bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3141: Evolution of core archetypal phenotypes in progressive high grade serous ovarian cancer
Authors: Nath Aritro , Cosgrove Patrick , Copeland Benjamin , Mirsafian Hoda , Christie Elizabeth , Pflieger Lance , Majumdar Sumana , Cristea Mihaela , Han Ernest , Lee Stephen , Wang Edward , Fereday Sian , Traficante Nadia , Salgia Ravi , Werner Theresa , Cohen Adam , Moos Phillip , Chang Jeffrey , Bowtell David , Bild Andrea
Published: July 2021

The evolution of resistance in high-grade serous ovarian cancer (HGSOC) cells following chemotherapy is only partially understood. To uncover phenotypic changes associated with chemotherapy resistance, we profiled single-cell RNA-sequencing (scRNA-seq) transcriptomes of HGSOC tumors collected longitudinally during patient treatment. Analysis of scRNA-seq data from two independent patient cohorts revealed that HGSOC is driven by three core archetypal phenotypes, defined as oncogenic tasks that describe the majority of the transcriptome variation. A multi-task learning approach to identify the biological tasks of each archetype identified metabolism and proliferation, cellular defense response, and DNA repair signaling. The metabolism and proliferation archetype evolved during treatment and was enriched in cancer cells from patients that received multiple-lines of treatment and had elevated tumor burden indicated by CA-125 levels. The emergence of archetypes was not consistently associated with specific whole-genome driver mutations. However, archetypes were closely associated with subclonal populations at the single-cell level, indicating that subclones within a tumor often specialize in unique biological tasks. Our study reveals the core archetypes found in progressive HGSOC and shows consistent enrichment of subclones with the metabolism archetype as resistance is acquired to multiple lines of therapy.

Citation Format: Aritro Nath, Patrick Cosgrove, Benjamin Copeland, Hoda Mirsafian, Elizabeth Christie, Lance Pflieger, Sumana Majumdar, Mihaela Cristea, Ernest Han, Stephen Lee, Edward Wang, Sian Fereday, Nadia Traficante, Ravi Salgia, Theresa Werner, Adam Cohen, Phillip Moos, Jeffrey Chang, David Bowtell, Andrea Bild. Evolution of core archetypal phenotypes in progressive high grade serous ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3141.
Abstract 3151: β Hydroxy β Methylbutyrate synergizes with cytotoxic chemotherapy and immunotherapy in a mouse model of pancreatic cancer
Authors: Coleman Michael F. , Liu Kristyn A. , Etigunta Suhas K. , Pfeil Alex J. , Tang Xiaohu , Fabela Salvador , Lashinger Laura M. , Cui Zhengrong , Hursting Stephen D.
Published: July 2021

Background: Pancreatic ductal adenocarcinoma (PDAC) remains the 4th most deadly cancer in the US. Current chemotherapeutic regimens poorly control PDAC progression and no approved immunotherapeutic options currently exist. PDAC incidence and progression are increased by obesity, which promotes a potently protumor PDAC microenvironment characterized by inflammation, fibrosis, and immunosuppression. β Hydroxy β Methylbutyrate, found to be safe and well-tolerated in clinical trials over many years, blunts cancer cachexia; however it has not been tested in PDAC. Further, the role of HMB in modifying existing therapies has been understudied. We sought to determine if HMB treatment would alter therapeutic response of lean and obese mice to gemcitabine or anti-PD1 immunotherapy.

Methods: Lean and obese C57BL/6 mice were treated with HMB alone or in combination with gemcitabine or anti-PD1 immunotherapy following subcutaneous injection of Panc02 PDAC cells. In addition, bone marrow-derived macrophages (BMDM) were treated with HMB in vitro with or without LPS. Tumor/macrophage transcriptomic analysis, followed by gene set enrichment analysis and enrichment mapping identified transcriptional responses. CIBERSORTx determined microenvironment immune cell composition.

Results: a) HMB reduces obesity-associated PDAC progression; b) HMB treatment and gemcitabine together suppress PDAC growth in obese mice; c) HMB mitigates PDAC immunosuppression and promotes tumor immune surveillance; d) HMB enhances the efficacy of anti-PD1 immunotherapy; e) HMB promotes M1 polarization of BMDM.

Conclusion: Overall, HMB evokes antitumor immune responses in the context of pro-inflammatory stimuli such as obesity or immunotherapy. Moreover, HMB promotes immune surveillance in PDAC, synergizing with both cytotoxic chemotherapy and immunotherapy. HMB has been found to be safe and well tolerated in clinical trials for many years. Hence, HMB-induced suppression of PDAC tumor growth and promotion of immune surveillance may offer significant synergy with chemotherapies or immunotherapies in PDAC. This research was supported by R35CA197627 to S. Hursting.

Citation Format: Michael F. Coleman, Kristyn A. Liu, Suhas K. Etigunta, Alex J. Pfeil, Xiaohu Tang, Salvador Fabela, Laura M. Lashinger, Zhengrong Cui, Stephen D. Hursting. β Hydroxy β Methylbutyrate synergizes with cytotoxic chemotherapy and immunotherapy in a mouse model of pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3151.
Abstract 3156: Over-expression of CEACAM6 negatively modulates the tumor microenvironment in pancreatic cancer
Authors: Pandey Ritu , Chen Yuliang , Patil Mihir , Conway Patrick J. , Al-Khinji Aisha , Mahadevan Daruka
Published: July 2021

Background: CEACAM6 is a cell adhesion receptor of the Ig-family over-expressed exclusively in primates and human pancreatic ductal adenocarcinoma (PDA). It is present in all subtypes and is enriched classical activated stroma subtype. CEACAM6 has multifaceted roles in KRAS mutant PDA and is a poor prognostic maker (Pandey et al. Sci Rep 2019). We report here in PDA, high CEACAM6 modulates the tumor microenvironment with an immunosuppressive landscape.

