Mouse Model Of Pancreatic Cancer Research Articles

Non‐Natural MUC1 Glycopeptide Homogeneous Cancer Vaccine with Enhanced Immunogenicity and Therapeutic Activity

AbstractGlycopeptides derived from the glycoprotein mucin‐1 (MUC1) have shown potential as tumor‐associated antigens for cancer vaccine development. However, their low immunogenicity and non‐selective conjugation to carriers present significant challenges for the clinical efficacy of MUC1‐based vaccines. Here, we introduce a novel vaccine candidate based on a structure‐guided design of an artificial antigen derived from MUC1 glycopeptide. This engineered antigen contains two non‐natural amino acids and has an α‐S‐glycosidic bond, where sulfur replaces the conventional oxygen atom linking the peptide backbone to the sugar N‐acetylgalactosamine. The glycopeptide is then specifically conjugated to the immunogenic protein carrier CRM197 (Cross‐Reactive Material 197), a protein approved for human use. Conjugation involves selective reduction and re‐bridging of a disulfide in CRM197, allowing the attachment of a single copy of MUC1. This strategy results in a chemically defined vaccine while maintaining both the structural integrity and immunogenicity of the protein carrier. The vaccine elicits a robust Th1‐like immune response in mice and generates antibodies capable of recognizing human cancer cells expressing tumor‐associated MUC1. When tested in mouse models of colon adenocarcinoma and pancreatic cancer, the vaccine is effective both as a prophylactic and therapeutic use, significantly delaying tumor growth. In therapeutic applications, improved outcomes were observed when the vaccine was combined with an anti‐programmed cell death protein 1 (anti‐PD‐1) checkpoint inhibitor. Our strategy reduces batch‐to‐batch variability and enhances both immunogenicity and therapeutic potential. This site‐specific approach disputes a prevailing dogma where glycoconjugate vaccines require multivalent display of antigens.

Rac1 GTPase Regulates the βTrCP-Mediated Proteolysis of YAP Independently of the LATS1/2 Kinases

Background: Oncogenic mutations in the KRAS gene are detected in >90% of pancreatic cancers (PC). In genetically engineered mouse models of PC, oncogenic KRAS drives the formation of precursor lesions and their progression to invasive PC. The Yes-associated Protein (YAP) is a transcriptional coactivator required for transformation by the RAS oncogenes and the development of PC. In Ras-driven tumors, YAP can also substitute for oncogenic KRAS to drive tumor survival after the repression of the oncogene. Ras oncoproteins exert their transforming properties through their downstream effectors, including the PI3K kinase, Rac1 GTPase, and MAPK pathways. Methods: To identify Ras effectors that regulate YAP, YAP levels were measured in PC cells exposed to inhibitors of oncogenic K-Ras and its effectors. Results: In PC cells, the inhibition of Rac1 leads to a time-dependent decline in YAP protein, which could be blocked by proteosome inhibitor MG132. This YAP degradation after Rac1 inhibition was observed in a range of cell lines using different Rac1 inhibitors, Rac1 siRNA, or expression of dominant negative Rac1T17N mutant. Several E3 ubiquitin ligases, including SCFβTrCP, regulate YAP protein stability. To be recognized by this ligase, the βTrCP degron of YAP (amino acid 383–388) requires its phosphorylation by casein kinase 1 at Ser384 and Ser387, but these events must first be primed by the phosphorylation of Ser381 by LATS1/2. Using Flag-tagged mutants of YAP, we show that YAP degradation after Rac1 inhibition requires the integrity of this degron and is blocked by the silencing of βTrCP1/2 and by the inhibition of casein kinase 1. Unexpectedly, YAP degradation after Rac1 inhibition was still observed after the silencing of LATS1/2 or in cells carrying a LATS1/2 double knockout. Conclusions: These results reveal Rac1 as an oncogenic KRAS effector that contributes to YAP stabilization in PC cells. They also show that this regulation of YAP by Rac1 requires the SCFβTrCP ligase but occurs independently of the LATS1/2 kinases.

All-trans retinoic acid-mediated ADAR1 degradation synergizes with PD-1 blockade to suppress pancreatic cancer.

