bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5152: SJP1604, a nucleolin-targeted therapeutic agent for relapsed/refractory acute myeloid leukemia
Authors: Um Jihyun , Lee Dohyeong , Park Yongbin , Moon Sung Hwan , Lee Soo Jin , Ki Min-Hyo
Published: September 2020

Acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) are cancers characterized by the rapid growth of abnormal white blood cells. Although the standard first-line therapy leads to high rates of remission, approximately 30% of the treated patients are refractory and more than 50% of them face a relapse because of the occurrence of drug resistance and high toxicity. Therefore, there is a strong unmet medical need for the next-generation targeted therapy to overcome drug resistance and reduce toxicity in AML/MDS. Here, we designed an aptamer-drug conjugate (SJP1604) specific for nucleolin, which is highly expressed only on the cell membrane of cancer cells including AML cells. We identified that SJP1604 (APTA-16) inhibited the growth of AML cells and drug-resistant AML cells in vitro. SJP1604 presented outstanding anti-leukemic efficacy in vivo via syngeneic mouse model and xenograft mouse model. In addition, the synergistic effects were observed upon combination treatmeant with venetoclax in vitro. We also figured out that nucleolin as a clinical biomarker was significantly highly expressed on relapsed/refractory (R/R) AML patient-derived bone marrow cells ex vivo. SJP1604 selectively targeted cancer cells by nucleolin-binding on cell membrane with its high targeting ability and plasma stability owing to its unique conformational property, therefore, it demonstrated less cytotoxicity in vitro/in vivo and a superior therapeutic index based on AUC comparison in pharmacokinetic studies. The mechanism of the anti-leukemic efficacy of SJP1604 was also studied. First of all, SJP1604 decreased the expression of nucleolin and inactivated NFκB signaling pathway via dephosphorylation of NFκB, which resulted in diminished expression of DNMT1 followed by restored p15 expression. In addition, SJP1604 increased p53 expression and decreased bcl-2 expression in the protein and mRNA level. These could be one of potential mechanism of anti-leukemogenesis of SJP1604. In conclusion, these findings suggest that SJP1604 targeting nucleolin could be developed as a first-in-class drug for relapsed/refractory AML and MDS. GLP safety/toxicology studies of SJP1604 have been completed and clinical trials will be initiated in 2020.

Citation Format: Jihyun Um, Dohyeong Lee, Yongbin Park, Sung Hwan Moon, Soo Jin Lee, Min-Hyo Ki. SJP1604, a nucleolin-targeted therapeutic agent for relapsed/refractory acute myeloid leukemia [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 5152.
Abstract 5182: Examining the effects of honokiol on EGFR-positive glioblastoma multiforme
Authors: Smith Keller , Johnson Blake
Published: September 2020

Glioblastoma (GBM) is a highly aggressive and lethal form of primary brain tumor. Systemic suppression of the immune system, specifically T-cell impairment, significantly contributes to GBM development and progression. Recent advances in cancer immunotherapies have shown great promise in alleviating such tumor-associated immunosuppression, namely programmed cell death 1 (PD-1)/programmed death ligand 1 (PD-L1) checkpoint inhibitors. Regrettably, early clinical studies evaluating the efficacy of checkpoint inhibition in recurrent GBM have rendered only modest improvements in overall survival rates, due partially to insufficient inhibition of AKT-STAT3 pathways which are often dysregulated downstream of hyperactive epidermal growth factor receptors (EGFR) in GBM. In the current study, we aim to evaluate a newly discovered small molecule inhibitor of AKT-STAT3, honokiol, and its ability to suppress AKT-STAT3 activity in the context of hyperactive EGFR in GBM. Using U87-MG cells that stably overexpress a common hyperactivating mutation of EGFR, EGFRvIII, we have preliminary evidence that honokiol is an effective inhibitor of U87-EGFRvIII cellular migration as a result of its robust AKT-STAT3 pathway suppression. Thus, honokiol may provide a novel means of pharmacological inhibition of AKT-STAT3 hyperactivity in GBM and therefore greater clinical efficacy of immune checkpoint inhibitors in this deadly disease.

Citation Format: Keller Smith, Blake Johnson. Examining the effects of honokiol on EGFR-positive glioblastoma multiforme [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 5182.
Abstract 5204: Targeting OGG1 arrest cancer cell proliferationby mitochondrial dysfunction and replication stress
Authors: Benitez-Buelga Carlos , Visnes Torkild , Sanjiv Kumar , Mortusewicz Oliver , Berglund Ulrika Warpman , Helleday Thomas
Published: September 2020

Altered oncogene expression in cancer cells causes lost redox homeostasis resulting in oxidative DNA damage, e.g., 8-oxoguanine (8-oxoG), repaired by base excision repair (BER). PARP1 coordinates BER, and relies on the upstream 8-oxoguanine-DNA glycosylase (OGG1) to recognise and excise 8-oxoG. Here we hypothesize that OGG1 may represent an attractive target to exploit reactive oxygen species (ROS) in cancer. Although OGG1 depletion is well tolerated in non-transformed cells, we report here that OGG1 depletion obstruct A3 T-cell lymphoblastic acute leukemia growth in vitro and in vivo, validating OGG1 as a potential anti-cancer target. In line with this hypothesis, we show OGG1 inhibitors (OGG1i) target a wide range of cancer cells, with a favourable therapeutic index to non-transformed cells. Mechanistically, OGG1i and shRNA depletion cause lost mitochondrial function leading to S-phase damage, replication stress and lost proliferation or cell death, representing a novel mechanistic approach to target cancer. This study adds OGG1 to the list of BER factors, e.g., PARP1, as potential targets for treatment of cancer.

