bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5141: Using single cell gene expression to develop treatment response predictions in multiple myeloma
Authors: Scolnick Jonathan , Xu Stacy , de Mel Sanjay , Soekojo Cinnie Yentia , Chng Wee Joo
Published: June 2022

The promise of precision medicine in oncology is to identify the ideal treatment for each patient, eliminating failed treatment cycles and reducing treatment burdens on patients and payors. While gene sequencing can predict effective treatment options in some cases, studies have estimated that only 10-15% of cancer patients are treated with genotype matched drugs. Additional treatment selection tools for predicting patient response thus remains a critical clinical need.

While gene sequencing has clear utility in predicting therapeutic response, gene expression is less commonly used in clinical tests to guide treatment selection. Gene expression is theoretically attractive to predicting therapeutic response because, unlike mutational analysis, it provides a direct readout of the dysregulated biochemical activities in the cells which may be targeted by particular therapies. One of the major problems with conventional approaches to using gene expression to predict therapeutic response is tumor heterogeneity; that is, a biopsied tumor contains multiple cells types and even different tumor clones that can confound a conventional bulk gene expression analysis. More recently, single-cell RNA expression analysis technologies have grown in popularity. These technologies enable the fine characterization of tumors and their cellular makeup and hold the promise of increasing the power of gene expression measurements for identifying therapy response prediction tools.

Here we show the application of single cell gene expression analysis with Proteona’s MapResponse™ machine-learning algorithms to developing a predictive classifier for patient response to Daratumumab treatment in multiple myeloma. Starting with publicly available data (Cohen et al. Nat. Medicine 2021), we developed a classifier that accurately predicted the response of 94% of subjects in the published study. This classifier was able to accurately predict response in an independent cohort of Daratumumab-treated patients. In contrast, the classifier performed poorly in a cohort of patients who did not receive Daratumumab. Together these data suggest the classifier has specificity to predicting response to Daratumumab and is not a general prognostic signature. These early findings point to the potential value of combining single-cell RNA sequencing with machine-learning methods such as MapResponse™ to bring precision medicine to more patients.

Citation Format: Jonathan Scolnick, Stacy Xu, Sanjay de Mel, Cinnie Yentia Soekojo, Wee Joo Chng. Using single cell gene expression to develop treatment response predictions in multiple myeloma [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 5141.
Abstract 5144: Using proteomic approach to identify potential tumor-associated antigens (TAAs) as biomarkers in Hispanic hepatocellular carcinoma
Authors: Ma Yangcheng , Qiu Cuipeng , Zhang Xiaojun , Wang Xiao , Zhang Jianying
Published: June 2022

Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide with high prevalence and poor prognosis. The incidence rate and mortality rate in Hispanic HCC is relative higher than other ethnic groups. Autoantibodies against tumor associated antigens (TAAs) have long been proved as useful biomarkers in cancer diagnosis. In the present study, sera from 25 Hispanic patients with HCC and 25 sera from normal individuals were used in a proteomic approach to identify HCC-related TAAs. Twenty-four immunoreactive protein spots (16 from HepG2 cells, and 8 from SUN449 cells) were excised from the two-dimensional electrophoresis (2DE) gels, digested with trypsin, and subsequently analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Of 24 immunoreactive protein spots, 21 were identified with compatible molecular weights and isoelectric points. After further analysis, a total of 9 proteins were found to be potential TAAs, including endoplasmic reticulum chaperone BiP (HSPA5), 60 kDa heat shock protein (HSP60), triosephosphate isomerase (TPI1), protein disulfide-isomerase (PDI), Actin-cytoplasmic 2 (ACTG1), calreticulin (CRP55), alpha-enolase (ENO1), heat shock 70 kDa protein (HSP70), tropomyosin alpha-3 chain (TPM3). Among these proteins, HSPA5 was corresponded to 5 protein spots on the 2DE gels, while 4 proteins (HSPA5, HSP60, TPI1 and PDI) were found in both HepG2 and SNU449 cell lines. Enrichment analysis show that these proteins are mostly responsible for protein folding in endoplasmic reticulum as well as ubiquitin ligase binding function. In the subsequent study, autoantibodies against these proteins will be detected by immunoassay, and further evaluated as biomarkers in detection of Hispanic HCC.

Citation Format: Yangcheng Ma, Cuipeng Qiu, Xiaojun Zhang, Xiao Wang, Jianying Zhang. Using proteomic approach to identify potential tumor-associated antigens (TAAs) as biomarkers in Hispanic hepatocellular carcinoma [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 5144.
Abstract 5175: Novel approach for glioblastoma characterization via tumor specific extracellular vesicles
Authors: Hsia Tiffaney , Yekula Anudeep , Batool Syeda M. , Carter Bob S. , Balaj Leonora
Published: June 2022

INTRODUCTION: The utility of photosensitizers (PS), such as 5-Aminolevulinic acid (5-ALA), has been limited to fluorescence-guided surgery (FGS) and photodynamic therapy. The downstream metabolism of 5-ALA results in tumor-specific, highly selective accumulation of PpIX, an endogenous porphyrin, in tumor cells. We have previously reported evidence of PpIX in EVs derived from the plasma of glioblastoma (GBM) patients. Here, we characterize the tumor-specific nature of PpIX fluorescent EVs through quantification of their cargo, demonstrating their utility in the advancement of liquid biopsy for GBM diagnosis and management.