As a double-stranded RNA-editing enzyme and an interferon-stimulated gene, double-stranded RNA-specific adenosine deaminase (ADAR1) suppresses interferon signaling and contributes to immunotherapy resistance. Suppression of ADAR1 overcomes immunotherapy resistance in preclinical models, but has not yet been translated to clinical settings. By conducting a screening of a subset of the FDA-approved drugs, we found that all-trans retinoic acid (ATRA, also known as tretinoin) caused ADAR1 protein degradation through ubiquitin-proteasome pathways and concomitantly increased PD-L1 expression in pancreatic and breast cancers. In addition, the combination of ATRA and PD-1 blockade reprogrammed the tumor microenvironment and unleashed antitumor immunity and thereby impeded tumor growth in pancreatic cancer mouse models. In a pilot clinical trial, a higher dose of ATRA plus the anti-PD-1 antibody nivolumab prolonged median overall survival in patients with chemotherapy-resistant pancreatic cancer compared to a lower dose of the same regimen. In this study, ATRA was the first drug to be found to cause ADAR1 degradation. We propose translation of a promising 2-pronged antitumor strategy using ATRA and nivolumab to convert immunologically "cold" into "hot" tumors susceptible to immune checkpoint blockade.

Contemporaneous Inflammatory, Angiogenic, Fibrogenic, and Angiostatic Cytokine Profiles of the Time-to-Tumor Development by Cancer Cells to Orchestrate Tumor Neovascularization, Progression, and Metastasis.

Cytokines can promote various cancer processes, such as angiogenesis, epithelial to mesenchymal transition (EMT), invasion, and tumor progression, and maintain cancer stem-cell-like (CSCs) cells. The mechanism(s) that continuously promote(s) tumors to progress in the TME still need(s) to be investigated. The data in the present study analyzed the inflammatory, angiogenic, fibrogenic, and angiostatic cytokine profiles in the host serum during tumor development in a mouse model of human pancreatic cancer. Pancreatic MiaPaCa-2-eGFP cancer cells were subcutaneously implanted in RAG2xCγ double mutant mice. Blood samples were collected before cancer cell implantation and every week until the end point of the study. The extracted serum from the blood of each mouse at different time points during tumor development was analyzed using a Bio-Plex microarray analysis and a Bio-Plex 200 system for proinflammatory (IL-1β, IL-10, IFN-γ, and TNF-α) and angiogenic and fibrogenic (IL-15, IL-18, basic FGF, LIF, M-CSF, MIG, MIP-2, PDGF-BB, and VEGF) cytokines. Here, we find that during cancer cell colonization for tumor development, host angiogenic, fibrogenic, and proinflammatory cytokine profiling in the tumor-bearing mice has been shown to significantly reduce host angiostatic and proinflammatory cytokines that restrain tumor development and increase those for tumor growth. The proinflammatory cytokines IL-15, IL-18, and IL-1β profiles reveal a significant host serum increase after day 35 when the tumor began to progress in growth. In contrast, the angiostatic cytokine profiles of TNFα, MIG, M-CSF, IL-10, and IFNγ in the host serum revealed a dramatic and significant decrease after day 5 post-implantation of cancer cells. OP treatment of tumor-bearing mice on day 35 maintained high levels of angiostatic and fibrogenic cytokines. The data suggest an entirely new regulation by cancer cells for tumor development. The findings identify for the first time how pancreatic cancer cells use host cytokine profiling to orchestrate the initiation of tumor development.

Targeting Human Pancreatic Cancer with a Fluorophore-Conjugated Mucin 4 (MUC4) Antibody: Initial Characterization in Orthotopic Cell Line Mouse Models.

Background/Objectives: Pancreatic cancer is the third leading cause of death related to cancer. The only possible cure presently is complete surgical resection; however, this is limited by difficulty in clearly defining tumor margins. Enhancement of the visualization of pancreatic ductal adenocarcinoma (PDAC) tumor margins using near-infrared dye-conjugated tumor-specific antibodies was pioneered by using anti-CEA, anti-CA19.9, and anti-MUC5AC in orthotopic mouse models of pancreatic cancer. Recently, an antibody to Mucin 4 (MUC4) conjugated to a fluorescent probe has shown promise in targeting colon tumors in orthotopic mouse models. Methods: In the present study, we targeted pancreatic cancer using an anti-MUC4 antibody conjugated to IRDye800 (anti-MUC4-IR800) in orthotopic mouse models. Two pancreatic cancer human cell lines were used, SW1990 and CD18/HPAF. Results: Anti-MUC4-IR800 targeted the two pancreatic cancer cell line tumors in orthotopic mouse models with high tumor-to-pancreas ratios and high tumor-to-liver ratios, with greater targeting seen in SW1990. Conclusions: The present results suggest anti-MUC4-IR800's potential to be used in fluorescence-guided surgical resection of pancreatic cancer.

