bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5569: Discovery of tumor-specific T cell receptors (TCRs) by reverse genetic engineering platform
Authors: Han Yanyan , Liang Xiaoling , Ou Shudan , Chen Xihe , Ma Minjun , Li Jun , Ma Yifan , Zhou Xiangjun
Published: September 2020

TCR-T therapy is an ideal cell therapeutic strategy against solid tumors. However, the traditional method of TCR cloning has two major problems. First, we could only develop TCRs against previously discovered tumor antigen epitopes. Second, the affinity maturation of TCR sometimes leads to mortal toxicities due to the unexpected cross-reactivities. Therefore we hereby present a reverse genetic engineering platform to clone tumor specific TCRs from immunotherapies benefit cancer patients. Multi-antigen stimulating cell therapy (MASCT) is a cell-based immunotherapy combining therapeutic DC vaccine and adoptive T cell transfer, the IND application of which was approved in China. We observed dynamic T cell responses against multiple tumor-associated antigens used in MASCT-treated HCC patients, which showed a significant correlation with clinical outcome. Based on this observation, we propose that expanded tumor-specific T cells in MASCT-benefit cancer patients could be good sources to isolate clinical relevant TCRs. A HPV positive metastatic cervical cancer patient has shown partially response after MASCT treatment alone, and survived over 5 years. Moreover, a metastatic lung cancer patient has also shown partially response and survived for longer than 2 years after combination treatment of MASCT and anti-PD1 antibody. Enhanced and persist T cell responses against several TAAs as well as HPV antigens were detected in PBMCs of both patients. We successfully expended and isolated tumor antigen-specific T cells by IFNγ-secreting cell enrichment, and obtained alpha/beta chains paired TCR sequences using single T cell NGS technology. We selected 14 pairs of TCRs isolated from cervical cancer patient for further validation. TCRs from lung cancer patient were currently under synthesis or sequencing. Among 14 TCRs, a TAA-specific TCR and a HPV18E7-specific TCR showed superior antigen specificity and optimum avidity, and would be applied for TCR-T development. Of notes, our reverse genetic engineering of TCR-T (ReGET) platform.can not only identify both HLA class I and II-restricted TCRs, but also identify novel epitopes of tumor antigens, which may provide new targets for TCR-T therapy against solid tumors.

Citation Format: Yanyan Han, Xiaoling Liang, Shudan Ou, Xihe Chen, Minjun Ma, Jun Li, Yifan Ma, Xiangjun Zhou. Discovery of tumor-specific T cell receptors (TCRs) by reverse genetic engineering 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 5569.
Abstract 5674: Discovery of AZD9833, an oral small molecule selective degrader of the estrogen receptor (SERD)
Authors: Scott James S. , Moss Thomas , Stokes Steve , Nissink Willem M. , Morrow Christopher J. , Lawson Mandy , Cureton Natalie , Gangl Eric , Gutierrez Pablo Morentin , Mather Richard , Lindemann Justin P. , Sykes Andrew , Fisher David , Polanski Radoslaw , Carroll Danielle , Twomey Aisling Barton , Klinowska Teresa
Published: September 2020

The estrogen receptor alpha (ERα) is expressed in >70% of breast cancers and is a clinically validated target in oncology. Anti-hormonal therapies that block ER function directly (e.g. tamoxifen) or therapies that block the production of estrogen itself (e.g., aromatase inhibitors) have proven to be effective treatments of the disease. Further advances in disease control have been made with the development of the Selective Estrogen Receptor Degrader (SERD) fulvestrant, which both antagonizes ERα-driven tumor cell growth and causes degradation of the ERα receptor. The first-generation oral SERDs, including AZD9496, showed full ER degradation in MCF-7 cells but incomplete ER degradation in other cell lines, and partial agonism. Building on that learning, extensive structure-enabled chemical optimization has resulted in the identification of a next generation oral SERD AZD9833, a potent degrader and antagonist of the ERα receptor (both EC50 <1 nM in MCF7 cells) with favorable physicochemical properties (logD7.4 <3). AZD9833 delivers maximal ERα degradation equivalent to fulvestrant in all ER+ cell lines tested, including MCF7, CAMA1, T47D and BT474. In addition, and in contrast to AZD9496, AZD9833 does not cause ER agonism in the endometrial carcinoma cell line Ishikawa in vitro and does not cause an increase in the thickness of the endometrium in juvenile rats. In several patient-derived and cell line xenograft models, including models with clinically relevant ESR1 mutations, AZD9833 completely suppresses tumor growth and gives equivalent pharmacodynamic activity to supra-clinical levels of fulvestrant. Furthermore, in an ESR1 wild-type and an ESR1 D538G PDX model, AZD9833 demonstrated combinatorial benefit with palbociclib. Based on this preclinical data AZD9833 has progressed into a multi-stage monotherapy and palbociclib combination First Time in Patient clinical trial, SERENA-1 (NCT03616587).

