Tolero Pharmaceuticals (United States)

Myelofibrosis (MF) is the deadliest form of myeloproliferative neoplasm (MPN). The JAK inhibitor Ruxolitinib can reduce constitutional symptoms but it does not substantially improve bone marrow fibrosis. Pim1 expression is significantly elevated in MPN/MF hematopoietic progenitors. Here, we show that genetic ablation of Pim1 blocked the development of myelofibrosis induced by Jak2V617F and MPLW515L. Pharmacologic inhibition of Pim1 with a second-generation Pim kinase inhibitor TP-3654 significantly reduced leukocytosis and splenomegaly, and attenuated bone marrow fibrosis in Jak2V617F and MPLW515L mouse models of MF. Combined treatment of TP-3654 and Ruxolitinib resulted in greater reduction of spleen size, normalization of blood leukocyte counts and abrogation of bone marrow fibrosis in murine models of MF. TP-3654 treatment also preferentially inhibited Jak2V617F mutant hematopoietic progenitors in mice. Mechanistically, we show that TP-3654 treatment significantly inhibits mTORC1, MYC and TGF-β signaling in Jak2V617F mutant hematopoietic cells and diminishes the expression of fibrotic markers in the bone marrow. Collectively, our results suggest that Pim1 plays an important role in the pathogenesis of MF, and inhibition of Pim1 with TP-3654 might be useful for treatment of MF.

ObjectivesTo assess the impact of patient age and weight on the pharmacokinetics (PK), testosterone (T) suppression and safety from four fixed dosing regimens (7.5, 22.5, 30, or 45 mg for 1‐, 3‐, 4‐, or 6‐months, respectively) of subcutaneous in situ gel delivered leuprolide acetate (Gel‐LA) injected via the ATRIGEL Delivery System in patients with prostate cancer (PCa).Patients and methodsTwo patient populations were specified for analysis: a small cohort of surgically castrated PCa patients and a large, pooled population of PCa patients from four pivotal trials of Gel‐LA. Two separate analyses of the impact of age and weight on study endpoints were conducted: (1) PK and safety of a single monthly dose of Gel‐LA in a Phase 1 study with PCa patients who had undergone bilateral surgical orchiectomy (“Bilaterally orchiectomized male study”); (2) PK/pharmacodynamic (PD) effects and safety using pooled data from four pivotal trials assessing 1‐, 3‐, 4‐, and 6‐month dosing of Gel‐LA in patients with advanced PCa, stratified by age and body weight (pivotal trials).ResultsEight orchiectomized patients from the “Bilaterally orchiectomized male study” and 438 patients from the pivotal trials were included in the analyses. Age and body weight did not appear to affect the PK results in the orchiectomized patient population. Pooled pivotal trial data showed that serum T levels did not appear to be influenced by age or weight; ≥90% of patients across all age groups and ≥92% of patients across all weight groups achieved T ≤ 50 ng/dL by week 4. Median T levels for castration (T ≤ 50 ng/dL) were maintained from week 3 until the end of the study and all subgroups achieved median T ≤ 20 ng/dL by week 4. Patients from the orchiectomized patient study did not report any serious treatment‐related adverse events (AEs) and there were no AE‐related withdrawals from the study. The most common AEs were hot flashes and injection site events. The safety profiles from pivotal trials have been previously described and, as expected, were consistent with known effects of LHRH agonist therapy and suppression of T levels.ConclusionPK and PD of Gel‐LA appear to be unaffected by age and body weight, as demonstrated by persistence of effective drug levels through the dosing period and consistent T suppression across different ages and body weights.