A Novel Pan-RAS Inhibitor with a Unique Mechanism of Action Blocks Tumor Growth in Mouse Models of GI Cancer.

ADT-007 has unique pharmacological properties with distinct advantages over other RAS inhibitors by circumventing resistance and activating antitumor immunity. ADT-007 prodrugs and analogs with oral bioavailability warrant further development for RAS-driven cancers.

Dronedarone hydrochloride (DH) induces pancreatic cancer cell death by triggering mtDNA-mediated pyroptosis

Pancreatic cancer is one of the leading causes of cancer-associated mortality, with a poor treatment approach. Previous study has shown that inducing pyroptosis in pancreatic ductal adenocarcinoma (PDAC) slows the growth of PDACs, implying that pyroptosis inducers are potentially effective for PDAC therapy. Here, we found that Dronedarone hydrochloride (DH), an antiarrhythmic drug, induces pyroptosis in pancreatic cancer cells and inhibits PDAC development in mice. In PANC-1 cells, DH caused cell death in a dosage- and time-dependent manner, with only pyroptosis inhibitors and GSDMD silencing rescuing the cell death, indicating that DH triggered GSDMD-dependent pyroptosis. Further work revealed that DH increased mitochondrial stresses and caused mitochondrial DNA (mtDNA) leakage, activating the cytosolic STING-cGAS and pyroptosis pathways. Finally, we assessed the anti-cancer effects of DH in a pancreatic cancer mouse model and found that DH treatment suppressed pancreatic tumor development in vivo. Collectively, our investigation demonstrates that DH triggers pyroptosis in PDAC and proposes its potential effects on anti-PDAC growth.

210 (PB198): KRASmulti inhibition and checkpoint targeting therapy show strong combination potential in KRAS wild-type and KRASG12V-driven syngeneic mouse models of pancreatic cancer

210 (PB198): KRASmulti inhibition and checkpoint targeting therapy show strong combination potential in KRAS wild-type and KRASG12V-driven syngeneic mouse models of pancreatic cancer

133 (PB121): A novel pan-RAS inhibitor (ADT-1004) with superior efficacy over mutation-specific KRAS inhibitors in mouse models of pancreatic cancer

133 (PB121): A novel pan-RAS inhibitor (ADT-1004) with superior efficacy over mutation-specific KRAS inhibitors in mouse models of pancreatic cancer

Mesenchymal stem cell-delivered paclitaxel nanoparticles exhibit enhanced efficacy against a syngeneic orthotopic mouse model of pancreatic cancer

Pancreatic cancer is considered the deadliest among various solid tumors, with a five-year survival rate of 13 %. One of the major challenges in the management of advanced pancreatic cancer is the inefficient delivery of chemotherapeutics to the tumor site. Even though nanocarriers have been developed to improve tumoral delivery of chemotherapeutics, less than 1 % of the drugs reach tumors, rendering inadequate concentration for effective inhibition of tumors. As a potential alternative, mesenchymal stem cells (MSCs) can effectively deliver their cargo to tumor sites because of their resistance to chemotherapeutics and inherent tumor tropism. In this study, we used MSCs for the delivery of dibenzocyclooctyne (DBCO)-functionalized paclitaxel (PTX)-loaded poly(lactide-co-glycolide)-b-poly (ethylene glycol) (PLGA) nanoparticles. MSCs were modified to generate artificial azide groups on their surface, allowing nanoparticle loading via endocytosis and surface conjugation via click chemistry. This dual drug loading strategy significantly improves the PTX-loading capacity of azide-expressed MSCs (MSC-Az, 55.4 pg/cell) compared to unmodified MSCs (28.1 pg/cell). The in vitro studies revealed that PTX-loaded MSC-Az, nano-MSCs, exhibited cytotoxic effects against pancreatic cancer without altering their inherent phenotype, differentiation abilities, and tumor tropism. In an orthotopic pancreatic tumor model, nano-MSCs demonstrated significant inhibition of tumor growth (p < 0.05) and improved survival (p < 0.0001) compared to PTX solution, PTX nanocarriers, and Abraxane. Thus, nano-MSCs could be an effective delivery system for targeted pancreatic cancer chemotherapy and other solid tumors.

