bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4508: Modulating the release rate of polyethylene glycol (PEG) polymers from interleukin-2 (IL-2) PEG conjugates to optimize anti-tumor efficacy
Authors: Li Hui , Song Yuntao , Zhou Haiping , Liao Chuan , Xu Meng , Li Chuang
Published: September 2020

The cytokine interleukin-2 (IL-2) is an endogenous agonist of the IL-2 pathway that activates and expands tumor killing lymphocytes. High doses of IL-2 are needed to activate CD8 T cells and NK cells, through binding on low-affinity IL-2 receptor beta gamma subunits (IL-2Rβγ). Severe toxic side effects associated with high doses, as well as the poor PK profile of IL-2, have limited its clinical usage. In addition of the toxicities, IL-2 also stimulates proliferation and activation of regulatory T cells (Tregs) through binding on the high-affinity heterotrimeric IL-2 receptor alpha beta gamma subunits (IL-2Rαβγ), which may antagonize the intended anti-tumor immunity. We report on an IL-2 PEGylation strategy to improve the pharmacokinetic properties of IL-2. Utilizing our proprietary PEGylation technology, IL-2 was covalently connected with multiple polyethylene glycol (PEG) polymers through releasable linkers. Our technology enabled the conjugation of IL-2 with average of 5-6 polyethylene glycol (PEG) polymers in high efficiency and 80-90% isolated yield. Anti-tumor efficacy of IL-2 was optimized by modulating the hydrolysis rate of the releasable linkers connecting IL-2 and polymers. The releasable linkers used in the study had the hydrolysis rate in the range of 15 to 120 hours to release one PEG polymer from the conjugates. The conjugates with hydrolysis rate of 20-60 h showed the optimal anti-tumor efficacy in B16F10 animal model. These results suggest that our novel approach of IL-2 PEGylation may optimize IL-2 biological activity and anti-tumor efficacy. Modulating the releasable rate of the PEG polymer would optimize the pharmacokinetic properties of the released active IL-2 species and enable the optimized anti-tumor efficacy. We are currently exploring the usage of our technology platform to further engineer IL-2 as a non-IL2Rα binding conjugate for improved toxicity profile.

Citation Format: Hui Li, Yuntao Song, Haiping Zhou, Chuan Liao, Meng Xu, Chuang Li. Modulating the release rate of polyethylene glycol (PEG) polymers from interleukin-2 (IL-2) PEG conjugates to optimize anti-tumor efficacy [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 4508.
Abstract 4518: Pre-clinical development of NL-201: A de novo α-independent IL-2/IL-15 agonist
Authors: Walkey Carl D. , Hara Paul , Tatalick Laurie , Ulge Umut , Drachman Jonathan , Silva Daniel-Adriano
Published: September 2020

Interleukin-2 (IL-2) and interleukin-15 (IL-15) promote anti-tumor immune activity by stimulating effector T and NK cells expressing the β and γ subunits of the IL-2 and IL-15 receptors. However, preferential activation of off-target cell populations expressing the high-affinity α subunit (including regulatory T cells, eosinophils, and endothelial cells) has limited the safety, therapeutic benefit, and widespread clinical use of these cytokines. Recently, we used sophisticated computational methods to create Neoleukin-2/15 (Neo-2/15), an entirely de novo α-independent agonist of both the IL-2 and IL-15 receptors. Unlike strategies that attempt to sterically inhibit or mutate the α-binding region of IL-2 or IL-15, Neo-2/15 was computationally designed to have no α-binding interface, while also having increased affinity for the β and γ signaling subunits. As a result, Neo-2/15 stimulates effector T and NK cells more potently and selectively than IL-2 or IL-15. Here, we report on the preclinical development and pharmacology of NL-201, a long-circulating variant of Neo-2/15 that is intended for clinical applications. NL-201 was derived from Neo-2/15 via the site-specific conjugation of a single PEG molecule and is manufactured using a simple and scalable process. NL-201 has a prolonged blood half-life and a sustained pharmacodynamic effect while retaining the high potency, selectivity, and thermodynamic stability of Neo-2/15. Preclinical studies demonstrate the robust antitumor activity of NL-201 as a single agent across a wide range of syngeneic murine tumor models, including those that do not respond well to checkpoint inhibitors. Due to its stability, high potency, and selectivity, NL-201 may overcome the long-standing challenges associated with the clinical use of IL-2 and IL-15 for cancer immunotherapy.

Citation Format: Carl D. Walkey, Paul Hara, Laurie Tatalick, Umut Ulge, Jonathan Drachman, Daniel-Adriano Silva. Pre-clinical development of NL-201: A de novo α-independent IL-2/IL-15 agonist [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 4518.
Abstract 4543: Phenotypic screening of bispecific antibodies in 3D tumor immune cell cocultures
Authors: Daszkiewicz Lidia , Goverse Gera , Beztsinna Nataliia , Visser Benjamin , van der Drift Lois , Yan Kuan , Price Leo
Published: September 2020

INTRODUCTION

The co-culture of tumor and immune cells in 3D enables a better recapitulation of the complex interactions between cells and the tumor microenvironment. High content analysis can be used to evaluate the effects of diverse immunotherapeutics on anti-tumor immune responses in a 3D setting. Functional read-outs can include active migration of immune cells towards tumoroids, infiltration of immune cells into the tumoroids and tumor killing. Here, we have applied high content analysis to quantify the effects of bi-specific antibodies targeting both the tumor and the T cells in 3D culture in a 384 well plate format.

