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Mutations in the androgen receptor (AR) ligand-binding domain (LBD) can cause resistance to drugs used to treat prostate cancer. Commonly found mutations include L702H, W742C, H875Y, F877L and T878A, while the F877L mutation can convert second-generation antagonists such as enzalutamide and apalutamide into agonists. However, pruxelutamide, another second-generation AR antagonist, has no agonist activity with the F877L and F877L/T878A mutants and instead maintains its inhibitory activity against them. Here, it is shown that the quadruple mutation L702H/H875Y/F877L/T878A increases the soluble expression of AR LBD in complex with pruxelutamide in Escherichia coli. The crystal structure of the quadruple mutant in complex with the agonist dihydrotestosterone (DHT) reveals a partially open conformation of the AR LBD due to conformational changes in the loop connecting helices H11 and H12 (the H11–H12 loop) and Leu881. This partially open conformation creates a larger ligand-binding site for AR. Additional structural studies suggest that both the L702H and F877L mutations are important for conformational changes. This structural variability in the AR LBD could affect ligand binding as well as the resistance to antagonists.

Purpose To evaluate GT0918, a 2nd-generation AR antagonist, for its AR down-regulation activity among breast cancer patients. Methods The effect of GT0918 on AR protein expression was evaluated in AR expression breast cancer cells and in breast cancer xenograft model. A 3 + 3 phase I dose-escalation study was launched in Peking University Cancer Hospital. The endpoints included dose finding, safety, pharmacokinetics, and antitumor activity. Results GT0918 was demonstrated to effectively suppress the expression of AR protein and the growth of AR-positive breast cancer tumors in mouse xenograft tumor models. All patients treated with GT0918 were at a QD dose-escalation of five dose levels from 100 to 500 mg. The most common treatment-related AEs of any grade were asthenia, anemia, decreased appetite, increased blood cholesterol, increased blood triglycerides, decreased white blood cell count, and increased low-density lipoprotein. Grade 3 AEs were fatigue (2 of 18, 11.1%), aspartate aminotransferase increase (1 of 18, 5.6%), alanine aminotransferase increase (1 of 18, 5.6%), and neutrophil count decrease (1 of 18, 5.6%). Clinical benefit rate (CBR) in 16 weeks was 23.1% (3/13). Among 7 AR-positive patients, 6 can evaluate efficacy, and 2 completed 23.5- and 25-cycle treatment, respectively (as of 2020/1/20). PK parameters showed a fast absorption profile of GT0918 in the single-dose study. GT0918 and its major metabolite reached steady-state serum concentration levels at day 21 after multiple dosing. Conclusion GT0918 can effectively inhibit AR-positive breast cancer tumor growth. GT0918 was demonstrated well tolerated with a favorable PK profile. The suitable dose of GT0918 was 500 mg QD and may provide clinical benefits for AR-positive mBC.

Coronavirus disease 2019 (COVID-19) has caused the public health crisis in the whole world. Anti-androgens block severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry and protect against severe clinical COVID-19 outcomes. GT0918, a novel androgen receptor antagonist, accelerated viral clearance and increased recovery rate in outpatients by blocking SARS-CoV-2 infection though down-regulating ACE2 and TMPRSS2 expression. Further clinical study showed that GT0918 reduced mortality rate and shortened hospital stay in hospitalized COVID-19 patients. GT0918 also exhibits protective efficacy in severe COVID-19 patient in critical care. However, the mechanism of GT0918 treatment for severe COVID-19 disease is unknown. Here, we found GT0918 decreased the expression and secretion of proinflammatory cytokines through NF-κB signaling pathway. The acute lung injury induced by LPS or Poly(I:C) was also attenuated in GT0918-treated mice, compared with vehicle control group. Moreover, GT0918 elevated the NRF2 protein level but not mRNA transcription activity. GT0918 induced proinflammatory cytokines downregulation was partially dependent on NRF2. In conclusion, our data demonstrate that GT0918 reduced cytokine release and suppressed inflammatory responses through inhibiting NF-κB signaling and activating NRF2. GT0918 is not only effective for treatment of mild to moderate COVID-19 patients, but also a potential therapeutic drug for severe COVID-19 patients by reducing the risk of cytokine storm and acute respiratory distress syndrome.

