HLA‐G is known to inhibit NK‐cell- and CTL‐mediated cytotoxicity, therefore a reduction in HLA-G may release the inhibition of NK-cell and CTL recognition. Taken together, YF2 may modulate p300 and CBP mediated HLA and related gene expression to induce improved immune surveillance. YF2 is selectively cytotoxic in EP300 mutated B cell lymphoma lines. Since HDAC inhibitors have been approved in the clinic for patients with T cell lymphoma (TCL), we further tested the cytotoxic effect of YF2 in 6 TCL cell lines. Interestingly, YF2 is equivalently cytotoxic in TCL cell lines with IC50 ranging from 4.4 to 14 uM compared to the IC50 values in EP300 mutated BCL lines (5 to 11 uM). Additionally, YF2 induces apoptosis and G1 arrest in a dose dependent manner demonstrated by flow cytometry. We collected three primary lymphoma patient samples including one FL sample carrying CREBBP and p53 mutations and observed that YF2 induced cell death in patient samples, but not in PBMCs from 3 healthy donors, indicating a good therapeutic window. Robust synergy with combination of YF2 and romidepsin was observed in 100% (6/6) EP300 mutated BCL, 40% (2/5) EP300 wild-type BCL and 60% (3/5) TCL cells (EOB = 28). In vivo, SCID‐beige mice were xenografted with SUDHL6 and treated with either vehicle, YF2 40mg/kg, romidepsin 2mg/kg or the combination. Single agent YF2 led to equivalent tumor growth delay as FDA-approved drug romidepsin. The YF2 plus romidepsin combination led to significant tumor growth delay compared to the single agents and control. The efficacy of YF2 and romidepsin in PDX model will be determined in an ongoing study. Given the fact that the RING domain of p300 functions as an inhibitory domain to regulate p300 activity, we hypothesized that YF2 may bind to the RING domain, change the confirmation to facilitate access to the HAT substrate-binding pocket. As predicted by LBias, small molecule binding sites on p300 are on the RING domain and bromodomain. We further checked the potential binding site by docking YF2-like small molecules on the RING domain. We discovered the potential binding site of YF2 in the pocket of RING domain. Kinome screening showed YF2 has no off-target effects on 97 kinases. Ongoing experiments will determine the binding of YF2 to the p300 and CBP catalytic cores. In summary, our lead compound YF2 induces HAT enzymes by activating not only p300 but also CBP mediated acetylation of histone. Modulating HLA genes by YF2 may potentially offer an opportunity to reverse immune evasion adopted by lymphoma cells as a mechanism for survival. Disclosures O'Connor: Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Mundipharma: Consultancy, Membership on an entity's Board of Directors or advisory committees; ADCT Therapeutics, Affimed, Agensys, Merck, Seattle Genetics, Spectrum, Trillium, and Verastem Oncology.: Research Funding; TG Therapeutics: Other: Travel Support, Research Funding. Arancio:Appia Pharmaceuticals: Research Funding; Neurokine Therapeutics: Other: Founder. Amengual:Epizyme: Speakers Bureau; Appia Pharmaceuticals: Research Funding.

Suppression of BCL6 function by HDAC inhibitor mediated acetylation and chromatin modification enhances BET inhibitor effects in B-cell lymphoma cells

Multiple genetic aberrations in the regulation of BCL6, including in acetyltransferase genes, occur in clinically aggressive B-cell lymphomas and lead to higher expression levels and activity of this transcriptional repressor. BCL6 is, therefore, an attractive target for therapy in aggressive lymphomas. In this study romidepsin, a potent histone deacetylase inhibitor (HDACi), induced apoptosis and cell cycle arrest in Burkitt and diffuse large B-cell lymphoma cell lines, which are model cells for studying the mechanism of action of BCL6. Romidepsin caused BCL6 acetylation at early timepoints inhibiting its function, while at later timepoints BCL6 expression was reduced and target gene expression increased due to chromatin modification. MYC contributes to poor prognosis in aggressive lymphoma. MYC function is reduced by inhibition of chromatin readers of the bromodomain and extra-terminal repeat (BET) family, which includes BRD4. The novel combination of romidepsin and JQ1, a BRD4 inhibitor was investigated and showed synergy. Collectively we suggest that the combination of HDACi and BRD4i should be pursued in further pre-clinical testing.