Citation Format: James S. Scott, Thomas Moss, Steve Stokes, Willem M. Nissink, Christopher J. Morrow, Mandy Lawson, Natalie Cureton, Eric Gangl, Pablo Morentin Gutierrez, Richard Mather, Justin P. Lindemann, Andrew Sykes, David Fisher, Radoslaw Polanski, Danielle Carroll, Aisling Barton Twomey, Teresa Klinowska. Discovery of AZD9833, an oral small molecule selective degrader of the estrogen receptor (SERD) [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 5674.
Abstract 5681: Design, synthesis and study of small molecules with both AR degradation and AKR1C3 inhibitory activities
Authors: Xing Enming , Kong Xiaotian , Lou Wei , Shi Ruihua , Cheng Xiaolin , Gao Allen C. , Li Pui Kai
Published: September 2020

Prostate cancer is the most common cancer in men and it is estimated that 32,000 men in the US alone will die of this cancer in 2019. Androgen deprivation therapy (ADT) has been the mainstay of prostate cancer therapy. However, most of the patients will progress to castration resistance prostate cancer (CRPC) after several years of treatment. The current treatment of CRPC includes chemotherapeutic agents (docetaxel) and agents that target androgen signaling (abiraterone, enzalutamide and apalutamide). Ultimately, however, most patients develop resistance to the treatments. Two of the mechanisms of resistance involve the expression of constitutively active androgen receptor variant (AR-V7) and the increase in intra-tumoral biosynthesis of androgens. Several treatment strategies targeting AR-V7 have been reported. For the intra-tumoral biosynthesis of androgens, one of the enzymes, AKR1C3, plays a critical role in the development of CRPC and has been reported to be consistently elevated in CRPC patients who are resistant to abiraterone and enzalutamide.

We have been involved in the development of agents to treat CRPC and overcome drug resistance of CRPC patients. We reported that niclosamide, an FDA-approved anthelminthic agent, could degrade AR-V7. We also reported that niclosamide, as well as an AKR1C3 inhibitor, indomethacin, were able to overcome enzalutamide and abiraterone resistance in several prostate cancer cell lines, suggesting that niclosamide is also an AKR1C3 inhibitor. By examining the reported structures of AKR1C3 inhibitors, niclosamide was found to contain the core scaffold of 4-Hydroxy-1-(4-methoxybenzyl)-N-(3-(trifluoromethyl) phenyl)-1H-1,2,3-triazole-5-carboxamide. Using this knowledge, we designed a series compounds with dual AR degrading and AKR1C3 inhibitory activities. Preliminary studies showed the compounds caused AR-V7 degradation and to a lesser extent AR full length degradation. The compounds also inhibited AKR1C3 and caused a dose dependent growth inhibition of several prostate cancer cell lines.

Citation Format: Enming Xing, Xiaotian Kong, Wei Lou, Ruihua Shi, Xiaolin Cheng, Allen C. Gao, Pui Kai Li. Design, synthesis and study of small molecules with both AR degradation and AKR1C3 inhibitory activities [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 5681.
Abstract 5751: Patient-reported toxicities in head and neck cancer patients treated with immunotherapy
Authors: James Brian W. , Hoogland Aasha I. , Irizarry-Arroyo Nathaly , Muzaffar Jameel , Kirtane Kedar , Kish Julie A. , Jim Heather S. , Chung Christine H.
Published: September 2020

Background: Immune checkpoint inhibitors (ICIs) (i.e. nivolumab, pembrolizumab) are FDA-approved agents used in the treatment of various head and neck malignancies. While much is known about clinician-rated adverse clinical events of these immunotherapies from clinical trials, less is known about patient-reported toxicities. The goal of the current study was to describe patient-reported toxicities in patients receiving ICIs for head and neck cancer.

Methods: Patients with head and neck cancer were recruited as part of a larger study on quality of life on ICIs. Prior to initiation of ICIs, participants completed a sociodemographic questionnaire and a measure of symptomatic toxicities of ICIs developed based on input form providers, patients, and caregivers. Patients completed follow-up symptom assessments every 6-8 weeks until cancer progression. Severity of each symptom over the past 7 days was rated on a 5-point scale (i.e., 0 = “not at all,” 4 = “very much”). A score of 3 or 4 was considered moderate to severe symptomatology.

Results: There were 45 patients (22% female; M age = 62), who completed a least 2 symptom assessments before cancer progression. The five most common moderate to severe symptoms at baseline were fatigue (27%), changes in taste (18%), insomnia (16%), coughing (11%), and muscle aches (9%). At the assessment before cancer progression, the five most common moderate to severe symptoms were fatigue (38%), insomnia (24%), rash (13%), cough (13%), and numbness in hands and feet (13%). From the baseline assessment to the last symptom assessment preceding progression, most patients reported worsening skin dryness (49%), skin sensitivity (46%), weakness in arms of legs (46%), and itching (42%). Most patients also reported worsening fatigue (49%), although a large subset reported improvements in fatigue (36%).

Conclusion: Results indicate that fatigue and insomnia are the most common and severe patient-reported toxicities of ICIs for head and neck cancer. These preliminary findings warrant larger future studies with longer follow-up to better understand changes in patient-reported toxicities in this population.

