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Direct pharmacological inhibition of RAS has remained elusive, and efforts to target CRAF have been challenging due to the complex nature of RAF signaling, downstream of activated RAS, and the poor overall kinase selectivity of putative RAF inhibitors. Herein, we describe 15(LXH254, Aversa, R.; et al. Int. Patent WO2014151616A1, 2014), a selective B/C RAF inhibitor, which was developed by focusing on drug-like properties and selectivity. Our previous tool compound, 3(RAF709; Nishiguchi, G. A.; et al. J. Med. Chem.2017, 60, 4969), was potent, selective, efficacious, and well tolerated in preclinical models, but the high human intrinsic clearance precluded further development and prompted further investigation of close analogues. A structure-based approach led to a pyridine series with an alcohol side chain that could interact with the DFG loop and significantly improved cell potency. Further mitigation of human intrinsic clearance and time-dependent inhibition led to the discovery of 15. Due to its excellent properties, it was progressed through toxicology studies and is being tested in phase 1 clinical trials.

ABSTRACTAcross cultures and historical periods, people have attributed human traits to the divine. Because of the similarity between people’s mental representations of God and their mental representations of others, people’s perceptions of God may carry over to people’s perceptions of others, especially when people have recently thought about God. Two experiments examined whether priming God images through prayer leads people who believe in a benign God to view social targets in a more favorable light. In Experiment 1 (N = 57), Dutch Christians either prayed for or thought about a person, and then judged the emotions of others in the Reading the Mind in the Eyes Test. The results showed that prayer led participants to read fewer hostile emotions in others’ eyes, whereas prayer had no effect on perceiving positive emotions or non-hostile negative emotions. Experiment 2 (N = 50) extended this finding by showing that prayer only reduced social perceptions of hostility among participants with a positive God image. Thus, beliefs in a benign God may enhance interpersonal trust among believers, but only when God beliefs are cognitively accessible. These findings suggest that positive God beliefs may help to promote prosocial attitudes and cooperation within religious communities.

Currently, no approved monoclonal antibody (mAb) therapies exist for human multiple myeloma (MM). Here we characterized cell surface CS1 as a novel MM antigen and further investigated the potential therapeutic utility of HuLuc63, a humanized anti-CS1 mAb, for treating human MM. CS1 mRNA and protein was highly expressed in CD138-purified primary tumor cells from the majority of MM patients (more than 97%) with low levels of circulating CS1 detectable in MM patient sera, but not in healthy donors. CS1 was expressed at adhesion-promoting uropod membranes of polarized MM cells, and short interfering RNA (siRNA) targeted to CS1 inhibited MM cell adhesion to bone marrow stromal cells (BMSCs). HuLuc63 inhibited MM cell binding to BMSCs and induced antibody-dependent cellular cytotoxicity (ADCC) against MM cells in dose-dependent and CS1-specific manners. HuLuc63 triggered autologous ADCC against primary MM cells resistant to conventional or novel therapies, including bortezomib and HSP90 inhibitor; and pretreatment with conventional or novel anti-MM drugs markedly enhanced HuLuc63-induced MM cell lysis. Administration of HuLuc63 significantly induces tumor regression in multiple xenograft models of human MM. These results thus define the functional significance of CS1 in MM and provide the preclinical rationale for testing HuLuc63 in clinical trials, either alone or in combination.

Currently, no approved monoclonal antibody (mAb) therapies exist for human multiple myeloma (MM). Here we characterized cell surface CS1 as a novel MM antigen and further investigated the potential therapeutic utility of HuLuc63, a humanized anti-CS1 mAb, for treating human MM. CS1 mRNA and protein was highly expressed in CD138-purified primary tumor cells from the majority of MM patients (more than 97%) with low levels of circulating CS1 detectable in MM patient sera, but not in healthy donors. CS1 was expressed at adhesion-promoting uropod membranes of polarized MM cells, and short interfering RNA (siRNA) targeted to CS1 inhibited MM cell adhesion to bone marrow stromal cells (BMSCs). HuLuc63 inhibited MM cell binding to BMSCs and induced antibody-dependent cellular cytotoxicity (ADCC) against MM cells in dose-dependent and CS1-specific manners. HuLuc63 triggered autologous ADCC against primary MM cells resistant to conventional or novel therapies, including bortezomib and HSP90 inhibitor; and pretreatment with conventional or novel anti-MM drugs markedly enhanced HuLuc63-induced MM cell lysis. Administration of HuLuc63 significantly induces tumor regression in multiple xenograft models of human MM. These results thus define the functional significance of CS1 in MM and provide the preclinical rationale for testing HuLuc63 in clinical trials, either alone or in combination.

