bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1540: Accumulation of the long noncoding RNA Malat1 is a central driver and therapeutic target in metastatic lung cancer
Authors: Martínez-Terroba Elena , de Miguel Fernando J. , Robles-Oteiza Camila , Politi Katerina , Dimitrova Nadya
Published: June 2022

The identification and functional characterization of long noncoding RNAs (lncRNAs) have suggested a potential source of unexplored cancer therapy targets. The metastasis-associated lung adenocarcinoma transcript 1 (Malat1) was initially identified as a transcript significantly overexpressed in metastatic lung adenocarcinoma (LUAD) and strongly correlated with poor patient outcomes. Despite decades of effort to elucidate Malat1 functions and mechanisms of action in cancer, two key questions have remained unresolved: 1) Is Malat1 overexpression a driver of LUAD progression? and 2) What is the mechanism by which Malat1 overexpression promotes LUAD progression? We utilized CRISPR activation to model Malat1 overexpression in patient-derived LUAD cell lines and in the autochthonous KrasG12D/p53-deficient murine model of LUAD. We observed that Malat1 overexpression cooperates with p53 loss to promote LUAD progression and metastatic dissemination in vivo. Compared to controls, Malat1-overexpressing KrasG12D/p53-deficient tumors appeared poorly differentiated and were characterized by excessive stromal infiltration. Single-cell RNA sequencing of dissociated tumors revealed that Malat1 overexpression triggers pleiotropic immune and stromal responses that favor a metastasis-permissive tumor microenvironment through the secretion of cytokines and metalloproteinases and the recruitment of CAFs and tumor-associated macrophages. Consistent with the in vivo findings, we observed that Malat1 overexpression in vitro does not affect the proliferation but promotes the migration of tumor cells and cancer-associated fibroblasts (CAFs) through secreted factors, including inflammatory cytokines. Importantly, targeting the stabilizing ENE domain of overexpressed Malat1 by two independent approaches in vitro - direct mutagenesis and non-degrading antisense oligonucleotide (ASO) - led to Malat1 destabilization and rescued the increased cellular migration. Similarly, cytokine-targeted blocking antibodies reversed the effects of Malat1 overexpression. In sum, our results represent the first successful efforts to model the increased Malat1 levels observed in human LUAD and to dissect the pro-metastatic molecular consequences of overexpressed Malat1. Our findings indicate that Malat1 is both necessary and sufficient to rewire metastatic pathways and our preliminary data support further efforts to therapeutically target Malat1 and its downstream mediators in metastatic LUAD.

Citation Format: Elena Martínez-Terroba, Fernando J. de Miguel, Camila Robles-Oteiza, Katerina Politi, Nadya Dimitrova. Accumulation of the long noncoding RNA Malat1 is a central driver and therapeutic target in metastatic lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1540.
Abstract 1542: MALAT1 ablation dismantles homologous recombination repair machinery and sensitizes castrate resistant prostate cancer cells to PARP inhibitor
Authors: Yadav Anjali , Biswas Tanay , Praveen Ayush , Ateeq Bushra
Published: June 2022

Poly (ADP-ribose) polymerase (PARP) inhibitors have emerged as a promising target of intervention for metastatic castration-resistant prostate cancer (mCRPC), as approximately one-third of mCRPC patients harbor mutations in the genes associated with the homologous recombination (HR) pathway. Nonetheless, most patients inevitably develop resistance to PARP inhibition (PARPi), due to the induction of reversion mutations in the HR pathway. These mutations restore the function of HR genes and revert to the HR-proficient stage, which in turn decreases the vulnerability of cancer cells to PARP inhibitors. Here, we report that the clinical utility of PARPi could be efficaciously extended by targeting Metastasis associated lung adenocarcinoma transcript (MALAT1), a long non-coding RNA (lncRNA) often elevated in advanced-stage prostate cancer (PCa). In support of this, we show that mCRPC patients exhibit higher expression of genes involved in HR, which positively correlate with MALAT1 levels. Furthermore, RNA interference (RNAi)-mediated depletion of MALAT1 in CRPC cells perturb the expression of key HR genes, namely BRCA1/2, RAD51, EXO1, CHEK1/2, subsequently resulting in HR deficiency. This, in turn, escalates accumulation of DNA lesions in the MALAT1 ablated cells as noticed by abundance in γH2AX, a marker for DNA damage. As a consequence, MALAT1 ablated cells instigates G1/S phase arrest to fetch additional time to repair the damaged DNA or to induce apoptosis. Furthermore, we also provide evidence that the HR deficiency induced by MALAT1 depletion phenocopies “BRCAness” and exhibits synergy with clinically approved DNA repair inhibitors such as Olaparib. In particular, co-inhibition of MALAT1 and PARP1 exhibits a significant anti-proliferative effect reduced clonogenic survival and delays the resolution of γH2AX foci in CRPC cell lines. Taken together, our results establish that MALAT1 plays a central role in regulating the DNA damage response and provides a mechanistic rationale for dual targeting of MALAT1 and PARP in mCRPC patients.