Precision Targeting with EZH2 and HDAC Inhibitors in Epigenetically Dysregulated Lymphomas.

Both gain-of-function enhancer of zeste homolog 2 (EZH2) mutations and inactivating histone acetyltransferases mutations, such as CREBBP and EP300, have been implicated in the pathogenesis of germinal center (GC)-derived lymphomas. We hypothesized that direct inhibition of EZH2 and histone deacetyltransferase (HDAC) would be synergistic in GC-derived lymphomas. Lymphoma cell lines (n = 21) were exposed to GSK126, an EZH2 inhibitor, and romidepsin, a pan-HDAC inhibitor. Synergy was assessed by excess over bliss. Western blot, mass spectrometry, and coimmunoprecipitation were performed. A SU-DHL-10 xenograft model was utilized to validate in vitro findings. Pretreatment RNA-sequencing of cell lines was performed. MetaVIPER analysis was used to infer protein activity. Exposure to GSK126 and romidepsin demonstrated potent synergy in lymphoma cell lines with EZH2 dysregulation. Combination of romidepsin with other EZH2 inhibitors also demonstrated synergy suggesting a class effect of EZH2 inhibition with romidepsin. Dual inhibition of EZH2 and HDAC led to modulation of acetylation and methylation of H3K27. The synergistic effects of the combination were due to disruption of the PRC2 complex secondary to acetylation of RbAP 46/48. A common basal gene signature was shared among synergistic lymphoma cell lines and was characterized by upregulation in chromatin remodeling genes and transcriptional regulators. This finding was supported by metaVIPER analysis which also revealed that HDAC 1/2 and DNA methyltransferase were associated with EZH2 activation. Inhibition of EZH2 and HDAC is synergistic and leads to the dissociation of PRC2 complex. Our findings support the clinical translation of the combination of EZH2 and HDAC inhibition in EZH2 dysregulated lymphomas.

Latency reversal agents affect differently the latent reservoir present in distinct CD4+ T subpopulations.

Latency reversal agents (LRAs) have proven to induce HIV-1 transcription in vivo but are ineffective at decreasing the size of the latent reservoir in antiretroviral treated patients. The capacity of the LRAs to perturb the viral reservoir present in distinct subpopulations of cells is currently unknown. Here, using a new RNA FISH/flow ex vivo viral reactivation assay, we performed a comprehensive assessment of the viral reactivation capacity of different families of LRAs, and their combinations, in different CD4+ T cell subsets. We observed that a median of 16.28% of the whole HIV-reservoir induced HIV-1 transcripts after viral reactivation, but only 10.10% of these HIV-1 RNA+ cells produced the viral protein p24. Moreover, none of the LRAs were powerful enough to reactivate HIV-1 transcription in all CD4+ T cell subpopulations. For instance, the combination of Romidepsin and Ingenol was identified as the best combination of drugs at increasing the proportion of HIV-1 RNA+ cells, in most, but not all, CD4+ T cell subsets. Importantly, memory stem cells were identified as highly resistant to HIV-1 reactivation, and only the combination of Panobinostat and Bryostatin-1 significantly increased the number of cells transcribing HIV within this subset. Overall, our results validate the use of the RNA FISH/flow technique to assess the potency of LRAs among different CD4+ T cell subsets, manifest the intrinsic differences between cells that encompass the latent HIV reservoir, and highlight the difficulty to significantly impact the latent infection with the currently available drugs. Thus, our results have important implications for the rational design of therapies aimed at reversing HIV latency from diverse cellular reservoirs.

