bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1879: A novel dual CAB nectin-4 x CD3 bispecific antibody targeting solid tumors
Authors: Lucas Matthew , Cugnetti Ana Paula , Liu Haizhen , Xing Charles , Wang Jing , McNeeley Patricia , Wheeler Christina , Joyner Solmarie , Johnson Kyrie , Woodard Kathryn , Zhou Wei , Chang Cathy , Frey Gerhard , Boyle William J. , Short Jay M.
Published: April 2023

Nectin-4 is a Ca2+ independent immunoglobulin-like protein that is primarily involved in cell-cell adhesion. Nectin-4 is overexpressed in several tumor types, including breast, lung, urothelial, colorectal, pancreatic, and ovarian cancers. Overexpression of Nectin-4 is associated with multiple aspects of tumor progression such as proliferation, angiogenesis, epithelial to mesenchymal transition, metastasis, DNA repair and tumor relapse. This overexpression is correlated with poor prognosis, making Nectin-4 an attractive target for developing new therapeutic strategies for cancer. As a result, a substantial number of antibody drug conjugates targeting Nectin-4 are under development. Most of these therapies, however, display a high rate of drug related toxicities accentuating the need for alternative approaches for anti-cancer therapeutics targeting Nectin-4. We leveraged BioAtla’s Conditionally Active Biologic (CAB) platform to develop a dual CAB T cell engager (TCE) bispecific antibody targeting Nectin-4 and CD3 receptors (CAB-Nectin-4 x CAB-CD3 bispecific antibody). The dual CAB Nectin-4 x CD3 bispecific antibody was engineered to bind with high affinity to both Nectin-4 and CD3 receptor antigens under conditions that mimic the acidic tumor microenvironment, but with lower binding activity in physiological (alkaline) conditions. In vitro and in vivo efficacy data for a dual-CAB Nectin-4 x CD3 bispecific antibody will be presented. Our data will demonstrate that the dual-CAB bispecific antibody promotes cytotoxicity of Nectin-4-expressing cancer cells in vitro and induces complete tumor regression in vivo. Further, the dual CAB Nectin-4 x CD3 bispecific antibody has higher potency in inducing primary T cell activation measured by means of cytokine release and cancer cell cytotoxicity under acidic conditions compared to the non-CAB benchmark bispecific antibody.

Citation Format: Matthew Lucas, Ana Paula Cugnetti, Haizhen Liu, Charles Xing, Jing Wang, Patricia McNeeley, Christina Wheeler, Solmarie Joyner, Kyrie Johnson, Kathryn Woodard, Wei Zhou, Cathy Chang, Gerhard Frey, William J. Boyle, Jay M. Short. A novel dual CAB nectin-4 x CD3 bispecific antibody targeting solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1879.
Abstract 1967: Addressing breast cancer disparities: knowledge and attitudes towards cancer prevention through breastfeeding among African American women in Georgia
Authors: Vernon Marlo M.
Published: April 2023

Background: A qualitative needs assessment was conducted to inform the development of a culturally competent breast and ovarian cancer prevention intervention, through the promotion of breastfeeding (BF) and genetic screening for cancer risk among African American (AA) women. In Georgia, AA women are diagnosed with later-stage breast/ovarian cancer and have higher mortality rates. BF initiation and longer duration are associated with reduced risk of cancers, yet BF rates are lower among AA women and few BF promotion programs are culturally competent.

Methods: 20 AA Mothers (Age: 18-25y=40%, 25-29y=40%, 31-35y=20%; Edu: ≤HS=60%; # of children: 1=40%) who were either pregnant (n= 9, 45%) or within the first year postpartum (n= 11, 55%), were interviewed. Questions assessed BF intentions and prior experiences, knowledge of BF preventing cancer, familial breast/ovarian cancer risk, best promotion practices, and genetic screening attitudes.

Results: Mothers (70%) reported previous BF initiation, with 65% reporting 6wks as the longest BF duration. No mother reported knowing that BF can prevent cancer. When asked about their own risk of cancer, a quarter of mothers reported knowing they were at risk of breast or ovarian cancer; a large majority (85%) were open to genetic screening and wanted to know their genetic risk for cancer. Mothers identified two themes: education and help/peer support to improve BF initiation and duration. There were no differences in responses based on demographics, BF experience, parity, or pregnancy/postpartum.

Discussion: Results confirm a lack of knowledge of the cancer risk reduction conferred by BF. Women reported significant support for genetic screening and a need for education and peer social support for BF; these results will inform intergenerational cancer prevention education efforts.

Citation Format: Marlo M. Vernon. Addressing breast cancer disparities: knowledge and attitudes towards cancer prevention through breastfeeding among African American women in Georgia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1967.
Abstract 1976: Lung cancer screening disparities in Southeastern U.S
Authors: Rivers Desiree A.
Published: April 2023

Purpose: In Alabama, lung cancer is the leading cause of cancer death. High lung cancer incidence and mortality rates are attributed to high smoking rates among underserved, low income, and rural populations. Rural residents in Alabama tend to be older, engage in risky health behaviors, and have lower adherence to preventive care than do their urban and suburban counterparts. Disparities in mortality rates between rural and urban areas are particularly large for lung cancer. It can be explained by increased tobacco use and the preponderance of late-stage diagnoses following the lack of uptake of LCS. Therefore, we examined factors associated with LCS uptake among patients who were referred to screening at University of Alabama at Birmingham (UAB) from 2015-2020.

