bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2218: The molecular mechanism underlying KRAS regulation on STK31 expression in pancreatic ductal adenocarcinoma
Authors: Liu Yuting , Tang Shing Chun , Chen Yangchao
Published: April 2023

KRAS is frequently mutated in pancreatic ductal adenocarcinoma (PDAC). However, targeting mutant KRAS is still challenging. Kinase inhibitors are ideal targeted therapeutics for mutant KRAS-driven cancer. In our previous study, an esiRNA screening was prepared to identify some vital kinases that took part in KRAS mutant-driven pancreatic cancer. The STK31 was identified as a potential therapeutic target in KRAS mutant PDAC. This study aimed to investigate the underlying mechanism of STK31 in KRAS mutant PDAC as well as its therapeutic potential. The results demonstrated that STK31 was upregulated in KRAS mutant PDAC patients with poor survival. STK31 was also highly expressed in PDAC cell lines with KRAS mutant background. Inhibition of STK31 in KRAS mutant cell lines significantly reduced PDAC cell proliferation, colony formation, and migration. Gain and loss of function experiments revealed that STK31 was a downstream target of KRAS in PDAC. Pharmacological inhibition of several signaling pathways involved in KRAS showed MAPK/ERK signaling involved in STK31 regulation. Then, further mechanistic study validated that c-Jun, regulated by KRAS/MAPK/ERK signaling, directly regulated the transcription level of STK31 by binding to its promoter region. In summary, our results show that STK31 promotes PDAC cell proliferation under the control of the KRAS/MAPK/ERK/c-Jun transcriptional axis in PDAC. STK31 may act as a potential target in PDAC treatment.

Citation Format: Yuting Liu, Shing Chun Tang, Yangchao Chen. The molecular mechanism underlying KRAS regulation on STK31 expression in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2218.
Abstract 2226: Modeling the role of Tet-deficiency and restoration in Bcl2-driven B cell lymphoma progression
Authors: Lam Minh Quang , Cimmino Luisa
Published: April 2023

Loss of function in DNA methylation regulators such as the Ten-eleven translocation (TET1-3) proteins, have been implicated in the progression of lymphoma. TET enzymes constitute a family of iron and αKG-dependent dioxygenases that oxidize 5-methylcytosine in DNA to promote demethylation and the reversal of gene silencing. The TET enzymes require vitamin C for their optimal catalytic activity, however, many patients with hematological malignancies are vitamin C deficient. Enhancing and restoring TET function with high-dose vitamin C has been shown to block the progression of solid tumors and blood malignancies. A combination of genetic and epigenetic diversity in lymphoma can result in the aberrant overexpression of oncogenes, loss of tumor suppressor function, and genomic instability. Our research models Bcl2-driven B cell lymphoma with Tet-deficiency using transgenic RNAi for the knockdown of Tet1 or Tet2. In vitro, our results have shown that this model shows increased self-renewal capacity of early B cell progenitors and precursors that can be suppressed by genetic restoration of Tet expression or upon treatment with vitamin C. In vivo, the B cell compartment of mice with Bcl2 overexpression and Tet1 or Tet2 deficiency exhibit a block in B cell maturation during a long period of pre-malignancy, and a decreased frequency of follicular B cells in the spleen from 3 months of age. Our aim is to determine mechanistically the effects of Tet-deficiency on B cell lymphoma progression and whether vitamin C can restore Tet activity to suppress lymphoma progression.

Citation Format: Minh Quang Lam, Luisa Cimmino. Modeling the role of Tet-deficiency and restoration in Bcl2-driven B cell lymphoma progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2226.
Abstract 2233: Clinical validation of a methylation array-based diagnostic assay for improved classification of CNS tumors
Authors: Soucy Melissa , Renzette Nick , Zhuo Xinming , Nuni Prasanti , Herlth Kristi , Kelly Kevin , Omerza Gregory , Li Lei
Published: April 2023

Current cIMPACT-NOW and WHO guidelines state that the accurate diagnosis of many central nervous system (CNS) tumors must not only incorporate traditional histological findings but also include additional molecular data derived from sequence and methylation profiling techniques. To address this clinical need, our group has adapted and clinically validated the CNS tumor classifier that was pioneered and developed by the German Cancer Research Network (DKFZ) and University Hospital Heidelberg for both adult and pediatric CNS tumors. The classifier analyzes whole genome methylation data derived from the Illumina EPIC array system with a machine learning algorithm trained on a large reference set to provide a tumor family and class (a more granular classification than family) along with calibrated scores to aid in determining confidence in the assigned classifications. Around this core algorithm, we have generated a computational and reporting pipeline to generate a mid-throughput clinical diagnostic assay. We have validated this assay using a combination of adult and pediatric CNS tumor samples and have established reliable run-level and sample-level quality control metrics with empirically defined thresholds. With these thresholds, approximately 90% of samples with a neoplastic content of 70% or greater were “classifiable”, such that a result could be returned by the CNS tumor diagnostic assay. The performance of the assay was exceptional, with a sensitivity, specificity and accuracy each greater than 98% as determined with a validation sample set of 105 specimens. Further, 4 discordant calls between the original diagnosis and the output of the new methylation-based diagnostic were identified and underwent further histopathologic review. This additional analysis led to the re-classification of 2 specimens due to molecular analysis, highlighting the clinical utility of the assay. The new assay is currently offered for clinical testing under the name JAX OncoMethyl ArrayTM and addresses a clear need in the field of CNS tumor diagnostics.

