ALBERT

Description: A hallmark of prostate cancer progression is dysregulation of lipid metabolism via overexpression of fatty acid synthase (FASN), a key enzyme in de novo fatty acid synthesis. Metastatic castration-resistant prostate cancer (mCRPC) develops resistance to inhibitors of androgen receptor (AR) signaling through a variety of mechanisms, including the emergence of the constitutively active AR variant V7 (AR-V7). Here, we developed an FASN inhibitor (IPI-9119) and demonstrated that selective FASN inhibition antagonizes CRPC growth through metabolic reprogramming and results in reduced protein expression and transcriptional activity of both full-length AR (AR-FL) and AR-V7. Activation of the reticulum endoplasmic stress response resulting in reduced protein synthesis was involved in IPI-9119–mediated inhibition of the AR pathway. In vivo, IPI-9119 reduced growth of AR-V7–driven CRPC xenografts and human mCRPC-derived organoids and enhanced the efficacy of enzalutamide in CRPC cells. In human mCRPC, both FASN and AR-FL were detected in 87% of metastases. AR-V7 was found in 39% of bone metastases and consistently coexpressed with FASN. In patients treated with enzalutamide and/or abiraterone FASN/AR-V7 double-positive metastases were found in 77% of cases. These findings provide a compelling rationale for the use of FASN inhibitors in mCRPCs, including those overexpressing AR-V7.

Description: IPI-504 is a novel inhibitor of Hsp90 based on the geldanamycin pharmacophore. When placed in rat, monkey, and human blood, IPI-504 rapidly converts to the known and well-studied compound 17-allylamino-17-demethoxy-geldanamycin (17-AAG). 17-AAG is the subject of multiple clinical trials for the treatment of hematologic and solid tumors. However, 17-AAG suffers from poor aqueous solubility necessitating the use of sub-optimal formulations to deliver this agent to patients. IPI-504 is over 1000-fold more soluble than 17-AAG in aqueous solution. In vitro, both 17-AAG and IPI-504 bind tightly to, and selectively inhibit Hsp90 derived from cancer cells. The cytotoxic effect of IPI-504, as well as its ability to stimulate the degradation of Hsp90 client proteins and increase the intracellular levels Hsp70, were monitored in two human multiple myeloma cells lines (RPMI-8226 and MM1.S). The effects of IPI-504 were compared to 17-AAG. We demonstrate that the actions of IPI-504 are bioequivalent to 17-AAG and that both compounds induce apoptosis in these cells and stimulate the degradation of HER2 and c-Raf. In addition, both agents stimulate Hsp70 protein levels. In all cases the EC50s are virtually the same for both molecules (~200–400 nM). Furthermore, IPI-504 inhibits the secretion of immunoglobulin light chain from the RPMI-8226 multiple myeloma cells (EC50 ~300 nM). Importantly, IPI-504 is active in tumor xenograft models of multiple myeloma. The data indicate that active metabolites of IPI-504 accumulate in these xenografts long after these metabolites are cleared from the plasma compartment, suggesting that they preferentially accumulate in tumor cells based on their increased affinity to Hsp90 derived from tumor cells. In conclusion, we have developed IPI-504 as a novel, potent inhibitor of Hsp90 with greatly increased solubility over 17-AAG, and that IPI-504 is an active anti-tumor agent in vitro and in vivo.