Methods: RNA-Seq data of PDA patient samples from TCGA and GEO were obtained, normalized and log transformed for analysis. Tumor samples were stratified into cohorts of high and low CEACAM6. Pearson correlation coefficient is utilized to identify CEACAM6 correlated genes and enrichment of signaling pathways. The two cohorts were scored by ESTIMATE for stromal and immune population. Abundance of different cell populations were calculated utilizing MCPcounter and ConsensusTME methods. All analysis was performed using R (v 4.3).

Results: High CEACAM6 PDA samples had both stromal and immune components. Half of the high expressing samples had a higher immune score vs stromal score. The other half had both high or both low, immune and stromal score. Investigating the immune genes that are utilized for scoring, we found several immune genes differentially expressed in high CEACAM6 (up-regulated genes - ADAM8, IL2RG, LYZ, SELPLG involved in innate immunity) compared to low CEACAM6 samples. Genes under expressed in high CEACAM6 samples were CD27, NKG7, IL18RAP, LILRB1, KLKRK1 involved in promoting immune response and cell protection. Overall TCGA PDA samples have lower enrichment of immune cells compared to higher enrichment of fibroblasts, endothelial cells, macrophages, monocytes and dendritic cells. The NK cells are overall low in PDA but enriched in some lower CEACAM6 samples. Plasma cells, B-cells, CD8 cells, NK cells, Cytotoxic T-cells, Tγδ-cells have very poor representation in higher CEACAM6 samples but present in some low CEACAM6 expressing samples. Some of the TCGA PDA samples have T-reg cells but high CEACAM6 do not enrich of T-reg cells. High CEACAM6 samples are enriched for metabolic pathways.

Conclusions: CEACAM6 over-expressed in PDA have pathologic functions which include reshaping the stroma, suppressing host immunity and inhibiting apoptosis. An anti-CEACAM6 Mab increases apoptosis, necrosis and anti-tumor activity in PDA.

Citation Format: Ritu Pandey, Yuliang Chen, Mihir Patil, Patrick J. Conway, Aisha Al-Khinji, Daruka Mahadevan. Over-expression of CEACAM6 negatively modulates the tumor microenvironment in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3156.
Abstract 3178: A research tool for elucidating the interaction between human mucin 16 (CA125) and immune cells in ovarian cancer
Authors: Lakatos Kornel , Krauledat Petra , Serrano German Gonzalez , Hansen Peter , Cramer Daniel William , Elias Kevin , Patankar Manish , Hoballah Jawad
Published: July 2021

CA125 represents the circulating subunit of the large transmembrane glycoprotein MUC 16. Elevated levels of CA125 are suggestive of ovarian cancer and the magnitude of elevation correlates with burden of disease. A rising CA125 level after treatment suggests cancer recurrence. Ovarian tumor cells produce and shed MUC 16 which can bind to certain peripheral immune cells. To study this, current techniques use labelled-fluorescent markers that bind to CA125 on cells and flow cytometry to detect mean fluorescence. A limitation is that this technique may not be able to distinguish low-binding events from autofluorescence. Our new technology improves the resolution for studying CA125 bound to immune cells by employing antibody-labelled gold nanoparticles with far-greater sensitivity than fluorescent markers. The cells are then examined using darkfied microscopy rather than flow, which also permits automated slide scanning and focusing for 3-dimensional counting. This enables both simultaneous use of fluorescent markers to identify the immune cell populations as well as automated image analysis to quantify the amount of bound CA125 on individual cells. Preliminary data show that MUC 16 is preferentially bound to NK and T cells in women with ovarian cancer compared to binding in healthy controls. These insights may help elucidate the function of CA125 in ovarian cancer and improve our understanding of immune-tumor interactions.

Citation Format: Kornel Lakatos, Petra Krauledat, German Gonzalez Serrano, Peter Hansen, Daniel William Cramer, Kevin Elias, Manish Patankar, Jawad Hoballah. A research tool for elucidating the interaction between human mucin 16 (CA125) and immune cells in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3178.
Abstract 3183: Adrenergic signaling augments cytotoxic T-cell anti-tumor function in p53-deficient head and neck cancer
Authors: Silverman Deborah , Leahey Sara , Akhter Shamima , Xie Tongxin , Wang Yunfei , Doordjuin Elien , Calin George , Myers Jeffrey , Hwu Patrick
Published: July 2021