Citation Format: Melissa Soucy, Nick Renzette, Xinming Zhuo, Prasanti Nuni, Kristi Herlth, Kevin Kelly, Gregory Omerza, Lei Li. Clinical validation of a methylation array-based diagnostic assay for improved classification of CNS tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2233.
Abstract 2238: RAS activity-related features of metastatic pancreatic cancer
Authors: Mitsunaga Shuichi , Masuda Mari , Yoshimatsu Hiroki , Kamei Yoichi , Sasaki Mitsuhito , Watanabe Kazuo , Ikeda Masafumi
Published: April 2023

Background: RAS activity might play crucial roles in the aggravation of KRAS-mutated pancreatic cancer (PC), including systemic inflammatory reactions, tumor-driven inflammation, chemo-refractoriness and anorexia. We aimed to identify RAS activity-related features in PC patients with liver metastasis (LM), to create a murine xenograft model of human PC cells representative of RAS-driven aggravation.

Methods: RAS activities in 77 patients who underwent needle biopsy for LM of treatment-naïve PC were measured using an RAS signature score from a comprehensive cDNA microarray dataset of LM samples. Changes in the sum of the longest diameter of the target lesion in the first month of chemotherapy (ΔSLD), laboratory data and anorexia score at baseline were evaluated. Gene modules to be correlated with RAS activity and aggravation were evaluated using weighted co-expression network analysis (WGCNA). Human PC cells were classified as high and low activity according to in silico analysis of the DepMap CCLE expression dataset for profiling RAS activity and the best hub gene expression in the gene module. Subcutaneous xenograft models of human PC cells with high and low activity in severe combined immunodeficiency mice were compared to explore the RAS-related phenotype.

Results: KRAS-mutated LM was detected in 70 of the 77 PC patients (90.9%). RAS signatures were up-regulated in patients with KRAS-mutated LM (P < 0.01), high serum level of C-reactive protein (P = 0.01) as a systemic inflammatory marker, high expression of IL-6 mRNA by the tumor (P = 0.12) as an index of tumor-driven inflammation, and high level of phospho-Erk (P = 0.03). In 58 PC patients receiving first-line chemotherapy for KRAS-mutated LM, ΔSLD in patients with a high RAS signature was increased as compared to those in patients with a low RAS signature (P = 0.045). WGCNA methods revealed that the green module highly correlated with RAS activity and aggravation. Based on in silico analysis with classification of the human PC cell lines CFPAC-1 and SUIT-2 into high and low activity cells, respectively, CFPAC-1 tumors showed desmoplasia and high expression of tumor IL-6 and liver SAA1 as compared to SUIT-2 tumors (P = 0.046 and P = 0.016, respectively).

Conclusions: Tumor RAS activity is related to inflammation-related aggravation in PC patients. RAS-related inflammation can be accurately presented in a murine model of human PC cells, which might be an effective tool to test the inhibition of RAS-driven inflammation.

Citation Format: Shuichi Mitsunaga, Mari Masuda, Hiroki Yoshimatsu, Yoichi Kamei, Mitsuhito Sasaki, Kazuo Watanabe, Masafumi Ikeda. RAS activity-related features of metastatic pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2238.
Abstract 2239: Investigation of the molecular biological significance of YAP/TAZ expression in PNEN
Authors: Shiraishi Yuta , Hayashi Hiromitsu , Ogawa Daisuke , Yumoto Shinsei , Sato Hiroki , Matsumoto Takashi , Takematsu Toru , Maruno Masataka , Tsukamoto Masayo , Miyata Tatsunori , Nakagawa Shigeki , Mima Kosuke , Nitta Hidetoshi , Baba Hideo
Published: April 2023

[Background] Neuroendocrine neoplasms (NEN) developing in the digestive organs are relatively rare tumors, with 3 to 5 new cases per 100,000 population annually, most of which occur in the pancreas and digestive tract. In pancreatic neuroendocrine neoplasms (PNEN), the risk of metastasis and progression varies depending on the grade of the tumor, even when the tumor size is small. In clinical practice, it is often difficult to determine the indication for surgery or multidisciplinary treatment with chemotherapy. The mechanisms of tumor formation and progression in PNEN remain unknown. YAP/TAZ have been known to play an important role in pancreatic development. Here, we explore the biological and clinical significance of YAP/TAZ over-expression in PNEN.

[Methods] Immunostaining for YAP/TAZ was performed using 92 resected specimens in PNEN patients, and the association between YAP/TAZ overexpression and clinicopathological factors was elucidated. Furthermore, we generated a PNEN cell line with YAP and TAZ over-expression using QGP-1, and examined the intracellular signals regulated by YAP/TAZ expression.