MATERIAL and METHODS

Tumoroids generated from various cancer cell types were cultured in 3D in an extracellular matrix-rich protein hydrogel. Partially HLA-matching T cells isolated from healthy PBMCs donors were added with or without activation to the 3D culture and their infiltration into tumoroids and subsequent killing was visualized using high-content microscopy. Quantification of effects on immune cells and tumoroids was achieved with morphometric analysis with OMinerTM software.

RESULTS and DISCUSSION

Image-based analysis enabled the assessment of tumor-immune cell interactions and tumor killing in 3D co-cultures. The activation status of T cells determined the level of immune cell infiltration and killing of the tumoroids. T cell engaging bi-specific antibodies enhanced infiltration and tumor killing and was associated with increased IFNγ levels in culture supernatants.

CONCLUSION

Here we demonstrated the enhanced immune cell infiltration and tumor killing induced by bi-specific antibodies that simultaneously engage both T cells and tumors in a 3D setting. This assay represents a more physiologically relevant approach to enable cancer immunotherapy drug developers to evaluate the activity of novel immunotherapeutic molecules and better understand their mechanism of action.

Citation Format: Lidia Daszkiewicz, Gera Goverse, Nataliia Beztsinna, Benjamin Visser, Lois van der Drift, Kuan Yan, Leo Price. Phenotypic screening of bispecific antibodies in 3D tumor immune cell cocultures [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 4543.
Abstract 4569: DNA vaccine targeting BARF1 induces potent antitumor immunity against Epstein-Barr virus-associated carcinomas
Authors: Zhu Xizhou
Published: September 2020

Epstein-Barr Virus (EBV) is an oncogenic pathogen, with which more than 90% of world population has been infected. It is associated with several types of cancer, and no EBV-specific therapies are available for these diseases. Recent studies targeting EBV latent proteins have shown limited efficacy, for their moderate immunogenicity. Therefore, the development of effective anti-EBV immunotherapies is in need. In this study, we screened a group of EBV proteins and selected BARF1 as a vaccine candidate, for its strong immunogenicity and high expression in EBV positive nasopharyngeal and gastric carcinomas. The BARF1 vaccine was developed through a synthetic DNA approach and delivered in vivo by electroporation. The vaccine induced potent BARF1-specific antibody and polyfunctional effector T cell responses in both C57BL/6 and BALB/c mice. We next established two carcinoma models by generating BARF1-expressing CT26 and MC38 cell lines. In the therapeutic model where mice were challenged with cancer cells and then treated, BARF1 vaccine was able to suppress cancer progression alone and showed synergy when combined with anti-PD1 antibody. In the minimal residual disease model where mice were immunized and then challenged with cancer cells, one dose of BARF1 vaccine was able to generate 100% complete response in treated animals. CD8 T cell was shown to be necessary for this protection. Finally, using the in vivo imaging system, we found that the BARF1 vaccine-induced immune responses cleared tumor in 60% of subjects as soon as 48 hours post tumor challenge. Tumors were cleared in all animals by day 10. In summary, BARF1 DNA vaccine provides a novel tool of immunotherapy that is highly specific to EBV positive malignancies and combining it with checkpoint inhibitors might open up new avenues for precision and combination immunotherapy against EBV-associated cancer. In the future, we will also target other viral-specific cancer antigens with our DNA vaccine platform, exploiting its capability to induce potent cytotoxic T lymphocyte response.

Citation Format: Xizhou Zhu. DNA vaccine targeting BARF1 induces potent antitumor immunity against Epstein-Barr virus-associated carcinomas [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 4569.
Abstract 4576: Oncolytic vaccinia virus TG6010 expressing cytidine deaminase induces DNA damage and shows potent anti-tumor effects
Authors: Foloppe Johann , Kempf Juliette , Pichon Christelle , Findeli Annie , Farine Isabelle , Hortelano Julie , Moreau Baptiste , Klein Michèle , Cochin Sandrine , Brandely Renée , Koerper Véronique , Silvestre Nathalie , Machon Christelle , Guitton Jérôme , Quéméneur Eric , Erbs Philippe
Published: September 2020

The recombinant Vaccinia virus Copenhagen strain deleted in Thymidine Kinase and Ribonucleotide Reductase (VVCOPTK−RR−) is a potent and versatile oncolytic platform that has demonstrated strong activity in various preclinical models. The deletion of such VV genes inhibits viral replication in normal cells, while retaining therapeutic replication in tumor cells. TG6002, a VVCOPTK−RR− expressing the suicide gene FCU1, is under investigation in Phase I trials in patients with advanced gastrointestinal tumors.

We have developed a new product based on the VVCOPTK−RR− vector, named TG6010, expressing the human cytidine deaminase (CDA) that efficiently catalyzes the deamination of cytidine and deoxycytidine to uridine and deoxyuridine, respectively.

The tumor specific expression of CDA by the VV leads to a depletion of cytidine and deoxycytidine. This cytidine/deoxycytidine depletion resulting from CDA overexpression, activated a DNA damage response highlighted by an induction of H2AX phosphorylation.

Next, to validate the potential therapeutic use of TG6010, we analyzed the effects of the virus on human xenograft tumors implanted in mice. We observed, after systemic injection of TG6010, high expression of CDA in the tumors with a significant increase in DNA damage in vivo as revealed by the γH2AX foci assay. In addition, intraperitoneal administration of TG6010 in a xenograft model of human ovarian carcinomatosis resulted in significantly prolonged animal survival.

In conclusion, the oncolytic vaccinia virus expressing the cytidine deaminase (TG6010) has shown potent anti-tumor effects. Mechanistically, due to the cytidine deaminase overexpression, we observed induction of a DNA damage response. These are data encouraging of clinical development for this highly potent oncolytic Vaccinia virus TG6010 against ovarian cancer.