Few advances have been made against head and neck cancers, in part because treatment efforts have focused on the epithelial component of these tumors. However, therapies targeting the microenvironment, including immune checkpoint blockade (ICB), also remain ineffective in the majority of head and neck cancers. Like some other immunotherapy-nonresponsive tumors, head and neck cancers develop a complex microenvironment heavily infiltrated by nerve fibers arising from the peripheral nervous system. Studies in multiple models have shown that the ablation of different portions of the peripheral nervous system prevents cancer development and progression. We recently showed that existing nerves sprout and undergo autonomic reprogramming to an adrenergic-like phenotype as a result of orchestrated microRNA (miRNA) shuttling from cancer cells to neurons, resulting in activation of transcriptional programs that establish new neuronal identity. Given that activated T-cell subsets express high levels of adrenergic receptors, we hypothesized that adrenergic signaling may also influence tumor-infiltrating T-cell function. We used novel murine models of OCSCC to discern the effect of adrenergic nerve signaling on immune infiltration and immunotherapy response and correlated our findings with data from patient cohorts using bulk and single-cell RNA sequencing. We found that adrenergic signaling acutely enhanced cytotoxic T-cell activity in patient-derived tumor/T-cell pairs, and we elucidated a p53-dependent mechanism underlying this effect. Ultimately, these results provide a striking contrast to the known role of adrenergic signaling in cancer development: although nerve-cancer crosstalk drives cancer aggression and progression, tumor innervation may also better enable the host immune response. Therapeutic approaches targeting this critical component of tumor biology may be useful to improve patients' survival, treatment responses, and quality of life.

Citation Format: Deborah Silverman, Sara Leahey, Shamima Akhter, Tongxin Xie, Yunfei Wang, Elien Doordjuin, George Calin, Jeffrey Myers, Patrick Hwu. Adrenergic signaling augments cytotoxic T-cell anti-tumor function in p53-deficient head and neck cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3183.
Abstract 3190: PD-L1 protein expression in Middle Eastern breast cancer predicts favorable outcome in triple negative breast cancer
Authors: Al-Kuraya Khawla S. , Siraj Abdul K. , Parvathareddy Sandeep K. , Ahmed Saeeda , Benito Allianah , Iqbal Kaleem
Published: July 2021

Programmed cell death ligand 1 (PD-L1) has been shown to induce potent T-cell mediated anti-tumoral immunity. The significance of PD-L1 expression on prognosis of breast cancer remains controversial and its prevalence and prognostic value in breast cancer from Middle Eastern ethnicity is lacking. A total of 1003 unselected Middle Eastern breast cancers were analyzed for PD-L1 expression using immunohistochemistry. The association with clinico-pathological markers and impact on overall survival as well as recurrence free survival was determined. PD-L1 expression was seen in 32.8% (329/1003) cases. PD-L1 positivity was significantly associated with poor prognostic indicators such as younger patients (p = 0.0499), high grade tumors (p = 0025), ER negative (p < 0.0001) and PR negative (p = 0.0001) status as well as high Ki-67 index (p < 0.0001). We also found a significant association between PD-L1 expression and deficient mismatch repair protein expression (p = 0.0009). No association was found between PD-L1 expression and clinical outcome. However, on further subgroup analysis, PD-L1 expression was found to be an independent marker for favorable prognosis in triple negative breast cancer (TNBC) (Hazard ratio = 0.27; 95% confidence interval = 0.10 - 0.63; p = 0.0018). In conclusion, we demonstrated strong association between PD-L1 and MSI in Middle Eastern BC patients and showed that PD-L1 overexpression on tumor cells was an independent favorable prognostic factor in TNBCs from this ethnicity. Overall, these findings might help in the development of more appropriate treatment strategies for BC in Middle Eastern population.

Citation Format: Khawla S. Al-Kuraya, Abdul K. Siraj, Sandeep K. Parvathareddy, Saeeda Ahmed, Allianah Benito, Kaleem Iqbal. PD-L1 protein expression in Middle Eastern breast cancer predicts favorable outcome in triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3190.
Abstract 3192: PD-L1 is an independent prognostic marker in Middle Eastern PTC and its expression is upregulated by BRAFV600E mutation
Authors: Masoodi Tariq , Siraj Abdul K. , Parvathareddy Sandeep Kumar , Pratheeshkumar Poyil , Divya Sasidharan Padmaja , Begum Rafia , Diaz Roxanne , Saravanan Thangavel , Al-Obaisi Khadija , Siraj Sarah , Azam Saud , Qadri Zeeshan , Al-Kuraya Khawla S.
Published: July 2021

Background: PD-L1 inhibition is a promising therapeutic target whose efficacy has been demonstrated in several cancers. This study was conducted to evaluate the prognostic significance of PD-L1 expression in a large cohort of Middle Eastern PTC patients and to explore correlation of PD-L1 and BRAFV600E mutations in PTC tumors and cell lines.

Methods: Immunohistochemistry was performed to assess PD-L1 protein expression in PTC. We further conducted in vitro analysis to investigate the role of PD-L1 in regulating BRAFV600E in PTC cell lines.

Results: PD-L1 over-expression was noted in 32.4% (473/1458) of cases and significantly associated with adverse clinico-pathological parameters such as tall cell variant, extra-thyroidal extension and lymph node metastasis. Importantly, PD-L1 was found to be an independent poorer prognostic marker. We also found PD-L1 to be significantly associated with BRAF mutation and patients with co-existing PD-L1 over-expression and BRAF mutation had a poor disease-free survival compared to patients with BRAF mutation alone. In vitro analysis showed high expression of PD-L1 in BRAF mutated PTC cell lines compared to BRAF wild-type cell line. Inhibition of BRAF using vemurafenib induced PD-L1 expression in BRAF mutated cell lines without affecting cell growth. Knockdown of PD-L1 in BRAF mutated cell lines significantly decreased the cell growth and induced apoptosis.