Acute Myeloid Leukemia (AML) is the third hematological malignancies with the worst relative overall 5-year survival rate (11.7%) in hematological malignancies. AML is a heterogeneous disease with a broad spectrum of genomic changes and molecular mutations that lead to a poor prognosis and clinical outcome. Leukemic stem cells progress to myoblasts that continue to proliferate without differentiating, namely, immature blasts in AML. The hedgehog (HH)/glioma-associated oncogene homolog (GLI) signaling pathway is essential for embryonic and stem cell developments. This pathway has been one of the most promising targets for drug discovery and developments for AML. Although HH inhibitor Glasdegib in combo with low-dose cytarabine achieved FDA approval for AML, Venetoclax (BCL-2 inhibitor/ABT-199) plus a hypomethylating agent (HMA) have been dominating the regimens in AML recently. Here we reported GT1708, a HH inhibitor, improves ABT-199 (venetoclax)-induced apoptosis by down-regulating MCL-1 proteins in AML cells. GT1708 is a potent HH inhibitor (IC50=0.11 nM in HH pathway-driven cellular assay) and inhibited GLI expression with doses of 1, 3 and 10 mpk in a HH-dependent medulloblastoma animal models. In Molm-13 (AML) cells, GT1708 was shown to down-regulate the expression of MCL-1 proteins (anti-apoptotic proteins). In contrast, ABT-199 increased the expression of MCL-1. Furthermore, ABT+Aza (Azacidine/HMA drug) induced more MCL-1 expression than ABT-199. Importantly, GT1708 was shown to induce the expression of cleaved-PARP (c-PARP/apoptotic marker) and to increase c-PARP expression when combined with ABT-199. Due to ABT+Aza induced MCL-1 overexpression, the combination of both agents failed to induce c-PARP, suggesting MCL-1 overexpression conferring resistance to ABT+Aza therapy in Molm-13 cells. GT1708 was further evaluated in flow cytometry-based apoptotic assays. GT1708, ABT-199 and Aza were demonstrated to induce an early apoptosis by 9.98%, 47.5%, and 8.64%, respectively. In comparation, ABT+GT1708 were revealed to induce a 61% of apoptosis superior to 47% or 48.5% of apoptosis induced by either ABT or ABT+Aza in Molm-13 cells. These results confirm the role of MCL-1 overexpression in conferring resistance to ABT+Aza therapy, which can be overcome by ABT+GT1708 combo. GT1708/ABT combo were also shown a marginable superior antitumor activity than ABT along in Molm-13 animal models. GT1708 has been testing in a phase I study in AML patients with previous multiple lines of regimens. GT1708 has been shown to reduce blast counts in three of 13 AML patients treated with higher doses and demonstrated favorite PK and safety profiles. In brief, these results support the clinical development of GT1708 in combination with ABT-199 in AML patients. Citation Format: Liandong Ma, Qianxiang Zhou, Honghua Yan, Min Dong, Xiahe Han, Jiangwie Li, Jie Qu, Weidong Qian, Youzhi Tong. Combination of a clinical stage-hedgehog inhibitor, GT1708, improves Venetoclax-induced apoptosis by down-regulating MCL-1 proteins in AML cells. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4978.