Method: A retrospective cohort of individuals seen at UAB who were eligible for lung cancer screening from 01/01/2015 and 12/31/2020 was used to define the cohort. Included individuals were 55-80 years old, without diagnostic codes for lung cancer (ICD9 162.9 or ICD10 C34.90 within the past 10 years), and a history of smoking: Patient demographic variables included Age; Sex; Race/Ethnicity; distance from UAB and RUCA codes. Chi-square tests and Student t-tests was used to compare screening uptake across patient demographic and clinical variables. We used bivariate analyses to determine significant predictors of lung cancer screening uptake at UAB.

Results: 67355 were identified as eligible for LCS of whom 1147 were screened, 16 individuals were not AL residents and therefore were not included in further analysis. Of those 1129 individuals screened, mean age was 67.02, male (54.92%) and Non-Hispanic White (57.77%). When compared to those who were not screened(97.86%), those screened were more likely to have live in an urbanized area (2.14%). Additionally, when compared to those who were not screened (48.16 mi), those screened were more likely to live closer to UAB (28.67 mi).

Conclusion: LCS has not reached all subgroups; additional targeted efforts are essential to increase lung cancer screening uptake.

Citation Format: Desiree A. Rivers. Lung cancer screening disparities in Southeastern U.S [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1976.
Abstract 2054: Whole-genome sequencing based homologous recombination deficiency testing for precision oncology of breast cancers
Authors: Kim Ryul , Park Seongyeol , Lim Joonoh , Yi Boram , Koo Jaemo , Lee Sangmoon , Lee Jeong Seok , Park Yeon Hee , Ju Young Seok
Published: April 2023

Breast cancers (BCs) with defects in homologous recombination repair (HRR) pathway depend on the Poly (ADP-ribose) polymerase enzymes (PARP) for their survival and thus are sensitive to PARP inhibitors (PARPi). To screen out BCs with homologous recombination deficiency (HRD), germline mutations in BRCA genes have been tested, which constitute approximately 5% of unselected BC patients. However, a substantial proportion of BCs show HRD phenotypes despite the absence of BRCA inactivating mutations in germline. Herein, we apply whole-genome sequencing (WGS) and whole-transcriptome sequencing (WTS) in 332 samples from 325 BC patients for characterizing their HRD phenotypes for PARPi treatments. Of 325 BC patients, 75 cases (23.1%) showed remarkable HRD phenotypes, based on mutational signatures of HRR abrogation. Intriguingly, 51 cases do not harbor any germline mutations in HR-related genes including BRCA1 and BRCA2. The whole-genome portrait of these samples was similar to that of HRD-positive BCs with

germline mutation in HRR pathway, but showed markedly distinctive features from HRD-negative BCs. With a notable exception of BRCA1, the second most common somatic mutation in HRD-positive BCs without germline mutation, the mutational landscape of the whole study samples was primarily determined by hormone receptor and HER2 positivity, not by HRD status. Likewise, WTS-based clusters determined in an unsupervised manner were highly dependent on the histological subtype of BC, not on HRD status or germline mutations in the HRR pathway. Notably, differentially expressed gene and gene set enrichment analysis revealed that HRD-positive BCs had an increased inflammatory activity compared to BCs with proficient HR. Overall, our analysis demonstrate that WGS enables better classification of the HR status in BC than conventional BRCA germline testing, suggesting WGS as a rapid and sensitive tool for screening breast cancers for PARPi treatment.

Citation Format: Ryul Kim, Seongyeol Park, Joonoh Lim, Boram Yi, Jaemo Koo, Sangmoon Lee, Jeong Seok Lee, Yeon Hee Park, Young Seok Ju. Whole-genome sequencing based homologous recombination deficiency testing for precision oncology of breast cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2054.
Abstract 2088: gCNV-Seeker: A comprehensive germline CNV calling pipeline based on whole genome sequencing data
Authors: Sang Jian , Zhang Tongwu , Wedge David , Landi Maria Teresa
Published: April 2023