METHODS: Fluorescence activated cell sorting was adapted for nanoparticle isolation and PpIX fluorescent EVs were sorted from plasma collected from patients (N = 8) diagnosed with GBM, undergoing FGS. RNA cargo was extracted and quantified via low input RNA-Seq. Matching patient tumor and total plasma EVs along with healthy control plasma EVs (N = 8) were sequenced in parallel.

SUMMARY: Through sequencing analysis, we have demonstrated a distinct landscape of long RNAs (mRNA and lncRNA) in PpIX EVs, with isolation of this EV population resulting in an enrichment of lncRNAs. Considering the emerging diagnostic and prognostic potential of lncRNAs in GBM, exploration of such EVs provides valuable insight into the pathobiology of the tumor of origin. On a broader scale, gene expression analysis of PpIX EV cargo demonstrates at least a ten-fold enrichment of genes that are otherwise buried in the heterogeneous plasma EV background. Furthermore, downstream KEGG orthological analysis of PpIX EV cargo has revealed alignment of 78% of the significant pathways to processes implicated in cancer development and progression. We have identified a panel of genes unique to PpIX EVs, and reflective of the native tumor, that are implicated in GBM pathogenesis including GREM1, MAP4K4, and STAG2. Validation via quantitative and digital PCR of total plasma EVs from patient and healthy cohorts (N = 10, respectively) have yielded similar expression levels, as reflected in sequencing gene expression. Isolation of PpIX EVs, however, insinuates enriched genetic detection.

CONCLUSION: In summary, this preliminary cohort of GBM patients has demonstrated the utility of 5-ALA induced PpIX fluorescence outside of its original intentions: facilitating tumor-specific EV identification for the progression of liquid biopsy. These findings are an inviting breakthrough in the development of individualized disease detection and monitoring.

Citation Format: Tiffaney Hsia, Anudeep Yekula, Syeda M. Batool, Bob S. Carter, Leonora Balaj. Novel approach for glioblastoma characterization via tumor specific extracellular vesicles [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 5175.
Abstract 5243: 3-Bromopyruvate in combination with radiation inhibits pancreatic cancer growth by dismantling mitochondria and ATP generation in a preclinical mouse model
Authors: Roy Sanjit K. , Dukic Tijana , Bhandary Binny , Acharya Aneesh , Tu Kevin J. , Ko Young H. , Shukla Hem D.
Published: June 2022

3-Bromopyruvate (3 BP) is a promising anticancer drug against pancreatic cancer. It exerts potent anticancer effects by inhibiting hexokinase II enzyme of glycolysis pathway and ATP generation in cancer cells. In present investigation we have examined the effect of 3-BromoPyruvate employing panc-2 cancer cells, and syngeneic pancreatic cancer mouse model. The clonogenic survival assay demonstrated dose dependent inhibition of cell growth, and we observed 95% cell death at 15 uM conc. Further, ATP production was severely inhibited in cancer cells treated with same concentration of 3-BP. We also observed that 3-BP in combination with low doses of irradiation was more effective in killing cancer cells than 3-BP alone. Electron microscopy revealed that 3-BromoPyruvate severely damaged mitochondrial integrity which might have severely affected ATP generation in cancer cells. We further tested 3-BromoPyruvate (2.5-10.0 mg/kg) in syngeneic panc-2 tumor mouse model and observed that 3-BP at10.0 mg/kg dose significantly reduced tumor growth (volume) about 35-40%. Immunohistochemistry (IHC) data showed that 3-BP inhibits hexokinase II (HK2) and TGFbeta1 and enhanced active caspase-3 expression in tumor tissues as compared to untreated control. The disruption of the interaction between VDAC1 and HK2 due to 3BP treatment was confirmed by CO-IP. Interestingly, 3 BP at higher dose induces caspase-3 activation in tumor derived organoid without showing any effect on normal pancreatic organoid. Altogether, the results in this study demonstrated that 3 BP could be a promising anti-tumor drug against pancreatic cancer and its efficacy could be improved in combination with radiation therapy.

Citation Format: Sanjit K. Roy, Tijana Dukic, Binny Bhandary, Aneesh Acharya, Kevin J. Tu, Young H. Ko, Hem D. Shukla. 3-Bromopyruvate in combination with radiation inhibits pancreatic cancer growth by dismantling mitochondria and ATP generation in a preclinical mouse 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 5243.
Abstract 5259: Clinical feature of end-stage cerebral glioblastomas
Authors: Saito Norihiko , Hirai Nozomi , Kushida Naoki , Sato Sho , Hiramoto Yu , Fujita Satoshi , Nakayama Haruo , Hayashi Morito , Ito Keisuke , Sakurai Takatoshi , Iwabuchi Satoshi
Published: June 2022

Glioblastoma is the most common and aggressive primary brain tumor, and even with the best treatment, the tumor repeatedly recurs and grows, eventually invading the entire brain. There are few reports on the pathogenesis and pathological search in the terminal stage of glioblastoma. In this study, we report the pathological analysis of glioblastoma cases that we were able to perform from initial treatment to confirmation of death at our hospital. We analyzed 26 glioblastoma cases (including 18 autopsied cases) treated at our hospital from initial treatment to death. The mean age of the 26 patients was 60.7 years, and the mean overall survival was 16.7 months. The duration of clinical symptoms was 36.2 days from coma to death, and 12 days from the appearance of respiratory depression to death. Psychiatric symptoms and central fever were observed in patients with intrathecal dissemination. Steroids and antiepileptic drugs were often continued after completion of active treatment. Patients with intrathecal dissemination progressed rapidly and presented with a variety of symptoms, including psychiatric symptoms, headache, neck pain, and central fever. In addition, there was a case of diffuse infiltration from the brain parenchyma to the periventricular area in a patient treated with bevacizumab, suggesting the possibility of a change in the form of recurrence. In the terminal stage of glioblastoma, hypoxemia due to disturbance of respiratory center progresses from impaired consciousness to death. Convulsive seizures are rare in the dying stage, and continuation of antiepileptic drugs should be considered. Although many patients develop from local recurrence, it was suggested that new treatment may change the mode of recurrence or alter the trait of tumor cells. In recent years, the number of patients receiving home care and end-of-life care has been increasing due to the improvement of medical systems such as home care. Further investigation of the pathophysiology is necessary for better end-of-life care.