Introduction: HuLuc63 is a humanized monoclonal antibody that targets CS1 (CD2 subset 1, CRACC, SLAMF7, CD319), a cell surface glycoprotein that is highly and universally expressed on myeloma cells. In preclinical studies, we have shown that HuLuc63 treatment of mice with multiple myeloma (MM) xenograft tumors resulted in significant in vivoanti-tumor activity that is mediated at least in part by an antibody-dependent cellular cytotoxicity (ADCC) mechanism of action. The purpose of this study was to examine whether using HuLuc63 in combination with a panel of drugs having distinct modes of action (dexamethasone, thalidomide, bevacizumab (Avastin®), bortezomib (Velcade®)) could result in additional therapeutic benefit and provide a rationale for the design of future clinical trials.

Introduction: HuLuc63 is a humanized monoclonal antibody that targets CS1 (CD2 subset 1, CRACC, SLAMF7, CD319), a cell surface glycoprotein that is highly and universally expressed on myeloma cells. In preclinical studies, we have shown that HuLuc63 treatment of mice with multiple myeloma (MM) xenograft tumors resulted in significant in vivo anti-tumor activity that is mediated at least in part by an antibody-dependent cellular cytotoxicity (ADCC) mechanism of action. The purpose of this study was to examine whether using HuLuc63 in combination with a panel of drugs having distinct modes of action (dexamethasone, thalidomide, bevacizumab (Avastin®), bortezomib (Velcade®)) could result in additional therapeutic benefit and provide a rationale for the design of future clinical trials. Methods: HuLuc63 in combination with other agents was tested in vivo for anti-tumor activity using the human L363 and OPM2 xenograft models. SCID mice were implanted subcutaneously with myeloma cells and randomized into different groups (10–15 mice per treatment group) when the average tumor volume reached ∼100 mm3. HuLuc63 was administered via intra-peritoneal injection twice per week at doses of 1–10 mg/kg. Dosing for dexamethasone was 10 mg/kg twice weekly, thalidomide 50 mg/kg daily, bevacizumab twice weekly at 0.5 mg/kg, and bortezomib 1 mg/kg for two dosing cycles, each cycle consisting of twice weekly dosing for 2 weeks followed by a week of rest. Results: The combination of dexamethasone with HuLuc63 showed a statistically significant increase in anti-tumor activity over either agent alone (p < 0.04). Combination with thalidomide only showed a slight enhancement of tumor inhibition when dosed in combination with HuLuc63 but its anti-tumor activity did not reach a statistically significant increase in over that of HuLuc63 alone. Co-treatment of the anti-VEGF anti-angiogenic monoclonal antibody bevacizumab with HuLuc63 resulted in a significant increase in tumor inhibition (p < 0.05) over that observed with either antibody when used as a single agent. The strongest anti-myeloma activity was observed when HuLuc63 was combined with bortezomib, which appeared to result in a synergistic inhibition of tumor cell growth. None of the agents tested changed the CS1 expression level on the myeloma cells or diminished the anti-myeloma activity of HuLuc63. Conclusions: These results suggest that HuLuc63 may be combined with different classes of drugs to enhance its anti-myeloma effects. In particular, agents that may induce apoptosis of myeloma cells (dexamethasone and bortezomib) and anti-angiogenics (bevacizumab) may be of particular interest for future clinical testing. Further preclinical studies using HuLuc63 in combination with other agents are in progress. HuLuc63 is currently being evaluated in a phase I clinical study as monotherapy for the treatment of relapsed/refractory multiple myeloma.

Introduction: Current monoclonal antibody (mAb) therapies for multiple myeloma (MM) have had limited success due to narrow target expression across MM patient samples. A preferred strategy would be to develop cytotoxic human mAbs against novel antigens that are highly expressed in MM cells yet have limited expression in other cell types. CS1 (CD2 subset 1, CRACC, SLAMF7), a member of the CD2 family of cell surface glycoproteins, was found to be highly expressed in myeloma cells. In this study, we investigated the anti-myeloma activity of HuLuc63, a novel humanized anti-CS1 mAb. Methods: Microarray expression profiling was used to determine the CS1 mRNA levels in CD138-expressing myeloma cells from 101 MM patient samples. For detection of CS1 protein, flow cytometry was performed using the anti-CS1 mAb HuLuc63. Functional characterization of HuLuc63 was performed by assessing antibody-dependent cellular cytotoxicity (ADCC) and by assessing MM and bone marrow stromal cell (BMSC) interactions. Results: CS1 mRNA was expressed in CD138 cells from more than 96% (97/101) of MM patients. Flow cytometric analysis confirmed that protein expression mirrors the mRNA profile. Importantly, CS1 is also present in 12 MM cell lines that are either drug-sensitive or resistant. HuLuc63, but not an isotype control antibody, induced ADCC in a CS1-specific, dose-dependent manner against CD138-expressing MM lines and patient MM cells including dexamethasone (dex)-sensitive MM1S and dex-resistant MM1R cells. Significantly, HuLuc63 triggered autologous ADCC against CS1-expressing CD138-purified tumor cells from 11 MM patients resistant to conventional or novel therapies such as bortezomib (Velcade®) and an HSP90 inhibitor. Since CS1 may regulate cell adhesion, we next studied whether HuLuc63 alters MM cell adhesion to BMSCs. HuLuc63 inhibited MM cell adhesion to BMSCs in a dose-dependent manner, whereas human control IgG did not. However, the presence of BMSC appeared to reduce HuLuc63-induced cell lysis against MM1S and MM1R cells. Since the immunomodulatory drug lenalidomide (Revlimid®) enhances NK cell function, we further tested whether HuLuc63-induced ADCC against MM cells is augmented by lenalidomide. Pretreatment with lenalidomide markedly enhanced NK-cell-mediated lysis of autologous patient MM cells triggered by HuLuc63. Conclusions: We show that the new MM antigen, CS1, is expressed in myeloma cells from more than 96% of MM patients. The novel humanized anti-CS1 mAb, HuLuc63, induced significant cytotoxicity against MM cells including drug-resistant cells, and inhibited their interaction with BMSCs. These data suggest that HuLuc63 may have clinical utility in a spectrum of MM patients including those newly diagnosed with the disease as well as patients with late stage refractory disease.