Abstract Colorectal cancer (CRC) is the third leading cause of cancer-related mortality in the US (2017). Most patients present with stage II or III disease at diagnosis, with the 5-year survival rate between 53-89%. Survival in patients with stage IV CRC, however, is a discouraging 11%. Metastasis in CRC is linked to a mesenchymal phenotype, which is associated with chemoresistance. AXL, a receptor tyrosine kinase, promotes the mesenchymal phenotype in cancer cells and its expression is associated with drug resistance and poor outcomes. TP-0903, a clinical-stage, investigational small molecule inhibitor of AXL has been shown to reverse the mesenchymal phenotype and restore drug sensitivity in cells that no longer respond to standard agents in preclinical models. TP-0903 is hypothesized to be an active agent in CRC through reversal of the mesenchymal phenotype. In cell viability assays of CRC lines, TP-0903 treatment resulted in IC50 values ranging from 4.5 – 123 nM. Notably, cell growth inhibition by TP-0903 was independent of KRAS mutation status; the KRAS mutant HCT-116 line was the most sensitive CRC cell line tested. Mesenchymal markers, including Snail, were suppressed by 7.6-fold (mRNA) and 4.9-fold (protein) in the HCT-116 line at 500 nM. TP-0903 activity was also assessed in vivo using two KRAS mutant CRC models: HCT-116 and a patient-derived xenograft (PDX) model. In the HCT-116 xenograft model, single agent TP-0903 treatment achieved 69% tumor growth inhibition (%TGI) with an oral dosing schedule at 40 mg/kg. In a KRAS-mutant PDX model, TP-0903 achieved 44% TGI when mice were dosed at 40 mg/kg. Pharmacodynamic analyses were performed on tissues from the HCT-116 and PDX models. The ligand for AXL, GAS6, was significantly upregulated in tissues after TP-0903 treatment in both CRC in vivo models while soluble AXL and GAS6 were significantly downregulated in plasma in the PDX model. Furthermore, Axin2, a Wnt/β-catenin regulated gene, was downregulated by TP-0903 in tumor tissue from the PDX model, suggesting inhibition of the Wnt/βcatenin pathway. These data support a potential role for AXL in the promotion of the mesenchymal phenotype in CRC, and showed that AXL inhibition by TP-0903 suppressed the mesenchymal phenotype and was effective against CRC cells regardless of KRAS mutation status. These observations support further clinical investigation of TP-0903 as a potential therapeutic agent in metastatic CRC. A Phase I trial with this investigational agent is ongoing, including patients with KRAS mutant CRC (clincaltrials.gov, NCT02729298). Citation Format: Ryan Mangelson, Peter Peterson, Jason M. Foulks, Yuta Matsumura, Lars Mouritsen, Clifford J. Whatcott, David J. Bearss, Steven L. Warner. The AXL kinase inhibitor, TP-0903, demonstrates efficacy in preclinical models of colorectal cancer independent of KRAS mutation status [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2197.

Abstract The high rate of non-responders to immune checkpoint inhibitors (ICI) represents significant challenges in the field of immune-oncology. Breast cancer has emerged as a cancer type that responds poorly to ICI. Therefore, there is an urgent need to better understand how cancer cells escape immune surveillance and resist immune attacks. Cancer cell clones with mesenchymal features seem to be less susceptible to attacks by immune cells. Hence, epithelial-to-mesenchymal transition (EMT) targeting agents such as TP-0903 are emerging as potential combination candidates to enhance immune checkpoint inhibition. AXL is overexpressed in a variety of cancers, often driving the pathogenesis and progression of disease. AXL overexpression induces cancer growth, invasion, metastasis, drug resistance, and mesenchymal characteristics. Aside from cancer cells, AXL is also expressed on several types of immune cells including macrophages and dendritic cells. Recently, it has been reported that AXL plays an important role as a negative regulator of immune response, contributing to an anti-tumor immune suppression. We have previously identified TP-0903 to be a potent small molecule inhibitor of AXL (IC50=14 nM) capable of reversing EMT. In this study, we evaluated whether TP-0903 could enhance the ICB effect in an ICB-resistant triple negative, metastatic breast cancer mouse model (4T1). In the 4T1 syngeneic model, anti-PD-1 monotherapy failed to inhibit tumor growth indicating that this tumor is resistant to anti-PD-1 treatment. The combination of TP-0903 and anti-PD-1 resulted in statistically significant tumor growth inhibition versus TP-0903 monotherapy (p<0.05). We hypothesized that the combination effect was influenced by potential CD8+ T cell depletion, and further investigated the effects of TP-0903 on immune cells in spleen and tumors. Tumor growth inhibition was found to be associated with a decrease in myeloid-derived suppressor cells (MDSC) in the spleen and an increase in infiltration and activation of dendritic cells (DCs) in the tumor. Gene expression analysis revealed that TP-0903 treatment decreased multiple immunosuppressive cytokines and chemokines including IL-6 and G-CSF in vivo. These results suggest that TP-0903 modulates the immune-suppressive tumor microenvironment (TME) to reinvigorate T cell immunity in anti-PD-1 resistant 4T1 tumors. In conclusion, AXL inhibition with TP-0903 modulates TME and enhances the anti-tumor effects of ICBs in an anti-PD-1 resistant, metastatic breast cancer mouse model. These findings support additional clinical testing of TP-0903 in combination with ICBs, which is currently being investigated in patients with advanced solid tumors in an ongoing clinical study (NCT02729298). Citation Format: Yuka Kumagai, Jun Oishi, Megumi Nakamura, Jason M. Foulks, Clifford J. Whatcott, Steven L. Warner, David J. Bearss, Masashi Goto. TP-0903, a potent AXL receptor tyrosine kinase inhibitor, enhances the activity of anti-PD-1 therapy in a metastatic preclinical syngeneic model of breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3969.