Citation Format: Norihiko Saito, Nozomi Hirai, Naoki Kushida, Sho Sato, Yu Hiramoto, Satoshi Fujita, Haruo Nakayama, Morito Hayashi, Keisuke Ito, Takatoshi Sakurai, Satoshi Iwabuchi. Clinical feature of end-stage cerebral glioblastomas [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 5259.
Abstract 526: Novel methylation-based, tissue-free ctDNA assay accurately quantifies longitudinal tumor burden changes for precision treatment monitoring
Authors: Ye Patrick Peiyong , Viens Robb , Bower Xavier , Riku Shan , Tsao David , Atay Oguzhan
Published: June 2022

As novel cancer treatments become available, the need to quickly and accurately evaluate whether these treatments are effective remains unaddressed. Obtaining earlier feedback on the efficacy of a cancer therapy could prevent a poor treatment outcome by switching to a more effective therapy sooner. Levels of circulating tumor DNA (ctDNA) have been found to be prognostic of tumor progression, suggesting that a non-invasive liquid biopsy assay could provide longitudinal ctDNA measurements that accurately track tumor progression. However, while there is interest in using existing minimal residual disease (MRD) detection and treatment selection liquid biopsy assays for treatment monitoring applications, they both suffer from limitations in their ability to precisely and sensitively quantify trends in tumor progression over the course of treatment. In addition, tumor-informed MRD detection assays are often infeasible for treatment monitoring due to unavailability of the initial tissue sample.

We have developed and validated a novel methylation-based liquid biopsy assay for pan-cancer treatment monitoring without the need to obtain a sample from the tumor itself. Methylation has long been shown to be a strong and consistent biomarker for cancer, and because tumor tissue has widespread differential methylation across the genome compared to normal tissue, we are able to overcome molecule sampling limitations. In analytical validation, our assay could detect as small changes as 0.05%, e.g., an elevation of tumor fraction from 0.5% to 0.55%, a level of precision that is an order-of-magnitude better than any available assays. Next, we tested our assay on clinical specimens by measuring ctDNA in serially collected blood samples from subjects with cancer. As would be expected from the high level of precision, we were able to successfully demonstrate the predictive power of ctDNA measurements for clinical outcomes. These results from our methylation-based liquid biopsy assay open the path to earlier and extremely precise treatment monitoring for oncologists and their patients.

Citation Format: Patrick Peiyong Ye, Robb Viens, Xavier Bower, Shan Riku, David Tsao, Oguzhan Atay. Novel methylation-based, tissue-free ctDNA assay accurately quantifies longitudinal tumor burden changes for precision treatment monitoring [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 526.
Abstract 5279: Prognostic serum biomarkers in cancer patients with COVID-19: A systematic review
Authors: Lai Yun-Ju , Lee Te-An , Wang Shih-Han , Li Chia-Wei , McCullough Louise D. , Bello Dhimiter
Published: June 2022

Purpose: COVID-19 has led to 4 million deaths worldwide since 2019. COVID-19 patients with cancers likely express biomarker changes in circulation. While many biomarker studies focused on COVID-19 diagnosis and prognosis, the panel of biomarkers used in SARS-CoV-2 infected cancer patients for COVID-19 severity and prognosis are largely unclear. Therefore, this systematic review aims to determine what biomarkers have been measured in cancer patients with COVID-19 and their prognostic utility.

Methods: A systematic literature review in PubMed, Embase, and Scopus was performed on June 16th, 2021. The search keywords coronavirus, neoplasm, biomarkers, and disease progression were used to filter out 17 eligible studies, which were then carefully evaluated.

Results: A total of 4,168 patients from 17 eligible articles were included in this study. Sixteen types of cancer and 60 biomarkers were identified. The majority of changed biomarkers in the cancer patients with COVID-19 compared to the healthy group and non-cancer patients with COVID-19 were biochemical and inflammatory markers. The up-regulated markers, including CRP, D-dimer, ferritin, IL-2R, IL-6, LDH, and PCT, were identified in eligible studies. Albumin and hemoglobin were significantly down-regulated in cancer patients with COVID-19. Additionally, we observed that the SARS-CoV-2 infected cancer patients with lower levels of CRP, ferritin, and LDH successfully survived from antiviral drug and immunotherapy for COVID-19 treatments.

Conclusion: Several important clinical biomarkers, such as CRP, ferritin, and LDH, may serve as the prognostic markers to predict the outcomes following COVID-19 treatment and monitor the deterioration of COVID-19 in cancer patients.