Citation Format: Carlos Benitez-Buelga, Torkild Visnes, Kumar Sanjiv, Oliver Mortusewicz, Ulrika Warpman Berglund, Thomas Helleday. Targeting OGG1 arrest cancer cell proliferationby mitochondrial dysfunction and replication stress [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 5204.
Abstract 5206: Quantifying DNA-protein crosslinks induced by bifunctional alkylating agents in cultured human leukemia cells
Authors: Cerrato Hailey L. , Marchant Sam W. , Naccara Allie H. , LaJoie Thomas J. , Michaels Shawnya , Brown Dennis , Rice Kevin P.
Published: September 2020

Bifunctional alkylating agents, such as cisplatin and several nitrosoureas, have clinical utility as antineoplastic drugs largely due to their DNA alkylating and crosslinking activity. Other drug candidates such as laromustine (VNP-40101M) and dianhydrogalacticol (VAL-083) have similar DNA-alkylating activities and have shown success in clinical trials against hard-to-treat cancers such as acute myelogenous leukemia and glioblastoma multiforme. While these agents' interstrand and instrastrand DNA crosslinking activities are widely cited as the critical cytotoxic event, their capacity to crosslink DNA to chromosomally-affiliated proteins may also contribute to their anticancer profile. Indeed, DNA-protein crosslinks (DPCs) are highly toxic phenomena that are difficult for cells to repair and severely compromise gene transcription and DNA replication. Herein, we assess concentration-dependent DPC formation by a wide variety of bifunctional alkylating agents in HL-60 human leukemia cells using two parallel approaches. In one approach, protein-bound DNA was quantified using PicoGreen after precipitating proteins from lysates of drug-treated cells and compared to total DNA concentration. In another, chromosomal DNA was extracted from lysates of drug-treated cells then treated with fluorescein isothiocyanate, which modifies primary amines on proteins. Protein concentration was measured by fluorescence and compared to that of the extracted DNA. The nitrogen mustard mechlorethamine and the chloroethyl nitrosoureas carmustine and lomustine were the most potent agents, yielding significant DPC formation at clinically relevant doses (10−5 M). Cisplatin and laromustine yielded DPCs at similar concentrations, but to a lesser extent. Dianyhydrogalacticol, epichlorohydrin, and diepoxybutane, which all rely on epoxide functional groups for attachment to cellular nucleophiles, required at least an order of magnitude higher drug concentrations for significant DPC observation. These data suggest bifunctional alkylating agents featuring halide leaving groups are most apt to crosslink DNA to nuclear proteins and may offer insights into patterns of cytotoxicity and clinical utility for this important class of chemotherapeuric agents.

Citation Format: Hailey L. Cerrato, Sam W. Marchant, Allie H. Naccara, Thomas J. LaJoie, Shawnya Michaels, Dennis Brown, Kevin P. Rice. Quantifying DNA-protein crosslinks induced by bifunctional alkylating agents in cultured human leukemia cells [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 5206.
Abstract 5215: Inhibition of PIM kinases targets synthetic vulnerabilities and enhances antigen presentation in B cell lymphoma
Authors: Mondello Patrizia , Tarantelli Chiara , Cascione Luciano , Arribas Alberto J. , Rinaldi Andrea , Younes Anas , Bertoni Francesco
Published: September 2020

The PIM kinases are highly expressed in activated B-cell (ABC) diffuse large B-cell lymphoma (DLBCL). Oncogenic cooperation between PIMs and MYC has been demonstrated. Transgenic mice co-expressing Eμ-PIM and Eμ-MYC showed accelerated lymphomagenesis. Conversely, knockdown of PIMs dramatically decreased cMYC levels and lowered tumor incidence. Based on these preclinical data, a treatment strategy aiming at disrupting the oncogenic cooperation between PIMs and MYC may improve the outcome of DLBCL. Therefore, we treated a panel of DLBCL cell lines with increasing dose of the clinically relevant pan-PIM inhibitor (PIMi) AZD1208 (from 0.1 to 10μM) for 48 hours (Hrs), which resulted in a dose-dependent growth inhibition with a stronger efficacy in ABC DLBCL cell lines. The analysis of a CRISPR loss-of-function screening in three ABC (LY3, TMD8, HBL1) and three GCB (SUDHL-4, Pfeiffer, BJAB) DLBCL cell lines (Reddy et al, 2017) showed that PIM2 silencing led to significantly decreased viability irrespective of cell-of-origin, suggesting that this oncogene is essential for cell proliferation in DLBCLs. To identify the genes through which PIMs drive the lymphoma phenotype we performed gene expression profiling using 4 ABC DLBCL cell lines (RIVA, TMD8, SUDHL-2, U2932) treated with either DMSO or AZD1208 at 1μM for 4, 8 and 12 Hrs. We observed induction of 3,439 genes whereas 2,473 genes were downregulated. Gene pathway analysis showed that AZD1208 led to downregulation of genes regulated by MYC, including its known downstream p53 and NFKB target genes. On the other hand, AZD1208 treatment broadly induced MHC class II and antigen presentation genes as well as PI3K/AKT, cell cycle and glutaminase genes. Using a high-throughput screening approach, we found that the inhibitors of cell cycle (such as the BCL2 inhibitor venetoclax/ABT199 and the PLK4 inhibitor CFI-400945) and of glutaminase (CB839) enhanced the antiproliferative effect of AZD1208, whereas combinations with the PI3K/AKT/mTOR inhibitors had negligible synergistic effect. In conclusion, our study revealed previously unknown mechanisms of action of PIM inhibitors and provides a framework for future combination strategies.