[Results and Discussion] The overexpression of YAP, TAZ, and YAP/TAZ were observed in 55, 55, and 32patients, respectively. High YAP expression was significantly associated with positive lymphovascular invasion (p=0.04). YAP/TAZ-high group displayed significantly worse Overall survival (p=0.0023) and recurrence-free survival (p=0.007) compared to those in others. By Western-blotting, YAP/TAZ overexpression in the QGP-1 cell line enhanced phosphorylation of Akt and Erk1/2.

[Conclusions] High expression of YAP/TAZ in PNEN was implicated in poor prognosis, and was also associated with Akt, Erk1/2, and mTOR.

Citation Format: Yuta Shiraishi, Hiromitsu Hayashi, Daisuke Ogawa, Shinsei Yumoto, Hiroki Sato, Takashi Matsumoto, Toru Takematsu, Masataka Maruno, Masayo Tsukamoto, Tatsunori Miyata, Shigeki Nakagawa, Kosuke Mima, Hidetoshi Nitta, Hideo Baba. Investigation of the molecular biological significance of YAP/TAZ expression in PNEN [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2239.
Abstract 2246: Prevalence of mycobacterium tuberculosis among PAP smear samples of cervical cancer suspects
Authors: Awoke Wondimu Ashagre
Published: April 2023

Introduction: Cervical cancer is one of the most common cancers in women around the world. The Pap smear test is the most common method of detecting abnormal cells that may develop into cancer. In addition, we want to know how common mycobacterium tuberculosis is among those who provide Pap smear samples.

Objectives: The purpose of this study is to examine the knowledge, attitudes, and practices of working women in Addis Abeba, Ethiopia, regarding cervical cancer and Pap smear tests, as well as the associations between knowledge, attitudes, and practices and socio-demographic factors. In addition, we will investigate the prevalence of mycobacterium tuberculosis from smears among those who provide Pap smear samples.

Methods: This will be a cross-sectional questionnaire-based study of 100 Ethiopian females' attending at Arsho medical laboratories from September to December 2022, knowledge, attitudes, and practices, as well as the prevalence of tuberculosis. All women between the ages of 20 and 65 who wanted to be diagnosed with cervical cancer were eligible.

Expected outcome: To teach and recall common cervical cancer symptoms such as menstrual bleeding, the appropriate age for Pap smear tests, and the interval between tests. Examine their use of hormonal contraception and their attitude toward Pap smear testing over the last five years. It will also be useful to know the prevalence of cervical tuberculosis.

Citation Format: Wondimu Ashagre Awoke. Prevalence of mycobacterium tuberculosis among PAP smear samples of cervical cancer suspects [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2246.
Abstract 226: Targeted transcriptome sequencing enables exponential scaling of combinatorial barcoding in AML samples
Authors: Kim Grace , Papalexi Efthymia , Matulich Peter , Koehler Ryan , Schroeder Sarah , Tran Vuong , Roco Charles , Rosenberg Alexander
Published: April 2023

In the thirteen years since its inception, single-cell RNA-sequencing (scRNA-seq) has rapidly spread across multiple fields of research, leading to many new discoveries. As technologies have matured, the number of cells that can be processed in a single experiment has seen exponential growth with workflows now assaying up to one million cells in an individual experiment.

While high throughput sequencing methods have facilitated the discovery and characterization of various cell types, sequencing costs can be prohibitively high for routine use. Many applications of scRNA-seq are focused on cell type identification, gene regulatory networks, or biomarker discovery. These applications often do not require surveying the entire transcriptome, but rather require the interrogation of specific sets of well-characterized genes. In these cases, sequencing the entire transcriptome may be adding unnecessary project costs. To increase throughput and minimize sequencing costs, the development of a targeted gene enrichment method is required.

Here we extend our whole transcriptome split-pool combinatorial barcoding technology, to enable enrichment of a subset of genes in the final single cell sequencing libraries. To illustrate the power of our technology, we enriched a whole transcriptome library of human bone marrow mononuclear cells (BMMCs) from four acute myeloid leukemia (AML) donors and four control donors. From a library of 27,000 cells, we used our immune gene panel to enrich 1,000 genes representing canonical immune cell markers and pathways. Our method increased the percent of reads on target from as low as 1.6% in the whole transcriptome libraries to 75% in the targeted libraries. Furthermore, despite a nearly ten-fold reduction in sequencing reads between unenriched and enriched libraries, the resulting clustering yielded very high concordance of cell type identities and preserved AML-specific signatures such as FLT3, MKI67, and CD19.

Overall, we demonstrate our modular enrichment strategy preserves biological structure of the data and allows for deep characterization of gene signatures in health and disease. We envision our approach will enable researchers to simultaneously reduce sequencing costs while drastically scaling up the number of cells and samples across experiments.