Citation Format: Yun-Ju Lai, Te-An Lee, Shih-Han Wang, Chia-Wei Li, Louise D. McCullough, Dhimiter Bello. Prognostic serum biomarkers in cancer patients with COVID-19: A systematic review [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 5279.
Abstract 5289: Activating p53 through MDM2 and WIP1 inhibition effectively inhibits tumors in liver adenocarcinoma
Authors: Chen Chiao-Ping , Yeh Chun-Nan , Pan Yi-Ru , Hsiao Yu-Tien , Lo Chih-Hong , Jhuang Cai-Jhen , Wu Chiao-En
Published: June 2022

Intrahepatic cholangiocarcinoma (iCCA) is an adenocarcinoma arising from the intrahepatic bile duct and accounts for the second most cases of primary liver cancers after hepatocellular carcinoma. Lacking effective treatment leads to a poor prognosis for advanced iCCA so new targeted therapy is warranted. Impaired wild-type (WT) p53 tumor suppressor function by its negative regulators frequently occurs in iCCA. Therefore, restoration of WT p53 function by inhibiting its negative regulators is a therapeutic strategy being explored for cancer treatment. Both MDM2 inhibitor (MDM2i, RG7388) stabilizing p53 and WIP1 inhibitor (WIP1i, GSK2830371) increasing p53 phosphorylation lead to maximize p53 function. The combination of MDM2i/WIP1i has been reported in severe cancer types but no in vivo study has been done. In the current study, both liver adenocarcinoma cell lines, RBE and SK-Hep-1, were treated with the RG7388 alone and in combination with GSK2830371. Cell proliferation, clonogenicity, protein and mRNA expressions, and cell cycle distribution were performed to investigate the effect and mechanism of growth suppression. To evaluate the antitumor efficacy of RG7388 and GSK2830371 in vivo, xenografts with SK-Hep-1 cells were treated with combination therapy for two weeks in NOD-SCID mice. The combination of MDM2i and WIP1i significantly increased the growth inhibition, cytotoxic activity, increased p53 protein expression, and phosphorylation (Ser 15), leading to transactivation of downstream targets (p21 and MDM2). The in vivo test demonstrated that the combination treatment can significantly inhibit tumor growth. In this study, liver adenocarcinoma cell lines were inhibited by the combination via reactivation of p53 function evidenced by increased p53 expression, phosphorylation and its downstream targets. This efficacy was also validated by in vivo study. The current research provides a novel strategy for targeting the p53 pathway in liver adenocarcinoma.

Citation Format: Chiao-Ping Chen, Chun-Nan Yeh, Yi-Ru Pan, Yu-Tien Hsiao, Chih-Hong Lo, Cai-Jhen Jhuang, Chiao-En Wu. Activating p53 through MDM2 and WIP1 inhibition effectively inhibits tumors in liver adenocarcinoma [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 5289.
Abstract 5299: A therapeutic antibody against G-protein-coupled receptor class 5 member D to treat multiple myeloma
Authors: He Feng , Wu Kefei , Liang Jiabei , Li Jianhui , Peng Hao , Wu Guojin , Ning Jinying
Published: June 2022

Multiple myeloma (MM) comprised 0.9% of all new cancer cases and 1.2% of cancer death in 2020 and is the second most common hematological neoplasm in adults. Primary refractory to treatment and relapse resulted in a high death rate of MM patients. Bispecific antibodies (BsAbs) are attracting candidate drugs against multiple myeloma by targeting CD3 on T cells and MM specific cell surface markers such as B cell maturation antigen (BCMA), CD38, and G Protein-Coupled Receptor Class C Group 5 Member D (GPRC5D). In the current study, we developed a humanized antibody targeting GPRC5D (clone number G-02-9D7). The antibody was derived from hybridoma of mouse immunized with GPRC5D overexpressed mouse cell line. Compared to the GPRC5D antibody in clinical (JNJ-64407564), this antibody has a higher affinity to 293T cells overexpressing GPRC5D (EC50=1.9nM vs. 3nM), MM cell lines, and MM patient samples. In addition, this antibody did not bind to GPRC5A, B, and C and had weaker non-specific binding to human serum albumin than JNJ-64407564. Bispecific antibody derived from the GRPC5D antibody and CD3 antibody (clone number: sp34) induced specific cytotoxicity of MM cells in the presence of T cells from PBMC. This bispecific antibody stimulated the release of cytokines from T cells such as INF-γ, IL-2, IL-8, and TNF-α. In NCI-H929 prophylactic xenograft mouse model with PBMC transplantation, tumor growth could be prevented by administration weekly with this bispecific antibody as low as 0.1ug per mouse. In the presence of T cells, the bispecific antibody also resulted in complete remission of established MM.1S tumor in xenografted NSG mouse with the injection of 1ug antibody weekly. At last, MM patient bone marrow samples cultured in vitro with this bispecific antibody increased cell death of the CD138+ population. In summary, the GPRC5D antibody could efficiently target Multiple myeloma tumor cells.

Citation Format: Feng He, Kefei Wu, Jiabei Liang, Jianhui Li, Hao Peng, Guojin Wu, Jinying Ning. A therapeutic antibody against G-protein-coupled receptor class 5 member D to treat multiple myeloma [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 5299.
Abstract 5307: Critical platforms of PROTAC validation
Authors: Wang Ruifeng
Published: June 2022