Myc family members (MYC, MYCN and MYCL) are oncogenic transcriptional factors, which form dimers with Max to activate transcription activities to drive tumor initiation and progression. c-Myc deregulation has been identified in 80% of all tumor types, including B-cell malignancies, AML and a variety of solid tumors. N-Myc genetic alterations have been identified in small cell lung cancer, neuroblastoma, neuroendocrine prostate cancers and sarcoma. Myc-deregulation has been directly linked to the poor clinical outcome in these cancers, which makes Myc a therapeutic target for pharmacological inhibition. Here we described GT19630, a c-Myc/n-Myc degrader. GT19630 selectively degraded c-Myc proteins in c-Myc dependent blood cancer cells (IC50=1.5 nM) as compared to growth-factor regulated c-Myc in hematopoietic progenitor cells (TF-1) (IC50=52.5 nM). Similar selectivity of GT19630 has been demonstrated in cell proliferation and granulocyte–macrophage progenitor colony forming unit (GM-CFU) assays with the IC50s of 26.2 and 39.0 nM, respectively. GT19630 was shown to degrade c-Myc via proteasome degradation system and degraded CRBN-dependent endogenous neo-substrates of GSPT1, CK1α and IKZF1. GT19630 reduced transcriptional factors including c-Myc, Max, MXI1, SWF/SNF family members/associated proteins; ARID1A, SMARCE1, and SMARCC1 in transcriptional factor response element (TFRE) assays. Moreover, GT19630 inhibited the cell proliferation with IC50 Citation Format: Liandong Ma, Yuzhi Tong, Zhaohui Yang, Qianxiang Zhou, Honghua Yan, Ye Chen, Dong Chen, Ru Xu, Yini Wang, Jun Qin. Discovery and evaluation of GT19630, a c-Myc/n-Myc degrader, for targeting c-Myc-driven B-cell malignancies, acute myeloid leukemia (AML) and n-Myc driven cancers [abstract]. In: Proceedings of the Third AACR International Meeting: Advances in Malignant Lymphoma: Maximizing the Basic-Translational Interface for Clinical Application; 2022 Jun 23-26; Boston, MA. Philadelphia (PA): AACR; Blood Cancer Discov 2022;3(5_Suppl):Abstract nr A03.

c-Myc is an oncogenic transcriptional factor driving tumor initiation, progression and poor prognosis in 80% of all tumor types, especially in B-cell malignancies and small cell lung cancer (SCLC) with Myc genomic alterations. Myc dysregulation have been directly linked to the poor clinical outcome in these cancers. Therefore, it is highly warranted to discover and develop novel Myc therapeutical agents for targeting Myc driven cancers. Here we described GT19630, a GSPT1/Myc CELMoD. GT19630 was discovered through an SAR effort for c-Myc degrader by using c-Myc ELISA and Western blot assays in c-Myc driven HL60 AML cells. GT19630 selectively degraded c-Myc proteins in HL60 cells (IC50=1.5 nM) as compared to growth-factor regulated c-Myc erythroid progenitor cells (TF-1 cells) with IC50=52.5 nM. GT19630 also selectively inhibited HL60 cell proliferation (IC50= 0.33 nM), as compared to its IC50 (26.2 nM) in GM-CSF-TF-1 proliferation, as well as in bone marrow colony-forming cell assays (myeloid=40.2 nM), suggesting>100X selectivity for HL60 cells over normal blood cells. Through the SAR for Myc degrader, GT19630 has been evolved as a leading molecule sharing chemical properties to CELMoDs. Therefore, this compound was further evaluated by proteomics and western blot, GT19630 was confirmed to selective degradation of CELMoD targets, GSPT1/GSPT2 (translation termination factor G1 to S phase transition proteins 1 and 2) with IC90 Citation Format: Liandong Ma, Youzhi Tong, Zhaohui Yang, Qianxiang Zhou, Hongha Yan, Ru Xu, Jie Chen, Jie Pan, Huiyuan Wang, Jiangwei Li, Dong Chen, Xiang Cai, Jie Qu, Yini Wang, Jun Qin, Yuki Nishida, Michael Andreeff, Qilnli Guo, Yuki Nishida, Michael Andreeff. Discovery and evaluation of GT19630, a c-Myc/GSPT1 cereblon E3 ligase modulator (CELMoD), for targeting Myc-driven blood cancers and small cell lung cancers (SCLC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5479.