Citation Format: Anjali Yadav, Tanay Biswas, Ayush Praveen, Bushra Ateeq. MALAT1 ablation dismantles homologous recombination repair machinery and sensitizes castrate resistant prostate cancer cells to PARP inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1542.
Abstract 1578: Interaction of E-cadherin and EGFR in the collective cell migration of colon cancer cells
Authors: Winarso Viriya Adhiguna , Lu Yi-Wen , Weng Yu-Cheng , Hou Xiang-Ling , Koo Hui Min , Li Yu-n , Chao Wei-Ting
Published: June 2022

Collective cell migration is one of the most important cellular events where a cluster of cells moves together in a coordinated way. Many studies have found this phenomenon as the hallmark of cancer progression and metastasis. In epithelial single cell migration, downregulation of adherens junction (AJ) molecule E-cadherin occurs at the initial step known as the Epithelial to Mesenchymal Transition (EMT). However, in collective cell migration, E-cadherin junction is maintained to provide adhesion between cells. Moreover, it also mediates the cell-cell intercommunication that is critical, for instance, to alter the group polarization through actin cytoskeleton remodeling. Interestingly, Epidermal Growth Factor Receptor (EGFR) was found to colocalize with E-cadherin in the cell-cell contact region, implicating that E-cadherin and EGFR might interact in the cell junction of collective cell migration. To address this, a human colorectal cancer cell line exhibiting epithelial phenotype HT-29 was used. The cell density and calcium presence were modified to infer the effect of differential expression and stability of E-cadherin on the cellular signaling of proliferation and migration. The results showed that stable and enhanced expression of E-cadherin downregulates EGFR expression and reduces its activity. Moreover, the Src phosphorylation increased when the cell-cell contact was strengthened via calcium addition and decreased when the cell-cell contact was diminished via calcium chelation. Overall, this indicates that E-cadherin expression at the cell-cell junction and contact stability determine the EGFR expression, activity, and Src phosphorylation. With the goal of deciphering the relationship of E-cadherin and EGFR in cell contact, proliferation, and migration, we intend to finally implement the knowledge as a strategy or approach in promoting alternative methods to inhibit metastasis.

Citation Format: Viriya Adhiguna Winarso, Yi-Wen Lu, Yu-Cheng Weng, Xiang-Ling Hou, Hui Min Koo, Yu-n Li, Wei-Ting Chao. Interaction of E-cadherin and EGFR in the collective cell migration of colon cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1578.
Abstract 1600: Dose-dependent effects of interferon-gamma treatment on pancreatic ductal adenocarcinoma cell states
Authors: Liberman Deborah A. , Barkley Dalia , Yanai Itai
Published: June 2022

Interferon-gamma (IFNγ) is a canonical proinflammatory cytokine that has well-studied antitumor functions such as the upregulation of MHC Class I (MHC-I), increased Th1 development, macrophage activity, innate immune cell recruitment, and in tumor cells increased apoptosis, decreased angiogenesis and proliferation. However, IFNγ is also important in limiting tissue destruction due to inflammation, taking on an immunotolerant role and allowing cancer cells to evade immune surveillance. We previously identified an interferon response cell state across diverse cancer types, as defined by the relative expression of IFIT1, HLA-A, HLA-DRA, STAT1, and IRF1. The interferon response cell state is heterogeneously expressed in cancer cells and appears to function both by cancer cell extrinsic and intrinsic mechanisms. Several clinical trials have tested the efficacy of IFNγ as a therapeutic for immune system stimulation in patients with melanoma. However, these have failed to detect effective outcomes, and have even noted some pro-growth actions, perhaps consistent with IFNγ’s immunotolerant role in maintaining tissue homeostasis. We are interested in identifying cell populations that are vulnerable to the antitumor functions of IFNγ, as well as cell populations that may be susceptible to IFNγ’s pro-tumor actions. To test this, we treated a pancreatic ductal adenocarcinoma cell line with three doses of IFNγ in vitro every day for five days and analyzed expression of proteins associated with IFNγ signaling by flow cytometry each day. Distinct doses of IFNγ have been shown to affect varying degrees of antitumor and pro-tumor responses. For example, growth inhibition has been seen upon treatment with high dose IFNγ, while treatment with low dose IFNγ is consistent with increased metastatic capability such as increased resistance to cytotoxic lymphocytes and natural killer cells despite the upregulation of MHC-I. We thus treated cells with three doses of IFNγ: low, normal, and high. Preliminary results have shown a dose dependent effect of IFNγ including the upregulation of MHC-I and PDL1. Ongoing work includes identifying changes in cell states using single-cell RNA-sequencing to uncover how cancer cells respond to varying intensities of IFNγ exposure. Additionally, we are analyzing how release of IFNγ from cancer cells changes following treatment of IFNγ at different doses. This work will provide insight into the mechanisms by which cancer cells evade immune surveillance and how cell states can describe a tumor’s vulnerability to IFNγ’s antitumor and pro-tumor effects.

Citation Format: Deborah A. Liberman, Dalia Barkley, Itai Yanai. Dose-dependent effects of interferon-gamma treatment on pancreatic ductal adenocarcinoma cell states [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1600.
Abstract 1624: Immune cell killing assay for measuring the effects of immunomodulating agents in lung cancer cells in vitro
Authors: Mäki-Jouppila Jenni H. , Rissanen Jukka P. , Fagerlund Katja M.
Published: June 2022

Direct effects of cancer therapeutics on immune cell activity can be challenging to determine on system level in vivo. Therefore, it is important to develop and discover novel ways to study the effects of immune cells on cancer cells using also in vitro assays. Our aim was to validate an in vitro immune cell killing assay where activated peripheral blood mononuclear cells (PBMCs) target lung cancer cells and induce apoptosis.