Citation Format: Grace Kim, Efthymia Papalexi, Peter Matulich, Ryan Koehler, Sarah Schroeder, Vuong Tran, Charles Roco, Alexander Rosenberg. Targeted transcriptome sequencing enables exponential scaling of combinatorial barcoding in AML samples [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 226.
Abstract 2264: Role of first-line immune checkpoint inhibitors monotherapy for oncogene-driven non-small cell lung cancer
Authors: Chen Ying-Yin , Lin Chen-Wen , Hsein Yenh-Chen , Chen Chung-Yu
Published: April 2023

Though improvement of overall survival in non-small cell lung cancer (NSCLC) patients treated with immune checkpoint inhibitors (ICIs) was observed, their efficacy varies greatly among different immune and molecular profiles in tumors. Particularly, the clinical significance of ICIs for oncogene-driven NSCLC has been controversial. In this study, after excluding NSCLC patients harboring driven oncogenes as EGFR, ALK and ROS1 mutation, twenty NSCLC patents with higher Programmed cell death 1 ligand 1 (PD-L1) expression (PD-L1 ≧ 50%, immunohistochemical stained by SP263 or 22C3) were administered with Pembrolizumab alone as first-line immunotherapy. NSCLC patients with more higher PDL1 expression (PD-L1 ≧ 80%) had longer progression-free survival (PFS) PD-L1 ≧ 80% v.s. PD-L1 < 80%, PFS, median, 11.2 v.s 7.0 months, hazard ratio [HR]: 0.52, p = 0.03). 10 of them had other driven oncogenes (5 for KRAS, 3 for MET, 1 for BRAF and 1 for NRAS) were detected by next-generation sequencing (NGS) (Illumina iSeq 100 Sequencing System; AmpliSeq for Illumina Focus Panel). Patients harboring driven oncogene had shorter PFS by first-line immunotherapy as Pembrolizumab (With driven oncogene v.s. without, PFS, median, 4.2 v.s.10.8 months, HR: 2.76, p = 0.001). In conclusion, NGS analysis was recommended even the NSCLC patients with higher PD-L1 expression before immunnotherapy. First-line ICIs monotherapy should be cautious for oncogene-driven NSCLC patients.

Citation Format: Ying-Yin Chen, Chen-Wen Lin, Yenh-Chen Hsein, Chung-Yu Chen. Role of first-line immune checkpoint inhibitors monotherapy for oncogene-driven non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2264.
Abstract 227: In vivo functional genomics screen to uncover tumor intrinsic negative regulators of NK cell-mediated tumor cell killing
Authors: Duval Kayla , Krall Elsa , Piccioni Federica , Monberg Maria , Spatz Marianne , Ehrenberger Tobias , Follmer Nicole
Published: April 2023

Natural killer cells are innate immune cells whose activity is gated by activating and inhibitory receptors that bind ligands on potential target cells. NK cell gene signatures are associated with improved survival in several indications, and NK cell therapies have shown some encouraging results in the clinic. These data highlight the relevance of this cell type in anti-tumor immunity and has given rise to interest in amplifying NK responses to cancer as a complementary therapeutic strategy to current T cell based immunotherapies. The goal of this project is to identify tumor cell intrinsic negative regulators of NK cell-mediated tumor cell killing through in vivo CRISPR/Cas9 screening, with identification of both PD-1 blockade-dependent and -independent regulators. The syngeneic B16F10 model in C57Bl6 mice was used with or without NK depletion and with or without anti-PD1 treatment, with a custom library of 900 genes split into four subpools (one guide per gene per subpool). B16F10 cells were transduced in vitro and inoculated into mice ten days after spinfection. On day 14 after inoculation, tumors were harvested and subsequently the gDNA was isolated and sequenced. QC analysis indicated good guide recovery and screen performance. There were several hits from this screen that were depleted in the NK+ context but not in the NK depleted context, indicating their potential role in regulating NK mediated cell killing. Of the hits, several expected genes scored including B2M which is a known negative regulator of NK cell killing. Validation efforts include confirmation of dropout in other models as well as in vitro co-culture assays to assess enhanced NK killing of single gene KO cells. The hits identified in the screen will increase our understanding of anti-tumor immunity and serve as potential novel Oncology targets.

Citation Format: Kayla Duval, Elsa Krall, Federica Piccioni, Maria Monberg, Marianne Spatz, Tobias Ehrenberger, Nicole Follmer. In vivo functional genomics screen to uncover tumor intrinsic negative regulators of NK cell-mediated tumor cell killing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 227.
Abstract 2322: Precision immuno-oncology (IO) ISH/IF multiplex panel: Spatial detection of cytokines and IO biomarkers
Authors: Dennison Brenna , Allen Stephanie , Patel Bhavika , Stapleton Jacob , Archuleta Roni , Rath Mary Lou , Mehra Navi , Talwalkar Sameer S.
Published: April 2023