As of November 2021, there were 21 million confirmed active cases of COVID-19, including 77,016 patients in serious or critical condition (virusncov.com). However, there are no effective oral drugs for the treatment of severe COVID 19 patients. We here discuss the mechanism of action for Proxalutaminde to treat mild, moderate and severe COVID-19 Patients. Cellular entry and infection of SARS-CoV-2 virus are mediated by two key proteins in host cells, angiotensin converting enzyme 2 (ACE2), a host transmembrane protein, providing the binding sites for SARS-CoV-2 on the host cell surface, and transmembrane protease serine 2 protein (TMPRSS2), priming the S protein of SARS-Cov-2 to facilitate the viral entry into the host cells. Both ACE2 and TMPRSS2 proteins are regulated by androgen receptor (AR) signaling. Previously, Proxalutamide has been reported to downregulate the expression of ACE2 and TMPRSS2 in cells derived from prostate, lung cancer and normal lung epithelial cells. In this study, we demonstrate that Proxalutamide inhibited the infection of SARS-COV-2 wild type, alpha and delta variants, with IC50s of 69, 48 and 39 nM, respectively. Moreover, Proxalutamide reduced SARS-COV-2 viral load in outpatients with COVID-19 (82% viral RT-PCR negative rate in active group vs. 31% in placebo group after treatment for 7 days (p-value Severe COVID-19 disease leads to cytokine storm resulting in pulmonary inflammation and extensive damage in lung and other organs. Anti-inflammatory drugs, including Baricitinib and dexamethasone, have shown limited clinical benefit for hospitalized COVID-19 patients. Therefore, more effective drugs are in urgent need for patients suffering from severe COVID-19. Recently, Proxalutamide has been reported to reduce the mortality rate (HR=0.16) and lung injury (by 57%, active drug vs placebo groups) in hospitalized patients with COVID-19 in an IIT phase III study. We presented here the mechanism of action of Proxalutamide for targeting cytokine storm in severe COVID-19 patients. Proxalutamide was demonstrated to activate nuclear factor erythroid 2-related factor 2 (Nrf2) in macrophages, which stimulates the antioxidant response element (ARE) for reducing cytokine storm-induced organ damage in COVID-19. In addition, Proxalutamide inhibited TNF alpha and IL-6 expression and blocked INF gamma signaling by downregulating STAT1 expression in immune cells. Importantly, Proxalutamide reduced inflammatory cells in lungs in a Poly (I:C), pseudoviral induced-lung injury animal models. Further, Proxalutamide decreased C-reactive protein, D-Dimer and improved lymphocyte count, biomarkers for COVID-19 progression in clinical studies. Together, these results provide a strong rationale for the treatment of severe COVID-19 patients with Proxalutamide. Citation Format: Liandong Ma, Youzhi Tong, Qianxiang Zhou, Hongha Yan, Xiaodan Hou, Zhiha Ren, Jiangwei Li, Huiyuan Wang, Weidong Qian, Yu Zhang, Andy Goren, Arul Chinnaiyan. Mechanisms of action (MOA) for proxalutamide, an androgen receptor (AR) antagonist, for the treatment of mild, moderate and severe COVID-19 patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5274.

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes a worldwide pandemic named coronavirus disease 2019 (COVID-19). The disease progression and mortality rates of patients with COVID-19 are higher in males than in females. Hence, we explored the potential mechanism of the COVID-19 gender disparity and searched for potential alternative therapeutic agents as unmet medical needs for male patients with COVID-19. Methods: We collected clinical data from 1339 COVID-19 patients and analyzed androgen receptor (AR) chromatin binding and expression patterns by ChIP-seq and scRNA-seq data. Moreover, we studied the androgen-induced expression of ACE-2 and TMPRSS2, two key proteins for SARS-CoV-2 cellular entry and infection and the blockage of these two key proteins with our AR antagonist Proxalutamide (GT0918) in normal lung cells, cancer cells derived from prostate and lung cancers. Furthermore, we examined the effect of GT0918 on inducible nitric oxide synthase (iNOS) and Tumor necrosis factor-alpha (TNF-α), the macrophage-activation markers, in mouse macrophage cells. Results: We demonstrated higher rates of disease progression and mortality of male COVID-19 patients. The mechanism of the gender disparity may result from the androgen-AR activation in male patients with COVID-19. We revealed that androgen-AR activation induced the expression of ACE2 and TMPRSS2 under the androgen-dependent condition in cells derived from prostate and lung cancer, which was inhibited by the blockage of AR signaling with AR antagonist GT0918. Importantly, GT0918 also inhibited the expression of macrophage activation markers iNOS and TNF-α, the biomarkers for macrophage polarization. These results support the role of androgen-AR signaling in the disease progression and mortality in male patients with COVID-19.