Abstract Background: Non-small cell lung cancer (NSCLC) is a molecularly heterogeneous disease with a high propensity for drug resistance and metastasis. AXL, a member of the Tyro3-AXL-Mer family of receptor tyrosine kinases, is a central regulator of epithelial-to-mesenchymal transition (EMT) and enables tumor cells to invade and acquire drug resistance. AXL is overexpressed in lung tumors, correlates positively with tumor invasion, drug resistance, and negatively predicts overall survival. We mechanistically interrogate the effects of AXL inhibitor TP-0903 on EMT in lung adenocarcinoma cells using transcriptomic and proteomic profiling. Methods: Atomic force microscopy, Western blot analysis, RNA sequencing and mass cytometry (CyTOF) were all used to scrutinize the biomechanical properties, phenotypic, transcriptomic and proteomic profiles of A549 cells treated with 40nM TP0903 or shAXL knockdown. Results: TP-0903 attenuates total AXL/AXL phosphorylation and blunts transcriptional responses to TGFβ-Hippo signaling by disrupting the transcriptional complexes formed by SMAD2/3, SMAD4, YAP1 and TAZ. AXL knockdown or TP-0903 reverses EMT phenotype and reduces migration potential in A549 and H2009 adenocarcinoma cell lines. CyTOF data also identified resistant clones that overexpress TGFβ receptor II, TAZ protein and display hybrid EMT phenotypes. Conclusions: We are the first to report the interplay between AXL and TGFβ-Hippo signaling axis. TP-0903 study agent has excellent therapeutic promise in NSCLC and we speculate that TP-0903 drug can target epithelial to mesenchymal transitional states in lung cancer cells possibly through the inhibition of the AXL-TGFβ-Hippo signaling axis. Citation Format: Josephine Amalia Taverna, Chia-Nung Hung, Chun-Lin Lin, Pawel Osmulski, Meizhen Chen, Chiou-Miin Wang, Nicholas L. L. Lucio, Nameer Kirma, Chih-Wei Chou, Maria E. Gaczynska, Alia Nazarullah, Mark Wade, Lars Mouritsen, Tim Huang. AXL inhibitor TP-0903 attenuates AXL-TGFbeta Hippo signaling axis in lung adenocarcinoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2193.

Epithelial-to-mesenchymal transition (EMT) is a critical step in the progression of ovarian cancer (OVCA), which drives local invasion, metastasis and drug resistance leading to poor survival. AXL, a receptor tyrosine kinase, functions as a molecular driver of EMT in multiple cancers, including OVCA. The high mortality of this tumor is explained by the fact that the majority (75%) of patients present at an advanced stage, with widely metastatic disease within the peritoneal cavity often associated with ascites. The extent of ascites is a prognostic factor for survival, impacts quality of life, and correlates with the EMT phenotype. Approaches to block or reverse EMT are needed to improve the prognosis of OVCA patients, particularly those with refractory or early relapse (<6 months) disease following debulking surgery and conventional adjuvant treatments. TP-0903 is a potent, small molecule AXL inhibitor that has activity in reversing EMT in multiple models of disease and embryonic development. The objective of this study was to explore the anti-tumor activity in preclinical mesenchymal OVCA-derived models, which may provide rationale to evaluate TP-0903 in treatment-resistant or refractory OVCA patients. TP-0903 showed potent effects on cell viability in multiple OVCA-derived cell lines. The IC50 values in cell viability studies ranged from 33 nM (OVTOKO cells) to 840 nM (OVMANA cells). Kuramochi cells represent the most common ovarian cancer subtype, high-grade serous ovarian cancer and was particularly platinum-resistant (>5 μM), yet