Cytokines have a large impact on the tumor microenvironment by affecting cell growth, survival, inflammation, and differentiation. Understanding the spatial biology within the tumor microenvironment, including the cytokine organization within specific cell types, can lead to better treatment plans for patients. We developed a hybrid multiplex panel, using RNA in-situ hybridization (ISH) and immunofluorescence (IF) to investigate the spatial biology of the tumor microenvironment. This panel also enabled us to simultaneously detect the RNA of cytokines and protein of specific immuno-oncology (IO) biomarkers. This allows us to visualize spatially, not only the organization of cell types but also to identify the cellular sources of specific cytokines. In this study, we analyzed the tumor microenvironment of samples from non-small cell lung cancer (NSCLC) and breast cancer in two panels for cytokine RNAs and IO protein biomarkers. Panel 1 included TNFa and IL-2 cytokines combined with the CD68 and CD163 IO protein biomarkers. Panel 2 consisted of IL-6 and IL-2 cytokines with the PAX5 and CD3 IO protein biomarkers. Staining was completed on the Leica Bond RX autostainer, and slides were scanned using the PhenoImager™ Fusion. Visiopharm® software was used to analyze the tumor microenvironment of tissue samples and localize cytokines in specific cell types. CD163 positive macrophages (M2 phenotype) were seen predominantly within the tumors (NSCLC and breast cancer) whereas CD68 positive macrophages were identified towards the periphery in the non-tumor microenvironment. NSCLC tumor cells showed higher expression levels of IL-2, IL-6 and TNFa compared to breast cancer tumor cells. Predominance of CD3 positive T cells was noted with relative lack of PAX5 positive B cells within both NSCLC and breast cancer tumors. Multiplexing enabled assessment of relative expression levels of IL-2, IL-6 and TNFa in T lymphocytes and macrophages and TNFa in dendritic cells. Our hybrid multiplex panel can provide insights into the visualization of spatial organization of tumor microenvironment. It also enables semi-quantitative assessments for the expression levels of cytokines and their relative expression within tumor and immune cells, thereby helping to correlate response to treatment, especially with checkpoint inhibitors.

Citation Format: Brenna Dennison, Stephanie Allen, Bhavika Patel, Jacob Stapleton, Roni Archuleta, Mary Lou Rath, Navi Mehra, Sameer S. Talwalkar. Precision immuno-oncology (IO) ISH/IF multiplex panel: Spatial detection of cytokines and IO biomarkers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2322.
Abstract 2323: ZGGS18, a bispecific drug candidate of anti-VEGF and TGFbeta-trap, inhibiting tumor proliferation and improving tumor microenvironment with I/O therapy
Authors: Liu Ruifeng , Suarez Alfonso , Liban Tyler , Zhang Bin , Dai Tongcheng , Karow Margaret , Sheng Jackie , Sheng Zelin , Lv Binhua , Zhu Bing
Published: April 2023

ZGGS18 is a bifunctional fusion protein targeting both human vascular endothelial growth factor (VEGF) and transforming growth factor-β (TGF-β), which consist of a monoclonal antibody against VEGF and an engineered extracellular domain of TGF-β receptor II (TGF-βRII ECD). In vitro studies show ZGGS18 specifically and simultaneously binding towards various VEGFs and capturing different TGF-β isoforms by trap domains that play a synergistic role to block signaling of VEGF/TGF-β with their receptors. For instance, ZGGS18 inhibits human VEGF121 and 165 induced HUVEC proliferation; ZGGS18 also neutralizes TGF-β1 and -β3 related growth arrest of HT-2 cells. Further studies show that ZGGS18 does not induce ADCC- or CDC-dependent killing tumor cells expressing two soluble targets, suggesting its binding affinities towards Fc-gamma and complement C1q receptors without effector functions. These results support functional depletion of VEGF/TGF-β in tumor microenvironment (TME) by ZGGS18 essentially for its MOA to inhibit tumor growth. A dose-dependent in vivo anti-tumor efficacy of ZGGS18 (1, 3 and 15 mg/kg) has been found in PBMC humanized H460 lung cancer xenograft model. In addition, ZGGS18 has enhanced efficacies to kill tumor growth in MC38 xenograft models when it is combined with anti-PD-1 antibodies (keytruda), support the improvement of TME for I/O reagents by ZGGS18 in cooperation with its role in anti-proliferation and anti-angiogenesis. Toxicity and safety pharmacology studies with weekly repeat doses for four weeks in cynomolgus monkeys indicate that ZGGS18 is well tolerated over the dose range 13, 40 and 120 mg/kg without obviously toxic or adverse effects (NOAEL =120 mg/kg). PK/TK analyses indicate that ZGG18 has a prolonged half-life in plasma, attribute to its human neonatal Fc receptor (FcRn) binding and pH-dependent interaction for mAb recycling. The IND application of ZGGS18 has been approved by both FDA and Chinese NMPA for clinical trials in treatment of advanced tumors as monotherapy and/or in combined with PD-1/PD-L1 based I/O therapies.

Citation Format: Ruifeng Liu, Alfonso Suarez, Tyler Liban, Bin Zhang, Tongcheng Dai, Margaret Karow, Jackie Sheng, Zelin Sheng, Binhua Lv, Bing Zhu. ZGGS18, a bispecific drug candidate of anti-VEGF and TGFbeta-trap, inhibiting tumor proliferation and improving tumor microenvironment with I/O therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2323.
Abstract 2333: Verteporfin demonstrates antitumoral effect in murine cholangiocarcinoma models
Authors: Lee Jihye , Wang Xin , Bian Jing , Cam Maggie , Xie Changqing
Published: April 2023