To date, as of 17 November 2020, there have been 55 million confirmed cases of COVID-19, including 1,4 million deaths globally, as reported to WHO. It is, therefore, very critical to discovery and development of the available treatment options for COVID-19. We here discuss the repurposing 2nd generation androgen receptor antagonist Proxalutaminde to treat COVID19. We analyzed the gender disparity of disease severity and progression in 1339 patients with COVID-19 and investigated the mechanism of gender disparity in male vs female patients with COVID 19. As androgen-androgen receptor pathway may contribute to the difference in severity and disease progression in male and female patients with COVID-19, we used cell lines derived from lung, prostate cancer and normal lung epithelial cells to determine the effect of androgen and androgen receptor antagonist Proxalutamide on the expression of two key proteins for SARS-CoV-2 to infect and enter the host cells. Angiotensin converting enzyme 2 (ACE2), a host transmembrane protein provides the binding sites for SARS-CoV-2 on the host cell surface, transmembrane protease serine 2 (TMPRSS2), a cellular serine protease, prims the S protein of SARS-Cov-2 to facilitate the viral entry into the host cells. As cytokine store plays a major role in the disease progression of COVID-19, we examined the effect of GT0918 on inducible nitric oxide synthase (iNOS) and Tumor necrosis factor-alpha (TNF-α), the macrophage polarization/activation markers, in mouse macrophage cells. In this study, we demonstrated higher rates of disease progression and mortality of male COVID-19 patients than female patients. Furthermore, we revealed that androgen-AR activation induced the expression of ACE2 and TMPRSS2 under the androgen-dependent condition in cells derived from prostate and lung cancer, which was inhibited by the blockage of AR signaling with Proxalutamide. Importantly, Proxalutamide also inhibited the expression of iNOS and TNF-α, the biomarkers for macrophage polarization/activation. These results support the role of androgen-AR signaling in the disease progression and mortality in male patients with COVID-19. We are currently conducting clinical study in COVID-19 patients with Proxalutamide in Brazil (NCT04446429). Citation Format: Liandong Ma, Youzhi Tong, Qiang Guo, Yifeng Zhou, Qianxiang Zhou, Honghua Yan. Repurposing 2nd generation androgen receptor antagonist Proxalutamide to treat COVID-19 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 711.