A549 human lung carcinoma cells (ATCC) were used as target cells in the co-culture and were plated into 96-well plates. 24 hours later, human PBMCs (Lonza) were added to the culture as effector cells and activated with anti-CD3 and IL2. Apoptosis was detected using IncuCyte Annexin V reagent, which was added to the cells at the same time with immune cell activators. Cell proliferation was monitored with a live cell imager IncuCyte S3 (Sartorius) for three to four days before terminating the cultures. Cell viability was determined by CellTiter Glo (Promega) at the end of the study.

Treatment of cells with anti-CD3 was able to activate immune cells alone but the effects were observed with lower test concentrations in combination with IL-2. Therefore, IL-2 potentiates the effect of anti-CD3 on immune cell activation and decreases cancer cell viability in co-culture.

The immune cell killing assay based on live cell imaging can be used for studying the effects of immunomodulating agents on cancer cell viability and to evaluate the activity of immune cells in the presence of therapeutics.

Citation Format: Jenni H. Mäki-Jouppila, Jukka P. Rissanen, Katja M. Fagerlund. Immune cell killing assay for measuring the effects of immunomodulating agents in lung cancer cells in vitro [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1624.
Abstract 1631: Anti-cancer effects of vorinostat on 3D cultured pancreatic cancer cells
Authors: Emamzaden Fatemeh Nouri , Word Beverly , Hammons George , Lyn-Cook Beverly D.
Published: June 2022

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers, often having a low survival rate mainly due to late diagnosis and limited therapies because of drug resistance. Treatment results shown in cell models used in studying various drugs often fail in clinical trials. The lack of translation of the effects observed in vitro to patients continues to halt progress. Genetic mutations are known in PDAC; however, recent studies have suggested the role of epigenetics in its initiation and progression to a malignant phenotype. Histone deacetylase (HDAC) proteins are highly expressed in a variety of cancers, including pancreatic cancer. Recent studies have shown that targeting these proteins through the use of HDAC inhibitors may be an effective therapy for pancreatic cancer. Using a 3D cultured pancreatic cell model, anti-cancer effects by vorinostat, a HDAC inhibitor, were investigated. Although both pancreatic cell lines, PANC-1 and Mia, contain mutated K-ras and P53, a differential effect in response to vorinostat was observed in expression of cancer genes using a Human Pancreatic Adenocarcinoma TagMan Array 96 (Applied Biosystems). In the PANC-1 cell model, vorinostat decreased expression of STAT6, which is known to play a role in migration and invasion. It also decreased significantly expression of insulin growth factor 1 (IGF1), IL-6, EGF, VEGFA and TGFB1. CCND2 and CCND2 were also downregulated. In contrast, Mia cell model showed decreased expression of EGF and IGF1, but no change with the other genes. These data suggest that vorinostat has effects on several potential pathways in pancreatic cancer possibly through epigenetic mechanisms that affect certain genes involved in progression and invasion. Currently, this drug in combination with other drugs is being investigated for pancreatic cancer.

Citation Format: Fatemeh Nouri Emamzaden, Beverly Word, George Hammons, Beverly D. Lyn-Cook. Anti-cancer effects of vorinostat on 3D cultured pancreatic cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1631.
Abstract 1642: Evaluating in vivo efficacy of anti-VSIG4 antibodies in humanized B-hVSIG4 mice
Authors: Lyu Ruili , Shang Chengzhang , Kokabee Leila , Nie Yanhui , Yang Yi
Published: June 2022

V-set and immunoglobulin domain containing 4 (VSIG4) protein is an immune checkpoint molecule that is related to the B7 family of immune regulatory proteins. B7 family members have been implicated in both positive and negative regulation of T cell activation, particularly VSIG4 is a known negative regulator of T cell proliferation and IL-2 production. Pathological consequences of VSIG4 expression are associated with inflammatory disorders, lung cancer development, high-grade glioma, and multiple myeloma, suggesting VSIG4 is a potential target for drug development. Therefore, Biocytogen developed humanized VSIG4 mice (B-hVSIG4 mice) by replacing the murine Vsig4 gene with the human VSIG4 gene through homologous recombination. Flow cytometry analysis showed human VSIG4 was detectable in peritoneal exudate macrophages (PEMs) in homozygous B-hVSIG4 mice compared to wild type mice. Additionally, percentages of T cells, B cells, NK cells, DCs, granulocytes, monocytes, macrophages, CD8+ T cells, CD4+ T cells and Tregs in homozygous B-hVSIG4 mice were similar to those in wild type mice, demonstrating that introduction of human VSIG4 in place of its mouse counterpart does not change the overall development, differentiation or distribution of these cell types. For in vivo efficacy evaluation of an anti-human VSIG4 antibody (Ab-A, synthesized in house) using B-hVSIG4 mice, mouse colon adenocarcinoma MC38 cells were subcutaneously implanted into homozygous B-hVSIG4 mice (female, 7-week-old, n=5). Mice were grouped and treated when the tumor volume reached approximately 100 mm3. Results showed that an anti-hVSIG4 antibody was efficient at inhibiting tumor growth in B-hVSIG4 mice. In conclusion, B-hVSIG4 mice are as a promising model for preclinical in vivo studies to assess pharmacodynamic of anti-hVSIG4 antibodies. Keywords: VSIG4, humanized mice, immune checkpoint molecule