Citation Format: Johann Foloppe, Juliette Kempf, Christelle Pichon, Annie Findeli, Isabelle Farine, Julie Hortelano, Baptiste Moreau, Michèle Klein, Sandrine Cochin, Renée Brandely, Véronique Koerper, Nathalie Silvestre, Christelle Machon, Jérôme Guitton, Eric Quéméneur, Philippe Erbs. Oncolytic vaccinia virus TG6010 expressing cytidine deaminase induces DNA damage and shows potent anti-tumor effects [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 4576.
Abstract 4584: A novel combination of a multipeptide KRAS vaccine and an ACAT1 inhibitor to prevent lung cancer
Authors: Pan Jing , Zhang Qi , Palen Katie , Johnson Bryon , Wang Li , Sei Shizuko , Shoemaker Robert H. , Lubet Ronald A. , Wang Yian , You Ming
Published: September 2020

Kirsten rat sarcoma viral oncogene homolog (KRAS) gene encodes a small GTPase that cycles between GDP and GTP-bound states, which modulates important cell surface receptors essential for cell proliferation and survival. It is one of the most common drivers for many types of cancers, is present in up to 25% all human tumors, and 80% of mutation occurs in codon12. Efforts to target KRAS-driven cancers preventively or therapeutically have been unsuccessful. We recently formulated a multi-peptide KRAS vaccine (KVAX), which targets both wild-type and G12D mutant forms of KRAS and has 100% sequence homology between human and mouse. KVAX elicited strong immunologic response and exhibited striking cancer preventive efficacy when administered prior to KRAS mutation induction. However, clinical high-risk individuals may already express mutant KRAS, which has known to promote an immunosuppressive environment. Therefore, we combined KVAX with avasimibe (AVA), an ACAT1 inhibitor that modulates lipid metabolism in T cells and facilitates better TCR/peptide/MHC interactions, in syngeneic lung cancer mouse model and genetically engineered mouse model, in which mutant KRAS initiated lung tumorigenesis before vaccination. We found that the combination significantly decreased the presence of regulatory T cells in the tumor microenvironment, facilitated CD8+ T cell infiltration in tumor sites, and ultimately led to enhanced anti-cancer efficacy. Using 10X Genomics, we performed single cell RNA-seq on lung tumors from mice treated with KVAX and AVA and found significant increases in APC monocytes and CD4+ Th1 cells, especially increases in the effector/memory T cell population. Meanwhile, KVAX also decreased Tregs and M2 macrophages. These studies include the evaluation of tumor-infiltrating immune cells by single-cell RNA-seq (scRNA-seq) and provide a more detailed mechanisms of action of KVAX alone or in combination with AVA in modulating the tumor immune microenvironment.

Citation Format: Jing Pan, Qi Zhang, Katie Palen, Bryon Johnson, Li Wang, Shizuko Sei, Robert H. Shoemaker, Ronald A. Lubet, Yian Wang, Ming You. A novel combination of a multipeptide KRAS vaccine and an ACAT1 inhibitor to prevent 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 4584.
Abstract 4595: SERGE- the development of an optical sensor for the detection glioblastoma recurrence
Authors: Fischer Igor , Hänggi Daniel , Kahlert Ulf Dietrich
Published: September 2020

Objective: Total gross resection is the clinical most positive prognostic treatment of brain cancer. Tumor recurrence is a frequent event in patients undergone neurosurgical removal of the primary cancer. Current surveillance of the disease requires cost-intensive and infrastructural sophisticated means and is applied in an infrequent manner. Better tools to monitor the therapy success in order to increase the chances of total gross resection of the recurrent lesion are of high clinical need.

Concept and Results: Photodynamic diagnosis (PDD) is a clinical applied procedure that exploits metabolic alterations in brain tumor cells to discriminate of cancerous tissue from healthy brain parenchyma. This project aims to develop an implant-prototype that enables the surveillance of any tumor recurrence at the primary tumor resection cavity by applying the principles of PDD. We present initial electro-engineering details and the study design to probe its functionality in vitro and in vivo. Supported by the Volkswagen Foundation the project aims to create evidences for the technological feasibility and therapeutic benefit of SERGE as a future class III medical device.

Citation Format: Igor Fischer, Daniel Hänggi, Ulf Dietrich Kahlert. SERGE- the development of an optical sensor for the detection glioblastoma recurrence [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 4595.
Abstract 4601: Toward the development of a $100 screening test for 6 major cancer types
Authors: Xu Linhao , Wang Jun , Yang Ting , Tao Jinsheng , Liu Xin , Ye Zhujia , Zhao Dezhi , Jiang Zeyu , Chen Zhiwei , Fan Jian-Bing
Published: September 2020

Introduction: Early cancer detection remains a critical challenge to improve patients' survival rate and clinical outcomes. Alterations in DNA methylation play a key role in tumor initiation, development, progression and metastasis. Cell-free DNA (cfDNA) methylation analysis on liquid biopsies (e.g., blood and urine) holds a great promise for early detection of cancers even without symptoms. In this context, we are developing a non-invasive, cost-effective test that can detect several major cancer types, including lung (LnC), breast (BrC), colorectal (CRC), gastric (GC), esophageal (EC) and liver (LvC) cancers with high specificity (Sp) and sensitivity (Sn). These six cancer types represent two third of the newly diagnosed cancer patients and 72% of cancer deaths every year in China.