Romidepsin-Bendamustine Combination for Relapsed/Refractory T Cell Lymphoma

Background: The treatment of relapsed/refractory (R/R) peripheral T cell lymphoma (PTCL) is limited to a few agents. Romidepsin, a histone deacetylase inhibitor, was approved for PTCL treatment as a single agent in the R/R setting, yet with partial efficacy. Several attempts to combine romidepsin with other chemotherapy regimens have been reported, however, with significant toxicity. Objectives: To study the romidepsin-bendamustine combination in PTCL in an attempt to maximize efficacy while minimizing toxicity. Methods: We report on a series of 7 heavily pretreated PTCL patients (2–5 previous lines of therapy) treated with a romidepsin-bendamustine combination. Results: Four patients were not previously exposed to either drug. Of these, 2 achieved complete remission. Interestingly, 1 patient continued treatment with a prolonged progression-free survival of more than 4 years. Toxicity was minimal and no treatment-related deaths or discontinuation were noted. Significant nausea and vomiting were reported in over 50% of patients. Hematological toxicity was mild and lower than that reported for other romidepsin-chemotherapy combinations and was correlated with bone marrow involvement by lymphoma. Conclusions: Although reporting a small number of patients, our data suggest that the combination of romidepsin and bendamustine may be a feasible therapeutic option in R/R PTCL patients and merits further study.

Targeting mitochondrial hexokinases increases efficacy of histone deacetylase inhibitors in solid tumor models

Hexokinase 1 and 2 have been shown to inhibit Bak- and Bax-mediated apoptosis, leading us to combine the histone deacetylase inhibitor romidepsin with clotrimazole or bifonazole, two compounds that reportedly decrease mitochondrial localization of hexokinases. Cancer cell lines derived from breast, kidney, lung, colon or ovarian cancers were treated with a short-term exposure to 25 ng/ml romidepsin combined with either clotrimazole or bifonazole. The combination of romidepsin with 25 µM clotrimazole or bifonazole resulted in increased annexin staining compared to cells treated with any of the drugs alone. Cell death was caspase-mediated, as the pan-caspase inhibitor Q-VD-OPh was found to inhibit apoptosis induced by the combination. A549 lung cancer cells or HCT-116 cells deficient in Bak and Bax were also resistant to apoptosis with the combination implicating the intrinsic apoptotic pathway. We found that a 24 h treatment with clotrimazole or bifonazole decreased total hexokinase 2 expression, resulting in a 76% or 60% decrease, respectively, of mitochondrial expression of hexokinase 2. Mitochondrial hexokinase 1 levels increased 2-fold or less. Our work suggests that the combination of a short-term romidepsin treatment with bifonazole or clotrimazole leads to increased apoptosis, most likely due to decreased mitochondrial expression of hexokinase 2.

Romidepsin in Combination with Gemcitabine, Oxaliplatin, and Dexamethasone Shows Durable Responses in Aggressive Lymphomas Including AITL and DLBCL: Phase I Results

Romidepsin in Combination with Gemcitabine, Oxaliplatin, and Dexamethasone Shows Durable Responses in Aggressive Lymphomas Including AITL and DLBCL: Phase I Results

192 Gene expression profiling of mechlorethamine and romidepsin combination shows downregulation of the JAK/STAT pathway in cutaneous T-cell lymphoma

192 Gene expression profiling of mechlorethamine and romidepsin combination shows downregulation of the JAK/STAT pathway in cutaneous T-cell lymphoma

A Phase Ib/IIa Trial of the Combination of Romidepsin, Lenalidomide and Carfilzomib in Patients with Relapsed/Refractory Lymphoma Shows Complete Responses in Relapsed and Refractory B- and T-Cell Lymphomas

A Phase Ib/IIa Trial of the Combination of Romidepsin, Lenalidomide and Carfilzomib in Patients with Relapsed/Refractory Lymphoma Shows Complete Responses in Relapsed and Refractory B- and T-Cell Lymphomas

Autophagy blockade enhances the anti-cancer effect of Romidepsin in gastric cancer

Background: Autophagy, also known as macroautophagy, is a catabolic process that occurs in response to stress conditions to maintain cellular homeostasis. Inhibiting autophagy has been proposed as a new therapeutic strategy that could enhance the anticancer activity of histone deacetylase inhibitors. We previously demonstrated that romidepsin, a Food and Drug Administration-approved histone deacetylase inhibitor, acts as a potential anticancer agent. Therefore, this study aimed to develop effective combination treatment strategies to maximize the antitumor effects of romidepsin. Methods: Regulation of autophagy by romidepsin was assessed by western blot assay, transmission electron microscopy (TEM), flow cytometric analysis and immunocytochemistry. The autophagy-inducing effect of a combination of romidepsin and hydroxychloroquine (HCQ) in gastric cancer (GC) was assessed in vitro and in vivo . Results: Romidepsin induced autophagy in GC cells, possibly by regulating the extracellular signal-regulated kinase (ERK) and mammalian target of rapamycin (mTOR) signaling pathways. Furthermore, HCQ synergistically augmented the activity of romidepsin in vitro and in vivo . Conclusions: Our results suggest that combining romidepsin and HCQ may represent a novel therapeutic strategy for GC.