Citation Format: Patrizia Mondello, Chiara Tarantelli, Luciano Cascione, Alberto J. Arribas, Andrea Rinaldi, Anas Younes, Francesco Bertoni. Inhibition of PIM kinases targets synthetic vulnerabilities and enhances antigen presentation in B cell lymphoma [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 5215.
Abstract 522: Optimization of novel anti-4-1BB antibodies on Biocytogen's in vivo drug screening platform
Authors: Guo Chaoshe , Zhu Yu , Chen Yunyun , Yang Yi , Li Zhihong
Published: September 2020

4-1BB (CD137, tumor necrosis factor receptor superfamily 9) is an inducible costimulatory receptor expressed on activated T and natural killer (NK) cells. 4-1BB ligation on T cells triggers a signaling cascade that results in upregulation of anti-apoptotic molecules, cytokine secretion, and enhanced effector function. In dysfunctional T cells that have a decreased cytotoxic capacity, 4-1BB ligation demonstrates a potent ability to restore effector functions. On NK cells, 4-1BB signaling can increase antibody-dependent cell-mediated cytotoxicity. Agonistic monoclonal antibodies targeting 4-1BB have been developed to harness 4-1BB signaling for cancer immunotherapy.

We have interests in developing novel 4-1BB antibodies for boosting cancer patients' own immunity in light of other successful immune checkpoint modulators. We developed a cohort of 4-1BB specific antibodies using the classic hybridoma technology. Then, we took the advantages of Biocytogen's humanized mice and screened the purified antibodies in B-h4-1BB mice bearing MC-38 tumor cells. We found some antibodies that are as potent as Urelumab in vivo. We then humanized the antibody to obtain YH004. To determine the subtype of the antibody, we grafted the antigen-binding domain onto various human Fc backbones. Interestingly, we found that the human IgG1 isotype was more effective against tumors than the other isotypes in B-4-1BB mice with MC38 tumors and Treg depletion were noted in tumor-infiltrating lymphocyte analysis using the same murine model. In addition, we proved that the synergistic anti-tumor activity of YH004 in combination with pembrolizumab in B-hPD-1/h4-1BB mice. In brief, the 4-1BB antibody was validated in humanized mice to provide a favorable reference for future clinical evaluation.

Citation Format: Chaoshe Guo, Yu Zhu, Yunyun Chen, Yi Yang, Zhihong Li. Optimization of novel anti-4-1BB antibodies on Biocytogen's in vivo drug screening platform [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 522.
Abstract 523: Advanced engineering using CAR-T display libraries: Ultra-high throughput functional screen
Authors: Youssef Sawsan , Chou Joyce , Pineda Devin , Ives Sarah , Chiou Valerie , Buerckert Jean-Philippe , Newland Raymond , Daraeikia Shahrad , Liao Sindy , Jones Chelsea , Pettus Christina , Salas Jessica , Padilla Emelia , Smith Christopher , Day Giles , Maurer David , Waddell Ian , Glanville Jacob
Published: September 2020

CAR-T-cell therapies represent a new area of synthetic biology success in cancer immunotherapy. However, the complexity of their adverse events related to their activity are still a major challenge. Using our in-house fully human computationally optimized phage display library, we established a novel cell-based ultra-high throughput panning and screening CAR-T platform. Our proprietary CAR-T vector is designed as a series of cassettes incorporating our SuperHuman phage library with an optimal backbone. This allows us to design CAR-T with desired properties, including: Range of Affinities, Humanness, Epitope Diversity, Thermostability, Immunogenicity, Tonic activation, Toxicity, and Functional Activity. Our library's encoding ensures a single CAR-T construct per one reporter cell. Moreover, to confirm CAR-T-specific-activation, we incorporated three different markers that will be upregulated simultaneously. We used CD19 and BCMA to validate our platform selecting target-specific functionally active scFv candidates against both targets, which are selective to activation conditions such as tonic signaling, and tissue specific silencing. Collectively we show that our platform can be readily used to characterize CAR-T in high-speed and - throughput fashion.

Citation Format: Sawsan Youssef, Joyce Chou, Devin Pineda, Sarah Ives, Valerie Chiou, Jean-Philippe Buerckert, Raymond Newland, Shahrad Daraeikia, Sindy Liao, Chelsea Jones, Christina Pettus, Jessica Salas, Emelia Padilla, Christopher Smith, Giles Day, David Maurer, Ian Waddell, Jacob Glanville. Advanced engineering using CAR-T display libraries: Ultra-high throughput functional screen [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 523.
Abstract 524: Discovery of NKp46 antibody for NK cell engager using common light chain phage display approach
Authors: Yang Teddy , Zhu Seng , Gao Jing , Yao Jingyun , Gong Xumin
Published: September 2020