Citation Format: Brian W. James, Aasha I. Hoogland, Nathaly Irizarry-Arroyo, Jameel Muzaffar, Kedar Kirtane, Julie A. Kish, Heather S. Jim, Christine H. Chung. Patient-reported toxicities in head and neck cancer patients treated with immunotherapy [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 5751.
Abstract 5817: Anti-tropomyosin drugs prevent the rescue of vincristine-induced mitotic spindle defects
Authors: Gunning Peter , Wang Yao , Stear Jeff H. , Swain Ashleigh , Xu Xing , Bryce Nicole , Alieva Irina B. , Dugina Vera B. , Cripe Timothy , Stehn Justine , Hardeman Edna C.
Published: September 2020

Drugs targeting a major component of the actin filaments of cancer cells, tropomyosin Tpm3.1, synergize with anti-microtubule drugs in neuroblastoma and lung cancer models both in vitro and in vivo and a wide range of other cancer types in vitro. We have determined the mechanism of synergy in HeLa cells to gain insight into the potential interaction of actin filaments and microtubules in the survival and proliferation of cancer cells. HeLa cells exhibit a strong synergistic response to the combined treatment of vincristine (VCR) and anti-Tpm3.1 compounds, marked by an enhanced reduction in cell viability, apoptosis induction and mitotic cell cycle arrest. Tpm3.1 localizes to the cell cortex during mitosis, potentially associating with the microtubule network, particularly the dynein/dynactin complexes responsible for mediating cortical pulling forces during spindle assembly. VCR alone causes supernumerary NuMA organized acentrosomal microtubule organizing centers upon nuclear envelope breakdown, which can be resolved via a clustering mechanism to achieve bipolar cell division. The addition of anti-Tpm3.1 compounds inhibits NuMA-associated clustering in VCR-treated cells, leading to irreparable defects during spindle assembly and thus a largely increased number of cells with multi-polar spindles undergoing mitotic delay and catastrophe. We conclude that actin/Tpm3.1 filaments contribute to the formation of the bipolar spindle and play a critical role in the clustering of acentrosomal microtubule asters.

Citation Format: Peter Gunning, Yao Wang, Jeff H. Stear, Ashleigh Swain, Xing Xu, Nicole Bryce, Irina B. Alieva, Vera B. Dugina, Timothy Cripe, Justine Stehn, Edna C. Hardeman. Anti-tropomyosin drugs prevent the rescue of vincristine-induced mitotic spindle defects [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 5817.
Abstract 5822: Targeting an immortalization mutation to control glioblastoma
Authors: Iyer Jayashree S. , Tang Nanyun , Ferdosi Shayesteh R. , Gokhale Vijay , Hurley Laurence H. , Dhruv Harshil D. , Berens Michael E.
Published: September 2020

Glioblastoma multiforme (GBM), the most aggressive primary brain cancer, is a heterogeneous disease. Standard treatment of temozolomide and radiation therapy has remained unchanged for the past thirty years. However, over 80% of GBM tumors have a telomerase reverse transcriptase (TERT) promoter mutation, or TPM. Telomerase is an enzyme containing the catalytic subunit, telomerase reverse transcriptase (TERT), that mediates telomere elongation in the nucleus. In the absence of any effective molecular targeting therapy for GBM, the elucidation of oncogenic signaling of TERT could open new avenues in GBM treatment. Canonically, mutations of TERT, which result in TERT upregulation, maintain telomere length in the nucleus and promote indefinite proliferation of cancer cells. However, a non-canonical function of TERT in the mitochondria has recently been suggested. We screened GBM cell models against a novel small molecule inhibitor (RG1534, Reglagene Inc.) that interferes with the functionality of a mutated TERT promoter. RG1534 selectively suppresses glioma cell viability without affecting non-transformed normal human astrocytes. A dose dependent response was observed in glioma cell lines, resulting in a decrease in TERT mRNA expression. To validate this, TERT protein levels in glioma cells treated with RG1534 were measured and results showed a reduction in expression post treatment. More interestingly, RG1534 treatment leads to rapid induction of apoptosis in glioma cell line which does not correlate with the time course of the telomere shortening effect. Based on prior findings that mitochondrial TERT is involved in DNA damage, we conducted the fractionation of various glioma cell lines to measure the protein expression of TERT in subcellular compartments. We observed a higher expression of TERT in the mitochondria compared to the nucleus. This suggests that TERT may have non-canonical functions outside of telomeric elongation. In summary, our results demonstrate that non-canonical functions of TERT may play a critical role in glioma pathobiology and warrants further study.

Citation Format: Jayashree S. Iyer, Nanyun Tang, Shayesteh R. Ferdosi, Vijay Gokhale, Laurence H. Hurley, Harshil D. Dhruv, Michael E. Berens. Targeting an immortalization mutation to control glioblastoma [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 5822.
Abstract 5831: Clearance of therapy-induced senescent cancer cells by the senolytic ABT-263 in a model of non-small cell lung cancer
Authors: Carpenter Valerie , Saleh Tareq , Gewirtz David
Published: September 2020

While most chemotherapies will initially lead to tumor reduction in patients, residual tumor cells that enter a state of dormancy can re-emerge as aggressive, recurrent disease after treatment is terminated. One mechanism by which cancer cells may escape chemotherapy-induced death is senescence. Senescence is a specialized form of cell cycle arrest that is primarily characterized as a durable, prolonged loss of proliferative capacity. While senescence was traditionally considered to be irreversible, evidence has accumulated in recent years supporting the premise that senescence is instead a durable form of growth arrest from which a subpopulation of cells can ultimately escape. Moreover, the accumulation of senescent cells post cancer therapy has been linked with a plethora of unfavorable outcomes. A sequential approach in which chemotherapy is followed by a senolytic, an emerging class of drugs that selectively kill senescent cells, may therefore delay or circumvent recurrent disease by eliminating residual senescent cells. To this end, we tested whether etoposide- or radiation-treated non-small cell lung cancer (NSCL) cells become susceptible to the BH-3 mimetic ABT-263– an established senolytic– following senescence induction. Treatment with etoposide or radiation induced a robust senescent state, as evidenced by the development of multiple senescent phenotypes. Senescent tumor cells demonstrated increased sensitivity to ABT-263 compared to non-senescent cells, undergoing rapid apoptosis subsequent to ABT-263 exposure. We further determined that ABT-263's mechanism of action appears to depend upon the disruption of the BCL-XL/Bax interaction. ABT-263's capacity to eliminate senescent cells is conserved in vivo, as tumor-bearing animals treated with etoposide followed by ABT-263 undergo rapid tumor reduction followed by tumor maintenance, while etoposide-only treated animals develop progressive disease subsequent to treatment completion. These data support the use of senolytics as complementary therapy to eliminate residual tumor cells following chemotherapy or radiation, and thereby potentially delay or reduce recurrent disease.