Citation Format: Ruili Lyu, Chengzhang Shang, Leila Kokabee, Yanhui Nie, Yi Yang. Evaluating in vivo efficacy of anti-VSIG4 antibodies in humanized B-hVSIG4 mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1642.
Abstract 1644: Generation and validation of humanized GARP mice for testing novel anti-human GARP antibodies
Authors: Lyu Ruili , Li Huilin , Frame Jenna , Shen Yuelei
Published: June 2022

GARP is a type I transmembrane protein expressed on regulatory T cells (Tregs) and platelets. GARP expression has been shown to play an important role in the modulation of latent TGF-β1 activity by regulatory T lymphocytes (Tregs), and thus, GARP has emerged as an important regulator of peripheral immune tolerance. While GARP expression is critical for homeostasis, increased GARP expression on Tregs is associated with elevated immunosuppression in the context of tumor biology. Because of these immunomodulatory roles, targeting GARP in the context of oncologic or other diseases is of significant interest. Thus, the development of mouse models for safety and efficacy evaluation of GARP-targeted therapeutics prior to clinical trials is needed. To facilitate assessment of novel anti-human GARP antibodies, Biocytogen has generated a humanized GARP mouse model for in vitro functional validation and in vivo efficacy evaluation assays. Using CRISPR-based gene editing technology, the exons of the mouse Garp gene that encode the extracellular domain were replaced by the human GARP counterpart. mRNA Expression of the human GARP extracellular domain was confirmed in splenocytes isolated from homozygous humanized mice. Expression of human GARP was readily detected in FoxP3+ Treg cells isolated from the spleen of humanized GARP mice as assessed by flow cytometry. Importantly, the frequency of FoxP3+ Tregs and other mature leukocyte subpopulations, including T and B lymphocytes, Natural killer cells, dendritic cells, granulocytes, and monocytes/macrophages, was also similar in the blood and spleen of humanized GARP mice compared to wild-type mice. Subsequent in vivo efficacy assays using the MC38 tumor model demonstrated efficacy of the anti-human GARP antibody in slowing tumor growth in humanized GARP mice. Furthermore, combination of anti-human GARP and anti-mouse PD-1/PD-L1 blockade resulted in a significantly greater tumor inhibition than monotherapies. In summary, GARP humanized mice are a powerful platform for preclinical evaluation of anti-human GARP antibodies for cancer treatment.

Citation Format: Ruili Lyu, Huilin Li, Jenna Frame, Yuelei Shen. Generation and validation of humanized GARP mice for testing novel anti-human GARP antibodies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1644.
Abstract 1648: Evaluation of therapeutic drug candidates using a novel humanized B-hCSF1/hCSF1R mouse model
Authors: Lyu Ruili , Zhang Shujin , Soto Rebecca , Zhao Suman , Liu Chonghui , Yu Luke (Zhaoxue)
Published: June 2022

An often rate-limiting step for cancer immunotherapy is drug development. The generation of humanized mouse models can be used to assess the clinical utility of new drugs earlier in the discovery process. Tumor-associated macrophages (TAMs) represent an emerging therapeutic target, specifically high expression of the colony-stimulating factor 1 receptor (CSF1R) protein on tumors and TAMs facilitate a tumor permissive environment. Previous reports have demonstrated that blockade of the CSF1/CSF1R pathway reduced and reprogrammed intra-tumoral CSF1R+ TAMs, altogether alleviating immune suppression. To evaluate preclinical efficacy and toxicity of CSF1R targeted therapeutic candidates, Biocytogen has successfully generated a double humanized B-hCSF1/hCSF1R mouse model. The full-length encoding sequence of mouse Csf1 and the extracellular domain of Csf1r were replaced by human CSF1 and CSF1R genes, respectively. Evaluation of CSF1 and CSF1R protein expression in homozygous B-hCSF1/hCSF1R mice (H/H) was confirmed by ELISA and flow cytometry analysis. We next performed an in vivo efficacy screening study using genetically modified hCSF1 or wild-type MC38 cells inoculated into B-hCSF1/hCSF1R (H/H) mice and observed that anti-human CSF1R antibodies or combination therapy of anti-mouse PD-L1 and anti-human CSF1R antibodies were effective in controlling tumor growth. Our combined studies illustrate that Biocytogen’s novel B-hCSF1/hCSF1R mice offer a promising model for in vivo validation of CSF1 and CSF1R antibody therapeutics.

Citation Format: Ruili Lyu, Shujin Zhang, Rebecca Soto, Suman Zhao, Chonghui Liu, Luke (Zhaoxue) Yu. Evaluation of therapeutic drug candidates using a novel humanized B-hCSF1/hCSF1R mouse model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1648.
Abstract 165: Characterizing CLK3 in the colonic epithelium
Authors: Cabel Carly , Thorne Curtis
Published: June 2022

The purpose of this study is to determine the function of CLK3 in the colonic epithelium and colon cancer.

The intestinal epithelium undergoes constant renewal and is tightly regulated. Disruption of its homeostasis can result in the development of colorectal cancer (CRC). Glycogen synthase kinase 3 (GSK3) is a kinase that plays a central role in multiple signaling pathways including cellular proliferation. Our lab previously performed an siRNA screen to investigate hundreds of kinases to identify candidates which could interact with GSK3. Among the over 500 kinases that were tested, a protein kinase called CDC-like kinase 3 (CLK3) was identified as the strongest interaction with GSK3. We found that inhibition of GSK3 and CLK3 synergized to kill cancer cells. This led us to focus on CLK3 as a potential therapeutic target kinase in colon cancer.