Abstract 1048: Reactive oxygen species-mediated synergistic and preferential induction of cell death and reduction of drug resistance in oncogenic H-Ras-expressing bladder cancer cells by combined romidepsin and cisplatin with gemcitabine

Abstract Human urinary bladder cancer is the fifth most commonly diagnosed cancer in the United States and is often associated with induction of oncogenic H-Ras. Long-term survival of patients is suboptimal with the current chemotherapeutic regimen of combined cisplatin and gemcitabine and others due to acquired drug resistance and recurrence. Thus, it is urgent to develop new regimens effective in control of tumor growth, drug resistance, and recurrence to reduce the morbidity and mortality of this disease. We used our cellular system, consisting of human urinary bladder cancer J82 cells paired with its derived oncogenic H-Ras-expressing J82-Ras cells, to investigate the efficacy of a novel combination regimen in treating bladder cancer cells. Our studies revealed, for the first time, the ability of a combination of cisplatin and romidepsin with gemcitabine to synergistically and preferentially induced cell death and reduced drug resistance in J82-Ras versus J82 cells. The Ras-ERK-Nox pathway played an essential role in mediating signals to elevate reactive oxygen species (ROS), leading to enhanced caspase activation, DNA damage, and DNA oxidation, as well as reduced glutathione, to synergistically increased cell death and reduced drug resistance in cells treated with combined romidepsin, cisplatin, and gemcitabine. Preferentially induced Ras-ERK-Nox-ROS pathway, caspase activation, and DNA damage/oxidation, as well as reduced glutathione, contributed to the preferentially induced cell death and reduced drug resistance in J82-Ras versus J82 cells. Synergistically induced death and reduced drug resistance were also detected in human bladder cancer SW780 cells treated with combined romidepsin, cisplatin, and gemcitabine. Hence, our results lead us to suggest that a combination of romidepsin, cisplatin, and gemcitabine should be seriously considered as a new therapeutic regimen for controlling the development and recurrence of human urinary bladder cancer, especially Ras-ERK-activated cancers. Citation Format: Lenora A. Pluchino, Hwa-Chain R. Wang. Reactive oxygen species-mediated synergistic and preferential induction of cell death and reduction of drug resistance in oncogenic H-Ras-expressing bladder cancer cells by combined romidepsin and cisplatin with gemcitabine [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 1048. doi:10.1158/1538-7445.AM2017-1048

Abstract 155: Addition of RP6530, a dual PI3Kδ/γ inhibitor, accentuates Romidepsin activity in NHL cells in vitro

