bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5363: The preclinical selectivity and activity of APS03118, a highly selective and potent next-generation RET inhibitor
Authors: Drilon Alexander , Zhong Jun , Lu Ying , Liu Yongbo , Wang Hao , Chen Minchun , Chen Xiaohu , Zhu John , Lu Shun , Subbiah Vivek
Published: June 2022

Backgroud: Oncogenic RET is an actionable target across a variety of cancers. Selective RET inhibitors selpercatinib and pralsetinib were recently approved by the FDA and EMA for patients with RET-dependent NSCLC and thyroid cancers. The solvent front mutations (SFMs) RET G810C/S/R have been identified as mechanisms of acquired resistance to both drugs. The gatekeeper RET V804 mutation cannot be inhibited by selected next-generation RET inhibitors in development. APS03118 is a novel next-generation RET inhibitor which is potent against a range of RET fusions and mutations including both SFMs and gatekeeper mutations.

Methods: The selectivity, anti-RET activity, and intracranial efficacy of APS03118 were confirmed in vitro and in vivo in a variety of RET-dependent tumor models.

Results: APS03118 was highly selective against a panel of 468 kinases and demonstrated 130-fold selectivity over VEGFR-2. In enzymatic assays, APS03118 showed low nanomolar potency against wild type RET and 25 RET mutations/fusions, including the inhibition of RET G810R/C/S (IC50 0.04-5 nM) and RET V804M/L/E (IC50 0.04-1 nM). APS03118 inhibited RET phosphorylation (IC50 <15 nM) in Ba/F3 engineered RET cells (WT, G810R, V804M, M918T). In cell proliferation assays, APS03118 potently inhibited KIF5B-RET Ba/F3 (WT, V804M, V804L, M918T), CCDC6-RET Ba/F3 (WT, V804M, S904F), LC2/ad (CCDC6-RET), TT (RET C634W) (IC50 < 10nM); Ba/F3 RET G810R and G810S IC50 (8-65 nM). APS03118 demonstrated marked anti-tumor efficacy in vivo in RET-driven cell-derived (Ba/F3 KIF5B-RET, V804M, TT (C634W)) and patient-derived (KIF5B-RET, CCDC6-RET, CCDC6-RET V804M) xenograft tumor models. At 10 mg/kg BID, tumor regression was observed in these xenograft models (TGI 87-108%). Tumors completely subsided in CCDC6-RET orthotopic brain model with a 100% survival rate. In the Ba/F3 KIF5B-RET G810R xenograft model, APS03118 30 mg/kg BID showed 90% TGI and was well tolerated, and RET G810 mutations often drive clinical progression on current RET inhibitors.

Conclusions: APS03118 is a novel highly selective next-generation RET inhibitor that possesses potent in vitro and in vivo activity against a diverse range of RET alterations, including SFMs-mediated resistance. A first-in-human phase 1 trial for patients with RET-driven solid tumors with activating RET alterations is planned for 2022.

Citation Format: Alexander Drilon, Jun Zhong, Ying Lu, Yongbo Liu, Hao Wang, Minchun Chen, Xiaohu Chen, John Zhu, Shun Lu, Vivek Subbiah. The preclinical selectivity and activity of APS03118, a highly selective and potent next-generation RET inhibitor [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 5363.
Abstract 5387: Investigating the role of p53 in doxorubicin-mediated lysosome membrane permeabilization in breast cancer cells
Authors: Nicoletto Rachel , Ofner Clyde M.
Published: June 2022

Background: Doxorubicin (DOX) is a chemotherapeutic characterized by multiple mechanisms. Wild-type p53 (WTp53) may be activated in the presence of either DNA damage or ROS. Upon activation, WTp53 may upregulate the transcription of pro-apoptotic Bcl-2 family members capable of triggering mitochondrial membrane permeabilization or lysosome membrane permeabilization (LMP), ultimately resulting in apoptosis. We have previously reported of DOX-induced LMP between WTp53 and mutant p53 (MUTp53) breast cancer cells. The goals of this study were to establish (1) the role of p53 in the initiation of DOX-mediated LMP and (2) whether p53 is critical to the onset of cytotoxicity.

Methods: Studies were conducted using DOX at a range of concentrations and durations in both WTp53 (MCF-7 and MDA-MB-361) and MUTp53 (MDA-MB-231 and T47D) breast cancer cells. LMP was demonstrated using fluorescence microscopy as the cytosolic release of a 10 kDa AlexaFluor-488 Dextran conjugate that was preloaded into lysosomes. Viability was determined by MTT. Cytotoxicity was determined by trypan blue. Experiments were conducted in the presence of pifithrin-α p53 inhibitor to establish the importance of p53 towards DOX cytotoxicity.

Results: Significant LMP was determined as early as 0.5 h in both of the WTp53 cell lines at 10 μM. MDA-MB-361 cells demonstrated a greater degree of LMP with 1 μM DOX showing significant LMP by 2 h, an effect that was not observed in MCF-7 cells until 6 h. The earliest onset of notable LMP in MUTp53 MDA-MB-231 cells occurred at 6 h with 10 μM DOX only. T47D cells did not demonstrate notable LMP at this time. By 24 h, both WTp53 cell lines demonstrated LMP across all of the tested concentrations of DOX, while both MUTp53 cell lines only demonstrated notable LMP for 10 μM DOX. At this same time, notable reductions in cell viability for the tested range of concentrations were observed for all cell lines except for T47D cells. Overall, the inhibition of p53 reversed notable LMP in both MCF-7 and MDA-MB-361 cells across all of the tested conditions. Similarly, reduced viability was reversed following the inhibition of p53.

Conclusions: These results indicate a role of p53 in LMP in WTp53 cells. Furthermore, the combination of p53 and LMP seem necessary for the induction of cytotoxicity in WTp53 cells. Under circumstances of MUTp53 expression, cells promote a pro-oncogenic response with a lesser potential for LMP, MMP, and cytotoxicity.