In recent years, proteolysis targeting chimeric (PROTAC) technology has become an effective endogenous protein degradation tool in drug discovery. As a class of heterobifunctional small molecules, PROTACs recruit an E3 ubiquitin ligase to a given substrate protein, resulting in its targeted degradation through the endogenous ubiquitin-proteasome system (UPS). PROTAC consists of three chemical elements: a ligand that binds to a target protein, a ligand that binds to an E3 ubiquitin ligase, and a linker that connects the two ligands. Compared with traditional drug development, PROTAC technology possesses unique advantages including overcoming drug resistance and targeting the “undruggable”. Here we generated a kind of PROTAC target, like KRAS, BRD4, ESR1, associated cell lines to accelerate the discovery of new protein degradation drugs and the target protein degradation therapeutics by an optimized insertion system. In the beginning, we develop a system of different tag targeted cell lines, which can be easy to introduce HiBiT, Flag, GFP and also Halo Tag, et.al to target protein by CRISPaint or CRISPR Cas9 mediated HDR system. We also developed an arbitrary point mutation system based on the optimized PE3 mutation system, which can randomly modify any point mutation to simulate in vivo disease models in vitro. During this process, we have also optimized a series of validation system, like Targeted Protein Degradation (TPD), High-throughput flow cytometry (HTFC) and In-Cell Western (ICW) to shorten the detection time and improve detection stability. It is worth mentioning that we also develop a HiBiT and GFP dual-signal system, which can easily validate the target protein degradation and cell viability. We hope that our detection system can speed up the development of new drugs and contribute to human anti-tumor.Keywords: PROTAC, UPS, drug development, therapeutics

Citation Format: Ruifeng Wang. Critical platforms of PROTAC validation [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 5307.
Abstract 5310: Physiochemical and biological characterization of anti-EGFR based bispecific antibodies in triple negative breast cancer (TNBC) models
Authors: Mohan Nishant , Luo Xiao , Shen Yi , Olson Zachary , Agrawal Atul , Endo Yukinori , Rotstein David , Pelosof Lorraine , Wu Wen Jin
Published: June 2022

Triple-negative breast cancer (TNBC) accounts for approximately 10-20% of all diagnosed breast cancer. Both epidermal growth factor receptor (EGFR) and vascular endothelial growth factor receptor 2 (VEGFR2) frequently overexpress in TNBC and cooperate with each other in autocrine and paracrine manner to enhance tumor growth and angiogenesis. Therapeutic mAbs targeting EGFR (cetuximab) and VEGFR2 (ramucirumab) are approved by FDA for numerous cancer indications, but none of them are approved to treat breast cancers. TNBC cells secrete VEGF-A, which mediate angiogenesis on endothelial cells in a paracrine fashion and promote cancer cell growth in autocrine manner. To disrupt autocrine/paracrine loop in TNBC models in addition to mediating anti-EGFR tumor growth signaling and anti-VEGFR2 angiogenic pathway, we generated a bispecific antibody co-targeting EGFR and VEGFR2 (designated as anti-EGFR/VEGFR2 BsAb), in which cetuximab IgG backbone is connected to the single chain variable fragment (scFv) of ramucirumab via a glycine linker. Physiochemical characterization data shows that anti-EGFR/VEGFR2 BsAb binds to both EGFR and VEGFR2 in a similar binding affinity comparable to parental antibodies. Anti-EGFR/VEGFR2 BsAb demonstrates potent in vitro and in vivo anti-tumor activity in TNBC models. Mechanistically, anti-EGFR/VEGFR2 BsAb not only directly inhibits both EGFR and VEGFR2 in TNBC cells but also disrupts autocrine mechanism in TNBC xenograft mouse model. Furthermore, anti-EGFR/VEGFR2 BsAb blocks paracrine pathway mediated by VEGF/VEGFR2 in endothelial cells. Collectively, our novel findings demonstrate that anti-EGFR/VEGFR2 BsAb inhibits tumor growth via multiple mechanisms of action and has potential to be developed as an attractive targeted therapy for TNBC.

Citation Format: Nishant Mohan, Xiao Luo, Yi Shen, Zachary Olson, Atul Agrawal, Yukinori Endo, David Rotstein, Lorraine Pelosof, Wen Jin Wu. Physiochemical and biological characterization of anti-EGFR based bispecific antibodies in triple negative breast cancer (TNBC) 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 5310.
Abstract 5416: RNA-seq and proteomics to identify response to immunotherapy in advanced melanoma: a Spanish Melanoma Group Study
Authors: Trilla-Fuertes Lucia , Prado-Vazquez Guillermo , Gámez-Pozo Angelo , Lopez-Vacas Rocio , Herrera Maria Isabel Lumbreras , Soriano Virtudes , Garicano Fernando , Lecumberri Maria Jose , Rodriguez Maria , Majem Margarita , Perez Elisabeth , Gonzalez-Cao Maria , Oramas Juana , Magdaleno Alejandra , Fra Joaquin , Martin Alfonso , Corral Monica , Puertolas Teresa , Vara Juan Angel Fresno , Espinosa Enrique
Published: June 2022

Background: Prediction of response to immunotherapy remains an unmet need in the field of advanced melanoma.

Methods: Computational analyses, including probabilistic graphical models, sparse k-means, and consensus cluster, were used to characterize melanoma TCGA samples. The implication of the identified processes in response to immunotherapy was then studied in an independent cohort of 53 patients with advanced melanoma and treated with PD-1 inhibitors. Paraffin samples from this cohort were analyzed using RNA-seq and mass-spectrometry proteomics.

Results: In the TCGA cohort, there were two different layers of information: one related to molecular features of the tumor (based on keratinization, melanogenesis, and extracellular space), and one related to immune status. Therefore, two independent classifications of TCGA melanoma samples were established: molecular and immune. The immune classification distinguished between responders and not responders to immunotherapy in the second cohort (p= 0.0006, HR=6.52). Finally, high-throughput proteomics was used to characterize molecular mechanisms involved in response to immunotherapy, identifying several biological processes and proteins that may be relevant in treatment selection for patients who do not respond.