Germline Copy Number Variation (gCNV) is a type of genomic structural alteration including deletion or duplication of small genomic regions (50bp to 1MB) and can have an important role into cancer etiology. Whole genome sequencing (WGS) has been considered to be the most effective technology for genome-wide identification of gCNVs. However, an easy-to-use gCNV calling pipeline based on WGS data is still lacking. Here, we present the gCNV-Seeker, a user-friendly, rigorous computational pipeline to detect gCNV events based on WGS with standardized quality control and data visualization features. gCNV-Seeker initially adopts GATK probabilistic algorithms to detect a set of raw gCNV events and subsequently applies Binary Segmentation and Pruned Exact Linear Time (PELT) algorithms for re-segmentation and boundary revision of the gCNV candidates, respectively. In addition, gCNV-Seeker is built with several functionalities, including quality control, filtration, annotation and visualization to identify the gCNV regions (gCNVRs) of interest. We applied gCNV-Seeker to the WGS data from 872 lung cancers in never smokers from the Sherlock-Lung study and 3202 WGS data from the 1000 Genomes Project (1KGP) (reference) and identified CNVRs associated with lung cancer risk in never smokers. For example, in comparison to WGS data from 1KGP, we identified several CNVR candidates overlapping with known LC susceptible genes, e.g., the GSTM1/2 homozygous deletion (OR = 1.64, 95% CI=1.43-1.88, P < 0.01) and CYP2A6/7 homozygous or heterozygous deletion (OR = 1.59, 95% CI=1.24-2.03, P < 0.01). To evaluate its performance, we also carried out a comprehensive comparison of gCNV calling results between gCNV-Seeker and the 1KGP structural variant calling pipelines (PMID: 36055201) for common gCNVRs in the 1KGP WGS data. gCNV-Seeker showed a significant improvement on specificity and sensitivity for gCNVR detection compared with the 1KGP pipeline. In terms of specificity, 1315 out of 2692 (HGSV_9692) GSTM1/2 heterozygous deletion events (48.85%) originally identified by 1KGP were detected as homozygous deletions by gCNV-Seeker. These results were further confirmed by manual check. In terms of sensitivity, gCNV-Seeker detected total 299 deletion events in CYP2A6/7 regions, which were also manually confirmed. This corresponds to an increase of 48.02% compared to the 202 deletion events (HGSV_232739 & HGSV_232740) originally identified by the 1KGP pipeline. gCNV-Seeker will be a publicly available cross-platform (Linux and IOS) pipeline accessible in GitHub.

Citation Format: Jian Sang, Tongwu Zhang, David Wedge, Maria Teresa Landi. gCNV-Seeker: A comprehensive germline CNV calling pipeline based on whole genome sequencing data [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2088.
Abstract 2224: Applying optical genome mapping to detect genomic biomarkers and use for residual disease monitoring in hematologic malignancies
Authors: Chitsazan Alex , Pang Andy , Hastie Alex
Published: April 2023

Disease monitoring is important to detect cancer relapse, often as a result of a treatment-surviving cancer cell population. Detection of relapse is critical to determine the intensity and length of treatments and to predict the outcome of those treatments. Currently, karyotyping, fluorescent in situ hybridization (FISH), flow cytometry, PCR, and next-generation sequencing (NGS) are often used for this application. PCR is sensitive and highly effective, but as a targeted approach, it requires the identification of an appropriate biomarker. Karyotyping and FISH are used to follow structural variants, especially those detected at diagnosis and known to be drivers of carcinogenesis; however, karyotype has low sensitivity (~5-10% variant allele frequency (VAF)) and FISH is only targeted.We describe a novel workflow to find structural variants (SVs) using optical genome mapping (OGM) and to use those SVs as biomarkers for disease monitoring assessment. The initial SV profile from a cancer sample can be obtained by running the standard somatic workflow for OGM: 1.5 Tbp of high molecular genomic DNA is collected (without any amplification/selection), and molecules that can capture SV breakpoints are locally assembled into genome maps. These locally assembled genome maps are used to make high-confidence SV calls at least 5 kbp in size. Then, presumptive somatic variants are revealed by further comparison against an OGM control sample SV database (included in Bionano Access/Solve software). Subsequently, to detect residual cancer cells post-treatment, an ultra-deep coverage of approximately 5 Tbp of DNA can be collected, and these molecules are directly aligned to the genome maps in the initial cancer sample. This direct alignment approach is fast and sensitive in identifying remnants of pretreatment originator cells. We are currently applying this workflow to recover known leukemia-associated inter-chromosomal translocations, and our preliminary results indicate a sensitivity of detecting calls at a minimum of 0.5% VAF. Continual investigation on limit of detection is ongoing, but with the improvement on throughput and algorithms, we envision that this workflow to be essential in discovering low level clonal cancerous DNA.

Citation Format: Alex Chitsazan, Andy Pang, Alex Hastie. Applying optical genome mapping to detect genomic biomarkers and use for residual disease monitoring in hematologic malignancies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2224.
Abstract 2227: Streamlined workflow for analyzing and reporting hematological malignancies in Bionano VIATM software
Authors: Clifford Benjamin , Hauenstien Jen , Pang Andy Wing Chun , Burnside Rachel D. , Miller Neil , Hastie Alex , Chaubey Alka
Published: April 2023