Citation Format: Rachel Nicoletto, Clyde M. Ofner. Investigating the role of p53 in doxorubicin-mediated lysosome membrane permeabilization in breast cancer cells [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 5387.
Abstract 5397: Gene signatures for predicting patients with aggressive type of bladder cancer and shorter survivals
Authors: Sheu Jim Jinn-Chyuan , Lin Bo-Chen , Yang Oscar , Chang Chung
Published: June 2022

Bladder cancer is frequently associated with high recurrence and metastasis. With the success of cancer genome project in bladder cancer, we aimed to discover novel cancer drivers in muscle-invasive bladder cancer (MIBC), the most aggressive type of bladder cancer. A total of 155 genes were filtered out using RNAseq data from the TCGA cohort (n = 367), which are overexpressed (Z scores > 2.0) in more than 5% patients and correlated with poor overall survivals by Cox regression (p-value < 0.05) and FDR correction (q-value < 0.05). Using a unique HD-MAC (high-dimensional analysis of molecular alterations in cancer) algorithm developed by our team, 25 genes were further selected which show statistical significances as independent prognosis biomarkers for shorter overall survivals. Through clinical association study (including tumor stage, lymph node invasion, angiogenesis, and metastasis) by using CGDSR (Cancer Genomics Data Server in R), five most significant genes were further filtered which can be validated by another cohort (GSE13507). Although those genes show no direct gene-gene interactions, gene set enrichment analysis (GSEA) revealed common pathways governed by those driver genes, including cancer-related signaling, glycome alterations for TME remodeling, regulation in cell death, glutamate & neuronal signaling, and ER-Golgi transportation. The above findings can be also confirmed at the protein levels by IHC staining. In this study, how those key genes assist cell survival under nutrition crisis and their impacts on aggressive behaviors of bladder cancer will be discussed.

Citation Format: Jim Jinn-Chyuan Sheu, Bo-Chen Lin, Oscar Yang, Chung Chang. Gene signatures for predicting patients with aggressive type of bladder cancer and shorter survivals [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 5397.
Abstract 5404: Identification prognostic markers for patients with defective mismatch repair in colorectal cancer: MKI67, TPR,TCHH
Authors: Liu Zheng , Lv Jingfang , Zhao Hongmei , Wang Xishan , Wang Dongliang
Published: June 2022

Background: Deficiency in the mismatch repair system (dMMR/MSI-H) is found in approximately 15-20% of colorectal cancer. However, the prognostic evaluation of patients with dMMR CRC in stage II/III is still incomplete and lacks systematic studies.

Methods: In this study, we systematically analyzed the prognostic risk of 32 patients with dMMR CRC in stage II/III based on whole-exome sequencing and clinical characters. Differentially mutated genes (DMGs) were explored by fisher-test. For survival analysis, Kaplan-Meier survival curves were generated and compared using the log-rank test. Univariate and multivariate Cox regression analyses were performed to evaluate association DMGs with progression-free survival (PFS). Furthermore, these genes were verified in TCGA Pan Can Atlas 2018 cohort.

Results: The patients with dMMR CRC in stage II/III have the highest levels of MKI67, TCHH and TPR mutations (53.4%). Across all 73 patients with MSI-H, the mutational frequencies of MKI67, TCHH and TPR were (30%, 24% and 15%, respectively), which were associated with PFS (P=0.029, P=0.00261 and P=0.00298, respectively). That means the patients with MKI67, TCHH and TPR mutations have poor prognosis. Furthermore, MKI67, TCHH and TPR mutations were verified as independent factors for dMMR/MSI-H colorectal cancer both in training cohort and validation cohort.

Conclusions: MKI67, TCHH and TPR were potential prognostic markers for dMMR colorectal cancer. Based on these data and rationale, we suggest that mutations in all exons of these 3 genes should be integrated into predictive biomarker panels for patients with dMMR/MSI-H, aiming to provide more accurate guidance for the selection of personalized diagnosis.

Keywords: MKI67, TPR, TCHH, Defective Mismatch Repair, Colorectal Cancer, Prognostic Markers

Citation Format: Zheng Liu, Jingfang Lv, Hongmei Zhao, Xishan Wang, Dongliang Wang. Identification prognostic markers for patients with defective mismatch repair in colorectal cancer: MKI67, TPR,TCHH [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 5404.
Abstract 5443: BPI-421286: A highly potent small molecule inhibitor targeting KRASG12C mutation
Authors: Zhu Xiaoguan , Lu Yuan , Guo Jing , Yan Dan , Li Boyan , Chen Xiaoping , Chen Haibo , Lan Hong , Wu Hao , Ding Lieming , Wang Jiabing
Published: June 2022