Citation Format: Valerie Carpenter, Tareq Saleh, David Gewirtz. Clearance of therapy-induced senescent cancer cells by the senolytic ABT-263 in a model of 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 5831.
Abstract 5855: The genomic landscape of Mongolian hepatocellular carcinoma
Authors: Candia Julián , Bayarsaikhan Enkhjargal , Tandon Mayank , Budhu Anuradha , Forgues Marshonna , Lack Justin , Chao Ann , Chinburen Jigjidsuren , Wang Xin W.
Published: September 2020

Mongolia has the highest incidence of-and mortality from- hepatocellular carcinoma (HCC) in the world, which is between three and seven times higher than that observed in other high-incidence populations, such as South Korea, Thailand, and China. In Mongolia, where cancer is the second most common cause of death accounting for nearly a fifth of deaths, HCC is the most prevalent cancer type at about 40% of all cancers. Despite the daunting proportion of this longstanding health crisis, the molecular landscape of Mongolian HCC has not been studied yet. Filling this gap, we aim to identify robust molecular subclasses and driver features informative of the etiology and progression of the disease. Here, we describe molecular characteristics of 76 Mongolian HCC patients by whole-exome and whole-transcriptome sequencing. We present a comprehensive comparison of mutational signatures, driver genes and molecular subtypes of Mongolian HCC versus 373 HCC patients of different ethnicities and diverse etiologies. Mongolian HCC consists of several similar prognostic molecular subtypes in patients from other areas of Asia, Europe and North America as well as several unique types, suggesting potentially the presence of unique etiologies linked to Mongolian patients. Consistently, Mongolian HCC exhibits several common driver mutations (TP53, CTNNB1) frequently found in pan-cancer analyses but also a unique driver (SPTA1) that may be linked to hepatitis D viral infection. Furthermore, unique hotspot missense mutations were identified in driver genes GTF2IRD2B and PNRC2. These results indicate the existence of novel molecular mechanisms at play in Mongolian hepatocarcinogenesis.

Citation Format: Julián Candia, Enkhjargal Bayarsaikhan, Mayank Tandon, Anuradha Budhu, Marshonna Forgues, Justin Lack, Ann Chao, Jigjidsuren Chinburen, Xin W. Wang. The genomic landscape of Mongolian hepatocellular carcinoma [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 5855.
Abstract 5868: Cell barcoding, genetic screens and expression profiling at a single-cell level
Authors: Chenchik Alex , Makhanov Mikhail , Darst Russell , Deng Debbie , Tedesco Donato , Diehl Paul , Kobzik Lester
Published: September 2020

Novel single-cell methods for profiling clonal composition and phenotypes of genetically modified cancer cells may improve understanding of tumorigenesis and guide the development of new anti-cancer drugs. To facilitate these studies, we developed a panel of lentiviral barcoded sgRNA libraries to label and monitor cancer cells in time-course experiments in vitro and in mouse xenograft models. The sgRNA effectors and cellular barcodes are transcribed from lentiviral constructs and can be detected by NGS in a single-cell RNA expression profiling assay, allowing characterization of the subpopulation of cells descended from a single, barcoded knockout progenitor cell. To scale up genetic analysis, cell barcodes were incorporated in conjunction with sgRNA effector libraries to identify clonal phenotypic changes induced by specific genetic disruptions in progeny cells derived from a single progenitor cell. However, effective single-cell genetic screening designed for even only a few hundred genes requires expression profiling of one hundred thousand cells (or more) and is not practical using currently available instrumentation. Data will be presented showing how genetic screen technology combined with targeted RNA expression profiling of barcoded knockout cells can be applied for large-scale genetic screens and significantly improve phenotyping of distinct cell populations in several cancer model systems. Preliminary studies demonstrate the assay to have unparalleled throughput, sensitivity, and improved cost-effectiveness for high-throughput drug target discovery applications.

Citation Format: Alex Chenchik, Mikhail Makhanov, Russell Darst, Debbie Deng, Donato Tedesco, Paul Diehl, Lester Kobzik. Cell barcoding, genetic screens and expression profiling at a single-cell level [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 5868.
Abstract 5981: CAP70 suppresses the development of triple-negative breast cancer by inhibiting EGFR pathway activation
Authors: Zheng Junfang , Ma Yuanzhen , Qi Yijun
Published: September 2020

Background: Among all major subtypes of breast cancer, triple-negative breast cancer (TNBC) carries the worst prognosis for the patients. The poor clinical outcome of TNBC is further exacerbated by the lack of effective targeted therapies. Therefore, the identification of functional molecular targets for TNBC therapy is urgently needed. The goal of this study was to determine the role of CFTR-associated protein of 70 kDa (CAP70, also called PDZK1) in the development of TNBC and assess its value as a target for TNBC therapy.