NK cells express activating receptors such as CD16a (FcγRIIIA), NKG2D, SLAM family members and the natural cytotoxicity receptors (NCRs). Full activation of NK cells require co-engagement of different activating receptors. NKp46, a member of NCRs, is a 46 kDa glycoprotein and belongs to immunoglobulin (Ig) superfamily, NKp46 is frequently expressed on tumor infiltrating lymphocytes. Activation of NKp46 with antibody triggers not only NK cell cytotoxicity but also cytokine release. Taken together, NKp46 is a promising target for anti-tumor therapy. Here we described the discovery of fully human NKp46 antibody by phage display approach with common light chain and naïve human B cell library. Bispecific antibody has been researched for many years and very limited number of bispecific antibodies are currently in the market. Heavy and light chain miss pairing is the bottleneck of bispecific antibody discovery and development. To overcome the hurdle, we generated a fully human common light chain phage display library with two commonly used light chain sequences combined with heavy chain fragment from human naïve scFv library. Two light chain sequences were selected based on the approved antibodies sequences and the expression levels. The phage library was then used for panning against recombinant human NKp46 extracellular domain protein and CHOK1 cells overexpressing human NKp46. After conversion to full IgG format, 4 candidates bind specifically to NKp46 and cross react with both human and cyno NKp46. Affinities of the candidates were comparable to bench mark antibodies as determined by FACS. To summarize, we have generated fully human common light chain phage display library that can be used for bispecific antibody discovery. The anti-NKp46 antibodies discovered can be further developed for NK cell engager and NK cell redirecting bispecific antibody.

Citation Format: Teddy Yang, Seng Zhu, Jing Gao, Jingyun Yao, Xumin Gong. Discovery of NKp46 antibody for NK cell engager using common light chain phage display approach [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 524.
Abstract 5240: Effective inhibition of TMPRSS2-ERG positive prostate cancer cells by a new formula of ERGi-USU-6
Authors: Eldhose Binil , Xavier Charles P. , Pandrala Mallesh , Malhotra Sanjay V. , Dobi Albert
Published: September 2020

Background: Prostatic adenocarcinoma is among the leading cause of cancer-related deaths among men in the United States. The Erythroblast Transformation-Specific-Related Gene, ERG is dormant in normal prostate epithelium. Due to gene fusions, the ERG oncogene is frequently activated by androgenic signals in prostate cancer. ERG disrupts the normal differentiation, promotes epithelial-mesenchymal transition, migratory and invasive properties of cancer cells. Approximately 35% of metastatic castration-resistant prostate cancers harbor ERG oncogene. Due to the failure of androgen axis directed therapies, there is an urgent need to develop inhibitors to targeting prostate cancer-causing genes, such as ERG. We have identified a potent small-molecule inhibitor ERGi-USU that is remarkably selective for inhibiting the growth of ERG positive cancer cells through direct binding to the RIOK2 atypical kinase, a putative upstream regulator of ERG. We completed a structure-activity relationship (SAR) study and compound development resulting in the potent derivative, ERGi-USU-6. Our current objective is to improve the therapeutic properties of ERGi-USU-6, by new salt formulations.

Methods: Evaluation of the five selected salt formulations of ERGi-USU-6 were performed by assessing the growth of ERG positive prostate cancer cell line (VCaP) and by monitoring ERG and RIOK2 protein levels. Selectivity was assessed by monitoring the growth, endogenous ERG and RIOK2 levels in normal ERG positive human umbilical vein derived endothelial cells (HUVEC). The IC50 values for cell growth, compared to the parental compound were calculated in a 12-step dilution range performed in triplicates and independently repeated three times. Cell growth was measured by quantitative Cell Glow assay monitoring viable cells using Perkin Elmer Envision assay instrument and protein levels were quantified by measuring bioluminescence in IBright instrument in a wide dynamic range.

Results: One new salt formula with improved activity were identified, demonstrating improved cell growth (IC50=89nM vs. parental IC50=139), ERG protein and RIOK2 protein inhibition. None of the salt formulas of ERGi-USU-6 showed any effect on the primary cultures of the ERG positive normal endothelial cells (HUVEC) in the effective concentration range.

Conclusion: The first evaluation of salt formulas of ERGi-USU-6 may open the possibilities for preclinical assessments of this remarkably cancer-selective compound.

Citation Format: Binil Eldhose, Charles P. Xavier, Mallesh Pandrala, Sanjay V. Malhotra, Albert Dobi. Effective inhibition of TMPRSS2-ERG positive prostate cancer cells by a new formula of ERGi-USU-6 [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 5240.
Abstract 5262: Identifying the role of AXL in fusion kinase inhibitor resistant non-small cell lung cancer
Authors: Peters Tara , Dimou Anastasios , Le Anh , Doebele Robert
Published: September 2020

Chromosomal rearrangements involving ALK, ROS1, RET and others generate novel fusion kinases which comprise approximately 10% of the oncogenic drivers in lung adenocarcinoma. Several tyrosine kinase inhibitors (TKIs) are approved or in development for each of these oncogenes and have shown marked clinical improvement for patients, but resistance inevitably occurs, a significant proportion of which is not currently druggable. AXL is a receptor tyrosine kinase that is often upregulated in resistant cancers in response to both chemotherapy and targeted therapies such as EGFR inhibitors. AXL can promote resistance to targeted therapy by signaling through the MAPK and Akt pathways or heterodimerizing with other receptors to maintain growth and survival signaling. However, the role of AXL in mediating resistance to ALK, ROS1, and RET TKIs is not well understood. In screening our unique panel of patient-derived lung adenocarcinoma cell lines harboring ALK, ROS1 or RET gene fusions, we found elevated AXL expression was correlated with resistance to cognate TKIs, as well as mesenchymal features including spindle morphology and vimentin expression. Combining the AXL inhibitor R428 with TKI treatment, such as crizotinib for ALK and ROS1 driven cell lines, was effective at inhibiting colony formation and wound healing in fusion kinase inhibitor resistant cell lines. Combination treatment downregulates Akt phosphorylation in resistant cells, and we found the resistant cell lines were also more sensitive to PI3K inhibition than cell lines that are still sensitive to their specific TKI. Interestingly, we also identified AXL cross talk with other receptor tyrosine kinases. In a RET inhibitor resistant model expression of both AXL and EGFR are elevated and the receptors interact, which can be disrupted upon EGF stimulation, to drive resistance. Collectively, we show that AXL upregulation is a common marker of TKI resistance among ALK, ROS1, and RET fusion kinase driven lung adenocarcinoma models, and provide rationale for AXL as a clinical target for combination therapy to overcome drug resistance.