KRAS is a GTP-binding protein that links receptor tyrosine kinase activation to intracellular signaling. Mutation of KRAS favors the GTP-bound active state and constitutive activation of downstream effects. KRASG12C mutation has been identified as an oncogenic driver of tumorigenesis, which is found in approximately 14% of lung cancer, 4% of colorectal (CRC), and 1-3% of other solid tumors. Herein, we report BPI-421286, a highly potent and selective covalent small molecular KRASG12C inhibitor. In vitro, BPI-421286 showed a high Kinact/Ki value (256,000 M-1 s-1), and inhibited KRAS-GTP/GDP exchange with a lower IC50 (55 nM). In cell-based assays, BPI-421286 suppressed proliferation of KRASG12C mutant cell lines with sub-nanomolar potency, but did not affect cell lines with wide type or non-G12C KRAS mutations. BPI-421286 also inhibited phospho-ERK in MIA PaCa-2 cell with a low IC50 value (2.9 nM). In vivo, daily oral administration of 3-100 mg/kg BPI-421286 led to tumor growth suppression or regression in multiple KRASG12C mutant xenograft or PDX models of NSCLC, CRC and PDAC. The in vivo PKPD in MIA PaCa-2 xenograft model showed that durable pERK inhibition can be achieved without continuous drug exposure, featuring BPI-421286 as a covalent, non-reversible KRAS inhibitor. Combining BPI-421286 with several targeted agents, including a SHP2 inhibitor BPI-442096 and a CDK4/6 inhibitor BPI-16350, demonstrated enhanced tumor growth inhibition compared to either agent alone. Notably, for the acquired resistance mutations that are clinically observed after KRASG12C inhibitor treatment, such as G12C/H95D, G12C/H95Q, G12C/R68S and G12C/Y96C, BPI-421286 also exhibited strong inhibitory activities, suggesting that BPI-421286 could overcome the acquired resistance of currently approved KRASG12C inhibitors. Taken together, BPI-421286 is a highly potent and selective KRASG12C inhibitor, with favorable ADME properties and wide therapeutic index in pre-clinical toxicology studies. BPI-421286 is currently in Phase 1 clinical study.

Citation Format: Xiaoguan Zhu, Yuan Lu, Jing Guo, Dan Yan, Boyan Li, Xiaoping Chen, Haibo Chen, Hong Lan, Hao Wu, Lieming Ding, Jiabing Wang. BPI-421286: A highly potent small molecule inhibitor targeting KRASG12C mutation [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 5443.
Abstract 5468: A pharmacological inhibition of sterol O-acyltransferase 1improves response to enzalutamide in prostate cancer
Authors: Kim Sora Q. , Utturkar Sagar , Atallah Nadia M. , Kim Kee-Hong
Published: June 2022

Prostate cancer (PCa) growth is mediated by androgens via activation of androgen receptor (AR). Accordingly, androgen deprivation therapy (ADT) is the gold standard for the treatment of advanced PCa, but progression to castration-resistant PCa (CRPC) follows. Enzalutamide (ENZ) is an AR antagonist used for the management of CRPC. However, patients acquire resistance to the drug in a short period. Meanwhile, cholesterol is a precursor to androgen and its metabolism is dysregulated in PCa. In addition, cholesteryl ester (CE) accumulation aggravates tumor growth but its association with ENZ treatment has not been studied. Thus, we hypothesized that inhibition of CE formation by blockage of sterol-O-acyltransferase 1 (SOAT1) could enhance the response to ENZ. We found that SOAT1 mRNA is upregulated in clinical prostate tumors compared to normal/benign tissues in TCGA dataset. SOAT1 mRNA level was also highly correlated with AR expression in SU2CF/PCF cohorts and TCGA dataset, suggesting critical role of the cholesterol metabolism in prostate tumor progression and potential association to AR signaling pathway. We treated 22RV1 prostate cancer cells with ENZ in the presence or absence of SOAT1 inhibitor, avasimibe, and found the combination treatment effectively reduced the level of CEs compared to control, which accompanied inhibition of colony formation and cell proliferation. Combination index value was calculated and the relationship between the two drugs showed synergism. Furthermore, genetic knockdown of SOAT1 sensitized cells to ENZ treatment and also reduced the number of colonies formed. In TCGA dataset, patients who had previously undergone ADT were analyzed, and their Kaplan-Meier survival curve showed low SOAT1 expressing patients had a significantly longer progression-free survival rate than their counterpart, indicating that SOAT1 could act as a prognostic biomarker predicting PCa treatment outcome. Gene set enrichment analysis of patients with ADT history showed androgen response gen sets were enriched in high SOAT1 expressing PCa patient group. Inhibition of SOAT1 and the combination treatment did not change AR and ARV7 protein level, but downstream genes such as PSA were down regulated upon the combination treatment. In summary, our in vitro results showed blockage of SOAT1 enzyme sensitized PCa cells to ENZ treatment and analysis of PCa patient data suggested that SOAT1 gene expression has potential as a prognostic marker.

Citation Format: Sora Q. Kim, Sagar Utturkar, Nadia M. Atallah, Kee-Hong Kim. A pharmacological inhibition of sterol O-acyltransferase 1improves response to enzalutamide in prostate cancer [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 5468.
Abstract 5487: An auristatin-based peptide-drug conjugate targeting Kita-Kyushu lung cancer antigen 1 for precision chemoradiotherapy in gastric cancer
Authors: Chen Xiaotong , Liu Fangcen , Yu Xiaoxiao , Li Lin , Yan Jiayao , Liu Baorui
Published: June 2022

Gastric cancer is a worldwide health problem. Concurrent chemoradiotherapy (CRT) represents an important strategy to reduce recurrence and prolong survival for gastric cancer. However, non-targeted cytotoxic drugs capable of enhancing radiosensitization remain the main choice to be combined with radiotherapy, which has limited the anti-tumor efficacy of CRT by off-target side effects. Monomethyl auristatin E (MMAE) is a highly potent anti-tubulin agent and has been reported to act as an effective radiosensitizer recently. Kita-Kyushu lung cancer antigen 1 (KK-LC-1) is a cancer/testis antigen highly and exclusively expressed in multiple types of cancers including gastric cancer, representing a promising target for drug delivery. Previously, we identified a KKLC-1-targeting cyclic nonapeptide (1131 peptide) by screening a phage display peptide library. Here, we described a novel peptide-drug conjugate (PDC) 1131-MMAE by connecting the 1131 peptide with MMAE via a protease-sensitive linker for tumor-selective radiosensitization. We demonstrated that 1131-MMAE could be efficiently internalized by gastric cancer cells through KKLC1-mediating endocytosis and exhibited a comparable cytotoxicity profile with free MMAE. 1131-MMAE could arrest the cell cycle at the most radiosensitive G2/M phase in a dose-dependent manner and selectively enhance the radiation response of KK-LC-1 positive gastric cancer cells by increasing DNA double-strand break as indicated by γH2Ax immunofluorescence staining. Furthermore,1131-MMAE was able to inhibit tumor growth as a monotherapy at 2mg/kg with a good safety profile in vivo. Increased tumor control was achieved when 1131-MMAE was combined with radiotherapy at a relatively low dose of 0.3mg/kg. Overall, we proposed an MMAE-based PDC targeting a novel tumor marker KK-LC-1 and demonstrated its tumor-selective radiosensitization in gastric cancer models for the first time. This anti-KKLC1 PDC may help to promote the clinical progress in precision chemoradiotherapy for multiple types of cancer.