Methods: Bioinformatics analysis and immunohistochemistry were performed to determine the expression of CAP70 in TNBC. The functions of CAP70 were demonstrated in a series of in vitro and in vivo experiments. GST Pull-down, mass spectrometry, co-immunoprecipitation, bioinformatics analysis, pharmacologic treatment, and western blotting were performed to elucidate the potential mechanisms of CAP70 activities.

Results: CAP70 was specifically downregulated in TNBC tissues and was related to tumor stage, lymph node metastasis, and patient survival. Functional assays showed that CAP70 suppressed TNBC carcinogenesis, progression, and metastasis by inhibiting EGFR signaling. Moreover, inhibition of EGFR signaling by CAP70 enhanced the sensitivity of TNBC to EGFR inhibitor erlotinib.

Conclusions: CAP70 suppresses TNBC development and enhances its sensitivity to erlotinib by inhibiting EGFR signaling. CAP70 is a significant prognostic factor of TNBC and a potential molecular therapeutic target for the treatment of this subtype of breast cancer.

Citation Format: Junfang Zheng, Yuanzhen Ma, Yijun Qi. CAP70 suppresses the development of triple-negative breast cancer by inhibiting EGFR pathway activation [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 5981.
Abstract 5990: The potential role of artificial lactate influx in remodeling cancer-specific metabolism of colorectal cancer
Authors: Jeong Keun-Yeong
Published: September 2020

Cancer cells have a specific metabolic pathway that produces a large quantity of lactate from glucose by abnormal aerobic glycolysis, it can provide a secondary path that meets the metabolic needs for persistent survival. Accordingly, regulating cancer-specific metabolism has important implications for the discovery of novel therapeutics. In this study, the possibility of an impact on colorectal cancer was investigated that focused on the remodeling of cancer-specific aerobic glycolysis by the artificial influx of lactate. Intracellular lactate levels were assessed in the colorectal cancer cells following lactate calcium salt treatment. Expression of lactate dehydrogenase and pyruvate dehydrogenase were confirmed to investigate the effect of the increased lactate influx. The nicotinamide adenine dinucleotide ratio and concentration of α-ketoglutarate were measured to confirm the tricarboxylic acid coupling. The tube formation and wound healing assay were performed as well as the expression of hypoxia-inducible factor-1α and vascular endothelial growth factor receptor to confirm the loss of malignancy. Xenograft animal models were established to evaluate the antitumor efficacy by monitoring changes in the tumor volume. We demonstrated that an artificial influx of lactate was increased in colon cancer cells under a hypoxic condition thereby reversible activation of lactate dehydrogenase reacting to lactate. Then, the conversion to pyruvate was promoted, and as a result of increased stimulation of pyruvate dehydrogenase, the tricarboxylic acid was coupled, showing an oxidative metabolism similar to normal cells. The malignancy of cancer cells was

lost by undergoing remodeling of such cancer-specific, and a significant decrease in tumor growth was confirmed as a result of the anticancer effect of artificial lactate influx. These findings suggest the unique mechanisms in which the artificial influx of lactate would have anticancer effects by regulating cancer-specific aerobic glycolysis.

Citation Format: Keun-Yeong Jeong. The potential role of artificial lactate influx in remodeling cancer-specific metabolism of colorectal 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 5990.
Abstract 6001: 6-Phosphofructo-2-kinase inhibition to increase the efficacy of ER and CDK4/6 inhibitors against ER+ breast cancer
Authors: Lanceta Lilibeth , Lypova Nadiia , Chesney Jasson , Imbert-Fernandez Yoannis
Published: September 2020

Deregulation of the estrogen receptor (ER)-cyclin D1-CDK4/6 pathway is a hallmark of ER+ breast cancer that has prompted the development of CDK4/6 inhibitors. Although administration of CDK4/6 inhibitors to patients with ER+ advanced breast cancers have resulted in an improvement in progression free survival, almost all patients invariable develop resistance and relapse over time. Activation of glucose metabolism upon CDK4/6 inhibition is becoming increasingly evident. A key stimulator of glycolysis is the PFKFB3 enzyme which synthesizes fructose 2,6-bisphosphate (F2,6BP), a potent allosteric activator of the rate-limiting 6-phosphofructo-1-kinase (PFK1). F2,6BP controls flux throughout the entire glycolytic pathway and, as a result is required for the proliferation of cancer cells. In preliminary studies, we provide evidence that PFKFB3 expression and activity is increased in response to CDK4 inhibition in ER+ breast cancer cells and that a PFKFB3 inhibitor, PFK-158, increases the anti-tumor activity of anti-CDK4 targeted therapy in vitro. Importantly, we found that ER+ cancer cells with acquired resistance to palbociclib have increased levels of PFKFB3 and are highly susceptible to the effects of PFK-158. Our studies provide a clear rationale to propose the use of PFKFB3 inhibitors in combination with CDK4/6 inhibitors as an effective strategy to increase the efficiency and/or overcome resistance to CDK4/6 inhibition.