Citation Format: Tara Peters, Anastasios Dimou, Anh Le, Robert Doebele. Identifying the role of AXL in fusion kinase inhibitor resistant non-small cell lung cancer [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 5262.
Abstract 5264: Selective toxicity of MX-106-4C, a survivin inhibitor, in P-glycoprotein-mediated multidrug resistant colon cancer
Authors: Lei Zi-Ning , Wu Zhongzhi , Teng Qiu-Xu , Xiao Min , Li Wei , Wurpel John N. , Chen Zhe-Sheng
Published: September 2020

One of the major challenges in colon cancer chemotherapy is multidrug resistance (MDR), which is typically mediated by the overexpression of ATP-binding cassette (ABC) transporters, particularly P-glycoprotein (P-gp, ABCB1, MDR1). A number of P-gp inhibitors have been developed, however, none of these compounds have improved chemotherapeutic efficacy due to undesirable pharmacokinetic profiles or adverse effects, resulting in limited clinical success. Therefore, alternative approaches are urgently needed to circumvent MDR cancer. In previous study, a series of synthesized analogs of MX-106, as anti-cancer drugs targeting survivin, exhibited collateral sensitivity (CS) effect to P-gp overexpressing MDR colon cancer cells as well as ABCB1 gene transfected cells, reflected by more than 10-fold cytotoxic effect in P-gp positive MDR cell lines compared to drug sensitive cell lines. Among the analogs, MX-106-4C was identified as the leading compound with the most potent selective toxicity to P-gp overexpressing cells. MX-106-4C-induced CS effect was observed in both intrinsic and acquired P-gp overexpressing colon cancer cells, which was only partially reversed with the presence of a P-gp inhibitor. Nevertheless, this CS effect was abolished in ABCB1-knockout cells, indicating that the selective cytotoxicity was P-gp expression dependent, but only partially related to P-gp function. Furthermore, we found that MX-106-4C did not significantly affect P-gp ATPase activity or drug accumulation and efflux in P-gp-overexpressing cells. In P-gp overexpressing colon cancer cells, short-term (up to 72 h) incubation of MX-106-4C significantly down regulated P-gp expression at transcriptional level but not protein level, whereas long-term (14 d) incubation of MX-106-4C significantly down regulated P-gp protein expression and re-sensitized MDR colon cancer cells to doxorubicin. These findings suggested an indirect interaction and regulation between MX-106-4C and P-gp. Further study revealed that the selective cytotoxic effects of MX-106-4C were associated with cell cycle arrest at G1 phase and apoptosis through the downregulation of CDK4. Overall, this study demonstrates that MX-106-4C selectively kills P-gp positive MDR colon cancer cells and indirectly regulates P-gp, which provides a clue for CS compound design and a novel strategy to obviate P-gp-mediated colon cancer MDR by re-sensitizing heterogeneous tumors with CS agents.

Citation Format: Zi-Ning Lei, Zhongzhi Wu, Qiu-Xu Teng, Min Xiao, Wei Li, John N. Wurpel, Zhe-Sheng Chen. Selective toxicity of MX-106-4C, a survivin inhibitor, in P-glycoprotein-mediated multidrug resistant colon cancer [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 5264.
Abstract 5287: TGF-β-acetylated KLF5-Bcl-2 signaling mediates docetaxel resistance in prostate cancer cells
Authors: Li Yixiang , Zhang Baotong , Dong Jin-Tang
Published: September 2020

Purpose: In this study, we explored the role of KLF5 in TGF-β-induced drug resistance in prostate cancer cells to identify novel therapeutic targets.

Procedures: We used DU 145 and PC-3 prostate cancer cell lines in in vitro cell survival assay, colony survival assay, and xenograft mouse model to measure docetaxel (DTX) resistance. TGF-β1, the SB505124 TGF-β receptor I inhibitor, and the ABT-199 Bcl-2 inhibitor were used to determine if Bcl-2 mediates TGFβ induced docetaxel resistance. To measure the function of KLF5 and its acetylation at lysine 369 in drug resistance, stable cell lines were constructed with different forms of KLF5 after the endogenous KLF5 was deleted with the CRISPR-Cas9 system.

Results: We found that TGF-β induced DTX resistance depends on KLF5 expression. As a downstream molecule of TGF-β signaling, acetylated KLF5 at lysine 369 mediated TGF-β-induced DTX resistance. Moreover, acetylated KLF5 induced DTX resistance in xenograft mouse model. Mechanistically, we found that TGF-β and acetylated KLF5 upregulated Bcl-2 transcription, and inhibition of Bcl-2 sensitized cells to DTX. In addition, DTX promoted Bcl-2 degradation via a proteasome dependent pathway, and TGF-β signaling stabilized Bcl-2 via inhibition of Bcl-2 ubiquitination. Lastly, the TGF-β/acetylated KLF5/Bcl-2 signaling axis induced survival disadvantage in prostate cancer patients.

Conclusions: TGF-β induces transcriptional upregulation of Bcl-2 through acetylated KLF5 at lysine 369, and TGF-β stabilizes Bcl-2 via inhibition of DTX-induced Bcl-2 ubiquitination. The TGF-β-acetylated KLF5-Bcl-2 signaling axis is thus a novel mechanism mediating DTX resistance in prostate cancer.