Cytogenetic analysis for hematological malignancies most often includes karyotyping, fluorescence in situ hybridization, and rarely, chromosomal microarray (CMA). While considered the gold standard, karyotyping and FISH have significant limitations in resolution of structural variants (SVs). CMA has a much higher resolution, but cannot detect balanced SVs, and is less commonly used in the hematologic oncology setting. Optical genome mapping (OGM) can detect all classes of SVs at high resolution (duplications, deletions, inversions, insertions, translocations), and as such, detects many copy number variants (CNVs) and SVs that have not been previously described. Although these novel SVs will help better describe genetic contributions to disease, analysis, interpretation and curation can take considerable time. A streamlined workflow has been developed for use with Bionano VIA™ software for a comprehensive analysis of relevant SVs in a hematological malignancy genome. First, by employing disease-specific decision trees for variants published in guidelines for those conditions, the software automatically flags Tier 1A variants detected according to ACMG/AMP/CGC criteria. Second, the overall genome complexity is assessed by observing chromosomal abnormalities detected by OGM. Large events (>5Mb) are counted, and complex genomes are assigned if there are more than 3 or 5 events, depending on the cancer type. Third, calls are further refined by filtering on a pan-cancer specific list to capture those variants that are then manually classified as Tier 1B or 2 by the analyst. The Bionano VIA software allows case-specific and disease subtype-specific information to be added to various sections within the software, such as the Knowledgebase, Event Table, and Sample Info fields to facilitate autopopulation of reportable data and interpretations into a formatted report template. Bionano VIA software can also accommodate multiple platforms simultaneously (e.g., NGS panels) to provide users with a comprehensive view of genomic aberrations relevant to hematological malignancies. This workflow is intended to streamline analysis and automate reporting of oncology samples, which can be extremely complex and often require extensive research.

Citation Format: Benjamin Clifford, Jen Hauenstien, Andy Wing Chun Pang, Rachel D. Burnside, Neil Miller, Alex Hastie, Alka Chaubey. Streamlined workflow for analyzing and reporting hematological malignancies in Bionano VIATM software [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2227.
Abstract 2229: Cell state transitions shape the intratumoral composition of small cell lung carcinoma
Authors: Gopal Priyanka , Petty Aaron , Rogacki Kevin , Bera Titas , Bareja Rohan , Peacock Craig , Abazeed Mohamed
Published: April 2023

Introduction: Small cell lung carcinoma (SCLC) is characterized by rapid growth, early metastases, and initial response followed by almost invariable resistance to therapy. Studies to date have not determined the extent that diverse transcriptional programs drive SCLC and contribute to its lethality. We sought to characterize the intra-tumoral transcriptional heterogeneity of SCLC. We identify multivalent, distinct, and commutable transcriptional states that confer discrete functions in individual SCLC tumors.

Methods: We developed a biorepository of patient-derived xenografts (PDX) (n = 64) and matched PDX-derived ex vivo lines. We used multi-omic profiling (RNAseq, scRNAseq, and ATAC seq), single-cell fluorescence tracking of fate-defining transcription factor (TF)-driven states, and mathematical and statistical models (Markov chain) to study the topology of the SCLC transcriptional landscape and its plasticity. Human tumor material and associated clinical data were obtained after informed written consent on an IRB-approved prospective registry.

Results: We show that individual SCLC tumors are more heterogenous than previously appreciated, displaying distinctive equilibria in the proportion of cells within well-delimited cellular states (ASCL1, NEUROD1 and YAP1). We also show that transcriptional states undergo transitions, which we identified as a mechanism for maintaining cell state diversity. We measured the kinetics of state transitions using single-cell fluorescence tracking of ex vivo cultures and found that these measure were associated significantly with transition estimates using stochastic transition theory (i.e. Markov chains). ATAC-seq profiling indicated a role for the epigenome in the state diversity of SCLC. Namely, there was preferential promoter accessibility to Ascl1, NeuroD1, and Yap1 in a manner consistent with gene and protein expression in the respective subpopulations. Our results indicate that the transition rates between cell types in individual tumors were largely governed by tendencies to reach an equilibrium state that are critical for configuring intratumoral cell state proportions.

Conclusion: In conclusion, we demonstrate that TF driven cell states can transition to maintain an equilibrium in cell state proportions. Our work advances a model of cellular states and program diversity in SCLC and nominates new therapeutic strategies designed to limit the plasticity of this lethal cancer.

Citation Format: Priyanka Gopal, Aaron Petty, Kevin Rogacki, Titas Bera, Rohan Bareja, Craig Peacock, Mohamed Abazeed. Cell state transitions shape the intratumoral composition of small cell lung carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2229.
Abstract 2237: Inhibition of HSF1 demonstrates therapeutic efficacy in preclinical models of cholangiocarcinoma
Authors: Carlson Danielle M. , Abdelrahman Amro , Fogliati Alessandro , Lynch Isaac , Tomlinson Jennifer , Truty Mark , Smoot Rory
Published: April 2023

Introduction: Cholangiocarcinoma (CCA) is a highly lethal, and biologically heterogenous malignancy arising from the biliary tree. With only moderate responses to cytotoxic chemotherapy and despite the development of targeted therapies, treatment success has been hindered by primary or acquired resistance. New therapeutic strategies are necessary to improve outcomes in these patients. NXP800 is a heat shock factor 1 (HSF1) small-molecule inhibitor with activity previously noted in ovarian cancers with ARID1A mutations. ARID1A mutations are common, well-known players in cholangiocarcinoma. In preclinical exploratory studies, we examined treatment responses in validated cholangiocarcinoma patient derived xenograft (PDX) models.