Citation Format: Lilibeth Lanceta, Nadiia Lypova, Jasson Chesney, Yoannis Imbert-Fernandez. 6-Phosphofructo-2-kinase inhibition to increase the efficacy of ER and CDK4/6 inhibitors against ER+ breast 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 6001.
Abstract 6036: The clinicopathological significance of nerves and NGF receptors in esophageal cancer
Authors: Griffin Nathan , Gao Fangfang , Jobling Phillip , Wills Vanessa , Walker Marjorie M. , Faulkner Sam , Hondermarck Hubert
Published: September 2020

Recent developments in oncology have revealed that nerve outgrowth in the tumor microenvironment is essential to the progression of many human cancers. In esophageal cancer, the presence and role of nerves and is unclear. In this study, nerves were identified within the tumor microenvironment of esophageal cancer and were associated with the squamous subtype of the disease (p=0.044). Additionally, the surrounding or invasion of nerves by cancer cells (perineural invasion) was detected in 12% of esophageal cancers and was associated with poor patient survival (p=0.038). Nerves were found to express the TrkA and p75NTR receptors for nerve growth factor (NGF) and an association was observed between high production of NGF by cancer cells and the presence of nerves (p=0.022). In vitro, esophageal cancer cells were able to induce neurite outgrowth in PC12 neuronal cells and this neurotrophic activity was inhibited by anti-NGF blocking antibodies. In squamous cell carcinoma tissues, TrkA expression was found to be strongly increased in compared to both adenocarcinoma and normal oesophageal tissue (p<0.0001). The selective pharmacological inhibitor of NTRK1, GNF-5837, induced a decreased esophageal cancer cell proliferation, invasion and migration. Together, these data demonstrate that innervation is a significant feature in esophageal cancers that may be driven by cancer cell-released NGF. Therefore, this study reveals that interventions against the NGF-TrkA signaling pathway may have the potential to target cancer cells and inhibit cancer neurogenesis.

Citation Format: Nathan Griffin, Fangfang Gao, Phillip Jobling, Vanessa Wills, Marjorie M. Walker, Sam Faulkner, Hubert Hondermarck. The clinicopathological significance of nerves and NGF receptors in esophageal 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 6036.
Abstract 6072: MUC16-Cter cleavage increases the metastatic potential in pancreatic cancer
Authors: Marimuthu Saravanakumar , Lakshmanan Imayavaramban , Muniyan Sakthivel , Atri Pranita , Jain Maneesh , Ponnusamy Moorthy P. , Batra Surinder K.
Published: September 2020

Pancreatic cancer (PC) is one of the most lethal malignant tumors. Several membrane-bound mucins are overexpressed in PC, including MUC16. The role and molecular mechanism of MUC16 mucin, particularly carboxyl-terminal domain (MUC16-Cter) in metastasis of pancreatic cancer, are poorly understood. Therefore, we hypothesize that the “MUC16-Cter domain plays a vital role in facilitating metastasis in pancreatic cancer”. Here, we knockout MUC16 in Capan 1 PC cells and overexpressed MUC16-Cter (F114AA) domain in MiaPaCa2 PC cells to study its functional impact in PC. Capan-1 MUC16 knockout cells displayed a significant decrease in colony-forming by in vitro tumorigenicity assay, endothelial binding, and P-selectin binding ability compared to control cells. Whereas, MUC16-Cter domain overexpressed MiaPaCa2 cells showed a significant increase in colony formation as compared to vector control cells. These results suggest that MUC16 plays a role in promoting in vitro tumorigenicity and facilitating intravasation and circulation of PC cell by interacting with endothelial cells during the metastatic spread of PC cells. Interestingly, we observed that chemotherapeutic drug (gemcitabine) induces MUC16-Cter cleavage in PC cells. We generated a genetically engineered mouse model Muc16-/- with KrasG12D, Trp53R172H/+, Pdx1-Cre (KPCM) background to study the role of Muc16 in PC tumorigenesis and metastasis with existing KPC model. KPCM showed decreased the early onset of cancer formation compared to KPC. Further, the Kaplan Meier test showed an increased survival rate in Muc16 -/- with KPC background as compared to KPC mice. EMT markers showed decreased expression in KPCM compared to KPC tumors. KPCM cells showed a significant decreased in migratory potential, endothelial, and P-selectin binding ability KPC PC cells. In conclusion, our in vitro and in vivo findings confirm that MUC16 and its Cter domain might play an essential role during the PC tumorigenesis and metastasis. Therefore, targeting MUC16-Cter might be an ideal therapeutic strategy for inhibiting tumorigenesis and metastasis of PC.

Citation Format: Saravanakumar Marimuthu, Imayavaramban Lakshmanan, Sakthivel Muniyan, Pranita Atri, Maneesh Jain, Moorthy P. Ponnusamy, Surinder K. Batra. MUC16-Cter cleavage increases the metastatic potential in pancreatic 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 6072.
Abstract 6107: Stromal and immune changes in the aged lung microenvironment create a permissive niche for the metastatic outgrowth of melanoma
Authors: Fane Mitchell , Douglass Stephen , Alicea Gretchen , Webster Marie , Aguirre-Ghiso Julio , Weeraratna Ashani
Published: September 2020