Citation Format: Yixiang Li, Baotong Zhang, Jin-Tang Dong. TGF-β-acetylated KLF5-Bcl-2 signaling mediates docetaxel resistance in prostate cancer cells [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 5287.
Abstract 529: Oncostatin M receptor as a therapeutic target of radio-immune therapy in metastatic synovial sarcoma
Authors: Luelling Sarah
Published: September 2020

Synovial sarcoma is a soft tissue malignancy of the muscle tissue which primarily affects adolescents. While it is a rare disease, little advancement has been made in the treatment of this cancer. With an overall survival rate of roughly 40%, the need for new treatments for synovial sarcoma is evident. Oncostatin M Receptor (OSMR) is a type I cytokine receptor and is overexpressed in metastatic synovial sarcoma. OSMR does not have high expression in normal tissues, making it an ideal target for cancer therapy. We hypothesize that by using an anti-OSMR monoclonal antibody conjugated to a radioactive Cu67 isotope, synovial sarcoma can be targeted at both primary and metastatic locations through systemic therapy. Copper 67 is a gamma radiation emitting isotope which is tissue damaging and able to induce cell death in cancer cells. By conjugating the chelating molecule p-SCN-Bn-NOTA to an anti-OSMR antibody, Cu67 was able to be captured to the antibody. Capture efficiency of Cu67 was measured through TLC and found to be 64 % efficient. Binding studies through Surface Plasmon Resonance (SPR) found that the conjugation did not interfere with the binding affinity of the antibody. This data suggests that targeting OSMR through radio-immune therapy is a viable treatment and indicates further testing in animal models.

Citation Format: Sarah Luelling. Oncostatin M receptor as a therapeutic target of radio-immune therapy in metastatic synovial sarcoma [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 529.
Abstract 5293: A molecular classification of gastric cancer in Chilean patients
Authors: Pinto Mauricio P. , Muñoz-Medel Matias , Cordova-Delgado Miguel , Retamal Ignacio N. , Owen Gareth , Bravo Maria Loreto , Armisen Ricardo , Garrido Marcelo
Published: September 2020

Objective: Incidence and mortality rates of gastric cancer (GC) are characterized by their geographical heterogeneity. In Chile, GC is the leading cause of cancer death. To date, GC patients' response to standard therapies remains limited. A molecular classification of GCs may deliver better stratifications. Herein, we obtained clinical data, protein expression and genetic profiles in a cohort of Chilean patients and present a molecular classification that correlated with overall survival (OS) rates.

Methods: A total of 71 patients were included. Clinical data were obtained from medical records; protein expression was analyzed by a Tissue MicroArray. We also performed Next Generation Sequencing to assess p53 status (WT or Mut). Supervised clustering was used to generate a molecular classification. Kaplan-Meier method was used to calculate OS. Hazard ratio was calculated by Cox regression.

Results: We defined 5 GC subgroups: Epstein-Barr virus+ (EBV, n=9; 13%), Microsatellite Instable (MSI, n=9; 13%), E-cadherin loss (EMT-like, n=12; 17%). The remaining patients (those MSS-/EBV-/not EMT-like) were classified either as p53 WT (n=21; 30%) or p53 Mutated (n=20; 28%). According to subgroups, 5-year survival rates were: MSI=77,8%; EBV=43.2%; p53WT=43.5%; p53Mut=25% and EMT-like=16.7%. Hazard ratios for p53Mut and EMT-like were 5.1 (IC 95%: 1.16-22.41; p=0.031) and 6.81 (IC 95%: 1.48-31.24; p=0.014), respectively against the MSI group used as reference. This association is maintained in a multivariate model using age, gender and stage.

Conclusions: Our study defined 5 GC subgroups. These are associated to OS rates. EMT-like and p53Mut subgroups displayed poorer survival. Future studies should explore actionable targets in these subsets in order to improve their survival.

Citation Format: Mauricio P. Pinto, Matias Muñoz-Medel, Miguel Cordova-Delgado, Ignacio N. Retamal, Gareth Owen, Maria Loreto Bravo, Ricardo Armisen, Marcelo Garrido. A molecular classification of gastric cancer in Chilean patients [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 5293.
Abstract 5321: Novel imipridone ONC206 inhibits cell proliferation and induces apoptosis in uterine serous cancer through altering MAPK/AKT signaling network and metabolic reprogramming
Authors: Yeung Chi Lam Au , Hu Wen , Zhang Li , Ferri-Borgogno Sammy , Tarapore Rohinton S. , Allen Joshua E. , Lu Karen H. , Mok Samuel C.
Published: September 2020