sensitive to TP-0903 (210 nM). Baseline phospho-AXL (Y702) was exceptionally high in Kuramochi cells and TP-0903 inhibited this phosphorylation at concentrations as low as 100 nM. TP-0903 reversed EMT in OVCA cells as evident by the down-regulation of mesenchymal markers including Snail and Slug. Consistent with the reversal of the EMT, TP-0903 inhibited the migration of ES-2 OVCA cells in a scratch assay. TP-0903 suppressed ascites development in preclinical models of OVCA and inhibited metastatic intraperitoneal tumor dissemination. Treatment suppressed ascites development (ascites volume, vehicle: 3.34 mL, TP-0903: 0.28 mL; abdominal circumference, vehicle: 7.96 cm (day 15), TP-0903: 6.82 cm (day 15), p<0.05) in an orthotopic ES-2 model and extended overall survival compared to vehicle or cisplatin alone. Phospho-AXL modulation and changes in other mechanism-validating biomarkers were observed in the ES-2 model. Similar preclinical activities, including decreased ascites volume, were observed in A2780cis and OVCAR-3 xenograft models. These results support further clinical evaluation of TP-0903 in either single-agent or combination regimens treating recurrent or refractory OVCA. TP-0903 is investigated in a Phase I trial that includes an expansion cohort of this patient population (NCT02729298).Citation Format: Nozomi Tomimatsu, Ken Fujimura, Yuta Matsumura, Hiroki Umehara, Lars Mouritsen, Steven L. Warner, David J. Bearss. Targeting AXL kinase with TP-0903 successfully reverses the mesenchymal phenotype and extends survival in preclinical models of advanced ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2006.

The high rate of non-responders to immune checkpoint inhibitors (ICI) represents significant challenges in the field of immune-oncology. Breast cancer has emerged as a cancer type that responds poorly to ICI. Therefore, there is an urgent need to better understand how cancer cells escape immune surveillance and resist immune attacks. Cancer cell clones with mesenchymal features seem to be less susceptible to attacks by immune cells. Hence, epithelial-to-mesenchymal transition (EMT) targeting agents such as TP-0903 are emerging as potential combination candidates to enhance immune checkpoint inhibition. AXL is overexpressed in a variety of cancers, often driving the pathogenesis and progression of disease. AXL overexpression induces cancer growth, invasion, metastasis, drug resistance, and mesenchymal characteristics. Aside from cancer cells, AXL is also expressed on several types of immune cells including macrophages and dendritic cells. Recently, it has been reported that AXL plays an important role as a negative regulator of immune response, contributing to an anti-tumor immune suppression. We have previously identified TP-0903 to be a potent small molecule inhibitor of AXL (IC50=14 nM) capable of reversing EMT. In this study, we evaluated whether TP-0903 could enhance the ICB effect in an ICB-resistant triple negative, metastatic breast cancer mouse model (4T1). In the 4T1 syngeneic model, anti-PD-1 monotherapy failed to inhibit tumor growth indicating that this tumor is resistant to anti-PD-1 treatment. The combination of TP-0903 and anti-PD-1 resulted in statistically significant tumor growth inhibition versus TP-0903 monotherapy (p Citation Format: Yuka Kumagai, Jun Oishi, Megumi Nakamura, Jason M. Foulks, Clifford J. Whatcott, Steven L. Warner, David J. Bearss, Masashi Goto. TP-0903, a potent AXL receptor tyrosine kinase inhibitor, enhances the activity of anti-PD-1 therapy in a metastatic preclinical syngeneic model of breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3969.