Defining heritability, plasticity, and transition dynamics of cellular phenotypes in somatic evolution.

Single-cell sequencing has characterized cell state heterogeneity across diverse healthy and malignant tissues. However, the plasticity or heritability of these cell states remains largely unknown. To address this, we introduce PATH (phylogenetic analysis of trait heritability), a framework to quantify cell state heritability versus plasticity and infer cell state transition and proliferation dynamics from single-cell lineage tracing data. Applying PATH to a mouse model of pancreatic cancer, we observed heritability at the ends of the epithelial-to-mesenchymal transition spectrum, with higher plasticity at more intermediate states. In primary glioblastoma, we identified bidirectional transitions between stem- and mesenchymal-like cells, which use the astrocyte-like state as an intermediary. Finally, we reconstructed a phylogeny from single-cell whole-genome sequencing in B cell acute lymphoblastic leukemia and delineated the heritability of B cell differentiation states linked with genetic drivers. Altogether, PATH replaces qualitative conceptions of plasticity with quantitative measures, offering a framework to study somatic evolution.

αvβ3 integrin-targeted magnetic resonance imaging in a pancreatic cancer mouse model using RGD-modified liposomes encapsulated with Fe-deferoxamine.

Magnetic resonance (MR) imaging is a powerful imaging modality for obtaining anatomical information with high spatial and temporal resolution. In the drug delivery system (DDS) framework, nanoparticles such as liposomes are potential candidates for MR imaging. We validated that RGD peptides are possible targeting molecules for pancreatic cancer with αvβ3 integrin expression. This study aimed to evaluate RGD-modified liposomes loaded with ferrioxamine B for pancreatic cancer imaging. We synthesized four types of RGD-modified liposomes encapsulated with ferrioxamine B (SH-, H-, M-, and L-RGD-liposomes). The binding affinity of RGD-modified liposomes was evaluated in a competitive inhibition study using 125I-echistatin. To investigate the pharmacokinetics of RGD-modified liposomes, a biodistribution study using RGD-liposomes labeled with 111In was carried out in a human pancreatic cancer PANC-1 xenograft mouse model. Finally, MR was performed using ferrioxamine-B-loaded liposomes. RGD-liposomes inhibited the binding of 125I-echistatin to RGD. The biodistribution study revealed that 111In-RGD-liposomes accumulated significantly in the liver and spleen. Among the 111In-RGD-liposomes, 111In-H-RGD-liposomes showed the highest tumor-to-normal tissue ratio. In the MR study, H-RGD-liposomes loaded with ferrioxamine B showed higher tumor-to-muscle signal ratios than RKG-liposomes loaded with ferrioxamine B (control). We successfully synthesized RGD-liposomes that can target αvβ3 integrin.

Cutting-Edge Approach to Targeted Therapy: Repositioning of Old Drugs in Combination with Standard Clinical Chemotherapeutics Potentiates a Propitious Novel Targeted Therapy for Human Pancreatic Cancer

Metastatic pancreatic cancer leads to a fatal outcome, with a median progression-free survival of approximately six months when utilizing the most successful combination of chemotherapeutic regimens. When drug resistance develops, it facilitates an increase in primary tumor growth and new and growing metastases. Patients inevitably and quickly succumb to their disease and die. Notably, chemotherapy has an unintended impact on the development of drug resistance through the enhancement of EMT development and the enrichment of cancer stem cells (CSC). Recent report discovered that neuraminidase-1 (Neu-1) regulates EMT induction, angiogenesis, and cellular proliferation by the activation of several receptor tyrosine kinases. Here, the continual therapeutic inhibition of Neu-1 through intravenous administration of oseltamivir phosphate (OP) and aspirin (ASA) alongside GEM treatment significantly inhibits tumor progression, crucial compensatory signaling pathways, EMT program, CSC, and metastasis progression in a preclinical RAG2xCy double mutant BALB/c mouse model of human PANC-1 pancreatic cancer. The tumorigenic and metastatic potential of the xenotumors from the animals treated with the experimental protocols were significantly ablated when transferred into the mammary fat pads of NSG (NOD SCID gamma) branded mice. Keywords: pancreatic cancer; chemoresistance; drug repurposing; EMT.