CLK3 is part of a four-member family of CDC-like kinases involved in pre-mRNA splicing. They are dual-specificity kinases, meaning they can target both tyrosine and serine/threonine substrates. CLK3 is also a poor prognostic marker in colon cancer, with high expression of CLK3 leading to poor outcome of disease. In human colonic epithelial cells, we show that loss of CLK3 dampens Wnt response. This data, along with the strong genetic interaction with GSK3, suggests a role of CLK3 in the Wnt pathway. Because Wnt signaling is higher in the stem cell region of the crypt, and CLK3 shows a role in Wnt activity in human colonic epithelial cells, we hypothesize that CLK3 regulates mRNA splicing to promote Wnt signaling and stemness in colonic progenitor cells and carcinogenesis.

We are currently performing transcriptomics and phosphoproteomics in CLK3 CRISPR knockout cells to identify an mRNA-splicing and phosphorylation signature. We have found that the induction of Wnt-related pathway genes are lost in CLK3 knockout cells. Additionally, we are categorizing CLK3-dependent pre-mRNAs splicing events into functional groups controlling cell fate and validating in patient-derived normal and tumor organoid models. This work highlights CLK3 as a critical regulator of gut stem cells and potentially colorectal cancer stem cells.

Citation Format: Carly Cabel, Curtis Thorne. Characterizing CLK3 in the colonic epithelium [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 165.
Abstract 1665: Humanized mice for preclinical efficacy evaluation of drugs targeting CD73
Authors: Liu Xing , Ju Cunxiang , Sun Hongyan , Zhao Jing , Gao Xiang , Li Zhiying
Published: June 2022

CD73 expressed in Treg surface facilities the hydrolysis of AMP into adenosine which inhibits the activation, expansion and homing of T effector cells.Therapeutic efficiency of CD73 mAb had been studied in C57BL/6 and BALB/C mice xenografted with several murine cancer cell lines. However, for human CD73 mAb with no binding to murine CD73, CD73 humanized mouse models are essential for medicinal development.We established CD73 humanized mouse model (hCD73) on C57BL/6 and BALB/c background in which the mCD73 was replaced with hCD73. Similar expression level of hCD73 in peripheral blood was observed compared with mCD73 in wildtype mice. Besides, the proportion of T, B and NK cells in spleen of hCD73 mice were also analogy to that in wildtype mice. After treating with anti-hCD73 antibody, tumor growth was inhibited in our CT-26-hCD73 tumor model.Furthermore, hPD1/hPDL1/hCD73 mouse models were also established on C57BL/6 and BALB/c backgrounds based on hCD73 mice. However, a better anti-tumor effect of anti-CD73 antibody was found on BALB/c-hPD1/hPDL1/hCD73 bearing CT26 than that on B6-hPD1/hPDL1/hCD73 bearing MC38. Besides, Combination of anti-CD73 and anti-PD1 significant elevated anti-tumor effect comparing with anti-CD73 and anti-PD1 mono therapy in BALB/c-hPD1/hPDL1/hCD73 mice bearing CT26-hPDL1hCD73.In conclusion, BALB/c-hPD1/hPDL1/hCD73 is a well efficient mouse model for therapeutic effect study of anti-CD73 and relevant combination therapy.

Citation Format: Xing Liu, Cunxiang Ju, Hongyan Sun, Jing Zhao, Xiang Gao, Zhiying Li. Humanized mice for preclinical efficacy evaluation of drugs targeting CD73 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1665.
Abstract 167: Pre-clinical characterization of a novel series of FGFR2 selective inhibitors with potency against clinically relevant mutations
Authors: Fischer John , Bouhana Karyn , Chicarelli Mark J. , Dahlke Josh , Fell Brad , Fulton Jennifer , Guarnieri Anna , Jalluri Ravi , Johnson Amber , Mclean Brent , Mejia Max , Rieger Rob , Robinson John , Rodriguez Mareli , Sullivan Francis , Wang Yang , Winski Shannon , Zhou Yeyun
Published: June 2022

The fibroblast growth factor receptor (FGFR) family of protein tyrosine kinases plays a role in many physiological processes including angiogenesis and wound healing. FGFR mutations, fusions, rearrangements, and amplifications have been linked to the pathogenesis of multiple tumor types. For example, approximately 10-15% of patients with intrahepatic cholangiocarcinoma have FGFR2 fusions, and amplification of FGFR2 is prevalent in gastric cancer and implicated in tumor growth. Approved FGFR inhibitors produce responses in patients that harbor FGFR genetic alterations but show reduced activity against key mutations (e.g., V565, N550, etc.). These pan-FGFR inhibitors also frequently show dose-limiting toxicities including hyperphosphatemia which can be linked to the inhibition of FGFR1. To address these issues, a novel series of FGFR inhibitors that are potent and selective reversible inhibitors of FGFR2 with selectivity over FGFR1 have been identified. This series also maintains activity against clinically relevant mutations in the FGFR2 protein. Inhibition of cellular FGFR phosphorylation was measured using engineered cell lines to demonstrate that the inhibitors target both wild-type FGFR2 and activating FGFR2 mutations while sparing FGFR1. In addition, CDX tumor models showed in vivo target engagement and FGFR isoform selectivity. Herein, the in vitro and in vivo characterization of a representative selective reversible FGFR2 inhibitor is described.