Conclusions: We established that the immune information was independent of tumor molecular features in melanomas included in the TCGA. An immune classification of these tumors was established. This immune classification predicted response to immunotherapy in a new cohort of patients with advanced melanoma treated with PD-1 inhibitors. Finally, proteomics was used to identify possible targets in those patients who did not respond to immunotherapy.

Citation Format: Lucia Trilla-Fuertes, Guillermo Prado-Vazquez, Angelo Gámez-Pozo, Rocio Lopez-Vacas, Maria Isabel Lumbreras Herrera, Virtudes Soriano, Fernando Garicano, Maria Jose Lecumberri, Maria Rodriguez, Margarita Majem, Elisabeth Perez, Maria Gonzalez-Cao, Juana Oramas, Alejandra Magdaleno, Joaquin Fra, Alfonso Martin, Monica Corral, Teresa Puertolas, Juan Angel Fresno Vara, Enrique Espinosa. RNA-seq and proteomics to identify response to immunotherapy in advanced melanoma: a Spanish Melanoma Group Study [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 5416.
Abstract 5486: ENPP1 inhibitor ZX-8177 enhances anti-tumor activity of conventional therapies by modulating tumor microenvironment
Authors: Li Ying-ying , Peng Zhiyuan , Sun Shuaijun , Guo Kun , Kong Deming , Li Xiaomin , Xie Yuning , Shui Siyu , Wang Yin , Yang Jinfu , Hao Xiaolin Alan , Qin Xiaoli (Shelley)
Published: June 2022

Emerging studies revealed that tumor derived 2’3’-cGAMP could act as an immune transmitter and directly trigger anti-tumor immunity. Damage associated molecular patterns, e.g. damaged genomic DNA, released from injured cancer cells can be converted into 2’, 3’-cGAMP by cGAS. Subsequently, cGAMP triggers STING pathway activation to produce type I interferons (IFNs) and other related innate immunity in the tumor microenvironment. Ecto-nucleotide pyrophosphatase/phosphodiesterase-1 (ENPP1), a membrane-bound nucleotide hydrolase, has been discovered as the predominant enzyme hydrolyzing 2’3’-cGAMP (Li, et al). Hence, ENPP1 blockade in the tumor milieu is a potential strategy against tumors by improving innate immunity response. In this report, we present a potent ENPP1 inhibitor ZX-8177. ZX-8177 showed an IC50 of 9.5 nM in a biochemical assay and an IC50 of 11 nM in MDA-MB-231 cell-based enzymatic assay. In a functional assay for evaluating efficacy of ZX-8177, the production of IFN-β1 by THP-1 cells was measured after the cells were cultured in the conditioned medium collected from MDA-MB-231 cells treated with or without ZX-8177. The interferon production assay showed an EC50 of 15 pM, suggesting that ZX-8177 effectively facilitated IFN-β1 production via cGAMP-STING axis. In CT-26 syngeneic mouse model, ZX-8177 treatment at 2 mg/kg, BID for 14 days can achieve around 37-60% tumor growth inhibition (TGI). No body weight loss was observed during treatment. Flow cytometry analysis indicated that ZX-8177 treatment resulted in a 2-3-fold increase in tumor-infiltrating NK (CD314+CD335+) cells, cytotoxic CD8 T cells (GranzB+CD8+), and M1-macrophages (CD206-MHCII+); and a decrease in M2-macrophages (CD206+MHCII-) (p< 0.05). Combination of Mitomycin C (0.5 mg/kg, i.p., BIW) and ZX-8177 (2 mg/kg, i.p., BID) in CT-26 model yielded synergistic anti-tumor efficacy (increased >30% TGI). The level of cGAMP in tumor tissue harvested from the combination treatment group was around 2-fold higher than that in the vehicle group. Furthermore, synergistic tumor inhibition of ZX-8177 and anti-PD-L1 antibody was also observed in MC38 mouse model. TGI of anti-PD-L1 antibody vs. combo treatment is 53% vs. 75%, p < 0.05. ZX-8177 has a favorable DMPK and safety pharmacology profile to support it as a clinical candidate for further development.

Citation Format: Ying-ying Li, Zhiyuan Peng, Shuaijun Sun, Kun Guo, Deming Kong, Xiaomin Li, Yuning Xie, Siyu Shui, Yin Wang, Jinfu Yang, Xiaolin Alan Hao, Xiaoli (Shelley) Qin. ENPP1 inhibitor ZX-8177 enhances anti-tumor activity of conventional therapies by modulating tumor microenvironment [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 5486.
Abstract 5489: Utilization of cancer cell line screening and bioinformatic analyses to identify optimal developmental pathways for the novel anticancer agent BOLD-100
Authors: Raha Paromita , Park Brian , Carie Adam , Pankovich Jim , Bazett Mark
Published: June 2022