Methods: In our preliminary studies we evaluated response to HSF1 inhibition in a single, ARID1A wildtype PDX model. Xenografts were expanded into the flank of NOD/SCID mice and treatment begun when tumors averaged 120mm3. 16 tumor bearing mice were randomized and 8 mice were grouped into both the treatment and control arms. Mice were then treated with either NXP800 (35 mg/kg), in the treatment arm, or vehicle, in the control arm, via oral gavage 5 days on, 2 days off, for 28 days. Tumor volume and animal weight were collected twice weekly.

Results: After screening of our Human CCA PDX library of 57 patients, we identified an established and histologically validated PDX from a patient with intrahepatic cholangiocarcinoma obtained at surgical resection. Molecular characterization of the tumor identified NRAS Q61 mutation, FGFR1 and FGFR2 overexpression, androgen receptor overexpression, microsatellite stability, and a total tumor mutational burden of 2.6. RNA sequencing was completed and an evaluation of a 21-gene signature of YAP activity had identified this PDX as having an elevated activity signature compared to a cohort of 27 additional PDX tumors. Treatment with NXP800 was associated with a statistically significant decrease in tumor size compared to control tumor bearing mice, with a mean difference of the delta in tumor volume (852.6 mm3; 95% CI: 321 - 1384; P-value = 0.0057).

Conclusion: NXP800, a HSF1 inhibitor, demonstrated significant therapeutic activity in a cholangiocarcinoma PDX. Further studies are needed and being performed to determine the role of HSF1 inhibition in human cholangiocarcinoma.

Citation Format: Danielle M. Carlson, Amro Abdelrahman, Alessandro Fogliati, Isaac Lynch, Jennifer Tomlinson, Mark Truty, Rory Smoot. Inhibition of HSF1 demonstrates therapeutic efficacy in preclinical models of cholangiocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2237.
Abstract 2241: Transcriptomic analysis of tetraspanins in colorectal cancer
Authors: Suwatthanarak Thanawat , Tanjak Pariyada , Chaiboonchoe Amphun , Acharayothin Onchira , Thanormjit Kullanist , Suwatthanarak Tharathorn , Tangkullayanone Watsaphon , Pithukpakorn Manop , Chinswangwatanakul Vitoon
Published: April 2023

Introduction: Transmembrane proteins and their overexpression have been highly involved in malignancies. In particular, Tetraspanins (TSPANs)—a family of four-span transmembrane proteins—have been proposed as “master organizers” of multi-molecular membrane complexes by associating with specific partners and thus contributed to the progression of many cancer types. To date, transcriptomic analysis of all human TSPANs in colorectal cancer (CRC) has remained unclear. Here, we focused on utilizing next-generation RNA sequencing (RNA-Seq) to explore the expression level of TSPANs in primary tumor tissues of CRC, which is a complex disease and the second cause of cancer mortality worldwide.

Experimental Procedures: Resected primary tumor tissues of CRC were collected from 100 patients along with the written informed consents. Later, total RNA was extracted from all frozen patient tissues. RNA-Seq of those RNA samples was next done before the transcriptomic analysis of TSPANs in CRC was performed. Immunofluorescence (IF) and Immunohistochemistry (IHC) stains were also employed to confirm the target protein expression in formalin-fixed paraffin-embedded tissue (FFPE) slides from CRC patients.

Results: As a result of transcriptomic analysis of TSPANs in CRC tissues, among 33 human TSPANs, the high expression levels (transcript per million above the average of all TSPANs) of TSPAN1, TSPAN8, TSPAN24, TSPAN29 and TSPAN30 were observed in the patient-derived CRC tissues. Notably, the expression level of TSPAN8 was comparable with that of EPCAM, which is a well-known tumor marker. Moreover, the high expression of TSPAN8 protein could be clearly observed in both IF and IHC-stained FFPE slides from CRC patients. Our result highlights the possible relevance of these overexpressed TSPANs in CRC development.

Conclusions: This study is the first RNA-Seq-based transcriptomic analysis of all human TSPANs in CRC. Our finding suggests the significant overexpression of particular TSPANs, especially TSPAN8, in CRC. Further analyses will be beneficial for translational research in CRC. The overexpression of TSPANs may suggest a framework to further explore diagnostic, prognostic, and predictive surface biomarkers, therapeutic targets, and other applications for CRC.

Citation Format: Thanawat Suwatthanarak, Pariyada Tanjak, Amphun Chaiboonchoe, Onchira Acharayothin, Kullanist Thanormjit, Tharathorn Suwatthanarak, Watsaphon Tangkullayanone, Manop Pithukpakorn, Vitoon Chinswangwatanakul. Transcriptomic analysis of tetraspanins in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2241.
Abstract 2340: Development of a CCR8 monoclonal antibody which blocks CCR8 signaling and abolishes the immunosuppressive function of Treg for the treatment of cancer
Authors: Fan Guohuang , Wang Jianfei , Wang Na , Zhang Yuanyuan , Zhang Yong , Tian Binle , Jia Xuebing
Published: April 2023