Citation Format: John Fischer, Karyn Bouhana, Mark J. Chicarelli, Josh Dahlke, Brad Fell, Jennifer Fulton, Anna Guarnieri, Ravi Jalluri, Amber Johnson, Brent Mclean, Max Mejia, Rob Rieger, John Robinson, Mareli Rodriguez, Francis Sullivan, Yang Wang, Shannon Winski, Yeyun Zhou. Pre-clinical characterization of a novel series of FGFR2 selective inhibitors with potency against clinically relevant mutations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 167.
Abstract 1691: The landscape of intra-tumor heterogeneity (ITH) and its association with actionable mutation and blood tumor mutation burden (bTMB) in patients with non-small cell lung cancer (NSCLC) assessed by circulating tumor DNA (ctDNA)
Authors: Kim Leeseul , Lee Yeun Ho , Kim Hye Sung , Choi Horyun , Kim Jinah , Chae Young Kwang
Published: June 2022

Background: ITH, which is caused by subpopulations of cells with distinct genomic alterations within the same tumor is known to have an impact on the biological profile and natural history of the tumor. However, assessment of ITH is mostly done tissue-based. Herein, we investigated the correlation of ITH assessed by ctDNA with bTMB and the presence of actionable mutations.

Method: Blood samples were collected from consecutive 53 patients with NSCLC seen at Northwestern Memorial Hospital. For each sample, SNV, indel, and CNV results generated by sequencing of all plasma ctDNA (Guardant360) were formatted for PyClone. ITH scores were calculated using the number of clusters identified in the sample divided by the number of variants detected in the sample.

Result: bTMB was evaluable for 22 patients (41%). 7 patients (13%) had actionable mutations associated with FDA-approved therapies. Cluster number was positively correlated with bTMB (Pearson r 0.69, p-value 0.0004, CI 0.38 to 0.86) and was negatively correlated with the presence of actionable mutations (median 2 vs 5, p-value 0.01). ITH scores were not significantly correlated with bTMB (Pearson r 0.03, p-value 0.89, CI -0.39 to 0.44).

Conclusion: Higher cluster number was inversely correlated with the presence of actionable mutations and positively correlated with bTMB. Our study illustrates the importance of utilizing liquid biopsy to assess ITH with tissue-based ITH in clinics for a better understanding of the biological profile of tumor.

Citation Format: Leeseul Kim, Yeun Ho Lee, Hye Sung Kim, Horyun Choi, Jinah Kim, Young Kwang Chae. The landscape of intra-tumor heterogeneity (ITH) and its association with actionable mutation and blood tumor mutation burden (bTMB) in patients with non-small cell lung cancer (NSCLC) assessed by circulating tumor DNA (ctDNA) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1691.
Abstract 1701: Single-cell multi-omic profiling and clonal tracing of the human gynecological tumor microenvironment
Authors: Liu Vincent , Sandor Katalin , Daniel Bence , Berthoin Lionel , Sabri Shan , Panagiotopoulou Sofia , Yin Yajie , Hiam-Galvez Kamir , Sit Rene , Fan Zi , Galvin Brendan , Khan Omar , Bezman Natalie , Grogan Jane , Howitt Brooke , Zheng Grace , Lareau Caleb , Satpathy Ansuman
Published: June 2022

Tumor initiation and progression are the result of a close interplay between malignant cells and immune cells. While single-cell genomics has enabled the molecular dissection of the cellular heterogeneity underlying complex cancers, many facets of tumor microenvironments remain unresolved. In particular, the ontogeny of myeloid cells have not been fully understood in human tumors, although the source and expansion of macrophages have been suggested to underlie tumor development and metastasis in mice. In an effort to comprehensively characterize these tumor microenvironments, we profiled endometrial, ovarian, and metastatic tumors from 16 patients using CITE-seq (an atlas of 258,810 cells) and mitochondrial single-cell ATAC-seq (mtscATAC-seq; consisting of 173,820 cells). Our multi-omic analyses reveal the heterogeneity of tumor-infiltrating immune cells at clonal, epigenetic, transcriptomic, and proteomic levels. Notably, all tumor types harbored diverse exhausted T cell subsets, including terminal and precursor exhausted T cells, and gamma/delta T cells. Clonal somatic variant analysis between tumor tissues and peripheral blood mononuclear cells identified blood-derived monocytes, rather than embryonically derived cells, as the origin of macrophages within gynecological tumors. Further, our analyses show that macrophages expand minimally within the tumor microenvironment, suggesting that they may be continuously replenished by blood, an observation that may facilitate new approaches for cancer immunotherapy. In total, our dataset represents the first multi-modal single cell atlas of gynecologic tumors and reveals distinct features of T cell heterogeneity and myeloid expansions in complex tumor microenvironments.