Abstract The ‘7+3' induction regimen has been the standard of care for frontline acute myeloid leukemia (AML) for more than 30 years. There are still significant unmet needs for frontline patients, particularly those with high-risk features. Alvocidib, a potent CDK9 inhibitor, has shown clinical activity in multiple studies of frontline and relapsed/refractory AML. In some clinical studies, alvocidib has been administered as part of the ACM (alvocidib, cytarabine, and mitoxantrone) regimen. The ACM regimen has demonstrated greater clinical responses compared to 7+3 in frontline intermediate and high-risk AML patients. The ACM regimen is now being evaluated in a multi-institution, Phase II study using a MCL-1 dependency biomarker to select patients likely to be sensitive to the alvocidib-containing regimen. Alvocidib activity is primarily driven by its inhibition of CDK9/RNA-polymerase II and the subsequent suppression of key anti-apoptotic proteins, including MCL-1. Alvocidib induces apoptosis and potentiates the activity of apoptosis-inducing agents, such as cytarabine and mitoxantrone. The synergy of the ACM regimen led us to hypothesize that alvocidib may enhance the anti-leukemia activity of the 7+3 regimen in preclinical models as well. Cell viability and apoptosis measurements were made following treatment using the CellTiter-Glo and Caspase-Glo assays according to manufacturer protocol. MCL-1 mRNA expression and the expression of additional markers was determined using RT-qPCR. Protein expression was determined using standard immunoblotting techniques. Alvocidib and the 7+3 regimen were also tested in vivo in the MV4-11 AML xenograft model. Here, it was observed that the 72-hour viability assays yielded single agent IC50 values of alvocidib, cytarabine, or daunorubicin in cultured AML cells ranging from 2.2 nM to 567 nM. In apoptosis assays, a modest induction was observed with cytarabine, daunorubicin, or alvocidib. However, the three-drug combination more than doubled the induction of apoptosis compared to any of the agents alone. We also observed that MCL-1 was suppressed in the combination treatment. In a xenograft study, 21.1 and 48.5% tumor growth inhibition (TGI) was observed following either single agent treatment of daunorubicin (0.5 mg/kg) or cytarabine (60 mg/kg), respectively. Treatment of alvocidib (1.25 mg/kg) yielded 60.0% TGI. Combination treatment, however, resulted in tumor shrinkage, yielding 116.2% TGI. While we have reported previously that alvocidib synergizes with cytarabine and mitoxantrone preclinically in the ACM regimen, our data here demonstrates that alvocidib potentiates the activity of cytarabine and daunorubicin as well. Our results here provide additional scientific rationale for an investigational Phase Ib clinical study employing alvocidib in combination with 7+3 in frontline AML patients, which is currently ongoing. Citation Format: Wontak Kim, Clifford J. Whatcott, Adam Siddiqui-Jain, David J. Bearss, Steven L. Warner. Alvocidib enhances the efficacy of cytarabine and daunorubicin (7+3) in non-clinical models of acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4823.

Abstract Alvocidib is a potent inhibitor of cyclin-dependent kinase-9 (CDK9) and induces apoptosis in cancer cells by reducing the expression of short-lived, anti-apoptotic proteins such as MCL-1. Alvocidib, as a part of a sequential combination regimen with cytarabine and mitoxantrone (ACM), is currently in a Phase II clinical trial in relapsed/refractory acute myeloid leukemia (AML). Patients with AML that have a high dependence on MCL-1 are considered more likely to benefit from the alvocidib-containing regimen. MCL-1 has emerged as a key protein in drug resistance of multiple solid tumor types including breast, prostate and lung cancers. The use of alvocidib in clinical settings beyond the ACM regimen is somewhat limited by the current intravenous route of administration. An orally administered form of alvocidib would allow prolonged repression of MCL-1 through chronic dosing and scheduling. Alvocidib itself is highly permeable in CACO-2 monolayers and is soluble at acidic pHs but solubility is strikingly reduced at neutral or basic conditions, which could hamper the development of an oral formulation. We hypothesized that a phosphate prodrug of alvocidib would improve solubility under neutral or basic conditions and enable the efficient systemic delivery of alvocidib via oral administration. We synthesized a phosphate prodrug of alvocidib, TP-1287, in three steps from the parent compound. The solubility of TP-1287, was determined at various pH levels. It was found to be highly soluble under acidic, neutral, and basic conditions (1.5 mg/mL at pH 2.2; 1.8 mg/mL at pH 4.5; 9.5 mg/mL