Methods: In order to identify highly effective DNA methylation markers for early detection, we have conducted genome-wide methylation profiling on more than 2,000 benign and malignant tissue samples, including large number of early stage (Stage I/II) tumors and pre-cancerous lesions from lung, breast, colorectum, liver, stomach and esophagus. We then developed a high performance assay, Aurora, to capture cancer-specific DNA methylation signatures in plasma. Specifically, cfDNA was extracted from each of the plasma samples, bisulfite-converted, and analyzed by NGS on Illumina MiSeq system.

Results and Discussion: To date, methylation profiles of plasma samples derived from, 1) 58 normal and 106 malignant pulmonary nodule patients, 2) 31 normal and 85 malignant CRC patients, and 3) 81 normal and 78 malignant breast nodule patients, were analyzed. Overall performances (Area Under Curve, AUC) of the Aurora test are 90%, 98% and 92% for LnC, BrC and CRC, respectively. Further clinical validation and evaluation are being conducted in larger independent cohorts of these three types of cancers as well as additional cohorts for the other cancers (GC, EC and LvC). The superior performance and low cost (~$100) make the Aurora test very promising for simultaneous screening of multiple major cancers in symptom-free, average risk populations.

Citation Format: Linhao Xu, Jun Wang, Ting Yang, Jinsheng Tao, Xin Liu, Zhujia Ye, Dezhi Zhao, Zeyu Jiang, Zhiwei Chen, Jian-Bing Fan. Toward the development of a $100 screening test for 6 major cancer types [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 4601.
Abstract 4626: TBX1 functions as a potential oncogene in breast cancer
Authors: Huang Shuya , Shu Xiang , Wu Jie , Wang Jifeng , Guo Xingyi , Ping Jie , Bauer Joshua A. , Long Jirong , Shu Xiao-Ou , Zheng Wei , Cai Qiuyin
Published: September 2020

We have previously identified a genetic variant, rs34331122 in the 22q11.21 locus, to be associated with breast cancer risk in genome-wide association studies. This SNP is located in the intronic region of the T-box transcription factor 1 (TBX1) gene. TBX1 is a member of the T-box transcription factor gene family, which plays important roles in embryogenesis and organogenesis. Previous studies have indicated that TBX1 exerted potential tumor suppressor functions in mouse skin tumors and human thyroid cancer. However, the biological functions and molecular mechanisms of TBX1 in breast cancer remain largely unknown. In this study, we investigated whether the association of the 22q11.21 locus with breast cancer risk was mediated by the TBX1 gene. Using The Cancer Genome Atlas (TCGA) data, we found that TBX1 expression was significantly upregulated in breast cancer tissues compared to adjacent normal breast tissues. To investigate the biological functions of the TBX1 gene in breast cancer, we knock-downed TBX1 gene expression using siRNAs in MDA-MB-231, MCF-7 and T47D breast cancer cell lines. Our results showed that knocking down TBX1 expression significantly inhibited breast cancer cell proliferation, migration and invasion. We further performed RNA sequencing in both TBX1 knocked-down and control MDA-MB-231 cells. Our differential gene expression and gene set enrichment analyses showed that knocked-down TBX1 expression significantly downregulated cell cycle-related genes, including CDC6, CDK6, DHFR, CIT, ORAI1 and TOP2A (P < 10−30). These six downregulated genes in TBX1 knocked-downed MDA-MB-231 cells were further confirmed by quantitative real-time PCR (qPCR). Furthermore, using the JASPAR database to predict TBX1 binding motif, we found that TBX1 may bind to the promoter regions of these six genes. In summary, our results suggest that TBX1 acts as a potential oncogene in breast cancer through regulating the expression of cell cycle-related genes, and the associations between genetic variants in the 22q11.21 locus and breast cancer risk may be mediated through altered expression of the TBX1 gene.

Citation Format: Shuya Huang, Xiang Shu, Jie Wu, Jifeng Wang, Xingyi Guo, Jie Ping, Joshua A. Bauer, Jirong Long, Xiao-Ou Shu, Wei Zheng, Qiuyin Cai. TBX1 functions as a potential oncogene in 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 4626.
Abstract 4647: Diet quality indexes and risk of hepatocellular carcinoma: Findings from the Singapore Chinese Health Study
Authors: Luu Hung N. , Neelakantan Nithya , Geng Ting-ting , Wang Renwei , Adams-Haduch Jennifer , Jin Aizhen , Van Dam Rob M. , Koh Woon-Puay , Yuan Jian-Min
Published: September 2020

Background. There is a limited research on the association between combined dietary measures and liver cancer risk. We investigated the association between the four diet-quality index (DQI) scores, the Alternative Health Eating Index-2010 (AHEI-2010), Alternate Mediterranean Diet (aMED); Dietary Approaches to Stop Hypertension (DASH) and Heathy Diet Indicator (HDI) with liver cancer risk, using data from the Singapore Chinese Health Study, a prospective cohort of 63,257 Chinese men and women aged 45-74 at enrollment in 1993-1998 with up to 25 years of follow-up.

Methods. The Cox proportional hazard regression method was used to estimate hazard ratio (HR) and its 95% confidence interval (CI) of HCC with each DQI after adjustment for multiple potential confounders. The unconditional logistic regression method was used to evaluate the associations between the DQIs and HCC risk among a subset of individuals who tested negative for hepatitis B surface antigen (HBsAg).