Uterine serous carcinoma (USC) is the most aggressive subtype of endometrial cancer. Patients with advanced stage USC have poor survival rates and new treatment regimens are lacking. ONC201 is the first clinical bitopic antagonist of dopamine receptor D2 (DRD2), that is well tolerated and currently being investigated in several clinical trials for oncology. ONC206 is a chemical derivative of ONC201 with the same impridone core structure, which is also a DRD2 antagonist that exhibits distinct receptor pharmacology and nanomolar potency in various preclinical cancer models. However, the effects of ONC206 on USC progression and the mechanism of action have not been thoroughly explored. Using both in vitro and in vivo models, and multiple USC cell lines, the effects of ONC206 on cell proliferation and apoptosis were determined. Reverse phase protein arrays (RPPAs) and Western blot analyses were used to determine the effect of ONC206 on the expression of key proteins in various signaling networks in ONC206 treated USC cells. The results showed that ONC206 suppressed USC cell proliferation and induces apoptosis in a dose-dependent manner. Luciferase labeled USC cell ARK1-bearing mice treated with 100mg/kg ONC206 had significantly lower chemiluminescent signals than those treated with the control buffer. RPPA data showed that ONC206 treated USC cells had markedly lower expression signals in p38MAPK, p-AKT, p-S6, and multiple mitochondrial proteins associated with mitochondrial ATP synthesis including MTCO1, and TFAM than the control cells did. Significantly lower ATP levels and cytochrome c oxidase activities in ONC206 treated USC cells than in control cells were demonstrated by luminescent ATP detection assay kit and the cytochrome c oxidase assay kit. These data suggest that ONC206 suppresses USC progression through inhibiting MAPK/AKT network, which subsequently leads to metabolic reprogramming and increased apoptosis. Moreover, by knocking-out DRD2, USC cells became more resistant to ONC206 treatment. This suggested that the effect of ONC206 is likely mediated through its binding to DRD2. ONC206 also showed a synergistic effect with paclitaxel in vitro. Further studies which demonstrate the optimal dosage and the efficacy of treatment of USC using ONC206 are warranted.

Citation Format: Chi Lam Au Yeung, Wen Hu, Li Zhang, Sammy Ferri-Borgogno, Rohinton S. Tarapore, Joshua E. Allen, Karen H. Lu, Samuel C. Mok. Novel imipridone ONC206 inhibits cell proliferation and induces apoptosis in uterine serous cancer through altering MAPK/AKT signaling network and metabolic reprogramming [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 5321.
Abstract 5324: Structure-based design of CDC42/RHOJ effector inhibitors for the treatment of cancer
Authors: Flesher Jessica L. , Jahid Sohail , Ortega Jose A. , La Sala Giuseppina , Brindani Nicoletta , Arencibia Jose M. , Manigrasso Jacopo , Hachey Stephanie , Chen Chi-Fen , Hughes Chris , De Vivo Marco , Ganesan Anand K.
Published: September 2020

The CDC42 family of GTPases (RHOJ, RHOQ, CDC42) control both the ability of tumor cells to invade surrounding tissues and the ability of endothelial cells to vascularize tumors. While recent studies have developed small molecules that target RAS GTPases, little progress has been made in targeting the related CDC42 family of GTPases for cancer treatment. Here, we use computer-aided drug design to identify a novel class of inhibitors that act on an allosteric pocket in the active form of the CDC42 GTPase, RHOJ. These allosteric inhibitors prevent RHOJ and CDC42 from binding to their downstream effector PAK, while having no effect on the interactions between the closely related GTPase RAC1 and PAK or RAS and its downstream effector RAF. Our lead compound ARN22089 has a druglike profile and can block both tumor growth and tumor angiogenesis in a three-dimensional vascularized microtumor (VMT) model, indicating that ARN22089 blocks RHOJ/CDC42 signaling in both the tumor cell and the tumor endothelium. Short term treatment of nascent melanoma tumors with ARN22089 halted the growth of BRAF mutant autochthonous mouse melanoma tumors, slowed the growth of melanoma patient-derived xenografts, and induced tumor necrosis in PDX models. In summary, we describe a multidisciplinary structure-based drug discovery platform that can identify new RHO family allosteric inhibitors and use this system to identify RHOJ inhibitors that block tumor growth in vivo.

Citation Format: Jessica L. Flesher, Sohail Jahid, Jose A. Ortega, Giuseppina La Sala, Nicoletta Brindani, Jose M. Arencibia, Jacopo Manigrasso, Stephanie Hachey, Chi-Fen Chen, Chris Hughes, Marco De Vivo, Anand K. Ganesan. Structure-based design of CDC42/RHOJ effector inhibitors for the treatment of cancer [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 5324.
Abstract 5339: Paradoxical androgen receptor regulation by small molecule enantiomers
Authors: Patsch Katherin , Liu Chao , Kashemirov Boris , Sura Harish , Zapotoczny Grzegorz , Sun Yuanye , Shah Pavan P. , Katz Jonathan E. , Chilakala Sujatha , Joo Ah Young , Ung Nolan , Aljehani Maya , Sun Ren X. , Agus David B. , Ruderman Daniel , McKenna Charles E.
Published: September 2020

Small molecules that target the androgen receptor (AR) are the mainstay of therapy for lethal castration-resistant prostate cancer (CRPC), but they ultimately lead to resistance and progression to terminal disease. While individual resistance-associated AR mutations have been identified, the underlying mechanisms remain poorly understood. Here, we report a new class of AR-targeting compounds derived from the anti-androgen BMS-641988 (N-((3aR,4R,5R,7R,7aS)-2-(4-cyano-3-(trifluoromethyl)phenyl)-4,7-dimethyl-1,3-dioxooctahydro-1H-4,7-epoxyisoindol-5-yl)ethanesulfonamide) that reveal unexpected biological behavior: as expected, the C-5 (R)-enantiomers are antagonists, however the corresponding (S)-enantiomers, including the previously unknown C-5 (S)-enantiomer of BMS-641988 itself, are potent agonists of AR, with ED50 values comparable to dihydrotestosterone and R1881. A series of in vitro assays that interrogate different steps of AR functionality–including AR binding via fluorescence polarization, nuclear translocation and hyperspeckling via live cell confocal microscopy, ARE-luciferase, and RT qPCR arrays–confirm that the (R)-enantiomers are potent antagonists of AR in multiple cell lines, including AR T877A mutation-expressing LNCaP and AR overexpressing VCaP. While these cells are virtually resistant to BMS-641988, two of the novel derivatives, (R)-USC-1702 and (R)-USC-1707, inhibited their growth with IC50 values in the lower nM range. Importantly, these compounds do not produce the toxic metabolite that led to the clinical failure of BMS-641988. This is the first documented evidence of small molecule enantiomer pairs showing opposite functional effects while targeting AR. This new AR regulation duality augments our understanding of AR function and may have implications for anti-androgen drug development.