Abstract Background: Relapsed or refractory Non-Hodgkin’s Lymphoma (NHL) represents a significant unmet medical need. Novel treatment options currently being explored include Romidepsin, a histone deacetylase inhibitor, which has demonstrated significant preclinical and clinical activity. Because the PI3K pathway is often activated in NHL, combining Romidepsin with a PI3K δ/γ inhibitor may be a viable alternative to chemotherapy. RP6530 is a novel, potent, and selective PI3K δ/γ inhibitor that demonstrated high potency against PI3Kδ (IC50=25 nM) and γ (IC50=33 nM) enzymes with selectivity over α (>300-fold) and β (>100-fold) isoforms. Dose-escalation trials to evaluate the safety and efficacy of RP6530 in patients with hematological malignancies are currently underway at several sites in US and Europe. The objective of this study was to evaluate the effect of a combination of Romidepsin and RP6530 in NHL cells. Methods: Combination of Romidepsin and RPRP6530 was evaluated in the following NHL cell lines: OCI-LY-1, OCI-LY-10, RAJI, DAUDI, TOLEDO, JEKO, MAVER, and SU-DHL-1. Synergism or additivity between Romidepsin and RP6530 was determined using different concentrations of the compounds in a 5 x 5 grid and calculated based on the BLISS score. Effect on apoptosis and cell cycle were estimated by flow cytometry. Phosphorylated AKT and PARP were determined by Western Blotting. Results: Synergism or additivity between Romidepsin and RP6530 for anti-proliferative activity was noticed in all the cell lines tested with effect being most pronounced in OCI-LY-1, OCI-LY-10, JEKO, and SU-DHL-1. Incubating cells with 3-5 nM Romidepsin and 3 or 5 μM RP6530 resulted in a 2 to 5-fold increase in the total apoptotic population. Cell cycle analysis indicated that the combination of Romidepsin (1-5 nM) and RP6530 (1-5 μM) caused a G0/G1 arrest (2 to 3-fold) with a corresponding increase in percent of apoptotic cells. While Romidepsin had minimal effect of pAKT, treatment with RP6530 reduced expression by 40-90% across cell lines. Further, Romidepsin + RP6530 enhanced expression of cleaved PARP (8 to 60-fold) compared to either agent alone Conclusions: Addition of RP6530, a potent and selective dual PI3Kδ/γ inhibitor, potentiated Romidepsin activity in NHL cells. Findings provide a rationale for use of the combination in future clinical trials involving naïve or relapsed NHL patients thereby providing a safer alternative to currently available therapy. Citation Format: Srikant Viswanadha, Satyanarayana Eleswarapu, Seeta Nyayapathy, Swaroop Vakkalanka. Addition of RP6530, a dual PI3Kδ/γ inhibitor, accentuates Romidepsin activity in NHL cells in vitro [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 155. doi:10.1158/1538-7445.AM2017-155

Romidepsin alone or in combination with anti-CD20 chimeric antigen receptor expanded natural killer cells targeting Burkitt lymphoma in vitro and in immunodeficient mice

ABSTRACTFacilitating the development of alternative targeted therapeutic strategies is urgently required to improve outcome or circumvent chemotherapy resistance in children, adolescents, and adults with recurrent/refractory de novo mature B-cell (CD20) non-Hodgkin lymphoma, including Burkitt lymphoma (BL). Romidepsin, a histone deacetylase inhibitor (HDACi), has been used to treat cutaneous T-cell lymphoma. We have demonstrated the significant anti-tumor effect of anti-CD20 chimeric antigen receptor (CAR) modified expanded peripheral blood natural killer (exPBNK) against rituximab-sensitive and -resistant BL. This study examined the anti-tumor activity of romidepsin alone and in combination with anti-CD20 CAR exPBNKs against rituximab-sensitive and -resistant BL in vitro and in vivo. We found that romidepsin significantly inhibited both rituximab-sensitive and -resistant BL cell proliferation in vitro (P < 0.001) and induced cell death in rituximab-sensitive Raji (P < 0.001) and cell cycle arrest in rituximab-resistant Raji-2R and Raji-4RH (P < 0.001). Consistent with in vitro observations, we also found romidepsin significantly inhibited the growth of rituximab-sensitive and -resistant BL in BL xenografted NSG mice. We also demonstrated that romidpesin significantly induced the expression of Natural Killer Group 2, Member D (NKG2D) ligands MICA/B in both rituximab-sensitive and -resistant BL cells (P < 0.001) resulting in enhancement of exPBNK in vitro cytotoxicity through NKG2D. Finally, we observed the combination of romidepsin and anti-CD20 CAR exPBNK significantly induced cell death in BL cells in vitro, reduced tumor burden and enhanced survival in humanized BL xenografted NSG mice (p < 0.05). Our data suggests that romidepsin is an active HDAC inhibitor that also potentiates expanded NK and anti-CD20 CAR exPBNK activity against rituximab-sensitive and -resistant BL.