Description: Background CD47 is over-expressed by many tumor types and protects tumor cells from destruction via tumor-intrinsic and -extrinsic means. The fully human anti-CD47 monoclonal antibody (mAb) SRF231 has previously been shown to block the "don't eat me" CD47/signal regulatory protein alpha (SIRPα) interaction and induce macrophage-mediated phagocytic uptake of CD47-expressing tumor cells, either alone or in the presence of anti-CD20 mAb. Furthermore, SRF231 inhibited tumor growth in preclinical models of aggressive non-Hodgkin lymphoma (Holland P, et al. ASH 2016). Here, we explored the activity of SRF231 against CLL cells for the first time, both as monotherapy and in combination with rituximab or venetoclax (VEN). Methods Peripheral blood mononuclear cells from 24 CLL patients were evaluated for CD47 surface expression by flow cytometry. Primary CLL or Jurkat target cells were treated ex vivo with SRF231 or isotype control and evaluated in phagocytosis and cell death assays. Human monocyte-derived macrophages were cocultured with fluorescently-labeled target tumor cells and exposed to SRF231 and/or rituximab (commercial supply). BH3 profiling was performed by gently permeabilizing primary CLL cells and measuring the release of cytochrome C (Cyto-C) in response to BH3-only peptides by flow cytometry. Priming for apoptosis was measured by Cyto-C release in response to BIM BH3 peptide, and pro-survival protein dependencies were measured by response to specific BH3-only sensitizer peptides. Statistical analyses were by unpaired and paired t-test with a two-tailed nominal p ≤ 0.05 considered as significant. In vivo antitumor activity was assessed using tumor xenograft studies in CB17 SCID mice. Mice with established, subcutaneous Ri-1 tumors were randomized and treated with either isotype control, SRF231, VEN (Medkoo), or combination of SRF231 and VEN. Results CD47 was expressed in all primary CLL cells (n = 24, median mean fluorescence intensity [MFI] 7913, range 3575-18,329) with a slightly higher expression in unmutated CLL (U-CLL) vs mutated CLL (M-CLL) samples (U-CLL median MFI = 9106, n = 8 vs M-CLL, median MFI = 7713, n = 14, 2 unknown, p = 0.047). Primary CLL cells were significantly more susceptible to phagocytosis upon ex vivo treatment with SRF231 in combination with rituximab (median % increase in phagocytosis over isotype control of 32.28% in the combination vs 11.78% with rituximab alone, n = 24, p 〈 0.0001). Upon coculture of Jurkat cells with macrophages, SRF231 not only induced phagocytosis (EC50, 332 ± 65 ng/mL, n = 3), but also induced cell death of non-phagocytosed target tumor cells (EC50, 295 ± 43 ng/mL, n = 3). While soluble SRF231 did not induce significant target tumor cell killing, immobilized SRF231 induced Jurkat cell and primary CLL cell death (median % alive of 34.6% in SRF231 treated cells vs 64.4% in controls, n = 24, p 〈 0.0001). To assess the mechanism of cell death induction, tumor cells were pretreated with a pan-caspase inhibitor, Z-VAD-FMK, which revealed that SRF231-mediated tumor cell death is caspase-independent. In primary CLL cells, BH3 profiling confirmed that SRF231 did not alter mitochondrial priming for apoptosis or pro-survival Bcl-2 family protein dependencies. Pre-treatment with the phospholipase C (PLC) inhibitor U73122 prior to SRF231 exposure partially blocked the ability of SRF231 to kill CLL cells (median % alive of 45.4% in pre-treated cells vs 25.4% in controls, n = 6, p = 0.0029). In addition to these in vitro studies, SRF231 displayed profound antitumor activity in a xenograft model of B-cell lymphoma as a single agent, and led to complete and durable tumor regression in combination with VEN. Conclusion Ex vivo treatment of primary CLL cells with SRF231 led to dual antitumor effects of tumor cell-extrinsic plus -intrinsic mechanisms by augmenting rituximab-induced phagocytosis and inducing tumor cell death. SRF231 induced death of tumor cells through a caspase-independent mechanism that depends at least partially on PLC. In vivo, SRF231 in combination with VEN led to complete and durable tumor regression in a xenograft model. SRF231 is currently being evaluated across multiple tumor types in a Phase 1 clinical trial (NCT03512340). Disclosures Valentin: Roche: Other: Travel reimbursement; AbbVie: Other: Travel reimbursement. Peluso:Surface Oncology: Employment, Equity Ownership. Adam:Surface Oncology: Employment, Equity Ownership. Zhang:Surface Oncology: Employment, Equity Ownership. Armet:Surface Oncology: Employment, Equity Ownership. Guerriero:GSK: Research Funding; Eli Lilly: Research Funding. Lee:Surface Oncology: Employment, Equity Ownership. Palombella:Surface Oncology: Employment, Equity Ownership. Holland:Surface Oncology: Employment, Equity Ownership. Paterson:Surface Oncology: Employment, Equity Ownership. Davids:Surface Oncology: Research Funding; Celgene: Consultancy; Verastem: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; MEI Pharma: Consultancy, Research Funding; Pharmacyclics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche/Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Research Funding; Astra-Zeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie, Inc: Consultancy, Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Merck: Consultancy; TG Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees.

Description: The transmembrane protein CD47 is an immunoglobulin superfamily member involved in multiple cellular processes, including cell migration, adhesion and T cell function. The interaction between CD47 and signal regulatory protein alpha (SIRPα), an inhibitory protein expressed on macrophages, prevents phagocytosis of CD47-expressing cells. CD47 was originally identified as a tumor antigen on human ovarian cancer and was subsequently shown to be expressed on multiple human tumor types, including both hematologic and solid tumors. In many hematologic cancers, high CD47 expression is associated with poor clinical outcomes. CD47 is also expressed at low levels on virtually all non-malignant cells, and loss of expression or changes in membrane distribution may serve as markers of aged or damaged cells, particularly on red blood cells (RBC). High expression of CD47 on tumors blocks phagocytic uptake, subsequent antigen cross-presentation and T cell activation, which collectively contribute to tumor immune evasion. Agents that block the CD47-SIRPα interaction can restore phagocytic uptake of CD47+ target cells and lower the threshold for macrophage activation, which can enhance the efficacy of therapeutic antibodies with ADCC-enabling activity. We developed and characterized a CD47 blocking antibody and evaluated its activity in multiple hematologic models. SRF231 is a fully human monoclonal antibody that binds with high affinity to human CD47 and blocks the CD47-SIRP▢ interaction. SRF231 promotes macrophage-mediated phagocytic clearance of several hematologic primary tumor samples and cell lines in vitro. For example, SRF231 increases phagocytosis of Raji tumor cell line targets with an EC50 of ~300 ng/ml. Enhanced phagocytosis is preferential for tumor cells over normal leukocytes and RBC. Tumor cell phagocytosis can be enhanced in the presence of opsonizing antibodies (e.g., anti-CD20 Ab) when co-administered with SRF231. In vivo efficacy of SRF231 was assessed in several preclinical murine xenograft models of hematologic malignancies. Notably, SRF231 administration led to profound tumor growth inhibition in models of multiple myeloma, diffuse large B cell lymphoma and Burkitt's lymphoma as a single agent and in combination with opsonizing antibodies. In the Raji xenograft model, single agent therapy leads to 112% tumor growth inhibition. This anti-tumor activity is at least partially dependent on macrophages, as depletion of macrophages via clodronate administration leads to reduced tumor growth inhibition. Tumor-associated macrophage (TAM) numbers and polarization status are also modulated by SRF231 treatment. In summary, the CD47 mAb SRF231 induces robust tumor cell phagocytosis and tumor clearance both alone and in combination with opsonizing antibodies in pre-clinical models of myeloma and lymphoma. These properties warrant further development of SRF231 in hematologic malignancies. SRF231 is currently in IND-enabling studies and is expected to enter clinical trials in 2017. Disclosures No relevant conflicts of interest to declare.