Within the tumor microenvironment (TME), regulatory T (Treg) cells are one attractive cell type for targeting as they play a critical role in immunosuppression. Therapeutic strategies that specifically inhibit tumor-infiltrating Tregs while sparing peripheral and normal tissue Tregs are highly desirable. The tumor-associated Tregs express the chemokine receptor CCR8, which plays a role in the induction, expansion, chemotactic migration, and immunosuppression of tumor-associated Tregs. Thus, targeted inhibition of CCR8 is a potential therapeutic strategy against cancer. We have developed a monoclonal antibody antagonist of CCR8, IPG0521, which potently inhibited the receptor-mediated signaling and chemotaxis in Treg cells with single digital nanomolar IC50. RNA sequencing (RNA-seq)analysis indicated that CCL1-induced up-regulation of immunosuppressive genes, including PD-1, CTLA4, IL-10, TIGIT etc., were reversed by IPG0521 in tumor-derived Treg cells. IPG0521 potently inhibited the growth of multiple tumor types in both syngeinic and immune-humanized mouse models, including lung cancer, liver cancer, breast cancer, via inhibiting Treg and activating CD8+ T cells. These data pave the way for the development of IPG0521, which is under IND filing, as a novel therapeutic agent against cancer.

Citation Format: Guohuang Fan, Jianfei Wang, Na Wang, Yuanyuan Zhang, Yong Zhang, Binle Tian, Xuebing Jia. Development of a CCR8 monoclonal antibody which blocks CCR8 signaling and abolishes the immunosuppressive function of Treg for the treatment of cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2340.
Abstract 2341: Group 2 innate lymphoid cells influence pancreatic stromal cell composition in homeostasis and pancreatic cancer
Authors: Yip Thomas , Lee Sheng , Stockis Julie , Garcia Celine , Raghunathan Shwetha , Lim Bram , Pinaud Silvain , Png Shaun , Halim Timotheus
Published: April 2023

Background: Group 2 Innate Lymphoid Cells (ILC2s) are tissue-resident innate immune cells critical in orchestrating type-2 immune responses. Cancer-associated fibroblasts are increasingly recognized as having important pro- and anti-tumorigenic roles. While type-2 immunity has been linked to fibrosis and tumor development, how ILC2s influence the stromal microenvironment in these settings is poorly understood. Here, we investigated the role of ILC2s on pancreatic stromal cell (PSC) behavior in healthy and tumor-bearing mice.

Methods: Naive, inflamed, and tumor-bearing pancreata of WT or ILC2-deficient mice were profiled by flow cytometry, histology, or multiplex immunofluorescence imaging. We assessed two-way communication between ILC2s and stromal cells using in silico analysis of single-cell RNAseq or in vitro coculture transcriptomic data. Putative crosstalk mechanisms were tested with in vitro and in vivo assays. Additionally, we performed two-photon microscopy of reporter mice pancreata and multiplex immunofluorescent imaging of tumor sections.

Results: Imaging studies show that ILC2s co-localize with IL-33+ stromal cells in the naïve, inflamed and tumor-bearing pancreas. In vivo and in vitro studies indicate that ILC2s are critical mediators of stromal cell proliferation and density in homeostatic and inflamed conditions, while influencing stromal cell-driven immune modulation in cancer. Single-cell transcriptomic analysis reveal distinct Il33+ and Il33- stromal cell clusters, which may engage in different crosstalk mechanisms with ILC2s, as determined by in silico inferential analysis, in vitro co-culture and in vivo studies.

Conclusion: ILC2s strongly influence stromal cell composition in both the healthy pancreas and pancreatic tumors. Given the importance attributed to the characteristic desmoplasia of pancreatic tumors, ILC2-driven modulation of the stromal compartment may affect tumor progression.

Citation Format: Thomas Yip, Sheng Lee, Julie Stockis, Celine Garcia, Shwetha Raghunathan, Bram Lim, Silvain Pinaud, Shaun Png, Timotheus Halim. Group 2 innate lymphoid cells influence pancreatic stromal cell composition in homeostasis and pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2341.
Abstract 2386: Label free imaging for rapid assessment of tumor viability in live tumor fragments
Authors: Ouellette Jonathan , Wargowski Ellen , Wait Eric , Zahm Chris , Johnson Scott , Oliner Jon
Published: April 2023

Live tumor fragments (LTF) that maintain a patient's relevant tumor microenvironment provides an opportunity to study treatment response. To verify viability of the fragments over time, we have devised and applied a label-free quantitative microscopy-based approach to monitor LTF health. This method directly assesses intrinsic fluorescence from metabolic co-factors and allows the spatial and temporal visualization of cell status. Using the label-free method of multiphoton fluorescence lifetime microscopy (MP-FLIM), we can assess excised tumor samples over 48 hours to determine the health of individual cells. Specifically, with MP-FLIM, we can analyze signals generated by the intrinsically fluorescent metabolic co-factor NAD(P)H which correlates with mitochondrial outer membrane permeabilization and the irreversible cascades leading to cell death. Our findings using MP-FLIM were confirmed using a standard caspase 3/7 live apoptosis assay. These data demonstrate MP-FLIM can detect and quantify cell viability without the use of potentially toxic dyes, thus enabling longitudinal multi-day studies assessing the effects of therapeutic agents on LTF.