Conclusion: Our data suggests that PD-L1 might represent a useful prognostic marker in Middle Eastern PTC and PD-L1 inhibition could be a potential therapeutic option for aggressive PTC cancers, such as tall cell variant, BRAF mutation positive patients that are unresponsive to standard treatment.

Citation Format: Tariq Masoodi, Abdul K. Siraj, Sandeep Kumar Parvathareddy, Poyil Pratheeshkumar, Sasidharan Padmaja Divya, Rafia Begum, Roxanne Diaz, Thangavel Saravanan, Khadija Al-Obaisi, Sarah Siraj, Saud Azam, Zeeshan Qadri, Khawla S. Al-Kuraya. PD-L1 is an independent prognostic marker in Middle Eastern PTC and its expression is upregulated by BRAFV600E mutation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3192.
Abstract 3193: Development and functional characterization of NC762, a novel therapeutic antibody targeting B7-H4, for the treatment of malignancies
Authors: Archer Kristina , Ceradoy Justine , Coupet Tiffany , Bosiacki Jason , Song Chang , Weiss Ido , Niu Sue , Obot Tete , Xu Jinglin , O'Neill Tom , Ferrell Alison , Abukharma Hasan , Cusumano Zac , Flies Dallas , Langermann Sol , Liu Linda N.
Published: July 2021

Human B7 homolog 4 (B7-H4) is a transmembrane protein in the B7 family of molecules that is expressed on tumor cells in the tumor microenvironment (TME). High protein expression on tumors and low expression on healthy tissue makes B7-H4 an attractive molecule for direct targeting with low off-target toxicity. NC762 is a humanized immunoglobulin gamma 1, kappa (IgG1κ) monoclonal antibody specific for human B7-H4 that is being developed for the treatment of cancer and demonstrates an excellent safety profile in IND-enabling studies. The IgG1κ region of NC762 contains three-point mutations (S239D/A330L/I332E a.k.a. DLE) which enhances binding to CD16a (FcγRIIIa) in order to increase antibody-dependent cellular cytotoxicity (ADCC) activity. NC762 binds to human B7-H4 on SKBR3 cells, a breast cancer cell line that endogenously expresses B7-H4 and induces in vitro ADCC activity. Despite the high homology, NC762 does not cross-react with rodent B7-H4, therefore, in vivo testing was performed in a humanized xenograft murine model in NSG mice, where NC762 restricted tumor growth in a dose-dependent manner. Depleting human natural killer (NK) cells prior to tumor inoculation reduced anti-B7-H4 parent antibody activity, supporting a role for NK-cell mediated ADCC in tumor growth retardation. However, in the absence of human PBMCs, NC762 also achieved significant anti-tumor activity. Furthermore, a mutant of NC762 with a low FcγR binding IgG1 domain showed similar activity as the DLE IgG1, demonstrating an ADCC-independent mechanism of tumor growth restriction mediated by B7-H4 mAb treatment. Together, these results indicate that even with the ADCC-enhanced IgG1κ region, NC762 is able to mediate ADCC-independent anti-tumor activity for the eradication of B7-H4 expressing tumors; and is expected to provide benefit in multiple oncology indications.

Citation Format: Kristina Archer, Justine Ceradoy, Tiffany Coupet, Jason Bosiacki, Chang Song, Ido Weiss, Sue Niu, Tete Obot, Jinglin Xu, Tom O'Neill, Alison Ferrell, Hasan Abukharma, Zac Cusumano, Dallas Flies, Sol Langermann, Linda N. Liu. Development and functional characterization of NC762, a novel therapeutic antibody targeting B7-H4, for the treatment of malignancies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3193.
Abstract 3199: In vitro killing assays for immuno oncology candidates
Authors: Janss Thibaut J. , Lamy Juliette , Arnold Johan , Boelen Ellen , Pattijn Sofie
Published: July 2021

The need to better understand the tumor microenvironment (TME) dictates the characterization of the cell types involved, the roles they play and how they respond to treatment. To develop better cancer immunotherapies, in vitro primary immune cell bioassays offer an early assessment of their effects on the various players of the TME. Cytotoxic cells are key players in the anti-tumor immune response. CD8+ T cells, also known as cytotoxic T lymphocytes, recognize and kill cells presenting antigens bound to MHC class I molecules, such as neoantigen present on the tumor cells. Activation of CD8+ T cells requires at least two signals from antigen presenting cells: binding of their T cell receptor (TCR) to the appropriate peptide presented on MHC I and co-stimulation by B7 binding to CD28. However, tumor environment developed different mechanisms to reduce this immune cell killing such as checkpoint inhibitors expression. In that case, the immune system will be dampened. A second type of cytotoxic cells, named Natural killer cells, are an innate immune cell type that plays an important role in anti-cancer immunity. NK cells target cells lacking MHC I expression, including cancer cells that have lost expression of MHC I. NK cells induce apoptosis in cancer cells via release of perforin and granzymes from granules, like CD8+ T cells. But here too, NK cells can be broken down by the tumor cells, notably by the PD1/PDL1 interaction. Increasing their efficacy as a therapeutic strategy has made the development of new therapeutics enhancing their anti-tumor response a priority. Those new therapeutics can have several shapes. The Antibody-dependent cell-mediated cytotoxicity (ADCC) activity of the IgG1 isotypes is one of the mechanisms that can be applied by immunoglobulin-based therapeutics. Another way to increase the killing of cancer cells is to target tumor antigens expressed by those cells using for example bispecific molecules that will enhance on the other side the T cell activation. Using primary immune cells, in vitro bioassays were developed to better screen the potential effect of new therapeutics on immune cell killing activity. Their ability to increase or induce a cytotoxic activity and facilitate the anti-tumor immune response can as a result be assessed early in the drug development process.