Abstract B007: Optimization of an Orthotopic Mouse Model of Pancreatic Cancer to Simulate Liver Metastasis and Cancer Cachexia

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive disease with a poor prognosis due to two key features. The first is metastasis at presentation, occurring in approximately 80% of PDAC patients, with 90% of these metastases occurring in the liver. The second is cachexia, which compromises treatment tolerance and reduces the quality of life for patients. Although various mouse models of PDAC exist, recapitulating both metastatic and cachectic features have been challenging. In this study, we optimized an orthotopic mouse model of PDAC by altering implantation sites, subcloning parental murine PDAC cells, adjusting the number of transplanted cells, and varying the age of recipient mice to study liver metastasis and cancer cachexia. These modifications significantly increased the latency of tumor development and the time course of the disease, concurrently increasing the rate of liver metastasis to approximately 70%. Furthermore, reliable cachexia endpoints were also achieved in both PDAC mice with and without metastases, mirroring the condition in patients. We also found that cachectic muscles from PDAC mice with metastasis exhibit a similar transcriptional profile to muscles derived from mice and patients without metastasis. Together, this model is likely to be advantageous in both advancing our understanding of the mechanisms of PDAC cachexia and in evaluating novel therapeutics. Citation Format: David J Wang, Victoria Spadafora, Benjamin R Pryce, Alexander Oles, Erin E Talbert, Micheal C Ostrowski, Denis C Guttridge. Optimization of an Orthotopic Mouse Model of Pancreatic Cancer to Simulate Liver Metastasis and Cancer Cachexia [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B007.

Abstract C001: Transforming Growth Factor-β Blockade in Pancreatic Cancer Enhances Sensitivity to Combination Chemotherapy in Mice

Abstract Background: TGFβ plays a critical role in promoting pancreatic tumor progression, attenuating CD8 T cell proliferation and cytokine production, enhancing the differentiation of myofibroblasts and the deposition of extracellular matrix. However, single agent TGFβ blockade has shown limited efficacy in mouse models of pancreatic cancer. Methods: We evaluated the TGFβ- blocking antibody NIS793 in combination with gemcitabine/nanoparticle (albumin-bound)- paclitaxel or FOLFIRINOX (folinic acid [FOL], 5-fluorouracil [F], irinotecan [IRI] and oxaliplatin [OX]) in orthotopic pancreatic cancer models. Single-cell RNA sequencing and immunofluorescence were used to evaluate changes in tumor cell state and the tumor microenvironment. Results: Blockade of TGFβ with chemotherapy reduced tumor burden in poorly immunogenic pancreatic cancer, without affecting the metastatic rate of cancer cells. Efficacy of combination therapy was not dependent on CD8 T cells, because response to TGFβ blockade was preserved in CD8-depleted or recombination activating gene 2 (RAG2-/-) mice. TGFβ blockade decreased total α-smooth muscle actin-positive fibroblasts but had minimal effect on fibroblast heterogeneity. Bulk RNA sequencing on tumor cells sorted ex vivo revealed that tumor cells treated with TGFβ blockade adopted a classical lineage consistent with enhanced chemosensitivity, and immunofluorescence for cleaved caspase 3 confirmed that TGFβ blockade increased chemotherapy-induced cell death in vivo. Conclusions: TGFβ regulates pancreatic cancer cell plasticity between classical and basal cell states. TGFβ blockade in orthotropic models of pancreatic cancer enhances sensitivity to chemotherapy by promoting a classical malignant cell state. This study provides scientific rationale for evaluation of NIS793 with FOLFIRINOX or gemcitabine/nanoparticle (albumin-bound) paclitaxel chemotherapy backbone in the clinical setting and supports the concept of manipulating cancer cell plasticity to increase the efficacy of combination therapy regimens. This study provided the scientific rationale for the evaluation of TGFβ with both chemotherapy backbones in the clinical setting. Ongoing evaluation of samples from the stopped clinical trials will inform as to the role of TGFb in human PDAC. Citation Format: Li Qiang, Megan Hoffman, Lestat Ali, Jaime Ivan Castillo Silva, Lauren Kageler, Ayantu Temesgen, Patrick Lenehan, Jennifer Wang, Elisa Bello, Victoire Cardot-Ruffino, Giselle Uribe, Annan Yang, Michael Dougan, Andrew Aguirre, Srivatsan Raghavan5, Marc Pelletier, Viviana Cremasco, Stephanie Dougan. Transforming Growth Factor-β Blockade in Pancreatic Cancer Enhances Sensitivity to Combination Chemotherapy in Mice [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr C001.