c-Myc is a transcriptional factor and an oncogene driver which plays a major role in tumor initiation, progression and poor prognosis in 80% of all tumor types, especially in c-Myc-additive B-cell and myeloid malignancies. c-Myc's critical role as an oncoprotein makes it an attractive target for pharmacological inhibition. However, c-Myc has been considered as “an undruggable” target since it is an intrinsically disordered protein and regulates key physiological functions in normal cells, which can be a liability for effectively pharmacological intervention. Thus, effective and selective c-Myc targeted molecules are highly warranted, but no such inhibitors are now in clinical testing. c-Myc is dependent on the heterodimerization with Max for the stability and activation of transcription. Here we describes a c-Myc/Max PPI small molecule inhibitor, GT-19077, selectively degrades c-Myc and inhibits the proliferation of c-Myc-addictive B-cell and myeloid malignant cells, but spares c-Myc and the cell proliferation in JAK2-STAT5 dependent erythroblast cells. Through the conformation generation/binding site predication, virtual screening of molecules (>200K), optimization and experimental validation, a leading c-Myc binding molecule series was identified. GT19077 was then discovered and optimized as a leading molecule that exhibited a direct c-Myc binding activity in HL-60 myeloid leukemia cells as shown in Cellular Thermal Shift Assay (CETSA). Importantly, GT19077 selectively degraded c-Myc in HL-60 cells with DC50 of 0.40 µM vs. DC50 of 2.31 µM in GM-CSF-JAK2-STAT5 dependent TF-1 erythroblast cells (>5 fold) as measured by ELISA assays. Consistently, GT19077 also selectively inhibited the proliferation of HL-60 cells with the IC50 of 0.34 µM vs. IC50 of 1.6 µM in GM-CSF stimulated TF-1 cells (>5 fold). Profiling on the proliferation of a selected 14 hematologic malignant cell panel with c-Myc overexpression, GT19077 selectively inhibited the proliferation of all seven B-cell malignant cell lines tested, which include three carrying Myc-IGH translocations and two expressing Kras G12A mutations, with IC50s as low as 160 nM. In addition, two myeloblast cell lines and one monoblast cell line were also sensitive to GT19077 with IC50 of 300-700 nM. In contrast, GT19077 showed almost no effect on the proliferation of four leukemia cell lines with known oncogene drivers, including two erythroblasts (JAK2-STAT5 dependent), one macrophage (FGFR1OP2-FGFR1) and one myeloblast (AML-ETO). These results suggest that c-Myc inhibitor GT19077 can selectively target c-Myc-addictive B-cell and myeloid malignancies, but spares the liability on JAK2-STAT5 driven erythroblasts. Moreover, GT19077 has almost no effect on hematologic malignancies with different oncogene drivers that can be used as biomarkers for patient stratification. Further, GT19077 demonstrated a favorable PK profile with the bioavailability of 57.3% in rat by intraperitoneal (IP) administration. Currently GT19077 is being evaluated in B-cell and myeloid malignant animal efficacy models. Since this novel c-Myc inhibitor demonstrates a superior potency and a unique electivity profile, as compared to many experimental c-Myc inhibitors, it has potential for the drug development for targeting c-Myc-addictive B-cell and myeloid malignancies. Citation Format: Liandong Ma, Youzhi Tong, Zhaohui Yang, Qian Xiang Zhou, Jie Chen, Ye Chen, Honglei Bi, Jie Pan, Qi Sun, Ru Xu, Luhua Lai. GT19077, a c-Myc/Max protein-protein interaction (PPI) small molecule inhibitor, selectively targets c-Myc-additive B-cell and myeloid malignancies [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-111.

Reactivation of androgen receptor (AR) plays a key role in prostate cancer growth, especially in castration-resistant prostate cancer (CRPC) progression. Accumulated data have demonstrated that the AR reactivation is resulted from AR amplification, AR overexpression and mutations in AR ligand binding domain (LBD) as well as AR splicing variants (AR-V7). This AR reactivation also drives the resistance to AR targeted therapies, including 1st and 2nd generations of AR antagonists, such as Bicalutamide (Bica) and Enzalutamide (Enza, MDV3100). Therefore, the development of novel and effective AR antagonists is in urgent need for overcoming AR reactivation for patients with metastatic castration-resistant prostate cancer (mCRPC). Here we report a novel AR binding pose of Proxalutamide (GT0918), an AR antagonist, in silico of drug’s mode of action. We also reveal the effects of GT0918 on the transactivation of the wild type AR (wtAR) and AR mutants with comparisons to 1st and 2nd generations of AR antagonists (Bicalutamide and Enzalutamide). GT0918 was reported to have higher binding affinity than Bica (11.4X) and Enza (3.5X) in AR ligand binding assays previously. To gain detail insights into the mode of action of GT0918 and Enza in AR LBD, structure-based computer modeling was performed. The mode of drug action shows that GT0918 binds to AR LBD pocket. This compound not only fits in the AR LBD pocket very well, but also its structure pushes AR Helix 12 away, resulting in the decreased interaction between AR Helix 12 and LBD pocket, which creates a unique and significant difference from other AR blockers. GT0918 was further assessed in luciferase-based AR transactivation assays. The results showed GT0918 significantly inhibited the androgen-induced transactivation of the wtAR. Importantly, it also blocked the transcriptional activity of tested mutant ARs arising to targeted AR therapies, including the AR mutant F877L that converts the Enzalutamide and Apalutamide (ARN-509) from antagonist to agonist activity. In clinical trial setting, some mCRPC patients progressed on Enzalutamide and bicalutamide showed stable disease and durable responses after received GT0918 from various doses. In conclusion, our results support the clinical development of GT0918 in prostate cancer patients who progressed on Enzalutamide or/and AR blockers. Citation Format: Liandong Ma, Qianxiang Zhou, Yang He, Jie Chen, Karl Zhou, Qingqing Zhou, Chunyun Chen, Jason Shaoyun Xiang, Xiaobing Deng, Luhua Lai, Youzi Tong. Proxalutamide (GT0918), a novel androgen receptor (AR) antagonist, targeting resistance mechanisms to AR signaling-directed castration-resistant prostate cancer (CRPC) therapies [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 3861.