Citation Format: Xiaotong Chen, Fangcen Liu, Xiaoxiao Yu, Lin Li, Jiayao Yan, Baorui Liu. An auristatin-based peptide-drug conjugate targeting Kita-Kyushu lung cancer antigen 1 for precision chemoradiotherapy in gastric cancer [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 5487.
Abstract 5524: AK127, a novel monoclonal antibody (mAb) targeting T cell immunoreceptor (TIGIT)
Authors: Huang Zhaoliang , Pang Xinghua , Zhong Tingting , Jin Chunshan , Chen Na , Xia Dennis , Zhang Peng , Wang Max , Xia Michelle , Li Baiyong
Published: June 2022

Introduction: Immune checkpoint blocking antibodies achieved great clinical success, yet a large fraction of cancer patients receive minimum benefit from current immunotherapies targeted PD-1 and CTLA-4. As a new multifunctional immune checkpoint molecule, TIGIT holds promise to become an important cancer immune therapy target. AK127 is a novel humanized immunoglobulin (Ig) G1 monoclonal antibody (mAb) targeting T cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain (TIGIT). AK127 binds to TIGIT specifically and blocks the interaction of TIGIT with its ligands, thus relieves immune suppression and promotes anti-tumor immune response.

Method: The effect of AK127 on secretion of IL-2 in mixed culture of Jurkat-TIGIT and HT1080-aCD3scFv cells (ELISA) was determined in cellular assays. Binding kinetics of AK127 to C1q, FcγR were measured by Fortebio. ADCC, ADCP and CDC activities were determined in cellular assays. In preclinical in-vivo pharmacology studies, synergistic anti-tumor activity of AK127 and a bi-specific anti-PD1/CTLA4 antibody Cadonilimab (AK104) was investigated in BALB/c-hPD-1/hTIGIT transgenic mice with CT26.WT tumor model.

Result: AK127 blocked the immune inhibitory signal mediated by TIGIT, resulting in enhanced IL-2 secretion. AK127 exhibited binding to FcγIa, FcγRIIIa and C1q, and eliciting ADCC, ADCP and CDC. In animal models, both AK104 (0.5 mg/kg) and AK127 (4 mg/kg) significantly inhibited tumor growth. The combination of AK127 and AK104 produced a significantly enhanced anti-tumor effect than either monotherapy, demonstrating the synergistic anti-tumor activity of AK127 combined with AK104.

Conclusion: AK127 binds to human TIGIT with high affinity, competitively blocks the binding of TIGIT with its ligand CD155 and CD112, relieves immune suppression in both cellular and animal model systems, and promotes anti-tumor immune response.

Citation Format: Zhaoliang Huang, Xinghua Pang, Tingting Zhong, Chunshan Jin, Na Chen, Dennis Xia, Peng Zhang, Max Wang, Michelle Xia, Baiyong Li. AK127, a novel monoclonal antibody (mAb) targeting T cell immunoreceptor (TIGIT) [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 5524.
Abstract 5541: BGB-15025, a potent and selective HPK1 inhibitor, is efficacious as a single agent or in combination with PD-1 antibody in multiple tumor models
Authors: Liu Ye , Li Jing , Li Zhuo , Wang Qin , Zhou Xing , Liu Xiaoxin , Zhang Bo , Wang Xitao , Xu Sanjia , Sun Hanzi , Song Xiaomin , Yuan Xi , Wang Zhiwei , Liu Xuesong
Published: June 2022

Hematopoietic progenitor kinase 1 (HPK1), a hematopoietic cell-restricted serine/threonine protein kinase, has been reported to serve as a critical negative feedback regulator of T lymphocytes by phosphorylating the adaptor protein SLP76 in the TCR complex. The negative feedback role of HPK1 in TCR signaling makes it a promising target for immuno-oncology therapy. It has been recently demonstrated that the kinase activity of HPK1 is essential for its antitumor activity, and HPK1 inhibitors can be potentially combined with immune checkpoint inhibitor therapy (such as PD-1 antibody) for effective cancer treatment. Here, we report a potent and selective HPK1 inhibitor BGB-15025 which has been identified through in-house high throughput screening and structure-guided drug design. BGB-15025 potently inhibits HPK1 kinase activity in biochemical assay, with a 50% inhibitory concentration (IC50) of 1.04 nM under Km concentration of ATP. Consistently, BGB-15025 can potently reduce SLP76 phosphorylation and increase downstream ERK phosphorylation in a concentration dependent manner in T cells-based assay. As a consequence, BGB-15025 induces TCR activation and IL-2 production in T cells. Based on in-house profiling, BGB-15025 shows good selectivity profile among MAP4K family and does not affect ZAP70 phosphorylation up to 1 μM. Oral administration of BGB-15025 demonstrates dose-dependent pSLP76 inhibition in splenic T cells and induces serum IL-2 in mouse model. In efficacy studies, BGB-15025 exhibits anti-tumor activity in GL261 tumor model as single agent. Moreover, BGB-15025 demonstrated combination effect with anti-PD-1 antibody in CT26 and EMT-6 syngeneic tumor models. BGB-15025 is currently in phase I clinical trial to treat patients with advanced solid tumor, both as a single agent and in combination with anti PD-1 antibody tislelizumab.