Results. During an average 17.7 years of follow-up, 561 incident HCC cases were identified. High scores of AHEI-2010, aMED and DASH, representing higher dietary quality, were associated with lower risk of HCC (all Ptrend<0.05). Compared with the lowest quartile, HRs (95% CIs) of HCC risk for the highest quartile of AHEI-2010, aMED and DASH were 0.71 (0.55-0.91), 0.73 (0.54-0.98), and 0.68 (0.52-0.89), respectively. These inverse associations were stronger in men than in women. No significant association between HDI and HCC risk was observed. Among HBsAg-negative individuals, similar inverse associations were observed, and the strongest inverse association was for aMED that reached statistically significance level (HRQ4vsQ1=0.48, 95% CI: 0.24-0.98).

Conclusion. Higher scores of AHED, aMED, and DASH were significantly associated with reduced risk of HCC in this Asian population. The findings support the notion that adherence to a healthier diet may lower the risk of HCC, suggesting that dietary modification may be an effective approach for primary prevention of HCC.

Funding: The Singapore Chinese Health Study was supported by the National Institutes of Health (NIH) of the United States (grants # R01 CA144034 and UM1 CA182876).

Citation Format: Hung N. Luu, Nithya Neelakantan, Ting-ting Geng, Renwei Wang, Jennifer Adams-Haduch, Aizhen Jin, Rob M. Van Dam, Woon-Puay Koh, Jian-Min Yuan. Diet quality indexes and risk of hepatocellular carcinoma: Findings from the Singapore Chinese Health Study [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 4647.
Abstract 4682: Influence of PI3K/AKT pathway on imatinib mesylate treatment outcome in chronic myeloid leukemia patients
Authors: Banapuram Swathi , Gorre Manjula , Digumarti Raghunadha Rao , Satti Vishnupriya , Palle Komaraiah , Annamaneni Sandhya
Published: September 2020

Chronic myeloid leukemia (CML) is a common myeloproliferative disorder characterized by cytogenetic abnormality [t(9;22)(q34;q11)] that generates BCR-ABL oncogene with aberrant tyrosine kinase activity, the driver event in CML origin. Tyrosine kinase inhibitors (TKIs) are remarkably effective; yet, treatment failure is common in some cases due to the accumulation of mutations in BCR-ABL itself and other genes causing drug resistance/relapse, which indicates that BCR-ABL inhibition alone may not be sufficient to combat CML. The PI3K/Akt/mTOR signaling pathway downstream of BCR-ABL has been proven to be an important survival mechanism in leukemias. In vitro studies demonstrated that targeting this pathway could be a viable strategy, but needs further validation in CML patients. In this study, we examined the critical PI3K/AKT/mTOR pathway genes (PIK3CA, PTEN, AKT1, SHIP1, mTOR, and FOXO3A) in CML patients (n=51) using qPCR method at diagnosis and after treatment, and analyzed with respect to BCR-ABL status, and JAK2, STAT5, and STAT3 levels. Interestingly, significant upregulation of AKT1 (p=0.012) and PIK3CA (p=0.026) and downregulation of PTEN (P=0.06) and SHIP1 (p=0.039) was observed in samples at diagnosis compared to their post imatinib treated. Remarkably, these elevated AKT1 and PI3KCA transcript levels were significantly associated with increased BCR-ABL and JAK2 transcript levels, but not STAT3 and STAT5 suggesting PI3K pathway as a prominent target in CML. However, with respect to the clinical phase and imatinib response, none of the genes showed significant differences except for PTEN, which was significantly downregulated in advanced phase (p<0.042) and in cases with poor imatinib response. Overall, these results indicate a critical role for PI3K pathway in CML and its maintenance.

Citation Format: Swathi Banapuram, Manjula Gorre, Raghunadha Rao Digumarti, Vishnupriya Satti, Komaraiah Palle, Sandhya Annamaneni. Influence of PI3K/AKT pathway on imatinib mesylate treatment outcome in chronic myeloid leukemia patients [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4682.
Abstract 4686: Core transcriptional regulatory circuitry FOXA1-MAX-SP1 promotes malignancy of hepatocellular carcinoma
Authors: Peng Li , Yuan Xiaoqing , Chen Zhen , Zhang Chaoyang , Tian Duanqing , Yin Dong , Lin De-Chen
Published: September 2020

Background: Super-enhancers (SEs) are large clusters of transcriptionally active enhancers. Functionally, they are capable of driving the expression of key genes that control cell identity, which are also particularly important for tumor biology. This study is aimed at probing the biological function and mechanisms of the hyper-activation of the SEs associated core transcriptional regulatory circuitry (CRC) constituted by FOXA1-MAX-SP1 in hepatocellular carcinoma.

Results: ChIP-seq analysis of H3K27ac showed that the expression of three transcription factors FOXA1, MAX and SP1 is driven by super-enhancers in hepatocellular carcinoma cells. Notably, these three transcription factors also regulate large number of super-enhancers, whose characteristics fit the mathematical model of the core transcriptional regulatory circuitry (CRC). ChIP-seq analysis of FOXA1, MAX, SP1, H3K27ac, H3K4me1 and H3K4me3 confirmed the occupancy of FOXA1, MAX and SP1 at super-enhancers and promoters of some genes, including FOXA1, MAX and SP1. Also, FOXA1, MAX and SP1 have a positive correlation with each other in human hepatocellular carcinoma tissue. In addition, CCK8 assay showed that FOXA1, MAX and SP1 could promote cell growth of HCC cells. Relative open chromatin at FOXA1, MAX and SP1 genes mirrored their more active transcriptional state in HCC patients from TCGA ATAC-seq data. The mRNA expression of these three TFs were increased, which predict a worse prognosis of HCC patients.