Cholangiocarcinoma (CCA) is a heterogenous malignancy that is derived from the biliary epithelium. There is growing concern as CCA incidence in United States has steadily increased. Additionally, CCA disproportionately affects different populations, and overall prognosis remains poor with a 5-year survival rate of 10%. Moreover, the Hippo/yes-associated protein (YAP) pathway has been reported to affect various aspects of tumorigenesis and recent reports showed that higher expression of YAP is negatively associated with the survival rate in CCA patients. Thus, we investigated the antitumoral effect of verteporfin, a YAP pathway inhibitor, in YAP/Akt and Notch/Akt hydrodynamic tail vein injected murine models. Our results demonstrated reduced liver weight in verteporfin treated groups compared to the vehicle treated group. Single cell RNA sequence analysis noted the verteporfin treated group had reduced stemness-related expressing cells in the malignant cell population. Furthermore, B cell, CD8+ T cell, and Tumor-Associated Macrophages M1 (TAM M1) populations increased following verteporfin treatment. In summary, this study indicates that verteporfin reduces tumorigenesis in CCA by the regulation of cancer stemness and modulation of immune cells, which includes anti-tumoral/pro-inflammatory TAM M1 in CCA animal models.

Citation Format: Jihye Lee, Xin Wang, Jing Bian, Maggie Cam, Changqing Xie. Verteporfin demonstrates antitumoral effect in murine cholangiocarcinoma models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2333.
Abstract 2337: Overcome solid tumor immunosuppression for T cells-mediated cancer elimination
Authors: Shi Lei , Bian Zhen , Zen Jane , Stylli Harry , Liu Yuan
Published: April 2023

T cell immunotherapy to solid tumor requires not only sufficient numbers of cancer cell-specific cytotoxic T cells, but also their infiltration into tumor tissues and the ability to resist hostile TME immunosuppression.

Through a proprietary process, we have identified a unique reagent, termed KX147, of which treatment of tumor potently repolarizes the TME, resulting in abrogation of multi-axes immunosuppressive signal transduction pathways reprograme of the TME for innate and adaptive immunity against cancer. We found that immunotherapy-refractory solid tumors under therapies generally fail to induce intratumoral macrophages for proinflammatory phenotypic change, nor did they reprogram the TME towards anti-tumor immunogenicity. Rather, these tumors arbitrate strong resistance through increasing IL-10 production, TGFβ signaling (TGFBR1 and TGFBR2), and CCL2 chemokine that attracts MDSC, leading to wound healing and reinforcement of tumor immunosuppression. In stark contrast, tumors treated with KX147 exhibit completely different TME responses to checkpoint blockade, RT and other immune-modulatory therapies, and reshape intratumoral macrophages from immunosuppression to potent proinflammation, featuring high expressions of TNFα, IFNγ, IL-1β, IL-6, IL-12, IL-17, IL-18, etc., while abating IL-10 and TGFβ. Moreover, KX147 also induces a prominent immunogenic antigen presentation machinery with elevated cell surface MHC-I, MHC-II, and costimulatory molecules CD80, CD86, CD40, OX40L on macrophages, which through antigen presentation induces large expansion of tumor-specific T cells for tumor elimination. Together, our studies demonstrate a novel immunotherapy strategy with KX147 combination, which holds promises for rapid and systemic elimination of solid tumors and metastases to achieve cure.

Citation Format: Lei Shi, Zhen Bian, Jane Zen, Harry Stylli, Yuan Liu. Overcome solid tumor immunosuppression for T cells-mediated cancer elimination [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2337.
Abstract 2343: Targeting EPHB2/ABL1 restores anti-tumor immunity in a preclinical model of ependymoma
Authors: Ren Jun , Amoozgar Zohreh , Zhao Yuhui , Lei Pinji , Ho William W. , Robergea Sylvie , Huang Peigen , Duda Dan G. , Xu Lei , Jain Rakesh K.
Published: April 2023

Ependymoma is a common type of brain tumor in children, often resistant to available cytotoxics. Molecular profiling studies have led to a better understanding of ependymoma pathology and revealed a critical role of EPHB2. However, the immune system's role in tumor progression and treatment response remains poorly understood. Here, we show that the multikinase inhibitor dasatinib potently inhibits EPHB2 and ABL1 and the growth of ependymoma both in vitro and in vivo. Using an immunocompetent mouse model of ependymoma that faithfully recapitulates its immune microenvironment, we found an increased frequency of M2-like tumor-associated macrophages (TAMs), which proportionally increased with tumor size. Treatment with dasatinib reprogrammed the immune microenvironment of ependymoma by polarizing the TAMs toward an M1-like phenotype and increasing CD8 T cell activation. Dasatinib treatment induced complete regression of established mEPN-Ephb2 tumors in 78% of the animals and protected survivors against tumor recurrence. These studies indicate that dasatinib may be an effective therapy for this molecular subgroup of pediatric brain tumors and should be tested in clinical trials.