Citation Format: Thibaut J. Janss, Juliette Lamy, Johan Arnold, Ellen Boelen, Sofie Pattijn. In vitro killing assays for immuno oncology candidates [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3199.
Abstract 3201: Single-cell trajectory analysis reveals a melanoma-driven distinct hematopoietic response in murine spleen
Authors: Kara Nihan , Samusik Nikolay , Shi Xiaoshan , Lomas Chip , Widmann Stephanie , Tyznik Aaron J.
Published: July 2021

Growing evidence suggests that tumor progression can interfere with normal hematopoiesis and skew the host system to undergo myeloid biased changes. Hematopoietic stem and progenitor cells (HSPCs) are a rare population of precursor cells giving rise to progeny that replenish blood cells throughout life. HSPCs are mainly localized in the bone marrow niche in healthy adults, however pathophysiological conditions such as cancer can cause extramedullary hematopoiesis in organs like the spleen. In this study, we investigated tumor-driven phenotypic and molecular alterations in the HSPC compartment at a single-cell level during extramedullary hematopoiesis using single-cell multiomics. We used a B16-F10 melanoma mouse model to uncover the tumor-mediated changes. Mice with tumor-burden showed splenomegaly and alterations in the composition of the HSPC compartment in the spleen consistent with extramedullary hematopoiesis. We then isolated four HSPC populations, including Lin- Sca1+ cKit+ (LSK), common myeloid progenitor (CMP), granulocyte-macrophage progenitor (GMP) and megakaryocyte-erythrocyte progenitor (MEP) cells from the spleen and bone marrow of melanoma-burdened mice using the BD FACSMelody™ Cell Sorter. Next, we used an integrated workflow on the BD Rhapsody™ Single-Cell Analysis System for the downstream single-cell multiomics analysis. We utilized oligo-conjugated antibodies for NGS-based sample multiplexing and protein detection (BD® Single-Cell Multiplexing Kit, BD® AbSeq Assay), along with whole transcriptome analysis for a comprehensive analysis of mRNA expression. Data analysis using the X-shift algorithm paired with single-cell force-directed layout visualization enabled delineation of the developmental trajectories of HSPC populations and exploration of phenotypic and transcriptomic changes across different hematopoietic subpopulations and across distinct anatomic sites. The analysis also identified unique hematopoietic cell clusters present during melanoma-driven extramedullary hematopoiesis in the spleen. Overall, our findings highlight melanoma-driven perturbations in hematopoiesis by utilizing advanced single-cell multiomics technology.

Disclaimers: For Research Use Only. Not for use in diagnostic or therapeutic procedures. Class 1 Laser Product. BD-23277 (v1.0) 1120 BD, the BD Logo, FACSMelody and Rhapsody are trademarks of Becton, Dickinson and Company or its affiliates. © 2020 BD. All rights reserved.

Citation Format: Nihan Kara, Nikolay Samusik, Xiaoshan Shi, Chip Lomas, Stephanie Widmann, Aaron J. Tyznik. Single-cell trajectory analysis reveals a melanoma-driven distinct hematopoietic response in murine spleen [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3201.
Abstract 322: Protein expression shift and potential diagnostic markers through proteomics profiling of A549 lung cancer cells
Authors: Olajuyin Ayobami Matthew , Olajuyin Adefunke Kafayat , Wang Ziqi , Xu Zhiwei , Raji Hayatu M. , Okeke Malachy I. , Zhang Xiaoju
Published: July 2021

Highlights:-Adenocarcinoma lung cancer is common but difficult to diagnose. - Little is known for its proteome when methanol extract of Calotropis procera was used for the treatment groups. -In the treatment versus control groups, about 413 proteins were upregulated while about 626 proteins were downregulated. -Some of the proteins are potential diagnostic markers.

Objectives: Adenocarcinoma lung cancer is a common type of lung cancer but the therapeutic and diagnosis are problematic. We focused on the protein expression of Adenocarcinoma lung cancer using A549 cancer cells and unveiled novel diagnostic makers and mechanisms in which methanol extracts of Calotropis procera ameliorate the disease.

Methods: A549 lung cancer cell lines were obtained and we have 3 control groups and 3 treatment groups. The treatment groups were treated with 80 ug/ml of methanol extracts of Calotropis procera leaf and proteins were extracted for tandem mass tag mass spectrometry. Differential protein expression was performed between treatment groups and controls and the identified proteins were analyzed for pathway enrichment. Selected proteins were further validated in another set of samples using a more quantitative method.

Results: In the treatment versus control, about 413 proteins were upregulated while about 626 proteins were downregulated. Conclusion: This balanced proteomics method not only offers important understandings of protein expression and pathways but also discover potential diagnostic markers for adenocarcinoma lung cancer. It also reveals the molecular mechanisms in which the methanol extracts of Calotropis procera leaf prevent carcinogenesis.