Abstract A066: nSMase2-mediated exosome secretion shapes the tumor microenvironment to immunologically support pancreatic cancer

Abstract The pleiotropic roles of nSMase2-generated ceramide include regulation of intracellular ceramide signaling and exosome biogenesis. We investigated the effects of eliminating nSMase2 on early and advanced PDA, including its influence on the microenvironment. Employing the KPC mouse model of pancreatic cancer, we demonstrate that pancreatic epithelial nSMase2 ablation reduces neoplasia and promotes a PDA subtype switch from aggressive basal-like to classical. nSMase2 elimination prolongs survival of KPC mice, hinders vasculature development, and fosters a robust immune response. nSMase2 loss leads to recruitment of cytotoxic T cells, N1-like neutrophils, and abundant infiltration of anti-tumorigenic macrophages in the pancreatic preneoplastic microenvironment. Mechanistically, we demonstrate that nSMase2-expressing PDA cell small extracellular vesicles (sEVs) reduce survival of KPC mice; PDA cell sEVs generated independently of nSMase2 prolong survival of KPC mice and reprogram macrophages to a proinflammatory phenotype. Collectively, our study highlights previously unappreciated opposing roles for exosomes, based on biogenesis pathway, during PDA progression. Citation Format: Audrey M Hendley, Sudipta Ashe, Atsushi Urano, Martin Ng, Tuan A Phu, Xianlu Laura Peng, Changfei Luan, Anna-Marie Finger, Gun Ho Jang, Natanya R Kerper, David I Berrios, David Jin, Jonghyun Lee, Irene R Riahi, Oghenekevwe M Gbenedio, Christina Chung, Jeroen P Roose, Jen Jen Yeh, Steven Gallinger, Andrew V Biankin, Grainne M O’Kane, Vasilis Ntranos, David K Chang, David W Dawson, Grace E Kim, Valerie M Weaver, Robert L Raffai, Matthias Hebrok. nSMase2-mediated exosome secretion shapes the tumor microenvironment to immunologically support pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A066.

Abstract B041: Fibroblast STAT3 Signaling in Pancreatic Cancer: Implications for Immunotherapy and Tumor Microenvironment Modulation

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive cancer distinguished by a dense stromal microenvironment. The current investigation delves into the fibroblast-specific STAT3 signaling in the PDAC tumor microenvironment (TME) and its effects on immune cell infiltration and extracellular matrix (ECM) modulation. An association was identified between heightened levels of phosphorylated STAT3 (pSTAT3) in tumor-associated fibroblasts (CAFs) and a more unfavorable prognosis, particularly among male patients. Comparative analysis of gene expression patterns in normal fibroblasts and those deficient in STAT3, extracted from mouse models of pancreatic cancer, unveiled distinct gene expression profiles related to the inflammatory signaling pathway (IL6, ADM, CXCL11, CD80, VEGFA, TGFB1, STC1, STC2). Utilizing ChIP-Seq analysis with an anti-pSTAT3 antibody, it was confirmed that STAT3 binds to the regulatory regions of these genes, supporting its regulatory role in the inflammatory signaling pathway. Moreover, the absence of STAT3 in CAFs resulted in a decline in M2 macrophages and an upsurge in M1 macrophages within the TME, indicating a potential function of STAT3 in influencing macrophage polarization. Noteworthy observations also revealed that STAT3 signaling in CAFs impacts ECM remodeling, as illustrated by reduced collagen deposition. Furthermore, the deficiency of STAT3 in CAFs led to a notable reduction in the quantity of neutrophils within the TME in our mouse model. Additionally, a plausible involvement of STAT3 in neutrophil recruitment and activation through the CXCL1/CXCR2 axis was indicated. Alterations in the infiltration of neutrophils and macrophages due to the absence of STAT3 could profoundly influence the immune response against PDAC. The modified composition of ECM following STAT3 deletion might also impact the infiltration and functioning of immune cells. These outcomes underscore the significance of STAT3 signaling in CAFs as a promising therapeutic target in PDAC, offering potential for modulating the tumor microenvironment, augmenting anti-tumor immunity, and advancing patient outcomes. Citation Format: Samaneh Saberi, Cameron Bumbleburg, Caroline Everett, Julia E Lefler, Victoria Jordan, Lu Han, Sudarshana Sharma, Michael C. Ostrowski. Fibroblast STAT3 Signaling in Pancreatic Cancer: Implications for Immunotherapy and Tumor Microenvironment Modulation [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B041.