Citation Format: Vincent Liu, Katalin Sandor, Bence Daniel, Lionel Berthoin, Shan Sabri, Sofia Panagiotopoulou, Yajie Yin, Kamir Hiam-Galvez, Rene Sit, Zi Fan, Brendan Galvin, Omar Khan, Natalie Bezman, Jane Grogan, Brooke Howitt, Grace Zheng, Caleb Lareau, Ansuman Satpathy. Single-cell multi-omic profiling and clonal tracing of the human gynecological tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1701.
Abstract 1762: A novel antibody drug conjugate linker enabling production of ADCs with high drug to antibody ratios and fast payload release for improved efficacy
Authors: Luong Noemie , Bellocq Nathalie , Marx Leo , Pantin Mathilde , Tchumi Benjamin , Roubaudi-Fraschini Marie-Claude , Riff Camille , Mutter Manfred , Attinger Antoine
Published: June 2022

Antibody drug conjugates (ADCs) represent an attractive approach to bring very cytotoxic drugs into tumors while avoiding systemic toxicities. Currently, most commercially available ADCs come with toxicities associated with the nonspecific release of the payload in the circulation. We have developed a novel drug multi-linker technology that enables the generation of stable and potent ADCs. It is selectively recognized and cleaved by the carboxydipeptidase activity of Cathepsin B that has been shown to be overexpressed in cancer cells. The technology allows for a fast and specific cleavage, which results in efficient drug release in the tumor. Different payloads can be attached to antibodies, and the drug to antibody ratio (DAR) can be modulated up to DAR16, resulting in customed cytotoxic activity. Here we characterize the properties of the multi-linker using different payloads, different DARs, and trastuzumab as the targeting antibody. Trastuzumab based multi-linker ADCs were generated and tested in vitro and in vivo. The linker-payload kinetic release and the cytotoxicity of these ADCs were assessed in vitro. Pharmacokinetic (PK) and pharmacodynamic properties of selected multi-linker ADCs were then investigated in relevant in vivo models and compared to trastuzumab emtansine (T-DM1) and trastuzumab deruxtecan (T-Dxd). Trastuzumab based multi-linker ADCs were shown to efficiently target Her2 expressing cells. In vitro, increasing the DAR had a positive effect on cytotoxicity. In a breast cancer model expressing low levels of Her2, a trastuzumab based multi-linker ADC with mertansine (DM1) at DAR8 showed superior activity, including complete tumor regressions, as compared to T-DM1 and T-Dxd and was well tolerated. In addition, the trastuzumab based multi-linker ADCs showed total antibody PK profiles comparable to T-DM1 and good stability in plasma. In conclusion, these results demonstrate that this novel linker technology can be used to produce stable and high DAR ADCs with improved efficacy.

Citation Format: Noemie Luong, Nathalie Bellocq, Leo Marx, Mathilde Pantin, Benjamin Tchumi, Marie-Claude Roubaudi-Fraschini, Camille Riff, Manfred Mutter, Antoine Attinger. A novel antibody drug conjugate linker enabling production of ADCs with high drug to antibody ratios and fast payload release for improved efficacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1762.
Abstract 1797: Bcl-2 up-regulation mediates taxane resistance downstream of APC loss
Authors: Maloney Sara , Zabala Sarah , Richmond Grace , Prosperi Jenifer R.
Published: June 2022

Patients with triple negative breast cancer (TNBC) are treated with traditional chemotherapy, such as paclitaxel (PTX). Despite the efficacy of taxanes, many tumors will recur due to drug resistance. Therefore, the need to understand the mechanism behind drug resistance is critical to improve patient outcome and survival. Our lab was the first to show that loss of the Adenomatous Polyposis Coli (APC) tumor suppressor caused resistance to PTX in the MDA-MB-157 human TNBC cell line. In the absence of APC, apoptosis induction was decreased, as measured through cleaved caspase 3 and annexin/PI staining. To understand the molecular mechanisms behind APC-mediated PTX response, we analyzed the BCL-2 family of proteins and found a robust increase of the pro-survival family member, Bcl-2. The BH3 mimetic, ABT-199 (Venetoclax), which specifically targets Bcl-2, has been used as a single or combination therapy in multiple hematologic malignancies. In addition, ABT-199 has shown promise in multiple subtypes of breast cancer. Therefore, we used ABT-199 in combination with PTX to address the hypothesis that APC-induced Bcl-2 increase is responsible for PTX resistance. Combination treatment in three CRISPR-mediated APC knockout TNBC cell lines (MDA-MB-157, MDA-MB-231, and SUM159) caused changes in cell proliferation and apoptosis. Combined, these data suggest restored sensitivity to PTX using ABT-199. Our studies are the first to show that Bcl-2 inhibition can restore PTX-sensitivity in APC mutant breast cancer cells. These studies are critical to advance better treatment regimens in patients with TNBC.