Introduction: We have recently shown that CS1 (CD2 subset 1, CRACC, SLAMF7), a cell surface glycoprotein of the CD2 family, is uniformly expressed on myeloma cells from multiple myeloma (MM) patients. Based on its high expression in MM and limited expression in normal cells, we propose CS1 as a novel and specific antibody target for the treatment of MM. Methods: A panel of monoclonal anti-CS1 antibodies (mAbs) was generated to identify a potential therapeutic candidate. MAb clones MuLuc63 and MuLuc90 were selected for testing in CS1 positive MM xenograft models in vivo in severe combined immunodeficient mice. HuLuc63, a humanized IgG1 version of MuLuc63, was generated as the potential therapeutic candidate for the treatment of MM. HuLuc63 and Fc-modified versions of HuLuc63 were tested for anti-tumor activity in mouse models vivo. In vitro antibody-dependent cellular cytotoxicity (ADCC) assays were performed to define the potential mechanism of action for HuLuc63. Results: Both MuLuc63 and MuLuc90 exhibited significant in vivo anti-tumor activity compared to isotype control antibodies in the L363 MM xenograft model. MuLuc63 was significantly more potent, resulting in rapid tumor eradication in most of the animals for the length of the study (~4 months). Based on these results, MuLuc63 was humanized to generate HuLuc63, which exhibited similar affinity for CS1 when compared to the mouse parent antibody. In two different MM xenograft models, L363 and OPM2, HuLuc63 exhibited significant anti-tumor activity resulting in tumor eradication in a high proportion of animals. To investigate the mechanism of action, two modified versions of HuLuc63 were tested in xenograft models. One version, HuLuc63-Ala,Ala, exhibits a mutation in the Fc region that decreases the ability to interact with the Fc receptor on natural killer (NK) cells. The second version, HuLuc63-LF, exhibits low levels of fucosylation in the Fc region that would result in increased binding to the Fc receptor. Compared to HuLuc63, the LF version exhibited significantly better in vivo anti-tumor activity towards, while the Ala,Ala mutant exhibited no anti-tumor activity. These data indicate that the Fc region of HuLuc63 is critical for its anti-tumor activity, and suggest ADCC as a possible mechanism of action. In vitro, HuLuc63 exhibits substantial ADCC towards L363 and OPM2 cells. The activity was dose-dependent, with increasing cytotoxicity being observed with concentrations ranging from 0.01µg/mL to10 µg/mL. Conclusions: These pre-clinical data support HuLuc63 as a new therapeutic for the treatment of MM and suggest that ADCC is part of the mechanism of action. HuLuc63 will be entering a phase I clinical study for multiple myeloma.

Introduction:We have recently shown that CS1 (CD2 subset 1, CRACC, SLAMF7), a cell surface glycoprotein of the CD2 family, is uniformly expressed on myeloma cells from multiple myeloma (MM) patients. Based on its high expression in MM and limited expression in normal cells, we propose CS1 as a novel and specific antibody target for the treatment of MM.

Introduction: Current monoclonal antibody (mAb) therapies for multiple myeloma (MM) have had limited success due to narrow target expression across MM patient samples. A preferred strategy would be to develop cytotoxic human mAbs against novel antigens that are highly expressed in MM cells yet have limited expression in other cell types. CS1 (CD2 subset 1, CRACC, SLAMF7), a member of the CD2 family of cell surface glycoproteins, was found to be highly expressed in myeloma cells. In this study, we investigated the anti-myeloma activity of HuLuc63, a novel humanized anti-CS1 mAb.