Citation Format: Jun Ren, Zohreh Amoozgar, Yuhui Zhao, Pinji Lei, William W. Ho, Sylvie Robergea, Peigen Huang, Dan G. Duda, Lei Xu, Rakesh K. Jain. Targeting EPHB2/ABL1 restores anti-tumor immunity in a preclinical model of ependymoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2343.
Abstract 2385: Targeting mannose receptor expression on macrophages for sentinel lymph node mapping in endometrial cancer: murine model
Authors: Cho Hyun-Woong , Jeon OK Hwa , Lee Sanghoon , Kim Hyun Koo , Lee Jae Kwan , Hong Jin Hwa
Published: April 2023

Background: Sentinel lymph node (SLN) mapping has been used in various cancer surgery including endometrial cancer as a targeted assessment tool for identifying lymph node metastases, an alternative for complete lymphadenectomy to minimize surgical morbidity. Neomannosyl human serum albumin (MSA) targets the macrophage mannose receptor CD206 which is highly expressed in metastatic lymph node. The aim of this study was to evaluate the effectiveness of indocyanine green-neomannosyl human serum albumin (MSA:ICG) which targets macrophage mannose receptor CD206 for SLN mapping.

Methods: LN targeting ability was examined on mouse footpad models of uterine cancer using MSA:ICG and ICG for comparison. Seven mice with uterine cancer and 2 mice without cancer as a control group were each assigned to MSA:ICG and ICG groups. After injection of tracers, fluorescence images were taken and SLN biopsy was done to confirm LN metastasis status. We compared the signal-to-background ratio (SBR) according to time.

Results: In both MSA:ICG and ICG group, 16 lymph nodes out of 18 were detected using fluorescence imaging (detection rate: 88.9%). Four lymph nodes in MSA:ICG group and 5 lymph nodes in ICG group were metastatic among 16 lymph node each group. Excluding lymph nodes that were not detected, the SBR after the injection of MSA:ICG group was significantly higher than ICG group in metastatic LN (MSA:ICG 4.36 vs ICG 3.07, p<0.001), non-metastatic lymph nodes (MSA:ICG 3.11 vs ICG 1.93, p=0.001), and normal control lymph nodes (MSA:ICG 1.38 vs ICG 0.54, p<0.001). Comparing signal intensity according to CD206 expression, there was no difference in SBR values between the two groups in patients without CD206 expression, but the MSA group had a statistically significantly higher SBR value than the ICG group (p<0.001) in patients with CD206 moderate or high expression.

Conclusions: This study showed that MSA:ICG may be an effective tracer for sentinel lymph node mapping. In addition, the metastatic lymph nodes expressing CD206 showed higher signal than normal lymph nodes in MSA:ICG group, unlike in ICG group. Further studies using larger animal models and clinical trials are needed to confirm this result.

Citation Format: Hyun-Woong Cho, OK Hwa Jeon, Sanghoon Lee, Hyun Koo Kim, Jae Kwan Lee, Jin Hwa Hong. Targeting mannose receptor expression on macrophages for sentinel lymph node mapping in endometrial cancer: murine model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2385.
Abstract 2394: Activatable fluorogenic probe for two-photon imaging of heparanase in vivo
Authors: Rabinowitz Zachary , Wang Zhishen , Somers Johnathan , Liu Jun , Jiang Joyce , Zhang Nina , Cui Lina
Published: April 2023

Heparanase, an endo-β-glucuronidase, is the only known enzyme that is responsible for the cleavage of heparan sulfate chains of heparan sulfate proteoglycans. Its enzymatic activity plays a key role in remodeling and degradation of the extracellular matrix, generation of biologically active carbohydrate fragments, and liberation of effector molecules, such as growth factors and chemokines, that are bound to the heparan sulfate side chains. Overexpression of heparanase has been implicated in most types of cancers and various inflammatory diseases that usually involve extensive heparan sulfate degradation and extracellular matrix remodeling. In particular, the role of heparanase in cancer is highly significant as it regulates multiple biological activities that fosters tumor progression, angiogenesis, metastases. Consequently, heparanase has been considered a biological target for various therapeutic applications including anti-cancer drug development. However, one of the major challenges in this research area is the development of robust and feasible heparanase probes to study the enzymes’ fundamental biology in the extracellular matrix, to provide heparanase-targeted diagnostics, and to aid in drug discovery efforts. Our lab has been developing various small-molecule-based heparanase activatable probes, and we have successfully developed chemical strategies that allow sensitive detection of heparanase activity in various contexts. Two-photon microscopy allows for deeper tissue penetration, excellent spatial resolution, and efficient light detection with reduced photobleaching effects. Herein, we extend our chemical strategies to develop a heparanase-activatable two-photon fluorogenic probe for real-time detection of heparanase activity in animal models of highly-aggressive brain cancer using two-photon imaging.

Citation Format: Zachary Rabinowitz, Zhishen Wang, Johnathan Somers, Jun Liu, Joyce Jiang, Nina Zhang, Lina Cui. Activatable fluorogenic probe for two-photon imaging of heparanase in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2394.
Abstract 2396: Longitudinal monitoring of stromal stiffness predicts immune escape in preclinical hepatocellular carcinoma
Authors: Minamiguchi Kiyoyuki , Cortes Andrea , Lopez Patricia Da Costa , Williams Malea , Anfossi Simone , Melancon Marites , Avritscher Rony
Published: April 2023

Background/Objectives: Increased tumor stiffness enhances disease progression and aggressiveness in several cancers. Stiffer extracellular tumor matrix hinders T cell trafficking and promotes immune escape. The interplay between liver tumor mechanical properties and hepatocellular immune cell landscape has not been fully investigated. The aim of our study is to correlate tumor stiffness quantified with shear wave elastography and tumor-infiltrating lymphocyte accumulation using a syngeneic orthotopic rat model of hepatocellular carcinoma (HCC).