Background: MET amplification (amp) is a resistance mechanism to EGFR TKI treatment. Emibetuzumab, a bivalent MET antibody (Ab) blocks HGF binding to MET and internalizes the receptor. Combination of emibetuzumab with EGFR TKIs (erlotinib, AZD9291, CO1686) or EGFR Ab (necitumumab, cetuximab) was evaluated in 3 ER xenograft models. Methods: Model 1: ER cell line HCC827ERL with high focal MET amp, high pMET, EGFR ex19 del (no T790M) was created from parental HCC827 NSCLC (EGFR ex19 del, EGFR amp, no MET amp) by increasing concentration of erlotinib in vitro over 7 months. Model 2: ER cell line HCC827-A8 was derived from HCC827 parental xenograft tumor serially passed in vivo with long term treatment of gefitinib and erlotinib. HCC827-A8 cells express high focal MET amp, high pMET/AXL (Western blot) while retaining EGFR ex19 del (no T790M). Model 3: LU0858 was an ER patient-derived NSCLC xenograft tumor, with focal MET amp, EGFR L858R (no T790M). MET amp and EGFRmt was determined by FISH and LNA-PCR sequencing respectively. Compound dosing: emibetuzumab 20 mg/kg qw; necitumumab 4 mg/kg or 20 mg/kg biw; cetuximab 4 mg/kg biw; erlotinib 25 mg/kg qd; 5 mg/kg AZD9291 qd; 30 mg/kg CO1686 bid. Results: EGFR inhibitors, but not emibetuzumab showed significant single agent anti-tumor effect in xenograft tumors derived from non-MET amp HCC827 parental cells. In MET amp ER models, single agent emibetuzumab resulted in tumor growth inhibition in Model 1 (T/C= 51.7%-61.0%, p Conclusion: The three erlotinib resistant models with MET amp and retaining sensitizing EGFRmt (ex19 del or L858R), and no acquired T790M were found resistant to other EGFR inhibitors (Abs and TKIs). Emibetuzumab in combination with either EGFR TKI or Ab showed anti-tumor activity in MET amp ER xenograft models including tumor regression in 2 out of 3 models. The combination of emibetuzumab with erlotinib is being evaluated in NSCLC patients with EGFR activating mutation (NCT01897480). Citation Format: Suzane L. Um, Victoria L. Peek, Jennifer R. Stephens, Jessica A. Baker, Holly K. Cannon, Joel D. Cook, Isabella H. Wulur, Roger Agyei, Sudhakar Chintharlapalli, Robert J. Evans, William J. Feaver, Lysiane Huber, Linda N. Lee, Ling Liu, Liandong Ma, Ruslan Novosiadly, Volker Wacheck, Sau-Chi Betty Yan. Antitumor activity of MET antibody emibetuzumab (LY2875358) in combination with EGFR inhibitors in erlotinib resistant (ER) xenograft mouse models [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 519. doi:10.1158/1538-7445.AM2017-519