Citation Format: Ayobami Matthew Olajuyin, Adefunke Kafayat Olajuyin, Ziqi Wang, Zhiwei Xu, Hayatu M. Raji, Malachy I. Okeke, Xiaoju Zhang. Protein expression shift and potential diagnostic markers through proteomics profiling of A549 lung cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 322.
Abstract 347: TET1 mutation as potential biomarker for immune checkpoint blockade in colorectal cancer
Authors: Yu Shaojun , Zhang Ding , Chen Shiqing
Published: July 2021

Background: TET1, a DNA demethylase, is encoded by TET1 gene. At present, TET1 mutation has been reported to be associated with enhanced tumor immunogenicity and activated anti-tumor immunity across multiple cancers. However, the clinical effect of TET1 mutation on immunotherapy in colorectal cancer is little known.

Methods: 876 patients with colorectal cancer were enrolled in this study. Genomic profiling of DNA was performed on formalin-fixed paraffin-embedded tumor samples by NGS with 733 cancer-related genes panel. The somatic and germline mutation data were both obtained. Benign and likely benign mutations were excluded from our analysis. Programmed death ligand-1(PD-L1) expression status was determined by immunohistochemistry, evaluated with the combined positive score (CPS) and tumor proportion score (TPS). The efficiency of immunotherapy was analyzed in a cohort of 85 patients with colorectal cancer from Morris et al.

Results: 40 of 876 patients (4.75%) harbored TET1 mutations, including nonsynonymous, frameshift and stopgain mutations. Totally, 66 TET1 single nucleotide mutation were identified. Median tumor mutation burden (TMB) value with TET1 mutations was 109.2 mutations/MB, significantly higher than 6.1 mutations/MB in TET1 wild-type group (p<0.001). Microsatellite instability-high (MSI-H) appeared more frequently in TET1 mutations group than TET1 wild-type group (50.0% vs 5.0%, p<0.001). For PD-L1 TPS, compare to TET1 wild type groups, the positive rate of PD-L1 expression in TET1 mutations was higher significantly (6.7% vs 2.7%, p<0.001). The same thing happened with PD-L1 CPS (50.0% vs 23.3%, p<0.001). Possibly, TET1 mutation was a potential marker for immunotherapy. The results were validated in a clinical cohort of immunotherapy, including 85 patients with colorectal cancer. The median overall survival was non-reached in 7 patients with TET1 mutations compare to 13 months in TET1 wild-type group (HR: 0.28, 95% CI 0.11-0.69, P=0.006).

Conclusions: TET1 mutation was strongly associated with higher TMB, MSI-H and PD-L1 expression and improved OS in patients receiving immune checkpoint blockade treatment, suggesting that TET1 mutation is a novel predictive biomarker for immunotherapy in colorectal cancer.

Citation Format: Shaojun Yu, Ding Zhang, Shiqing Chen. TET1 mutation as potential biomarker for immune checkpoint blockade in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 347.
Abstract 350: Co-expression and predictive value of HER family members for the response to therapy with cetuximab in patients with metastatic colorectal cancer
Authors: Khelwatty Said A. , Puvanenthiran Soozana , Essapen Sharadah , Bagwan Izhar , Seddon Alan M. , Modjtahedi Helmout
Published: July 2021

Background: Overexpression of the EGFR is common in patients with a wide range of cancers and the EGFR is an important target for therapy with monoclonal antibodies (mAbs) based drugs. Of these, cetuximab and panitumumab have been approved for the treatment of patients with metastatic colorectal cancer (mCRC). However, therapeutic benefits are seen in a subset of such patients and the duration of response can also be short. Indeed, there is currently no reliable predictive biomarkers for the selection of patients who benefit from therapy with the anti-EGFR antibodies. While KRAS mutation is currently used as an important negative predictive biomarker for the response to therapy with anti-EGFR mAbs, not all patients with wild type KRAS gain benefit from therapy with anti-EGFR mAbs and objective responses have been reported in mCRC patients with KRAS mutations.

Aim: The aim of this study was to investigate the predictive value of EGFR and its dimerization partner (i.e. all other members of the HER family) for the response to treatment with cetuximab in patients with wild-type KRAS status mCRC.

Methods: Using immunohistochemistry (IHC) we investigated the co-expression and predictive value of all four members of the HER family, for the response to cetuximab in 144 mCRC patients and in 21 paired primary tumours and their metastatic sites.

Results: Co-expression of all four members of the HER family were found in 10% of the cases examined. The expression of wtEGFR was an indicator of poorer overall survival and the membranous expression of HER2 and HER3 3+ intensity was associated with a shorter progression free survival (PFS). In contrast, the cytoplasmic expression of HER2 was associated with better PFS. In 48% and 71% of the cases examined, there were discordance in the expression of EGFR or one or more HER family members in paired primary and related metastatic tumours, respectively.

Conclusion: Our results implicate the importance of large prospective investigation of the expression level and predictive value of not only the therapeutic target (i.e. EGFR protein) but also its dimerization partners for the response to therapy with anti-EGFR mAbs and other forms of HER inhibitors in both the primary tumours and metastatic sites in colorectal cancer.