Citation Format: Sara Maloney, Sarah Zabala, Grace Richmond, Jenifer R. Prosperi. Bcl-2 up-regulation mediates taxane resistance downstream of APC loss [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1797.
Abstract 1915: Addressing cancer health disparities among US minority populations with patient-derived xenografts
Authors: Sanderson Brian J. , Lott Paul , Srivastava Anuj ,   PDXNet Consortium, Chuang Jeffrey H. , Carvajal-Carmona Luis G.
Published: June 2022

Despite the tremendous advances in cancer prevention and treatment strategies that have been achieved since the war on cancer plan in the 1970’s, it is clear that not all communities are equally benefiting from such advances and that minority populations nearly always have worse cancer health outcomes. These cancer health disparities are the result of complex interactions between social determinants of health, population-specific cultural behaviors and lifestyles, and unique biological differences that associate with ancestry. Two major limitations to the advancement of precision medicine in minority groups include the lack of: i) race/ethnic appropriate genomic data, which will facilitate identifying priority targets for therapeutic development, and ii) patient-derived models, such as patient-derived xenografts (PDXs) that recapitulate the unique interactions of germline and somatic genetics that exist in minority patients. In this presentation we describe the development of an analytical pipeline to quantify continental genetic ancestry that we have developed for the Cancer Genomics Cloud as part of the PDX Development and Trials Center Research Network (PDXnet). This pipeline leverages a novel reference panel for the program SNPweights, which includes a greater representation of genomic data from Native and Latin American populations than previously published reference panels, and thus provides improved assignment of American continental ancestry. We use this pipeline to characterize the diversity of the PDX models in PDXnet, which represent a manifold increase in models with African and American continental genetic ancestry compared with those available in other public resources. Finally, we present power analyses to demonstrate that additional PDX models derived from patients from minority populations are critically needed to effectively address health equity goals in the US.

Citation Format: Brian J. Sanderson, Paul Lott, Anuj Srivastava, PDXNet Consortium, Jeffrey H. Chuang, Luis G. Carvajal-Carmona. Addressing cancer health disparities among US minority populations with patient-derived xenografts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1915.
Abstract 1916: Using systems medicine for comprehensive metabolic profiling of tumors: how tumor metabolism shapes prognosis and response to chemotherapy
Authors: Cook Daniel , Cole John A. , Carrigan Patricia , Peterson Joseph R. ,   The SimBiosys Team
Published: June 2022

Background: Metabolic disfunction is one of the hallmarks of cancer, and metabolism-based therapies are a key class of chemotherapeutics. Despite this, there has previously been no systematic way to identify metabolism-based targets for therapeutic intervention. This is especially concerning when considering that targeting tumor metabolism has been shown to increase the efficacy of standard chemotherapy in pre-clinical studies. We therefore developed a systems medicine approach to interrogate tumor metabolism systematically, identify the effects of metabolic heterogeneity on overall survival prognosis, and gain insights into new metabolism-associated targets or therapies to complement standard of care for cancer patients. Method: Our systems medicine framework statistically integrates metabolic network modeling with RNA-sequencing data (RNA-seq). This allows us to integrate molecular data about individual tumors (from RNA-seq) with a curated knowledge base of how these molecules interact within a patient’s tumor (using metabolic network models). This results in a mathematical description, or model, of a specific tumor’s metabolism that is able to be interrogated (i.e., it is high-dimensional like RNA-seq data) and able to be simulated (i.e., we can “drug” the model and investigate downstream effects). Results: We applied this systems medicine approach across a range of tumor types, including lung adenocarcinoma, pancreatic adenocarcinoma, metastatic melanoma, clear cell renal cell carcinoma, and salivary cystic adenoid carcinoma. Within each cancer type, we found significant patient-to-patient metabolic heterogeneity. Across cancer types, a consistent pattern that emerged was that the metabolic systems with the most patient-to-patient heterogeneity were also the systems most associated with patient overall survival. This result suggests that metabolic variability is a key driver of tumor response to therapy and overall patient survival. Furthermore, targeting metabolic pathways in a patient-specific manner could enhance chemotherapy in tumors with metabolic profiles associated with poor prognosis. Conclusion: This study shows that the specific metabolic profiles of tumors are key drivers of overall survival. We therefore believe that metabolic tumor characterization and intervention could be a useful strategy to enhance chemotherapy efficacy and overall survival in patients across tumor types.

Citation Format: Daniel Cook, John A. Cole, Patricia Carrigan, Joseph R. Peterson, The SimBiosys Team. Using systems medicine for comprehensive metabolic profiling of tumors: how tumor metabolism shapes prognosis and response to chemotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1916.
Abstract 1954: Circulating tumor cell kinetics and rare cell morphology monitor disease progression in patients with metastatic colorectal cancer
Authors: Narayan Sachin
Published: June 2022

As the world’s third most common cancer and second leading cause of oncology-related deaths, colorectal cancer (CRC) poses a variety of clinical challenges. CRC is marked by its extensive solid tumor heterogeneity and the difficulties in longitudinally monitoring a patient’s disease status. Being a minimally invasive prognostic tool, the liquid biopsy has immense potential to improve current clinical practice by providing real-time personalized information over the course of treatment. Utilizing the High-Definition Single Cell Assay (HDSCA) workflow, we employed a “no cell left behind approach” to analyze peripheral blood samples from metastatic CRC (mCRC) patients at various timepoints during treatment. We highlight the circulating rare cell population present in the liquid biopsy of mCRC patients, showing the spatial and temporal heterogeneities of CTCs. We observed a significant association between CTC subcategorization based on morphological differences and survival, emphasizing the heterogeneity within the CTC classification. Moreover, considering the time of sample collection in relation to the clinical scenario is critical in determining the prognostic relevance of static cellular enumerations as well as the change in cell populations over time. Through the integration of cellular morphology in a time-based analysis, the liquid biopsy can provide insight into the pathophysiology of mCRC by highlighting the complexity of the disease to monitor treatment response.

Citation Format: Sachin Narayan. Circulating tumor cell kinetics and rare cell morphology monitor disease progression in patients with metastatic colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1954.