Citation Format: Ye Liu, Jing Li, Zhuo Li, Qin Wang, Xing Zhou, Xiaoxin Liu, Bo Zhang, Xitao Wang, Sanjia Xu, Hanzi Sun, Xiaomin Song, Xi Yuan, Zhiwei Wang, Xuesong Liu. BGB-15025, a potent and selective HPK1 inhibitor, is efficacious as a single agent or in combination with PD-1 antibody in multiple tumor models [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 5541.
Abstract 5551: QLS31905, an anti-claudin 18.2/CD3 bispecific antibody for claudin 18.2-positive gastric cancer
Authors: Jiang Jiahua , Si Meimei , Luo Xiao , Wu Yun , Li Yongjun , Yan Xiaodan
Published: June 2022

Claudin 18.2 is a quadruple transmembrane protein with a complex structure and highly specific expression in gastric and pancreatic cancers. It is suggested that claudin 18.2 is an ideal target for cancer therapy. Anti-claudin 18.2 antibody IMAB362 has achieved remarkable success in the latest clinical trials. Multiple anti-claudin 18.2 monoclonal antibodies and chimeric antigen receptor T-cell therapy targeting claudin 18.2 have also entered phase I clinical trials. CD3 is a receptor that is present on the surface of T cells and is involved in the antigen recognition by T cells as well as in signal transduction. Anti-claudin 18.2/CD3 bispecific antibody can recruit T cells to tumor sites, activating T cells to kill tumor cells with high claudin 18.2 expression to improve efficacy. Here, we describe a new T cell-redirecting antibody (QLS31905) that is bispecific for claudin 18.2 and CD3. QLS31905 showed a potent in vitro cytotoxic activity against claudin 18.2-positive engineered cell lines HEK293-hCLDN18.2 and NCI-N87 CLDN18.2 as well as superior in vivo efficacy in humanized NCI-N87 CLDN18.2 model. Furthermore, QLS31905 was safe and well tolerated in a 4-week repeated dose (up to 10 mg/kg) toxicity study in cynomolgus monkeys. In vitro cytokine release was lower in the QLS31905 group than that in the AMG 910 (another anti-claudin18.2/CD3 bispecific antibody) group. Overall, these data suggested that QLS31905 has similar efficacy to AMG 910 and is safer. QLS31905 may be an effective and clinically valuable treatment for patients with claudin 18.2-positive gastric cancer.

Citation Format: Jiahua Jiang, Meimei Si, Xiao Luo, Yun Wu, Yongjun Li, Xiaodan Yan. QLS31905, an anti-claudin 18.2/CD3 bispecific antibody for claudin 18.2-positive gastric cancer [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 5551.
Abstract 5552: Novel set of OX40 targeted antibodies and VHHs include agonists and antagonists for future development
Authors: Cai Chenguang , Li Xiang , Hou Shin-Chen , Sun Guoqian , Li Ying , Yang Ting , Zhang Fengzhi , Sun Qi , Fu Shiying , Zhao Shuang , Lin You'you , Xue Feizheng
Published: June 2022

Immunotherapy is one of the most effective modern treatments for cancer. While immunotherapy operates through a broad class of mechanisms, modulating immune checkpoints have provided some of the most effective approaches, including inhibition of PD-1/PD-L1 pathway, CTLA-4, LAG-3, OX40, Tim-3, KIR and TIGIT to reverse tumor immune escape.OX40, also known as CD134 or TNFRSF4, is a membrane protein expressed in CD4+ and CD8+ T cells, neutrophils and NK cells. In the process of T cell activation, OX40 plays a synergistic stimulating function by binding with ligand OX40L. OX40 agonist immunotherapy has been previously shown to improve the survival rate of glioblastoma in mice, prevent tumor growth in ovarian cancer, increase the prognostic significance of CD8+ cell infiltration in colorectal cancer, and eliminate the inhibitory function of Foxp3+ Treg cells that constitutively Express OX40. On the contrary, blocking OX40 axis is a promising strategy to treat T cell hyperactivation in autoimmune diseases. As recently reported in the literature, a two-armed, random, double blinded phase 2b clinical trial demonstrated that telazorlimab, an antagonist OX40 antibody developed by Ichnos Sciences, met primary efficacy endpoints in treating atopic dermatitis.In this research, we developed a set of novel antibodies to OX40 using a hybridoma and VHH synthetic library; the set of antibodies includes both agonists and antagonists.We developed one hybridoma origin mAb into a human framework. Agonist activity was equal to MOXR0916 in NF- κB reporter-Jurkat assay, dependent on coupling by proteinA/cell-bound CD32b, non-competing with OX40L, and epitope mapped to domain 2+3.We identified dozens of binders from our synthetic single domain VH library. After construction into a IgG1Fc fusion form, 4 VHHs were obtained with significant agonistic activity, dependent on coupling by proteinA/cell bound CD32b. Another 5 VHHs were also obtained with significant antagonistic activity without coupling by proteinA/cell-bound CD32b. The 4 agonistic VHHs were confirmed with binding to domain 1+2, competitive to OX40L’s binding.These VHHs constructs represent a promising set of novel biologics to modulate OX40—as agonists or antagonists—and serve as the basis for development of potential new therapies in the clinic.