Methods: Rat hepatoma cell line McA-RH7777 were implanted in the left liver lobe of Buffalo rats. Maximum tumor size and stiffness were evaluated 3 weeks after implantation (baseline) and weekly for 14 days using ultrasound. The maximum tumor size of each rat was evaluated on transverse view of B-mode ultrasound. The elasticity of each rat was evaluated by shear wave elastography (SWE), and SWE measurements were obtained at the maximum tumor size in m/s. A circular region of interest was put manually on the SWE images to calculate tumor stiffness. SWE was performed 3 times for each rat at each time point, and the mean value of tumor stiffness was used for analysis. After 14 days, tumors were harvested for histology and immune cell isolation. Isolated cells were stained with CD3, CD4, CD8, CD25, CD45, CD103, CD161, CD279 (PD-1), FOXP3, IL17A, and INF-g and submitted to spectral cytometry.

Results: Shear wave speed at baseline and at 14 days ranged from 1.95 to 3.07 m/s and from 2.25 to 3.26 m/s, respectively. The rate of longitudinal increase or decrease in tumor stiffness measured by SWE is inversely correlated with intratumoral infiltration of CD45+/CD3+, CD3+/CD8+ T cells and IFN-producing CD3+/CD8+ T cells (P=0.015, r=-0.62[-0.857, -0.15], P<0.01, r=-0.74[-0.913, -0.346], P=0.02, r=-0.6[-0.859, -0.107]). Intratumoral CD3+/CD4+ T cells did not significantly correlate with the rate of change in stiffness but did correlate with the rate of change in tumor size measured by B-mode ultrasound (P=0.03, r=-0.58[-0.850, -0.0730]).

Conclusion: Our results confirm previous reports of immune escape in stiffer solid tumors in the context of HCC. Findings suggest that longitudinal monitoring of tumor stiffness can be useful for the assessment of immune cell infiltration.

Citation Format: Kiyoyuki Minamiguchi, Andrea Cortes, Patricia Da Costa Lopez, Malea Williams, Simone Anfossi, Marites Melancon, Rony Avritscher. Longitudinal monitoring of stromal stiffness predicts immune escape in preclinical hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2396.
Abstract 2403: Divergent evolutionary response to multiple fractions of radiation (MF2) therapy in cervical cancer cell lines (CCCL)
Authors: Kelley Johanna , Durmaz Arda , Kuznicki Michelle , Petty Aaron , Scott Jacob , Vargas Roberto
Published: April 2023

Introduction: Treatment for locally-advanced cervical cancer is not currently tailored based on genomic markers of treatment response. We aim to explore the potential for differential evolutionary trajectories of radiation (RT) resistance throughout treatment using two in vitro models.

Methods: Sixteen cervical cancer cell lines ( CCCL) were exposed to 1.8 Gy fractions of RT radiation for five sequential days and the surviving fraction (MF2) calculated. Differential gene expression (DGE) was performed on pre and post - treatment samples. Partial least squares (PLS) regression was completed with MF2 as the endpoint . Protein-protein interaction (PPI) network and overrepresentation analysis of genes was also performed. Four CCCL (BOKU, SKG II SF, ME180, and HeLa) were serially exposed to 50 Gy of gamma radiation RT given in 2Gy fractions. Five evolutionary replicates and three passage-controls were maintained. The mean integral survival as a function of dose (AUC) was calculated and ANOVA analysis performed.

Results: MF2 as the phenotypic endpoint identified 19 DE genes. PLS identified histones (H4C8, H2BC18, H2BC4) as positively associated and transcriptional regulators (NR2F1, KALRN) as negatively associated with MF2 (Figure 1AB). PPI and Reactome Pathways also highlighted histone deacetylase pathways, nucleosome assembly and regulation of membrane potentials (Figure 1BC). Longitudinal treatment RT identified differential evolutionary trajectories between CCCL replicates. Two BOKU (p = 0.005 and 0.0090), one SKG II SF (p = 0.0146), and one ME180 (p = 0.0384) replicates became resistant. Four SKG II SF (p < 0.0001), and two HeLa (p = 0.0005 and p = 0.0068) became more sensitive to radiationRT (1C).

Conclusion: Divergent evolutionary trajectories under therapeutic stress can be elicited using in vitro evolutionary experimentation with cervical cancer cell linesCCCL. Genomic correlation with divergent evolutionary trajectories may allow for the identification of real-time markers of resistance or sensitivity and allow for true RT radiotherapy personalization.

Citation Format: Johanna Kelley, Arda Durmaz, Michelle Kuznicki, Aaron Petty, Jacob Scott, Roberto Vargas. Divergent evolutionary response to multiple fractions of radiation (MF2) therapy in cervical cancer cell lines (CCCL) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2403.