Abstract B031: Alternate pain management strategies: targeting pancreatic cancer-related pain with Flavonoid

Abstract Purpose For up to 90% of cancer patients, pain is a substantial symptom, contributing to morbidity and, possibly, mortality. In this study we sought to test the hypothesis that a flavonoid, derived from Cannabis, and previously shown to suppress cancer cell growth in vitro and tumor progression in mice in vivo, could also reduce the pain caused by pancreatic tumors in vivo. Methods A subcutaneous tumor in the dorsum of C57BL/6 mouse was generated by sub- dermal injection of KPC mouse pancreatic cancer cell line (ATCC). All the mice were video recorded to observe and analyze their spontaneous pain-related behaviors, including facial washing, dorsal hunching, dorsal skin twitch, licking of dorsal skin and overall locomotion around the cage. Two five minute recording sessions were analyzed by a blinded observer during times before the cancer cell injection (Baseline), after tumor growth, to about 8mm diameter (30d and 37d post-injection) and just before and then 24h after intra-tumor injection of the flavonoid ( Group A: FBL-03G, 1 nmole in 100µL) or Vehicle(100µL; Group B). Pain-related behavioral changes were averaged from the two recording sessions and analyzed from Baseline, just before FBL-03G and 24h after and compared between test compound-and vehicle injected groups (n=7 in each), using non-parametric statistics. Results Facial washing (events/5min) was: reduced to 87% of Baseline in tumor-bearing mice, then increased by 150% at 24h post-injection in Group A (FBL-03G); reduced to 65% of Baseline in tumor-bearing mice of Group B, then reduced a further 9% by Vehicle. Dorsal hunching, which events occur only after tumor growth (Baseline =0), was reduced at 24h by 62% in Group A but increased by 75% in Group B. Dorsal twitch, also absent before tumor growth, was reduced by 64% in Group A, but increased by 19% in Group B. Dorsal skin licking was reduced from Baseline by 33% in tumor-bearing mice of Group A, then increased by 76% after FBL-03g injection whereas licking, reduced by 22% by tumors in Group B and then by an additional 4% by vehicle (Group B). Tumors reduced locomotion by 58% in Group A, which was reversed by 32% after FBL-03g (Group A); vehicle injection resulted in a 38% reduction in locomotion (Group B). Thus, in 4 of the 5 pain-related behaviors from tumors quantitated symptoms were reversed by FBL-03G and oppositely affected, or not at all, by Vehicle. Conclusions These results indicate a substantial relief of spontaneous pain-related pain in this mouse model of pancreatic cancer. Such an acute effect occurs much more rapidly than the previously published in vitro suppression of KPC cell growth (assayed after 9-12d of treatment) or the reduction of tumor size (1-7 weeks) in vivo and suggest that the anti-hyperalgesic affect is due to mechanisms that do not involve reduction in tumor mass. The findings, using a single dose and a small number of mice, appear promising for further studies that examine a range of doses or FBL-03G and its actions in intra-pancreatic tumors. Citation Format: Sayeda Yasmin-Karim, Suheera Haq, Mike Makrigiorgos, Gary Strichartz, Geraud Richards, Natasha Rayman. Alternate pain management strategies: targeting pancreatic cancer-related pain with Flavonoid [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B031.

Abstract B004: Targeting Metabolic Vulnerabilities in Pancreatic Cancer: The Synergistic Anti- Tumor Effects of a Ketogenic Diet and IDH1 Inhibition