Conclusion: FOXA1-MAX-SP1 regulate super-enhancers and may form a core transcriptional regulatory circuitry, so as to promote malignancy of hepatocellular carcinoma. They are potential therapeutic targets and prognostic biomarkers for hepatocellular carcinoma.

This work was supported by the National Natural Science Foundation of China (81802812 and 81972658), the Natural Science Foundation of Guangdong Province (2018A030313129 and 2019A1515012114).

Citation Format: Li Peng, Xiaoqing Yuan, Zhen Chen, Chaoyang Zhang, Duanqing Tian, Dong Yin, De-Chen Lin. Core transcriptional regulatory circuitry FOXA1-MAX-SP1 promotes malignancy of 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 4686.
Abstract 4721: Arid1a suppresses Akt/NRas-driven hepatocarcinogenesis in mice
Authors: Zhang Shanshan , Zhou Yufeng , Cao Di , Ren Qiannan , Zhang Meiyin , Mai Shijuan , Wang Huiyun
Published: September 2020

The SWI/SNF chromatin remodeling complexes are increasingly appreciated for their roles in carcinogenesis. Recent progress in human cancer genomic sequencing revealed that chromatin remodeling complexes are frequently targeted by cancer mutations, particularly in hepatocellular carcinomas (HCC). However, their precise role in HCC remains controversial and incompletely understood. AKT and NRAS are two major liver oncogenes that promote aberrant growth, metabolism and proliferation of hepatocytes, leading to initiation and development of HCC. Here, we investigated the role of Arid1a, one of the most mutated subunits of the SWI/SNF chromatin-remodeling complex across all human cancers, in hepatocarcinogenesis. To this end, we used Akt/NRas-driven mouse liver cancer model generated by hydrodynamic transfection (HDT). We found that overexpression of Arid1a strongly attenuated Akt/NRas-driven liver tumorigenesis and prolonged survival of animals. Conversely, knockdown of Arid1a accelerated Akt/NRas-driven liver cancer development and shortened survival in mice. We further showed that Arid1a antagonized activation of multiple molecular pathways downstream of Akt/NRas. Conclusion: Arid1a acts as a tumor suppressor in the Akt/NRas mouse HCC model. These new findings provide in vivo evidence for a tumor suppressive role of Arid1a-dependent chromatin remodeling in Akt/NRas-driven hepatocarinogenesis.

Citation Format: Shanshan Zhang, Yufeng Zhou, Di Cao, Qiannan Ren, Meiyin Zhang, Shijuan Mai, Huiyun Wang. Arid1a suppresses Akt/NRas-driven hepatocarcinogenesis in mice [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 4721.
Abstract 473: Preclinical model of radiation induced lymphopenia to identify abrogation strategies to enhance cancer therapy
Authors: Sharma Amrish , Neri Shinya , Venkatesulu Bhanu P. , Lin Steven H.
Published: September 2020

Radiation induced lymphopenia (RIL) is a common and yet often ignored side effect of radiation therapy that has been observed for over 100 years. Numerous studies have demonstrated the correlation between RIL and inferior overall survival outcomes in nearly all cancer types that are treated with radiation therapy. To assess the causative link between RIL and poor disease outcomes, as well as its impact on therapy response, we have developed a mouse model of RIL. Using cone beam CT based image-guided radiation targeting the mid-thorax, we showed that conventionally fractionated radiation of 2 Gy x 5 fractions caused severe and sustained lymphopenia in both C57BL/6 (Th1) and BALB/c (Th2). Increasing the radiation dose and radiation field resulted in more pronounced RIL. Enlarging the volume to include the spleen along with the thorax resulted in greater than 90% elimination of circulating lymphocytes (CD8/CD4/CD19/NK cells). However, radiation to the head resulted in only minimal lymphodepletion suggesting site specific susceptibility to the development of severe RIL. The severity of the lymphodepletion was also unperturbed by rate of blood flow as the administration of varying anesthetics that caused 5 fold differences in heart rate did not alter the severity of RIL. Functionally, RIL abrogated the immunotherapeutic effectiveness of anti-CTLA4 mAb as lymphopenic mice bearing CT26 tumors had complete lack of response to anti-CTLA4 treatment as compared to its control littermates. Similarly, animals treated with radiotherapy had nearly complete amelioration of radiotherapy effectiveness in the setting of severe RIL. However, exogenous administration of IL-15 super agonist (IL-15 SA) (IL-15 and IL-15Rα-Fc) therapy replenished cytotoxic CD8+ T killer cells and natural killer cells to control levels which restored the tumoricidal effect of radiation therapy. Taken together our study have generated the evidence for the causative link between RIL and poorer disease outcomes and presented a potential therapeutic strategy to overcome the adverse effect of RIL.

Citation Format: Amrish Sharma, Shinya Neri, Bhanu P. Venkatesulu, Steven H. Lin. Preclinical model of radiation induced lymphopenia to identify abrogation strategies to enhance cancer therapy [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 473.
Abstract 4743: Genetic characteristics of PanNETs, Rectal NETs, Thoracic NETs and its correlation with efficacy of chemotherapy
Authors: Chi Yihebali , Liu Weili , Zuo Lijie , Wang Yuehua , Cai Weijing , Shi Susheng , Ge Yanying , Li Rui , Song Lijie , Yang Yiqi , Liu Zheng , Dou Xiaowu , Zhao Hong
Published: September 2020

Background: Neuroendocrine tumors (NETs) are clinical heterogeneous. To date, no studies have focused on the genetic characteristics of NETs at different anatomical sites in the Chinese cohort. We aim to explore the genetic similarities or differences between NETs at different sites, and evaluate the predictive indicators of chemotherapy based on Temozolomide.