Citation Format: Katherin Patsch, Chao Liu, Boris Kashemirov, Harish Sura, Grzegorz Zapotoczny, Yuanye Sun, Pavan P. Shah, Jonathan E. Katz, Sujatha Chilakala, Ah Young Joo, Nolan Ung, Maya Aljehani, Ren X. Sun, David B. Agus, Daniel Ruderman, Charles E. McKenna. Paradoxical androgen receptor regulation by small molecule enantiomers [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 5339.
Abstract 534: Discovery of an anti-claudin18.2/anti-PD-L1 bispecific antibody SPX-301 in the SMARTOPTM format
Authors: Zhu Guidong , Ye Jingdong , Ma Jichun , Qin Jingdong , Cai Yi
Published: September 2020

This research aimed to produce anti-claudin18.2 (CLDN 18.2)/anti-PD-L1 bispecific antibodies to enhance the therapeutic index and expand the responsive population of the parent anti-CLDN 18.2 and anti-PD-L1 antibodies through the mechanisms of tumor-specific distribution and synergistic tumor killing. CLDN 18.2 is highly expressed in several tumors but is minimally expressed in normal tissues and a chimeric IgG1 CLDN 18.2 antibody zolbetuximab significantly improved PFS and OS in gastric cancer patients in a CLDN 18.2-dependent manner on the background of EOX. However, the response rate was moderate (~10%), leaving room for further improvement. Anti-PD-L1 antibodies showed remarkable efficacy in lung cancer and melanoma but in various settings of gastric cancer they showed weaker benefits, which warrant optimization. We designed anti-claudin18.2/anti-PD-L1 bispecific antibodies in the SMARTOPTM format, where a single-chain diabody containing optimized CLDN 18.2 and PD-L1 binding variable domains is connected to the Fc domain by linkers that can be optimized by phage display libraries. The bispecific antibodies were selected by affinity at low pH values and slow off-rate to improve the therapeutic window by enhancing tumor retention and preserving or improving binding affinity in the low pH tumor microenvironment. Immunogenicity was optimized by mutating the surface residues and disulfide bonds were inserted to improve stability. A representative bispecific antibody SPX-301 showed comparable binding affinity to the CLDN 18.2 and PD-L1 antigens to the parent antibodies (SPX-101 and durvalumab, respectively). SPX-301 represents a novel anti-claudin18.2/anti-PD-L1 bispecific antibody in the SMARTOPTM format. The format demonstrated excellence retention of antigen binding affinity, high expression levels, minimal aggregation, and straightforward purification. The preclinical characterizations of SPX-301 will also be presented.

Citation Format: Guidong Zhu, Jingdong Ye, Jichun Ma, Jingdong Qin, Yi Cai. Discovery of an anti-claudin18.2/anti-PD-L1 bispecific antibody SPX-301 in the SMARTOPTM format [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 534.
Abstract 535: T-cell recruitment tumor lysis via a novel c-MET/CD3 bispecific antibody
Authors: Huang Lei , Xie Kun , Wang Ruiqin , Gu Hua , Fang Jianmin
Published: September 2020

Anti-receptor tyrosine kinases (RTKs) therapy has been used for the treatment of a number of solid tumors in clinic. C-MET, also known as the hepatocyte growth factor receptor (HGFR), is a representative member of the RTK family. C-MET overexpression is commonly observed in tumor biopsies and associated with poor patient survival. Various kinase inhibitor and monoclonal antibody (e.g. Emibetuzumab and Onartuzumab) have been developed to block the HGF/c-MET interactions, however, those kinase-inhibiting-based therapeutics have shown limited success in clinical trials. There is an urgent need to develop new treatments targeting c-MET to overcome these limitations. Bispecific antibodies that recruit immune cells to the tumor microenvironment have been considered as effective therapeutic candidates. An example of this design strategy is the FDA-approved bispecific antibody blinatumomab. In this study, we have constructed an asymmetric bispecific antibody, which binds both c-MET and CD3, using a novel platform that is monovalent to epitope. Our c-MET/CD3 bispecific antibody showed potent T-cell mediated tumor cells killing in vitro. Meanwhile, this antibody inhibited the phosphorylation of c-MET and downstream signal transduction in tumor cells due to its monovalent nature. In c-MET-overexpressing lung cancer and ovarian cancer xenograft models, c-MET/CD3 bispecific antibody inhibited tumor growth and increased the proportion of activated CD56+ tumor infiltrating lymphocytes. Moreover, in vivo combination therapy with FDA-proved PD-1/PD-L1 antibodies showed higher tumor inhibition than monotherapy in above two tumor models. Collectively, the novel c-MET/CD3 bispecific antibody can provide immunotherapy for c-MET-overexpressing tumors which conventional antibody or small molecular inhibitor might not. These findings also support the immunotherapeutic effects on combination of T-cell dependent bispecific antibody and immune checkpoint blockade.

Citation Format: Lei Huang, Kun Xie, Ruiqin Wang, Hua Gu, Jianmin Fang. T-cell recruitment tumor lysis via a novel c-MET/CD3 bispecific antibody [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 535.