Citation Format: Chenguang Cai, Xiang Li, Shin-Chen Hou, Guoqian Sun, Ying Li, Ting Yang, Fengzhi Zhang, Qi Sun, Shiying Fu, Shuang Zhao, You'you Lin, Feizheng Xue. Novel set of OX40 targeted antibodies and VHHs include agonists and antagonists for future development [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 5552.
Abstract 5553: Anti-CD19-CD22 antibody drug conjugate to reduce tumor recurrence in B-cell hematologic cancers
Authors: Du Liang , Wan Tingting , Jin Yi , Yuan Jijun
Published: June 2022

With 1,279,000 new cases in 2020 and high incidence of relapse, hematologic cancers present significant opportunity for drug development. B-cell caners arise from various developmental stages of the B lymphocyte, and occur in several forms, including leukemia, lymphoma, and multiple myeloma. CD19 and CD22 are validated targets for B cell cancers, each with multiple drugs approved. However, single targeting therapies showed a poor median response likely due to loss of antigen expression upon treatment stress. CD19&CD22 dual targeting antibody drug conjugate can be a powerful killer of B-cell cancers, with more targeted patient populations due to the high expression of CD19, and good anti-tumor efficacy due to the high internalizing ability of CD22.To this end, we developed 7E11-13G9, which is a bispecific antibody designed for antibody-drug-conjugate for a larger market of B-cell hematological cancers.By the strong CD19 binding arm of 7E11 and the efficient CD22-mediated internalizing arm of 13G9, 7E11-13G9 BsAb kills more tumor cells with high potency and efficacy. In tumor cell killing assay, bispecific 7E11-13G9 showed significantly better anti-tumor efficacy than any of the benchmark antibodies, and even better than the combination of the anti-CD19 benchmark Loncastuximab with the anti-CD22 benchmark Inotuzumab. Significantly better anti-tumor effect and delayed tumor recurrence of CD19-CD22 bispecific compared with single target was demonstrated in Nalm6 acute lymphocyte leukemia model. 7E11-13G9 BsAb is stable and has good productivity in transient expression. Proof of concept study of antibody drug conjugate based on 7E11-13G9 was prepared, and showed great anti-tumor efficacy both in vitro and in vivo.

Citation Format: Liang Du, Tingting Wan, Yi Jin, Jijun Yuan. Anti-CD19-CD22 antibody drug conjugate to reduce tumor recurrence in B-cell hematologic cancers [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 5553.
Abstract 5572: T cell receptor modified with EDB-targeting ScFv and CD28 intracellular domain led to complete tumor regression in xenograft model
Authors: Zhang Zhijie , Qiu Wen , Liu Chang , Wang Muhan , Miao Yinghua , Ji Qingchao , Sun Rongchen , Yin Hongping , Yang Meijia
Published: June 2022

We explored use of chimeric antigen receptor T cell (CAR-T) for treatment of solid tumors, aiming at efficient killing of tumor cells while minimizing on-target off-tumor toxicities. We designed CAR targeting the extra domain B (EDB)-containing fibronectin, which is an oncofetal antigen widely expressed in tumor stroma and on the surface of cancer cells. Unlike membrane-spanning antigens, EDB-containing fibronectin is embedded in the extracellular matrix, and is enriched in new vasculatures and stromal tissues of tumor tissues. First, we made a second-generation EDB CAR (2G-EDB CAR) that killed EDB+ cells (U87MG, A549, and HUVEC) and showed tumor suppression in a U87MG xenograft tumor model. However, while in vitro the 2G-EDB CAR T cells were sufficiently cytotoxic, the in vivo inhibition of the tumor growth was moderate. Next we designed an EDB CAR that contained CD3ε to create 3ε-EDB CAR. We show that the 3ε-EDB CAR was incorporated into the T cell receptor (TCR) structure. The 3ε-EDB CAR completely displaced the native CD3ε in the TCR based on immune precipitation and Western analysis. The 3ε-EDB CAR was capable of inducing T cell activation and proliferation in the presence of EDB-fibronectin antigen. Further, the 3ε-EDB CAR T cells killed tumor cells containing EDB-fibronectin. In vivo, the 3ε-EDB CAR T cells demonstrated improved inhibition when compared to the 2G-EDB CAR T cells. To further enhance the EDB-targeting CAR, we added intracellular domain of CD28 to the 3ε-EDB CAR to create 3ε28-EDB CAR, and showed the 3ε28-EDB CAR was still incorporated into the TCR complex. The 3ε28-EDB CAR co-opted TCR complex and provided co-stimulatory signal upon binding of EDB antigens. Upon transduction of the 3ε28-EDB CAR into primary T cells, we observed upregulation of T cell activation markers CD25, CD69 and granzyme B in presence of the EDB antigen. We show that 3ε28-EDB CAR triggered the phosphorylation of CD3ζ, suggesting that EDB antigen stimulated co-opted TCR and signaled through the TCR-CD3 complex. T cells transduced with 3ε28-EDB CAR demonstrated stronger proliferation when activated by U87MG cells compared to 2G or 3ε-EDB CARs. Furthermore, 3ε28-EDB CAR T cells inhibited in vivo tumor growth in a dose-dependent manner, and most of the treated mice had no evidence of residual tumors. In conclusion, our results suggest that 3ε28-EDB CAR has great potential for treatment of solid tumors.