Methods: The patients were from a prospective clinical study STEM (from a single institute, NCT03204032, NCT03204019). This cohort included 47 patients with NETs, including 14 Pancreatic NETs (PanNETs), 11 Rectal NETs, 14 Thoracic NETs, and 8 Others. Whole exome sequencing (WES) was performed.

Results: A total of 76,057 somatic mutations were detected in 16,510 genes. The median tumor mutation burden (TMB) was 13.4, 10.5, 11.0 and 11.9 in PanNETs, Rectal NETs, Thoracic NETs and Others, respectively. After sequencing depth filtering, they were 3.5, 3.9, 3.3 and 3.6, respectively. Low intra-tumoral heterogeneity (ITH) was observed in NETs. The median ITH index was 2, 2.5, 2.5 and 2 in PanNETs, Rectal NETs, Thoracic NETs and Others, respectively; the median Shannon diversity index (SDI) was 0.23, 0.13, 0.23 and 0.24, respectively. The driver gene mutations varied in NETs at different sites. The recurrently mutated driver genes were MEN1, ATM, TSC2, PLXNB2, MUC6, AMER1 and USP9X for PanNETs; APC, APOB and FAT1 for Rectal NETs; KMT2B and CACNA1A for Thoracic NETs. Copy number analysis showed that copy number loss frequently occurred in all NETs. The median copy number variation burden (CNV burden) was 34.0, 10.9, 16.1 and 11.6 in PanNETs, Rectal NETs, Thoracic NETs and Others, respectively. The CNV burden of PanNETs was higher than others. Based on the results of CNVs, NETs at different sites could be distinguished by linear discriminant analysis (LDA). We analyzed the correlation between genetic characteristics and progression-free survival (PFS). As a result, NETs with higher TMB (>10 /Mb, P=0.043) or higher CNV burden (>20, P=0.047) showed statistically longer PFS. More patients should be included to find a correlation between genetic characteristics and PFS.

Conclusions: NETs at different sites harbored different somatic driver mutations, and copy number loss was widespread in NETs. NETs with higher TMB or higher CNV burden showed statistically longer PFS.

Citation Format: Yihebali Chi, Weili Liu, Lijie Zuo, Yuehua Wang, Weijing Cai, Susheng Shi, Yanying Ge, Rui Li, Lijie Song, Yiqi Yang, Zheng Liu, Xiaowu Dou, Hong Zhao. Genetic characteristics of PanNETs, Rectal NETs, Thoracic NETs and its correlation with efficacy of chemotherapy [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 4743.
Abstract 4747: Genomic and transcriptomic landscape of AFP-producing gastrointestinal adenocarcinoma
Authors: Li Jun , Li Xinlin , Ye Qiancheng , Tian Yu , Kong Xiangxing , Liu Yue , Xu Xiaoming , Zhang Lina , Xiao Qian , Hu Yeting , Xu Rongzhen , Li Jingsong
Published: September 2020

AFP-producing gastrointestinal adenocarcinoma (APA-GI) is a rare type of cancer, typically with hepatoid differentiation, chemotherapy resistance and poor prognosis. As the mechanisms of its AFP production and hepatoid differentiation remain unclear, the present study aims to reveal the pathogenesis of APA-GI through genomic and transcriptomic profile analysis. We firstly performed 200 × whole exome sequencing of 29 APA-GI and 8 non-APA-GI cases from our hospital, but found no significant difference in single nucleotide variants (SNV) or insertion/deletion(indels). Additionally, there was no overlap gene when compared with the known genes harbored at 20q11.21-13.12 reported in gastric hepatoid adenocarcinoma. We then performed single-cell mRNA sequencing in one APA-GI, and observed several hepatoid differentiation markers in AFP positive cells including HNF4A and HNF1A. To further determine the specific alterations of APA-GI, we divided 339 gastric cancer patients from TCGA database into AFP-low group and AFP-high group by the level of AFP transcription through a dynamic grouping method named grid search. We performed a series differentially expressed genes detections within different grouping strategies and no significant SNV or indels difference was recognized. We identified hepatoid differentiation markers (i.e. HNF4A, HNF1A and ALB, et al) in AFP-high patients in mRNA level. As the difference of hepatoid differentiation markers only occurred in the mRNA level without DNA alteration, we hypothesized that it was probably caused by epigenetic regulation. We further compared these genes with an epigenetic regulators database named EpiFactors. Finally, it showed CCCTC-Binding Factor Like (CTCFL) was stably related with AFP producing within the grid search. In conclusion, comprehensive genomic analysis of APA-GI shows significant hepatoid differentiation at the transcriptomic level without disease-specific DNA alteration. The pathogenesis of APA-GI may be associated with other mechanisms like epigenetic regulation. Further study was needed to confirm the role of epigenetic regulators we detected in the present study in the development of APA-GI.

Citation Format: Jun Li, Xinlin Li, Qiancheng Ye, Yu Tian, Xiangxing Kong, Yue Liu, Xiaoming Xu, Lina Zhang, Qian Xiao, Yeting Hu, Rongzhen Xu, Jingsong Li. Genomic and transcriptomic landscape of AFP-producing gastrointestinal adenocarcinoma [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 4747.
Abstract 4751: Brain and plasma choline compounds altered by cancer and cancer-induced cachexia
Authors: Bharti Santosh K. , Winnard Paul T. , Sharma Raj Kumar , Penet Marie-France , Krishnamachary Balaji , Bhujwalla Zaver M.
Published: September 2020

Introduction: Cancer-induced cachexia has lethal consequences. Cachexia results in lower tolerance to chemotherapy1. Cachectic patients experience a wide range of symptoms affecting the function of organs such as muscle, liver, brain, and heart, causing significant morbidity 2. An altered choline metabolism is one of the hallmarks of cancers. Here, for the first time, we identified changes in brain choline metabolism induced by cachexia-inducing pancreatic cancer xenografts and related these changes to plasma choline levels.