Cell line screening of unique compounds can provide mechanistic insights and identify optimal drug combination partners. Bioinformatics analysis of cell screen data and correlation to publicly available datasets can support identification of appropriate patient populations for subsequent preclinical and clinical development. BOLD-100 is a first-in-class ruthenium-based anticancer agent currently being tested in a Phase 1b/2a clinical trial in combination with FOLFOX in the treatment of advanced gastrointestinal cancers. BOLD-100 works by altering the unfolded protein response through selective GRP78 inhibition; and inducing reactive oxygen species which causes DNA damage and cell cycle arrest. Collectively, these result in cell death in a range of different cancer types, and in combination with many different classes of anticancer agents. To determine optimal indications for BOLD-100 development, BOLD-100 was tested against 316 cancer cell lines in 72-hour Cell Titer Glo assays with downstream bioinformatic analysis and validation experiments. Multiple cancer types showed preferential response to BOLD-100, including bladder, esophageal, pancreatic, multiple myeloma and ovarian cancers. Subtype analysis identified potential populations of increased responsiveness, including in bladder cancer where BOLD-100 had increased response in luminal and mixed subtypes, as compared to basal subtypes. Utilizing bladder cancer as a case study, subsequent combination testing of BOLD-100 in combination with fluorouracil or cisplatin demonstrated that BOLD-100 enhanced cell death across different bladder cancer cell lines through synergistic interactions with these standard-of-care agents. The pan-cancer response profile of BOLD-100 was compared against 449 other anticancer drug responses that are part of the GDSC database. BOLD-100 displayed limited correlation with existing drugs, suggesting a unique mechanism of action and clinical utility where standard-of-care agents have limited efficacy. Pharmacogenomic analysis of the cell screen data indicated potential pathways and genes of relevance to BOLD-100 response, including increased response in KRAS-mutant cancers. Collectively, BOLD-100 showed a unique sensitivity profile across a panel of over 300 cancer cell lines, identifying multiple potential indications for future development. Subsequent investigations into several cancer types of interest and drug combinations are ongoing.

Citation Format: Paromita Raha, Brian Park, Adam Carie, Jim Pankovich, Mark Bazett. Utilization of cancer cell line screening and bioinformatic analyses to identify optimal developmental pathways for the novel anticancer agent BOLD-100 [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 5489.
Abstract 5546: Inflamed lymph nodes mark active tumor immune response in colorectal cancer patients
Authors: Shen Dingcheng , Wang Xiaolin , Huang Meijin , Luo Yanxin , Yu Huichuan
Published: June 2022

Background Tumor-infiltrating lymphocytes (TILs) can be used to predict immune response in multiple tumors. Currently, the evaluation of TILs relies on pathological and molecular examine on resected tumor samples. Enlarged regional lymph nodes could be attributed to tumor involvement or local inflammation with active immune response. We therefore aimed to investigate the role of inflamed lymph nodes in distinguishing the hot tumor from cold tumor.

Methods Fresh-frozen tumor samples from pathologically-confirmed node-negative colorectal cancer patients with inflamed nodes (n= 22) and non-inflamed nodes (n= 23) were used to generate the genome-wide DNA methylation profile using Infinium Methylation EPIC BeadChip. The inflamed and non-inflamed nodes were defined as the pathologically-confirmed negative regional lymph nodes with enlarged size (≥ 5mm) and normal size (< 5mm) in radiological imaging, respectively. A MeTIL algorithm that integrates CD3+ T cell-specific differentially methylated positions was constructed and validated by immunohistochemical assay to evaluate TIL-based tumor immune response in bulk tumor samples.

Results The baseline characteristics were balanced between patients with inflamed and non-inflamed nodes. The high abundance of TILs was significantly associated with inflamed nodes (P= 0.025). In addition, we identified a subgroup of superhot tumors characterized by inflamed nodes, enriched TILs and favorable overall survival outcomes (HR 0.53, 95% CI: 0.36-0.97; P = 0.032).

Conclusions Inflamed regional lymph nodes in radiological imaging indicate potential active immune response in tumor. Our findings provide an easily accessible biomarker for cancer immunotherapy.

Citation Format: Dingcheng Shen, Xiaolin Wang, Meijin Huang, Yanxin Luo, Huichuan Yu. Inflamed lymph nodes mark active tumor immune response in colorectal cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5546.
Abstract 5557: PMC-309, a highly selective anti-VISTA antibody reverses immunosuppressive TME to immune-supportive TME
Authors: Park Cheon Ho , Byun Sang Soon , An Jin Yong , Han Hyerim , Lee Weon Sup
Published: June 2022

Purpose: We investigated the pharmacology of PMC-309 in ex vivo or in vivo characterization for the development of the preclinical study.

Experimental procedures: Target selectivity: it was measured by SPR assay. Human VISTA expressing CHO k1 cells or CHO k1 mock cells were used in cell binding assay. Target cell analysis: human peripheral blood mononuclear cells (PBMC) containing lymphocyte and myeloid lineage cells were used for target cells analysis with flow cytometry. In vitro T cell activity: human PBMC was employed for the evaluation of T cell activity (IRB #1041107-201703-BR-002-02) and was stimulated with or w/o PMC-309 in the presence of anti-CD3/28 Antibody. In vivo study: MC38 bearing human VISTA knock-in (KI) mice were employed for the assessment of anti-tumor activity of PMC-309. The tumor infiltrated immune cells: Immune cells in the TME were evaluated by immunohistochemistry (IHC) or flow cytometry (FACS) analysis. Statistics: All data were presented as the mean ± SEM and analyzed using one-way ANOVA followed by Dunnett’s multiple comparison tests in Graph pad prism®. ***p<0.001, **p<0.01 and *p<0.05 compared to Control group.

Summary of data: PMC-309 binding to VISTA expressing cells is highly selective and the selectivity is maintained even in the low pH conditions that mimic TME. PMC-309 enhances the secretion of IFN-gamma, TNF-alpha, and IL-2 in Mixed Lymphocyte Reaction (MLR) settings. In addition, PMC-309 promoted monocyte differentiation into M1 macrophage that stimulates proinflammatory cytokine secretion of T cells. For the in vivo study, PMC-309 was intravenously administrated in VISTA-KI mice. The tumor growth rate was suppressed in accompanied by a synergistic effect with an anti-PD1 antibody. The anti-tumor activity was associated with enhanced T cell activation, increased secretion of pro-inflammatory cytokines, and increased penetration of cytotoxic T cells, but lowing immune-suppressive MDSC cells into TME as demonstrated with IHC analysis.