Citation Format: Zhijie Zhang, Wen Qiu, Chang Liu, Muhan Wang, Yinghua Miao, Qingchao Ji, Rongchen Sun, Hongping Yin, Meijia Yang. T cell receptor modified with EDB-targeting ScFv and CD28 intracellular domain led to complete tumor regression in xenograft model [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 5572.
Abstract 5587: STING and TLR 7/8 Agonist combination can improve immune checkpoint blockade therapy efficiency
Authors: Revuri Vishnu , Bhatnagar Shubhmita , Schultz John , Larson Peter , Ferguson David M. , Panyam Jayanth
Published: June 2022

Despite significant advancements in immune checkpoint blockade (ICB) therapy, only few patients respond to the treatment. Non-immunogenic cold tumors lack T-cell infiltration, which results in reduced ICB therapeutic efficiency. Immune adjuvants can reprogram the non-immunogenic cold tumor microenvironment (TME) to inflamed hot TME by activating antigen presenting cells and improving T-cell homing. Here, we investigate the potentials of 558 (a novel TLR 7/8 agonist) and ADU-S100 (stimulator of interferon gene (STING) agonist in clinical trials) combination on improving the activity of ICB. Human TLR-specific reporter cell assay using HEK-Blue™-hTLR7 and 8, confirmed the activation of both TLR 7 and TLR 8 with EC50 of 0.18 µM and 5.34 µM respectively, when treated with 558. In addition, treatment of human PBMCs with 558 increased the IFN-γ and TNF-α cytokine secretions. Moreover, the levels of IL-10, an anti-inflammatory cytokine were not increased upon treatment with 558. Although treatment of bone marrow derived dendritic cells (BMDC) with 558 resulted in CD40 expression, a reduced expression of CD80 and CD86, costimulatory molecules that are required for T-cell activation, was observed. Interestingly, 558 in combination with ADU-S100 not only increased the CD40 expression but also improved the expression of CD80 (4-fold) and CD86 on BMDCs. Furthermore, the levels of pro-inflammatory cytokines TNF-α and IL-6 was significantly increased when BMDCs were treated with 558 and ADU-S100 combination compared to the individual treatments. TME constitute M2 polarized macrophages that promote secretion of anti-inflammatory cytokines and favor tumor progression. Interestingly, 558 and ADU-S100 combination reduced the expression of CD206, a mannose receptor that is highly expressed on M2 macrophages and improved the expression of CD80 (25-fold) and CD86 (6-fold) compared to 558 treatments alone. These results confirmed the polarization of M2 macrophages to immunogenic M1 macrophages. Overall, these studies indicate promising potential of 558 and ADU-S100 combination in reprogramming the TME to aid in effective cancer treatments.

Citation Format: Vishnu Revuri, Shubhmita Bhatnagar, John Schultz, Peter Larson, David M. Ferguson, Jayanth Panyam. STING and TLR 7/8 Agonist combination can improve immune checkpoint blockade therapy efficiency [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 5587.
Abstract 5622: Preclinical evaluation of humanized PD1/GITR combination therapy using double humanized mouse
Authors: Yuan Dongdong , zhang Dongdong , Sun Hongyan , Zhang Mingkun , Dong Liyou , Ju Cunxiang , Li Shuai , Wang Huiyi , Zhao Jing , Gao Xiang
Published: June 2022

GITR (glucocorticoid-induced TNFR-related protein) is a member of tumor necrosis factor receptor superfamily, expressed on many kinds of cells including T cells, attracting attention as a co-stimulatory immune checkpoint molecule. GITRL, the ligand of GITR is mainly expressed on antigen presenting cells. GITR/GITRL interaction plays an important role on the development of immune response, which can serve as promising anti-tumor target. While more than twenty anti-GITR antibodies are at clinical stages, combination treatments of GITR agonist and PD-1 inhibitor may be a better choice. Thus, there is a need for pre-clinical mouse model to evaluate the anti-tumor effect of humanized PD1/GITR antibodies. Gempharmatech established PD1/GITR double humanized mouse line on a BALB/c background, in which the extracellular domain of mGITR was replaced with its human counterparts while the trans-membrane and cytoplasmic domain kept intact, as well as the same strategy of PD1. BALB/c-hPD1/hGITR mouse could successfully express similar levels of hGITR and hPD1 compared with BALB/c wild type mice. Moreover, no immune or developmental abnormalities were observed in the humanized mouse. Anti-GITR combination with anti-PD1 presented greater tumor growth inhibition than anti-PD1 or anti-GITR monotherapies in PD1/GITR double humanized mouse bearing CT26 tumors. In summary, BALB/c-hPD1/hGITR are valuable models for the pre-cilincal evaluation of candidate humanized PD1/GITR combination treatment or bispecific antibody therapies.

Citation Format: Dongdong Yuan, Dongdong zhang, Hongyan Sun, Mingkun Zhang, Liyou Dong, Cunxiang Ju, Shuai Li, Huiyi Wang, Jing Zhao, Xiang Gao. Preclinical evaluation of humanized PD1/GITR combination therapy using double humanized mouse [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 5622.
Abstract 5623: CD137 humanized mice for preclinical efficacy evaluation of therapeutic antibodies
Authors: Jiang Yunlong , Gu Tingting , Yu Weiwei , Sun Hongyan , Zhang Mingkun , Liang Juan , Li Shuai , Ju Cunxiang , Zhao Jing , Gao Xiang , Moore Mark W.
Published: June 2022

CD137 (41-BB, tumor necrosis factor receptor superfamily 9), expressed on activated T cells and NK cells, has gained increased attention as an anti-tumor target because of remarkable clinical efficacy using either a single agent or in combination with other immune checkpoint inhibitors. We constructed CD137 humanized mice based on two different backgrounds, C57BL6 and BALB/c mice, as well as dual-target and triple-target mice crossbred with PD1/PDL1 humanized mice to create an ideal model to study CD137 target drugs. The normal immune cell population in peripheral blood were not affected in CD137 humanized mice, and hCD137 was expressed on T cells after the stimulation with a anti CD3 antibody. We verified the efficacy of anti-CD137 drugs in BALB/c-hCD137 mice inoculated with different cancer cell lines (i.e. CT26, EMT6 and 4T1). The drug Urelumab showed a dramatic response in the hCD137 mice model inoculated with CT26 and EMT6 but not with 4T1 cell line. In a rechallenge study, no tumors grew in tumor-free mice previously treated with Urelumab, with effector memory T cells increased in peripheral blood, indicating Urelumab has a relatively long-lasting anti-tumor effect. The combination of Urelumab and anti-PD1 has a stronger tumor growth inhibition effect than that of anti-CD137 or anti-PD1 monotherapy on dual-target or triple target mice. These data show that the humanized CD137 mouse is a suitable model for evaluating the monotherapies or combination therapies targeting CD137.