Our previous data has indicated that while tumors grow more slowly in aged mouse skin, they form more lung metastases. We queried whether the lung microenvironment undergoes age-related changes that provide a permissive niche for metastatic outgrowth. Our previous data has implicated dermal fibroblasts as the drivers of age-related changes in the skin, and so we analyzed young vs aged healthy human lung fibroblasts. The secreted changes in aged lung fibroblasts suggest that the lung microenvironment evolves during aging to promote proliferation of melanoma cells. Specifically, these changes involve the secretion of factors such as sFRP1, that we find suppresses Wnt5A and AXL signaling, which typically drives a metastatic but slow-growing/dormant cancer phenotype. Interestingly, this switch promotes activation of the MER tyrosine kinase pathway, increasing proliferation. This switch is reinforced by the MER ligand PROS1, which is upregulated in the aged lung microenvironment. Consistent with this, our in vivo studies indicate that tumor cells disseminate to the lungs at the same rate in young and aged mice. However, they persist as single cell populations in the young mouse lungs, whereas they grow out rapidly in the aged mouse lung. We next queried whether the immune microenvironment also played a role in the differential outgrowth of lung metastases in young and aged mice. We find that aged lung fibroblasts secrete higher levels of immunosuppressive factors ARG-1 and CXCL1, which are both necessary for efficient metastatic outgrowth in vivo. Analysis of the aged lung microenvironment reveals that immunosuppressive MDSCs and Treg populations are significantly increased compared with the young lung, while effector CD4 and CD8 T cells are decreased. Importantly, depletion of MDSCs or Tregs in the aged lung both significantly decrease metastasis. Overall, we find that aged-induced changes in fibroblast secretory phenotypes and immune subpopulations promote efficient metastatic melanoma progression in the lung.

Citation Format: Mitchell Fane, Stephen Douglass, Gretchen Alicea, Marie Webster, Julio Aguirre-Ghiso, Ashani Weeraratna. Stromal and immune changes in the aged lung microenvironment create a permissive niche for the metastatic outgrowth of melanoma [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 6107.
Abstract 6133: Rats support cancer studies
Authors: Adedeji Bisoye Towobola , Fallon Noto K. , Moody Sam , Steffey Valeriya , Brenzel Chris , Crawford Jack , Jamling Tseten Yeshi , Narla Goutham
Published: September 2020

The use of the most predictive model is essential in transferring scientific knowledge from bench to bedside. In In vivo cancer research, immunodeficient mice models are being used, but have been found with limitations. They fall short in tumor take rates, growth kinetics, and their ability to support the growth of some human Cancer cell lines.

To address these limitations, we produced an immunodeficient rat, a Sprague-Dawley Rag2/Il2rg double knockout (SRG OncoRat) rat that lacks mature B cells, T cells, and circulating NK cells. Using a variety of human cell lines (VCaP, LNCaP, 22RV1) which have poor or highly variable growth kinetics in commercially available immunodeficient mouse models, we demonstrated that the SRG rat has superior tumor take rates and growth kinetics, providing a more efficient model for drug efficacy studies. Enhanced take rates enable the use of fewer animals for studies, allows for faster timelines to drug efficacy data, and overall, may provide a better platform for reproducible drug efficacy results. In addition, the SRG rat developed larger tumor volumes, allowing for serial fine needle aspirate biopsy for PK, PD and molecular analysis in the same animal throughout the course of the study without significantly affecting normal tumor growth. The SRG OncoRat is a valuable addition to current available rodent xenograft models for evaluating novel therapies.

Citation Format: Bisoye Towobola Adedeji, Noto K. Fallon, Sam Moody, Valeriya Steffey, Chris Brenzel, Jack Crawford, Tseten Yeshi Jamling, Goutham Narla. Rats support cancer studies [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 6133.
Abstract 6159: Blocking neuropeptide Y Y5 receptor prevents hypoxia-induced Ewing sarcoma metastasis
Authors: Adnani Mina T. , Galli Susana , Hong Sung Hyeok , Tilan Jason U. , Kitlinska Joanna
Published: September 2020

Ewing sarcoma (ES) is the second most prevalent malignant bone tumor after osteosarcoma in children and adolescents. While survival of patients with localized ES has improved, patients with metastatic ES still lack adequate therapies, which results in poor survival rates. Thus, there is an important need for new therapies targeting ES metastases and preventing its dissemnation. Previously, we have shown that ES cells constitutively express high levels of NPY and its receptors - Y1R and Y5R. However, tumor hypoxia changes expression pattern and functions of NPY system, leading to the activation of its Y2R and Y5R, which stimulate ES cell motility and invasiveness under these conditions. Hence, the goal of this study was to determine the role of NPY in hypoxia-induced ES metastases. Here, we focused on Y5R, since our data from other malignancies implicated this receptor as the major mediator of NPY's pro-metastatic effects. To this end, we used doxycycline (DOX)-inducible CRIPSR/Cas9 system to create an SK-ES1/Y5R knockout (KO) cell line. The Y5R KO significantly decreased motility of SK-ES1 cells. To determine the impact of this phenomenon on ES metastases, SK-ES1/Cas9/Y5RsgRNA cells were injected into gastrocnemius muscles of SCID/bg mice and the animals were fed with DOX+ or control diet. The Y5R KO did not affect primary tumor growth. Once the xenografts reached a volume of 0.25cm3, a femoral artery ligation (FAL) was performed in the tumor-bearing limbs to induce hypoxic conditions. Both primary tumors and their corresponding metastases were sequenced to confirm the efficiency of gene editing. Notably, majority of metastases arising in mice bearing SKES-1/Y5RsgRNA+DOX xenografts were initiated by the clones without Y5R gene modification, confirming the role of Y5R in ES dissemination. The frequency of metastases with efficient Y5R KO originating from hypoxic tumors was markedly decreased, as compared to the control group (-DOX). Moreover, Y5R KO prevented hypoxia-induced increase in osseous metastases previously reported by our group. Collectively, these data validate NPY/Y5R axis as a crucial mediator of the hypoxia-induced ES metastasis and implicate Y5R receptor as a novel target for therapies preventing ES dissemination. Importantly, the Y5R antagonist is already available and used in clinical trials.