Abstract Background: A ketogenic diet, characterized by high fat (90% kCal) and low carbohydrates (5% kCal), induces ketosis. Previous research suggests that a ketogenic diet shifts cancer cell metabolism from glycolysis to the tricarboxylic acid (TCA) cycle in the mitochondria. While the overall impact on cancer growth remains uncertain, most studies suggest an anti-cancer effect. We hypothesize that understanding the molecular and biochemical adaptations of pancreatic cancer cells to a ketogenic diet will reveal metabolic vulnerabilities and lead to rational therapeutic strategies that complement the diet. Methods: Mice were fed a ketogenic diet, with control mice on a normal diet. Pancreatic cancer xenografts were monitored for growth and subjected to biochemical analyses. Tumor metabolites were measured using LC/GC-MS, and oxidative stress was assessed by NADP+/NADPH, NAD+/NADH levels, and lipid peroxidation assays. Enzyme expression was determined by Western blot. In vitro, mitochondrial function was analyzed using the Seahorse FX Analyzer, and cell growth was measured by PicoGreen DNA quantitation. Results: A ketogenic diet slowed pancreatic cancer growth in multiple in vivo models, including MIA-PaCa2, KPC, and patient-derived xenografts. TCA metabolites increased in xenografts exposed to the ketogenic diet, with a significant upregulation of TCA cycle-associated enzymes. Specifically, the expression of BDH1 (a ketolytic enzyme), IDH1 (supports antioxidant defense and mitochondrial function), and IDH2 (mitochondrial homolog of IDH1) were elevated. Conversely, the glycolytic enzyme G3P was reduced. Additionally, a ketogenic diet induced oxidative stress in pancreatic cancer in vivo. Combining a ketogenic diet with an IDH1 inhibitor impaired antioxidant defense and mitochondrial function, resulting in significantly reduced tumor proliferation compared to either treatment alone. In vitro studies indicated that low glucose and fatty acids primarily drive the reduced proliferation of pancreatic cancer cells on a ketogenic diet, while exogenous ketone bodies are utilized for energy under glucose deprivation, generally promoting tumor growth. Conclusion: A ketogenic diet exerts a strong anti-tumor effect in multiple pancreatic cancer mouse models, likely due to low glucose and increased fatty acid load. The metabolic shift towards an oxidative phosphorylation (OXPHOS) phenotype makes pancreatic cancer cells particularly vulnerable to antioxidant and mitochondrial inhibitors. Citation Format: Omid Hajihassani, Mehrdad Zarei, Soubhi Tahan, Peter Gallagher, William Beegan, John Speers, James Choi, Aya Murray, Alexander Loftus, Helen Cheng, Anusha Mudigonda, Karen Ji, Jonathan Hue, Hallie Graor, Jordan Winter. Targeting Metabolic Vulnerabilities in Pancreatic Cancer: The Synergistic Anti- Tumor Effects of a Ketogenic Diet and IDH1 Inhibition [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B004.

Abstract A006: Generation of a flexible pancreatic cancer mouse model via somatic tissue engineering

Abstract Pancreatic cancer, recognized as one of the most lethal cancer types, exhibits a dismal five-year survival rate below 12%. The genetic and epigenetic heterogeneity in human cancer poses a tremendous challenge in understanding disease mechanisms and developing targeted therapeutics. Current mouse models in the field can recapitulate some features of human pathophysiology but have limitations. While tumor formation in patients is focal, most genetically engineered mouse models (GEMMs) express oncogenes throughout the pancreas. GEMMs also have limited flexibility to manipulate gene expression and model the heterogeneity of human cancer. Additionally, GEMMs require significant time for crossing and tumor formation. Tumor transplantation models offer greater flexibility, however, they often require immunodeficient mice, which limits their applications in modeling the tumor microenvironment. To overcome the limits of current mouse models, we combined the piggyBac transposon system, CRISPR/Cas9, and in vivo electroporation to engineer somatic pancreatic tissue to induce focal tumor formation in immune-competent mice. With this approach, we engineered wildtype pancreata to express oncogenic KrasG12D while knocking out multiple tumor suppressors. We optimized this protocol by generating multiplexed gRNA constructs and cell type-specific promoters to improve reproducibility. Low-grade tumors formed within 10 days after electroporation, progressed to higher grade within a month, and metastasized to peritoneal organs during disease progression. Mouse tumors also recapitulated histopathological features in human patients. Lastly, we used our novel mouse model to study how mutant Kras disrupts post-transcriptional regulation by microRNA to drive pancreatic tumorigenesis. This model offers a powerful platform for manipulating gene expression and functionality to model cancer heterogeneity, ultimately advancing precision oncology's prospects for treating human patients. Citation Format: Ziyue (Zoey) Yang, Mara Cardenas, Akihiko Miyauchi, Caroline Snider, Ronald Parchem. Generation of a flexible pancreatic cancer mouse model via somatic tissue engineering [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A006.