Citation Format: Yunlong Jiang, Tingting Gu, Weiwei Yu, Hongyan Sun, Mingkun Zhang, Juan Liang, Shuai Li, Cunxiang Ju, Jing Zhao, Xiang Gao, Mark W. Moore. CD137 humanized mice for preclinical efficacy evaluation of therapeutic antibodies [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 5623.
Abstract 5643: YK-2168, a potent and selective small molecule CDK9 inhibitor
Authors: Ding Charles Z. , Liu Yingchun , Zhou Wang
Published: June 2022

Objective: Cyclin-dependent kinase 9 (CDK9), which regulates transcriptional elongation, is an attractive therapeutic target for many cancers, especially for cancers driven by transcriptional dysregulation. Specifically, CDK9 mediated transcriptional regulation of short-lived antiapoptotic proteins is critical for the survival of transformed cells. Emerging evidence indicates that selective CDK9 inhibition may provide a therapeutic benefit against certain cancers. We disclose here for the first time a novel CDK9 inhibitor YK-2168 a clinical candidate and its preclinical results.

Method: The biochemical activities of YK-2168 for CDK9, CDK1 and CDK2 were measured by literature methods (Tinggui, 2014). The in vitro anti-proliferative activity was evaluated on selected leukemia cell line MV4-11, lymphoma cell line Karpas422 and gastric carcinoma cell line SNU16. The in vivo antitumor activity of YK-2168 was evaluated in cell-derived xenograft (CDX) models of Leukemia (MV4-11) and gastric carcinoma (SNU16), YK-2168 was administered by intravenous (IV) injection. PK/PD was assessed by western blot analysis of pS2-RNA Pol ll CTD expression and cleaved caspase 3 level in the treated tumor tissues.

Result: YK-2168 displayed potent selective inhibitory biochemical activity for CDK9 (IC50 : 7.5 nM), but that of CDK1 and CDK2 were modest (IC50 : 466.4 and 361.1 nM), indicative of good selectivity for CDK9. It also showed good anti-proliferative activities in tumor cell lines of different types, MV4-11 potency IC50 of 53.4 nM, Karpas422 potency IC50 of 91.1 nM, and SNU16 potency IC50 of 88.3 nM. YK-2168 showed in vivo antitumor activity in a MV4-11 CDX model with TGI =80 % @10 mpk QW (IV with 4 doses), compared with the same dose of BAY1251152 (TGI =65 % @10 mpk, QW). YK-2168 also showed good regressive antitumor activity in vivo in a SNU16 CDX model with TGI =115 % @10 mpk QW (while BAY1251152 totally inhibited growth with TGI =100%). Western blot analysis of the tumor samples showed that YK-2168 can also inhibit the expression of CDK9 downstream signal proteins and induced apoptosis in a dose-dependent manner.

Conclusion: We have discovered a potent and selective small molecule CDK9 inhibitor YK-2168, which showed potent CDK9 inhibition and strong antitumor activity in both in vitro and in vivo preclinical tumor models. These results together with GLP safety results are considered highly promising and warrant moving the compound forward to clinical investigation. In fact, YK-2168 was granted phase I clinical trials in both US and China.

Citation Format: Charles Z. Ding, Yingchun Liu, Wang Zhou. YK-2168, a potent and selective small molecule CDK9 inhibitor [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 5643.
Abstract 5665: LILRB2 mediates the immune escape of breast tumor cells by degrading HLA-A
Authors: Zhao Hui , jiang zhiyuan , wang zhiyu , chang yujie , ruan shunyi
Published: June 2022

Breast cancer has now risen to the number one cancer in the world, posing a great threat to human health. LILRB2 (leukocyte immunoglobulinlike receptor B2) is an important immunosuppressive receptor protein, which is mainly expressed in myeloid cells, such as macrophages and dendritic cells. At present, we have found clinically that patients with breast cancer bone and bone marrow metastasis have higher expression of LILRB2. In our research, we used Western blotting, real-time fluorescent quantitative Polymerase Chain Reaction (RT-PCR), flow cytometry, and other experimental methods to explore the expression and function of LILRB2 in breast cancer cells. In our research, we found that LILRB2does increase expression in breast cancer cells, therefore, we want to explore the role of LILRB2 in breast cancer cells. we find that LILRB2 will accelerate the proliferation of tumor cells and increase their migration ability. We tested the related signal pathways and found that LILRB2 can promote the enhancement of the NF-Kb signal pathway. Further, we have done related LILRB2 correlation analysis and found that LILRB2 can reduce human leukocyte antigen-A (HLA-A) Expression. Next, we tested the changes in the level of major histocompatibility complex (MHC-1) related mRNA and found that its mRNA expression increased, which was inconsistent with the decrease in its protein level we observed. Therefore, we used the proteasome inhibitor MG132 for treatment, and the results showed after adding MG132, the decrease in HLA-A expression caused by overexpression of LILRB2 was restored. Next, we knocked down the breast cancer cell line of membrane-associated ring finger (MARCH9). After we knocked down MARCH9, transfection of LILRB2 did not increase the ubiquitination of HLA-A. Therefore we show that LILRB2 can mediate the post-translational modification of HLA-A via MARCH9, which in turn mediates the degradation ofHLA-A. In conclusion, our research shows that LILRB2 maybe a candidate of clinical target for the treatment of breast cancer, preventing possible bone and bone marrow metastasis from breast cancer.

Citation Format: Hui Zhao, zhiyuan jiang, zhiyu wang, yujie chang, shunyi ruan. LILRB2 mediates the immune escape of breast tumor cells by degrading HLA-A [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 5665.