Citation Format: Said A. Khelwatty, Soozana Puvanenthiran, Sharadah Essapen, Izhar Bagwan, Alan M. Seddon, Helmout Modjtahedi. Co-expression and predictive value of HER family members for the response to therapy with cetuximab in patients with metastatic colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 350.
Abstract 36: ARAF mutations confer resistance to RAF dimer inhibitors in NRAS and BRAF mutant melanoma
Authors: Malek Shiva
Published: July 2021

Belvarafenib, a potent and selective RAF dimer inhibitor, exhibits clinical activity in BRAFV600E and NRAS mutant melanoma patients. Here, through generation of belvarafenib resistant NRAS mutant melanoma cells and through analysis of circulating tumor DNA from patients treated with belvarafenib, we identified novel recurrent mutations in ARAF, within the kinase domain. ARAF mutants conferred resistance to belvarafenib in both a dimer and kinase activity dependent manner. Belvarafenib induced ARAF mutant dimers, and dimers containing mutant ARAF were active in the presence of inhibitor. ARAF mutations may serve as a general resistance mechanism for RAF dimer inhibitors as the mutants exhibit reduced sensitivity to a panel of RAF dimer inhibitors. The combination of RAF plus MEK inhibition may be used to delay ARAF driven resistance and suggests a rational combination for clinical use. Taken together, our findings reveal specific and compensatory functions for the ARAF isoform and implicate ARAF mutations as a driver of resistance to RAF dimer inhibitors.

Citation Format: Shiva Malek. ARAF mutations confer resistance to RAF dimer inhibitors in NRAS and BRAF mutant melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 36.
Abstract 366: Identification of a clinically significant pan-cancer biomarker for homologous recombination deficiency
Authors: Takamatsu Shiro , Brown JB , Yamanoi Koji , Yamaguchi Ken , Hamanishi Junzo , Kobayashi Takashi , Mandai Masaki , Matsumura Noriomi
Published: July 2021

[Background] Genomic alterations in BRCA1/2 and the ‘genomic scar' signatures are associated with homologous recombination DNA repair deficiency (HRD) and serve as therapeutic biomarkers for sensitivity to platinum and PARP inhibitors in breast and ovarian cancers. However, the clinical significance of these biomarkers in other homologous recombination repair (HRR) pathway genes or in other cancer types is not fully understood.

[Method] From the TCGA and the AACR GENIE project databases, we obtained clinical and genomic data of patients with multiple solid cancers, evaluated both locus-specific LOH in HRR pathway gene mutations and two genomic scar signature scores (HRD score and mutational signature 3), identified potential HRD cases, and examined the efficacy of DNA-damaging drugs in these HRD cases.

[Results] Germline and somatic gene mutations in the HRR pathway genes with locus-specific LOH were significantly enriched in cases having high genomic scar scores, whereas those without the LOH were not enriched. Somatic mutations without the LOH were remarkably enriched in hypermutated cases and considered to be passenger mutations. The mutation rate and frequency with LOH in the HRR pathway genes varied widely by cancer type. When stratifying patients by gender and presence of TP53 mutation, the correlation between the two genomic scar scores and the resulting optimal cutoff values for biallelic HRR pathway gene alterations differed greatly. The potential HRD cases that were selected by the two genomic scar cutoffs and biallelic HRR pathway gene alterations showed significantly high gene expression-based HRD scores. In Cox proportional hazard multivariate analyses with covariates of age, stage, gender, HRD, and TP53 mutation, HRD was a good prognostic factor in the group with DNA-damaging agents, but a poor one in the group without the agents.

[Conclusion] Increased attention on locus-specific LOH in HRR pathway gene alterations and the differences in genomic scar signatures stratified by gender and TP53 mutation is newly warranted in order to properly assess HRD in a pan-cancer manner. The results in the present study proved the clinical significance of HRD across cancer types and may contribute to the development of personalized medicine based on HRD.

Citation Format: Shiro Takamatsu, JB Brown, Koji Yamanoi, Ken Yamaguchi, Junzo Hamanishi, Takashi Kobayashi, Masaki Mandai, Noriomi Matsumura. Identification of a clinically significant pan-cancer biomarker for homologous recombination deficiency [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 366.
Abstract 375: High tumor mutation burden predicts unfavorable clinical outcome in EGFR-mutated lung adenocarcinoma treated with targeted therapy
Authors: Lee Seung Hyeun , Sung Ji-Youn
Published: July 2021

Purpose: Although high tumor mutation burden (TMB) has shown association with benefit from immune checkpoint blockade therapies, its implications in lung cancer patients with driving mutations are still unclear. The objective of this study is to determine the association between TMB and treatment outcome in epidermal growth factor receptor (EGFR)-mutated lung cancer treated with tyrosine kinase inhibitors (TKIs).

Methods: We evaluated TMB in EGFR-mutant, lung adenocarcinoma patients who received first-line EGFR-TKIs. TMB was estimated by next-generation sequencing with a cancer gene panel (Oncomine™ Tumor Mutation Load Assay). We compared the response rate (RR), progression-free survival (PFS), overall survival (OS), and frequency of secondary T790M mutation according to the different TMB groups.

Results: Among the 131 patients who were treated with EGFR-TKIs, a total of 63 patients were eligible for the analysis. TMB was stratified by tertiles; low (≤2.13 mutations/Mb), intermediate (2.14-4.25 mutations/Mb), and high (>4.25 mutations/Mb). The TMB levels were not associated with any clinical parameters. The RR was significantly lower in the high TMB group than in the other groups (43.5% vs. 72.1% vs. 78.5%, all p = 0.01). In multivariate analysis, high TMB was independently associated with a shorter PFS in the overall population (hazard ratio [HR] = 2.64, p = 0.004), and associated with shorter OS in patients with exon 19 deletion (HR = 2.55, p = 0.041) compared with low TMB. The frequencies of secondary T790M mutation after TKI failure were not different among the different TMB groups.