Citation Format: Jonathan Ouellette, Ellen Wargowski, Eric Wait, Chris Zahm, Scott Johnson, Jon Oliner. Label free imaging for rapid assessment of tumor viability in live tumor fragments [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2386.
Abstract 239: Whole-genome sequencing based homologous recombination deficiency testing for precision oncology of breast cancers
Authors: Kim Ryul , Park Seongyeol , Lim Joonoh , Yi Boram , Koo Jaemo , Lee Sangmoon , Lee Jeong Seok , Park Yeon Hee , Ju Young Seok
Published: April 2023

Breast cancers (BCs) with defects in homologous recombination repair (HRR) pathway depend on the Poly (ADP-ribose) polymerase enzymes (PARP) for their survival and thus are sensitive to PARP inhibitors (PARPi). To screen out BCs with homologous recombination deficiency (HRD), germline mutations in BRCA genes have been tested, which constitute approximately 5% of unselected BC patients. However, a substantial proportion of BCs show HRD phenotypes despite the absence of BRCA inactivating mutations in germline. Herein, we apply whole-genome sequencing (WGS) and whole-transcriptome sequencing (WTS) in 332 samples from 325 BC patients for characterizing their HRD phenotypes for PARPi treatments. Of 325 BC patients, 75 cases (23.1%) showed remarkable HRD phenotypes, based on mutational signatures of HRR abrogation. Intriguingly, 51 cases do not harbor any germline mutations in HR-related genes including BRCA1 and BRCA2. The whole-genome portrait of these samples was similar to that of HRD-positive BCs with germline mutation in HRR pathway, but showed markedly distinctive features from HRD-negative BCs. With a notable exception of BRCA1, the second most common somatic mutation in HRD-positive BCs without germline mutation, the mutational landscape of the whole study samples was primarily determined by hormone receptor and HER2 positivity, not by HRD status. Likewise, WTS-based clusters determined in an unsupervised manner were highly dependent on the histological subtype of BC, not on HRD status or germline mutations in the HRR pathway. Notably, differentially expressed gene and gene set enrichment analysis revealed that HRD-positive BCs had an increased inflammatory activity compared to BCs with proficient HR. Overall, our analysis demonstrate that WGS enables better classification of the HR status in BC than conventional BRCA germline testing, suggesting WGS as a rapid and sensitive tool for screening breast cancers for PARPi treatment.

Citation Format: Ryul Kim, Seongyeol Park, Joonoh Lim, Boram Yi, Jaemo Koo, Sangmoon Lee, Jeong Seok Lee, Yeon Hee Park, Young Seok Ju. Whole-genome sequencing based homologous recombination deficiency testing for precision oncology of breast cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 239.
Abstract 2409: A synergistic effect of miR-21-5p inhibitor and PARP1 inhibitor to enhance the radiosensitivity of oral squamous cell carcinoma
Authors: Ma Lin , Hu Weitao , Fu Rui , Wu Anhui , Deng Qiangyong , Liu Qi
Published: April 2023

Radiotherapy is one of the standard treatments for oral squamous cell carcinoma (OSCC). Radiotherapy damages cancer cells mainly by inducing DNA double-strand breaks, but some advanced OSCC can effectively repair DNA damages, resulting in radiotherapy resistance. TCGA data showed that miR-21-5p was highly expressed in advanced OSCC and correlated with prognosis. However, the involvement of miR-21-5p in OSCC’s response to radiotherapy and the underlying mechanism are still unclear. When treated with a miR-21-5p inhibitor, we found OSCC cell lines (SAS and CAL33) became more sensitive to X-ray radiation at 2Gy (the commonly used single-dose in conventional radiotherapy), which was supported by the evidence that more resident 53BP1-marked-foci for non-repaired DNA double strand breaks (1.61±0.54 folder more in SAS, 1.47±0.49 folder more in CAL33, n=4, p<0.05) and less colony-forming capacity (14.62±2.36% in SAS, 18.65±4.32% in CAL33, n=3, p<0.05) than untreated control cells were observed at 24hr after radiation. However, the expression of PARP1, a DNA damage sensor for recognizing DNA damage and facilitating DNA repair, was significantly increased (1.38±0.12, n=4, p<0.05) in SAS cells treated with miR-21-5p inhibitor. In order to achieve a better tumor control, we tested the combined treatments with miR-21-5p inhibitor and PARP1 inhibitor (Olaparib) on SAS cells, which resulted in a synergistic effect in inhibition of the repair of DNA double strand breaks. The residual 53BP1 foci after 2Gy X-ray irradiation was increased 1.45±0.06 folder more when inhibiting both miR-21-5p and PARP1, while the colony-forming capacity of irradiated SAS cells was decreased by 39.55±5.89% (p<0.0001) in the same conditions. This work provided novel insights on the molecular mechanism of miR-21-5p regulated cell response to radiotherapy, which may give rise to potential therapeutic strategy for treatment of advanced OSCC.