at pH 6.8 and 9.3 mg/mL at pH 8.7) compared to alvocidib (4.4 mg/mL at pH 2.2; 1.3 mg/mL at pH 4.5; 0.02 mg/mL at pH 6.8 and 0.02 mg/mL at pH 8.7). Pharmacokinetic studies were conducted in mice in which TP-1287 was efficiently converted to the parent alvocidib (Cmax = 1922.7 ng/ml, t1/2 = 4.4 hr) with high oral bioavailability (%F = 182.3, compared to intravenous alvocidib). Efficacy and pharmacodynamic studies (measuring MCL-1 expression levels), were evaluated in tumor xenograft models. TP-1287 demonstrated significant anti-tumor efficacy in the MV4-11 AML mouse xenograft model and produced as much as a 61.7% inhibition of the pharmacodynamic biomarker MCL-1 in xenografted tumors, demonstrating a wide, 75-fold therapeutic dosing window. In addition, TP-1287 strongly inhibited tumor growth, achieving 109.1% tumor growth inhibition (%TGI) at the 7.5 mg/kg dose level. TP-1287 is highly soluble over a broader pH range than alvocidib and is efficiently metabolized to the parent compound in vivo, following oral administration. Tumor xenograft models and pharmacodynamic studies indicate that oral delivery of TP-1287 is efficacious in mice. Based on these results, we anticipate moving TP-1287, as an orally delivered CDK9 inhibitor, into a forthcoming clinical trial directed towards solid tumors vulnerable to the suppression of MCL-1. Citation Format: Wontak Kim, Hillary Haws, Peter Peterson, Clifford J. Whatcott, Steven Weitman, Steven L. Warner, David J. Bearss, Adam Siddiqui-Jain. TP-1287, an oral prodrug of the cyclin-dependent kinase-9 inhibitor alvocidib [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 5133. doi:10.1158/1538-7445.AM2017-5133

Abstract The proteasome inhibitor bortezomib is widely used in the treatment of patients with multiple myeloma (MM). The expression levels of many proteins increase as a result of bortezomib treatment, including the pro-apoptotic protein NOXA. NOXA functions to sequester the anti-apoptotic BCL-2 family member, MCL-1. High levels of MCL-1 and/or low levels of NOXA have been implicated in bortezomib resistance and negative patient outcomes, including short duration of treatment response. The BCL-2-specific BH3 mimetic venetoclax (ABT-199) has also been explored in multiple hematological malignancies, including the treatment of MM. Venetoclax induces apoptosis in a BCL-2 specific manner by directly inhibiting BCL-2 function. However, intrinsic resistance to venetoclax treatment observed in MM patient samples has been attributed to a low BCL-2-to-MCL-1 gene expression ratio, suggesting a central role for MCL-1 in cell survival in this context as well. Increased MCL-1 expression is a known resistance mechanism to venetoclax treatment in a variety of cell types including chronic lymphocytic leukemia and lymphomas. Considering the central role of MCL-1 to treatment efficacy in MM, we investigated the ability of an MCL-1-lowering agent, namely the CDK9 inhibitor alvocidib, to potentiate the activity of venetoclax in MM. Alvocidib suppresses MCL-1 expression via CDK9-mediated regulation of RNA polymerase II. Alvocidib has achieved robust improvements in the clinical response rates of high-risk, newly diagnosed acute myeloid leukemia (AML) patients as part of the time-sequential ACM regimen (alvocidib + cytarabine + mitoxantrone). We therefore hypothesized that alvocidib would potentiate the activity of venetoclax in MM through an MCL-1-dependent mechanism. In this report, we demonstrate that alvocidib inhibits the protein expression of MCL-1 in MM cells in a time-dependent fashion, up to 96 hours. In cell viability assays, the addition of up to 100 nM venetoclax resulted in a 2.8-fold reduction in the IC50 of alvocidib in the cultured OPM-2 cell line. Conversely, the potentiation of venetoclax activity with the addition of alvocidib resulted in a more than 500-fold decrease in IC50 in the relatively venetoclax-resistant OPM-2 cells. Additional studies are currently underway to investigate the efficacy of alvocidib and venetoclax in the context of bortezomib resistance where low NOXA may contribute to enhanced cell survival via MCL-1. Taken together, our data suggest that the combination of alvocidib with venetoclax may constitute a novel therapeutic regimen in the treatment of MM. Further, it suggests that CDK9-mediated targeting of MCL-1 may offer a clinical route to addressing intrinsic resistance in MM patients. Citation Format: Mark Livingston, Wontak Kim, Hillary Haws, Peter Peterson, Clifford J. Whatcott, Adam Siddiqui-Jain, Steven Weitman, David J. Bearss, Steven L. Warner. Alvocidib potentiates the activity of venetoclax in preclinical models of multiple myeloma [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 1106. doi:10.1158/1538-7445.AM2017-1106