Conclusion: High TMB was associated with unfavorable clinical outcome in patients with lung adenocarcinoma treated with EGFR-TKIs. Although further large-scaled studies are required, our data suggest that high TMB may be a predictive biomarker for adverse treatment outcomes and may constitute a distinct subgroup warranting tailored therapeutic approach in this clinical setting.

Citation Format: Seung Hyeun Lee, Ji-Youn Sung. High tumor mutation burden predicts unfavorable clinical outcome in EGFR-mutated lung adenocarcinoma treated with targeted therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 375.
Abstract 376: Prognostic significance of Glasgow prognostic score in NSCLC patients treated with immunotherapy after platinum-based cytotoxic chemotherapy
Authors: Kang Hye Seon , Kim In Kyoung , Kim Jin Woo
Published: July 2021

Introduction: The Glasgow prognostic score (GPS) reflects the host systemic inflammatory response and is validated prognostic factor in lung cancer. However, little is known about the prognostic role in non-small cell lung cancer (NSCLC) patients treated with immunotherapy after platinum-based cytotoxic chemotherapy.

Patient and Methods: This study used a lung cancer cohort of the Catholic Medical Center of Korea between January 2018 and June 2020. We included the patients who were diagnosed with unresectable advanced stage NSCLC or recurrent disease after pulmonary resection and had received at least one regimen of platinum-based chemotherapy before being administered immunotherapy. The patients with NSCLC treated with anti-PD1 or anti-PD-L1 (pembrolizumab, nivolumab, or atezolizumab) and assessed the prognostic value of the GPS. The GPS was calculated using C-reactive protein and albumin concentrations within 1 week before starting anti-PD1 or anti-PD-L1 treatment.

Results: A total of 78 patients with NSCLC treated with immunotherapy as 2nd or 3rd line therapy after platinum-based chemotherapy. Kaplan-Meier analyses revealed that higher GPS was significant predictors of shorter progression free survival (PFS) (Log rank < 0.001) and overall survival (OS) (Log rank < 0.001). In cox-regression multivariate analysis, the hazard ratios for disease progression were 2.146 (95% CI: 0.814-5.656; P = 0.123) for a GPS of 1 and 9.73 (95% CI: 2.931-32.298, P < 0.001) for a GPS of 2 relative to a GPS of 0. Older age (P = 0.033), lower PD-L1 expression (P = 0.036) and higher GPS (P = 0.007) were independently associated with shorter OS.

Conclusion: Higher GPS were identified as one of poor prognostic factor for OS and PFS in NSCLC patients received immunotherapy as 2nd or 3rd line therapy after platinum-based chemotherapy.

Citation Format: Hye Seon Kang, In Kyoung Kim, Jin Woo Kim. Prognostic significance of Glasgow prognostic score in NSCLC patients treated with immunotherapy after platinum-based cytotoxic chemotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 376.
Abstract 379: Effects on immunotherapy of EGFR mutation subtypes in non-squamous NSCLC
Authors: Tang Haibo , Li Xiaoning , Xie Wenzhuan , Huang Mengli
Published: July 2021

Background: Genetic alternations of Epidermal growth factor receptor (EGFR) in lung cancer had shown associated with response or resistance to anti-PD-1 therapy. However, there is a lack of understanding of the clinical significance of EGFR mutation subtypes in non-squamous non-small cell lung cancer (NSCLC).

Methods: We retrospectively analyzed 510 patients with non-squamous NSCLC treated with anti-PD-(L)1 monotherapy from four independent cohorts. The tumor tissues from the patients were subjected to MSK-IMPACT panel target sequencing or whole exome sequencing. The association between clinical features and specific EGFR alterations was analyzed.

Results: EGFR positivity was 13.33% (68/510), which is close to the proportion in CheckMate-057 (14.09%), while is higher than in KEYNOTE-010 (8.33%). Compared with 68 EGFR alteration lung cancers, DCR, ORR and durable clinical benefit (DCB) were better in patients with EGFR wide-type (Chi-square test, p = 0.0021, 0.0436 and 0.0014, respectively), while the PFS and OS in EGFR wide-type group was not significantly longer than the EGFR positive group (Log-rank test, p =0.1310 for PFS and 0.7730 for OS). Among EGFR positivity, the incidence of L858R only, T790M only, L858R/T790M mixed and others were 23.53%, 19.12%, 11.76% and 45.59%, respectively. A significant difference in survival among the four subtypes of EGFR positive group was observed, mutation of T790M only showed a worse PFS and OS probability compared to the other subtypes (p<0.001 and p = 0.002), whereas mutation of L858R only showed a poorer PFS (p=0.05).

Conclusion: Mutation of T790M in non-squamous NSCLC prospects worse survival than other EGFR mutations, while mutation of non-L858R and non-T790M subtypes was not a worse factor than EGFR wide-type group. The heterogeneity of EGFR alternations may be beneficial for establishing the uses of PD-(L)1 therapies for lung cancers.

Citation Format: Haibo Tang, Xiaoning Li, Wenzhuan Xie, Mengli Huang. Effects on immunotherapy of EGFR mutation subtypes in non-squamous NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 379.