A Phase Ib/IIa Trial of the Combination of Romidepsin, Lenalidomide and Carfilzomib in Patients with Relapsed/Refractory Lymphoma Shows Complete Responses in Relapsed and Refractory T-Cell Lymphomas

A Phase Ib/IIa Trial of the Combination of Romidepsin, Lenalidomide and Carfilzomib in Patients with Relapsed/Refractory Lymphoma Shows Complete Responses in Relapsed and Refractory T-Cell Lymphomas

The histone deacetylase inhibitor romidepsin synergizes with lenalidomide and enhances tumor cell death in T-cell lymphoma cell lines

ABSTRACTWe investigated the cytotoxic interactions of romidepsin, a histone deacetylase inhibitor, and lenalidomide, an immunomodulatory agent, in a T-cell lymphoma preclinical model. Hut-78 and Karpas-299 cells were treated with romidepsin and lenalidomide alone and in combination. The interaction between romidepsin and lenalidomide was evaluated by the Chou–Talalay method, and cell viability and clonogenicity were also evaluated. Apoptosis, reactive oxygen species (ROS) levels, and cell cycle distribution were determined by flow cytometry. ER stress, caspase activation, and the AKT, MAPK/ERK, and STAT-3 pathways were analyzed by Western blot. Combination treatment with romidepsin and lenalidomide had a synergistic effect in Hut-78 cells and an additive effect in Karpas-299 cells at 24 hours and did not decrease the viability of normal peripheral blood mononuclear cells. This drug combination induced apoptosis, increased ROS production, and activated caspase-8, −9, −3 and PARP. Apoptosis was associated with increased hallmarks of ER stress and activation of UPR sensors and was mediated by dephosphorylation of the AKT, MAPK/ERK, and STAT3 pathways.The combination of romidepsin and lenalidomide shows promise as a possible treatment for T-cell lymphoma. This work provides a basis for further studies.

Romidepsin and Azacitidine Synergize in their Epigenetic Modulatory Effects to Induce Apoptosis in CTCL.

Cutaneous T-cell lymphomas (CTCL) are a heterogeneous group of malignancies that despite available therapies commonly relapse. The emergence of combination epigenetic therapies in other hematologic malignancies have made investigation of such combinations in CTCL a priority. Here, we explore the synergistic antiproliferative effects of romidepsin, an HDAC inhibitor, and azacitidine, a demethylating agent, combination in CTCL. The growth inhibition under combination treatment and single agent was explored by the MTT cell viability assay and the Annexin V/propidium iodide (PI) apoptosis assay in different CTCL cell lines and tumor cells derived from Sézary syndrome patients. Quantitative analysis of a dose-effect relationship of romidepsin and azacitidine was done by the CompuSyn software. Investigation of mechanism of action was performed by flow cytometry, immunoblotting, qRT-PCR arrays, and chromatin immunoprecipitation. Global CpG methylation sequencing was utilized to study genome methylation alteration under the treatment modalities. The combination of romidepsin and azacitidine exerts synergistic antiproliferative effects and induction of apoptosis involving activation of the caspase cascade in CTCL cell lines and tumor cells derived from Sézary syndrome patients. We identified genes that were selectively induced by the combination treatment, such as the tumor suppressor geneRhoBthat is linked to enhanced histone acetylation at its promoter region in parallel with pronounced expression of p21. Global CpG methylation sequencing in a CTCL cell line and tumor cells demonstrated a subset of genes with a unique change in methylation profile in the combination treatment. The synergistic antiproliferative effects of romidepsin and azacitidine combination treatment justify further exploration in clinical trials for advanced CTCL.

Induction of MAPK- and ROS-dependent autophagy and apoptosis in gastric carcinoma by combination of romidepsin and bortezomib.

Proteasome inhibitors and histone deacetylase (HDAC) inhibitors can synergistically induce apoptotic cell death in certain cancer cell types but their combinatorial effect on the induction of autophagy remains unknown. Here, we investigated the combinatorial effects of a proteasome inhibitor, bortezomib, and an HDAC inhibitor, romidepsin, on the induction of apoptotic and autophagic cell death in gastric carcinoma (GC) cells. Isobologram analysis showed that low nanomolar concentrations of bortezomib/romidepsin could synergistically induce killing of GC cells. The synergistic killing was due to the summative effect of caspase-dependent intrinsic apoptosis and caspase-independent autophagy. The autophagic cell death was dependent on the activation of MAPK family members (ERK1/2 and JNK), and generation of reactive oxygen species (ROS), but was independent of Epstein-Barr virus infection. In vivo, bortezomib/romidepsin also significantly induced apoptosis and autophagy in GC xenografts in nude mice. This is the first report demonstrating the potent effect of combination of HDAC and proteasome inhibitors on the induction of MAPK- and ROS-dependent autophagy in addition to caspase-dependent apoptosis in a cancer type.