Citation Format: Mina T. Adnani, Susana Galli, Sung Hyeok Hong, Jason U. Tilan, Joanna Kitlinska. Blocking neuropeptide Y Y5 receptor prevents hypoxia-induced Ewing sarcoma metastasis [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 6159.
Abstract 6161: PARP inhibition enhances radiotherapy of SMAD4 deficient human head and neck squamous cell carcinomas in experimental models
Authors: Young Christian D. , Hernandez Ariel L. , Bian Li , Karam Sana D. , Raben David , Wang Xiao-Jing
Published: September 2020

Purpose: SMAD4 loss causes genomic instability and the initiation/progression of head and neck squamous cell carcinoma (HNSCC). Here, we study if SMAD4 loss sensitizes HNSCCs to olaparib (PARP inhibitor) in combination with radiotherapy (RT).

Experimental Design: We analyzed HNSCC TCGA data for SMAD4 expression in association with FANC/BRCA family gene expression. Human HNSCC cell lines were screened for sensitivity to olaparib. Isogenic HNSCC cell lines were generated to restore or reduce SMAD4 expression and treated with olaparib, radiation, or the combination. HNSCC pretreatment specimens from a Phase I trial investigating olaparib were analyzed.

Results: SMAD4 levels correlated with levels of FANC/BRCA genes in HNSCC. HNSCC cell lines with SMAD4 homozygous deletion were sensitive to olaparib. In vivo, olaparib or RT monotherapy reduced tumor volumes in SMAD4 mutant but not SMAD4 positive tumors. Olaparib with RT dual therapy sustained tumor volume reduction in SMAD4 deficient (mutant or knockdown) xenografts, which exhibited increased DNA damage and cell death compared to vehicle treated tumors. In vitro, olaparib or in combination with radiation caused lower clonogenic survival, more DNA damage-associated cell death and less proliferation in SMAD4 deficient cells than in SMAD4-positive (endogenous SMAD4 or transduced SMAD4) cells. Applicable to clinic, 5 out of 6 SMAD4-negative HNSCCs and 4 out of 8 SMAD4-positive HNSCCs responded to a standard treatment plus olaparib in a Phase I clinical trial, and SMAD4 protein levels inversely correlated with DNA damage.

Conclusion: SMAD4 levels are causal in determining sensitivity to PARP inhibition in combination with RT in HNSCCs.

Citation Format: Christian D. Young, Ariel L. Hernandez, Li Bian, Sana D. Karam, David Raben, Xiao-Jing Wang. PARP inhibition enhances radiotherapy of SMAD4 deficient human head and neck squamous cell carcinomas in experimental models [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 6161.
Abstract 6168: Characterization of ATM inhibitor (AZD1390) distribution and pharmacodynamic changes in brain and glioblastoma in preclinical models to inform on mechanism of action
Authors: Oplustil O'Connor Lenka , Hamm Gregory , Udriste Maria , Ling Stephanie , Wilson Joanne , Durant Steve , Hughes Gareth , Cosulich Sabina , Shao Wenlin , Pass Martin , Jones Gemma , Pierce Andrew , Goodwin Richard , O'Connor Mark , Barrett Carl , Harrington Elizabeth
Published: September 2020

Ataxia telangiectasia mutated (ATM) kinase is a key factor in DNA double strand break response and is constitutively activated in glioblastoma, leading to marked radioresistance [Bartkova, Carruthers] and therefore an attractive radiosensitisation target in this disease. AZD1390, a potent ATM inhibitor optimized to cross the blood brain barrier, is being evaluated with radiotherapy in Phase 1 (NCT03423628). Positron emission tomography (PET) studies demonstrated AZD1390 brain exposure in healthy human subjects. This study characterises ATM inhibitor penetration and distribution in brain and GBM, as well as pharmacodynamic biomarker changes in human orthotopic GBM patient-derived explant (PDX) models. Radiation +/- AZD1390 treatment was evaluated for compound concentration in plasma and relative abundance in brain/ GBM regions by mass spectrometry imaging (MSI). Parallel samples were analysed by immunohistochemistry for biomarkers of DNA damage response and ATM signalling. We show AZD1390 penetrates brain tissue with preferential compound accumulation into tumour. Higher ratios of AZD1390 in GBM versus normal brain correlate with improved efficacy and greater levels of DNA damage. Moreover, AZD1390 in the GBM region was heterogeneously distributed with MSI characterized specific metabolic clusters having varied AZD1390 intensities. As expected, radiation treatment activated ATM signalling (indicated by increase in pRAD50 and γH2AX positive cells) which was effectively inhibited by AZD1390 co-treatment. Our work provides novel insights into ATM inhibitor pharmacokinetics in brain tumours and demonstrates the benefit of using MSI in conjunction with pharmacodynamic markers to provide a more complete picture of AZD1390 mechanism of action for future translational studies on clinical GBM samples.

Citation Format: Lenka Oplustil O'Connor, Gregory Hamm, Maria Udriste, Stephanie Ling, Joanne Wilson, Steve Durant, Gareth Hughes, Sabina Cosulich, Wenlin Shao, Martin Pass, Gemma Jones, Andrew Pierce, Richard Goodwin, Mark O'Connor, Carl Barrett, Elizabeth Harrington. Characterization of ATM inhibitor (AZD1390) distribution and pharmacodynamic changes in brain and glioblastoma in preclinical models to inform on mechanism of action [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 6168.