Methods: Panc1 or Pa04C cells were implanted in the flank of SCID mice. Snap frozen brains were powdered under liquid nitrogen, weighed, and dual phase extraction was performed3. All 1H MR spectra were acquired on a Bruker Avance III 750 MHz MR spectrometer. Mouse plasma choline was quantified using a fluorometric assay (Choline Assay Kit, BioAssay Systems). Brain and plasma studies were performed with normal mice (n=10), mice with cachexia-inducing Pa04C tumors (n=9) and mice with non cachexia-inducing Panc1 tumors (n=10).

Results and Discussion: Cachexia-inducing Pa04C tumors induced significant weight loss in mice compared to Panc1 tumor bearing mice or normal mice3. A significant decrease of brain choline was observed in Pa04C tumor bearing mice compared to control mice as well as Panc1 tumor bearing mice, whereas brain phosphocholine (PC) levels were significantly lower compared to normal mice. Brain glycerophosphocholine (GPC) was significantly lower in both tumor bearing groups compared to normal mice. We observed a significant decrease of plasma choline in Pa04C tumor bearing mice. Plasma choline in Panc1 tumor bearing mice also decreased but not as drastically. The changes in plasma choline levels most likely contributed to the reduction of brain choline, PC and GPC. Reduced plasma choline may represent a risk factor for cancer as choline deficiency is known to result in lymphocyte apoptosis and DNA damage 4 that may compromise the immune response. Normalization of plasma choline using metabolic supplements merits investigation in reducing cancer morbidity and the onset of cachexia.

Acknowledgement: This work was supported by NIH R35CA209960 and R01CA193365.

References

1. Argiles JM et al., Nature reviews Cancer. 14(11):754-762.

2. Inui A. 2002. CA Cancer J Clin. 52(2):72-91.

3. Winnard PT, Jr., et al., Cancer Res. 76(6):1441-1450.

4. da Costa KA, et al., The American journal of clinical nutrition. 84(1):88-94.

Citation Format: Santosh K. Bharti, Paul T. Winnard, Raj Kumar Sharma, Marie-France Penet, Balaji Krishnamachary, Zaver M. Bhujwalla. Brain and plasma choline compounds altered by cancer and cancer-induced cachexia [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 4751.
Abstract 4808: ONC201 decreases protein chaperone ClpX to unleash mitochondrial protease ClpP activity, intregrated stress response and tumor cell death
Authors: Zhou Lanlan , Safran Howard P. , Glantz Michael J. , El-Deiry Wafik S.
Published: September 2020

Brain tumors are one of the most lethal cancers in children and young adults. We previously reported that first-in-class small-molecule imipridone ONC201 can antagonize dopamine receptor D2 (DRD2), inactivate ERK/AKT, induce the integrated stress response (ISR), upregulate pro-apoptotic TRAIL receptor DR5, deplete cancer stem cells, and induce growth arrest or cell death in tumor cells. ONC201 crosses the blood-brain barrier and has induced durable tumor regressions in adult and pediatric H3K27M-mutant glioma patients. We hypothesized that ONC201 may synergize with radiotherapy and temozolomide in brain tumors. Glioblastoma (GBM: SNB19, T98G and U251), diffuse intrinsic pontine glioma (DIPG: SU-DIPG-IV and SU-DIPG-36) and atypical teratoid rhabdoid tumor (ATRT: BT-12, BT-16) cell lines were tested using cell viability or colony formation assays with ONC201 up to 20 μM alone or in combination with radiotherapy up to 8 Gy and temozolomide up to 100 μM. We observed synergy between ONC201 and radiation and temozolomide by multiple assays with the best combination index of 0.31 under the treatment of 0.625 μM ONC201, 25 μM TMZ and 4 Gy irradiation in cell viability assay. We observed induction of PKA substrate phosphorylation as a marker of DRD2 antagonism, induction of ATF4 as a marker of ISR activation, and multiple markers of cell death in treated brain tumor cells. Mitochondrial dysfunction induced by ONC201 was greatly reflected in reduction of maximal cell respiration in brain tumor cell lines. ClpXP is a proteolytic machine in mitochondria comprising of the protein chaperone ClpX and protease ClpP, a recently described binding target of ONC201. We have observed that ONC201 induced cytotoxicity depends on ClpP and knockdown of ClpP strongly protects multiple human cancer cell lines from ONC201-mediated (but not other drugs, e.g. temozolomide) cytotoxicity. The ClpX protein level decreased dramatically upon ONC201 treatment. We are exploring the potential interplay between ClpP, dopamine receptors, the ISR and TRAIL signaling pathways after single agent or combinatorial treatments. Our data suggests that ONC201 may be combined synergistically with temozolomide or radiation to address gliomas, along with potential pharmacodynamic biomarkers. The results implicate ClpX and ClpP in downstream cell death induction following ONC201 treatment of brain tumor cells.

Citation Format: Lanlan Zhou, Howard P. Safran, Michael J. Glantz, Wafik S. El-Deiry. ONC201 decreases protein chaperone ClpX to unleash mitochondrial protease ClpP activity, intregrated stress response and tumor cell death [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 4808.