Conclusions: PMC-309 is a fully human anti-VISTA antibody that targets human VISTA specifically. PMC-309 enhanced both T cell activation and monocyte activation when monocytes were cultured with or without T cells by targeting VISTA. PMC-309 increased the number of T cell infiltration while a decrease of MDSCs in the TME. PMC-309 in combination with chemotherapy or other IO drugs could address highly medical unmet needs from patients with drug resistance to currently available IO treatment options.

Citation Format: Cheon Ho Park, Sang Soon Byun, Jin Yong An, Hyerim Han, Weon Sup Lee. PMC-309, a highly selective anti-VISTA antibody reverses immunosuppressive TME to immune-supportive TME [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 5557.
Abstract 556: Novel anti-TACI single and dual-targeting CAR T cells overcome BCMA antigen loss in multiple myeloma
Authors: Larson Rebecca C. , Castano Ana , Bouffard Amanda A. , Kann Michael C. , Schmidts Andrea , Gallagher Kathleen M. , Maus Marcela V.
Published: June 2022

Anti-BCMA CAR T cell therapy has had success clinically against multiple myeloma, recently leading to FDA approval. However, the median progression free survival is 8.8 months, suggesting that targeting BCMA alone may not be sufficient. Reports of BCMA-negative relapse have occurred, highlighting the need for therapeutics that can overcome antigen loss. We have generated a novel antibody against a second target in multiple myeloma, transmembrane activator and CAML interactor (TACI). This antibody specifically recognizes TACI-positive cells (with either endogenous or exogenous expression) and has no recognition of non-B cell peripheral blood mononuclear cells. We designed novel second-generation CAR T cells based on the single chain variable fragment (scFv) from this antibody. We observe that anti-TACI CARs are cytotoxic in vitro at comparable levels to anti-BCMA CAR against multiple myeloma cell lines MM1S and RPMI-8226. Anti-TACI CARs are also functional in vivo and expand in peripheral blood of xenograft multiple myeloma models. We generated BCMA-knockout MM1S cells and observe that while anti-BCMA CAR treated mice have outgrowth of tumor, anti-TACI CAR treated animals retain anti-tumor activity. To overcome potential antigen loss of either BCMA or TACI, we designed tandem bispecific CAR T cells based on both single-targeting CAR designs. These BCMA and TACI dual-targeting CARs are functional in vitro and in vivo, even in the context of single antigen loss. We monitor the activation (CD69), exhaustion (PD-1, TIM-3, LAG-3), and memory (CCR7, CD45RA) phenotype over time to investigate the differences of single versus dual-targeting CAR T cells. We also study the ability of these CAR T cells to bind soluble antigen, as well as their binding avidity to tumor cells. We observe that the anti-TACI scFv more proximal to the CD3ζ signaling domain has a stronger effect on CAR functionality than when this scFv is more distal. Our studies provide a potential superior therapeutic option which remains efficacious in the context of BCMA antigen loss.

Citation Format: Rebecca C. Larson, Ana Castano, Amanda A. Bouffard, Michael C. Kann, Andrea Schmidts, Kathleen M. Gallagher, Marcela V. Maus. Novel anti-TACI single and dual-targeting CAR T cells overcome BCMA antigen loss in multiple myeloma [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 556.
Abstract 5586: Demonstrating restoration of T cell function in exhausted T cells with IKZF3 small molecule inhibitor, Lenalidomide
Authors: Hay Joanne , Acklam Francis , Turner Darryl , Barbour Mark , Singh Preeti , Grant Courtney , Gooding Hayley , McPherson Rhoanne , Rzepecka Justyna
Published: June 2022

CD8+ T cell exhaustion as a result of prolonged antigenic stimulation in the tumour microenvironment is thought to be a major mechanism by which tumours can escape immune surveillance. T cell exhaustion is characterised by co-expression of checkpoint inhibitor molecules such as PD-1, LAG-3 and TIM-3; reduced proliferative capacity; and diminished production of pro-inflammatory cytokines such as IL-2, IFN-γ and TNF-α and treatment with checkpoint inhibitor blockade such as Nivolumab (anti-PD-1) have shown great promise in the treatment of a subset of individuals and tumour types. However, resistance to these approaches is prevalent and there is a need to identify novel targets to provide alternative options for the rescue of T cell exhaustion and cancer treatment for use alone or potentially in combination with checkpoint inhibitor modulation. One such target is IKZF3, a transcription factor that is increased in murine models of T cell exhaustion and reduces IL-2 expression. Previously we presented an in vitro human T cell exhaustion assay whereby in vitro stimulated T cells phenotypically and functionally recapitulate the exhausted T cells found within the tumour microenvironment and now we demonstrate that exhaustion can be partially rescued by treatment with the IKZF3 small molecule inhibitor, Lenalidomide, alone and in combination with checkpoint inhibitor blockade. The use of small molecule inhibitors offers new avenues for reversing T cell exhaustion in cases of checkpoint inhibitor resistance and this platform delivers a robust assay allowing for assessment of small molecules or checkpoint inhibitors both individually and in combination.

Citation Format: Joanne Hay, Francis Acklam, Darryl Turner, Mark Barbour, Preeti Singh, Courtney Grant, Hayley Gooding, Rhoanne McPherson, Justyna Rzepecka. Demonstrating restoration of T cell function in exhausted T cells with IKZF3 small molecule inhibitor, Lenalidomide [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 5586.