Citation Format: Lin Ma, Weitao Hu, Rui Fu, Anhui Wu, Qiangyong Deng, Qi Liu. A synergistic effect of miR-21-5p inhibitor and PARP1 inhibitor to enhance the radiosensitivity of oral squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2409.
Abstract 2481: Human osteoclast assays for studying the effects of bone metastasis treatments with different mechanisms of action
Authors: Fagerlund Katja M. , Ristola Mervi , Suominen Mari I. , Mäki-Jouppila Jenni H. , Rissanen Jukka P.
Published: April 2023

Metastasis to bone is one of the common complications associated with different types of advanced cancer. Zoledronic acid, a third generation nitrogen containing bisphosphonate, accumulates in bones where it binds to hydroxyapatite crystals. For years, zoledronic acid has been regarded as the standard of care for patients with advanced cancer and bone metastasis to prevent skeletal complications. As an alternative therapeutic option, denosumab has also been found to be effective in delaying skeletal-related events in advanced cancer. Denosumab, a human monoclonal antibody against receptor activator of nuclear factor kappa-B ligand (RANKL), blocks the RANKL/RANK signalling pathway. In addition to the skeletal effects, both zoledronic acid and denosumab have shown direct effect on cancer cells. In this study, effects of zoledronic acid and denosumab on osteoclast differentiation and bone resorption activity were studied using human CD34+ osteoclast precursor cells that were cultured on bovine bone slices in 96-well plates for 7 days in the presence of RANKL and M-CSF, allowing them to differentiate into mature osteoclasts. In the osteoclast differentiation assay, zoledronic acid (0.0001 - 1 000 µM) and denosumab (0.01 - 10 µg/ml) were added into the culture medium at day 0. Tartrate-resistant acid phosphatase 5b (TRACP 5b) activity was measured in the culture medium at day 7 as an index of osteoclast differentiation. In the osteoclast activity assay, the test compounds were added into the culture medium at day 7, and the formed osteoclasts were allowed to resorb bone for 3 days. C-terminal crosslinked telopeptides of type I collagen (CTX-I) was measured in the culture medium at day 10 as an index of bone resorption. Denosumab and zoledronic acid showed strong concentration dependent inhibition of osteoclast differentiation and resorption activity. We conclude that this osteoclast culture system can be used for identifying new potential compounds affecting the disease process of bone metastases with different mechanisms of action. The manifold effects of zoledronic acid and denosumab on cancer cells remain to be studied in more detail.

Citation Format: Katja M. Fagerlund, Mervi Ristola, Mari I. Suominen, Jenni H. Mäki-Jouppila, Jukka P. Rissanen. Human osteoclast assays for studying the effects of bone metastasis treatments with different mechanisms of action [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2481.
Abstract 2483: Attenuated Salmonella blocks cancer metastasis by activating NK cells in an IFN-γ-dependent manner
Authors: Rong Li , Lin Qiubin , Lin Shuhai , Fu Guo , Huang Jiandong
Published: April 2023

Introduction and Objectives: The recent remarkable clinical results of cancer immunotherapies, such as immune checkpoint blockade and chimeric antigen receptor T cells (CAR-T), has sparked a great deal of enthusiasm in manipulating the immune response in the fight against cancer. Bacterial cancer therapies represent an alternative approach to targeting solid tumors for evoking anti-tumor immune responses. As a facultative anaerobic pathogen that can penetrate and colonize solid tumors, Salmonella typhimurium gained lot of interest in the investigation of bacterial cancer therapies and can exert its intrinsic antitumor activity by directly inducing systemic immune responses. However, most bacterial cancer therapies, including salmonella-based cancer therapies, have mainly focused on the suppression of already established tumors rather than metastasis. Metastasis accounts for 90% of cancer related deaths and blocking of metastatic cascade has critical clinical impact. However, the clinical drug development for cancer treatment, including cancer immunotherapies, is evaluated largely depending on their ability to cause tumor shrinkage and ignores the effect on metastasis as it has proven challenging to target. Therefore, there is an urgent need for novel therapeutic strategies and agents targeting metastasis.

Experimental Procedures and Outcomes: Using an attenuated Salmonella typhimurium strain YB1 engineered by our lab, we have found a potent suppressive effect of attenuated salmonella on cancer metastasis, regardless of cancer types and genetic background, by evoking strong anti-metastatic immune response. Further study showed that suppression of cancer metastasis by attenuated Salmonella only requires the innate immune response. Among the many induced cytokines, we identified IFN-γ as an indispensable factor for inhibiting cancer metastasis. Based on CyTOF (mass cytometry or cytometry by time of flight) analysis of the innate immune responses after Salmonella treatment and antibody-mediated cell depletion, we further demonstrated that NK cells are the major factor involved in Salmonella-provoked metastasis suppression. We found that IFN-γ was mainly produced by NK cells during early Salmonella infection, and in turn, IFN-γ promoted the accumulation, activation and cytotoxicity of NK cells. The IFN-γ-dependent NK cells directly eliminated newly accumulated cancer cells in the lung to block the cancer metastasis cascade in response to the Salmonella treatment.

Citation Format: Li Rong, Qiubin Lin, Shuhai Lin, Guo Fu, Jiandong Huang. Attenuated Salmonella blocks cancer metastasis by activating NK cells in an IFN-γ-dependent manner [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2483.