Cachexia is the unintentional loss of skeletal muscle and adipose tissue. Over 80% of patients with pancreatic adenocarcinoma cancer (PDAC) suffer from cachexia, resulting in reduced survival and quality of life. The magnitude of adipose loss is often greater than muscle loss in patients and animal models. Our prior work in a mouse model of orthotopic PDAC cachexia revealed alterations of multiple pathways in adipose, predominantly inflammation, cytokine signaling, and metabolism. Deconvolution analysis predicts immune cell infiltration and activation. Inflammation-stimulated lipolysis results in fat pad wasting, while the resulting myosteatosis promotes skeletal muscle catabolism. Given this essential role for fat loss in the pathophysiology of cancer cachexia, we aimed to characterize the adipose tissue microenvironment in early and late PDAC cachexia. Methods: Male C57BL/6J (N=30) mice underwent orthotopic injection of KPC32098 pancreatic cancer cells derived from the LSL-KrasG12D:LSL-Trp53R172H:Pdx1-Cre mouse model or sham surgery. Body composition was determined by serial EchoMRI. Tissues were harvested every three days starting on day 6 after injection until euthanasia on day 18. Bulk RNA sequencing of the entire epididymal fat pad was performed. Single nucleus RNA sequencing of pooled sham (n=4) and pooled day 12 KPC (n=4) epididymal WAT nuclei was performed to identify changes in the adipose tissue microenvironment during cachexia development. Results: Tumors were palpable on day 9 and increased in size from day 12 (0.612 ± 0.092g) to day 18 (1.79 ± 0.309g, P<0.05). Total body fat loss was evident as early as day 9 (P < 0.001), although epididymal and inguinal white adipose tissues (WAT) were consistently reduced only at days 15 (P=0 and 18. Intrascapular brown adipose tissue (BAT) was reduced by day 18. Histology of skin biopsies showed subcutaneous adipose was reduced by day 12 and absent by day 18, with no difference in epidermis, dermis, or muscle layer thickness. Bulk RNAseq of epididymal WAT in late PDAC cachexia suggested de-differentiation but not browning, including reduced expression of pro-WAT genes Sfrp5, Slc2a4, and Fasn as well as increased expression of the anti-WAT marker Vdr. PPARg targets and other pro-BAT markers trended lower in expression. Early analysis of the snRNAseq data revealed cells positive for Krt18 and Pdx1 in KPC but not sham samples, suggesting that tumor cells are present in the epididymal WAT. Conclusions: Loss of percentage body fat precedes detectable wasting of individual fat pads in this model of PDAC cachexia. Subcutaneous adipose wasting contributes to this generalized loss of adiposity. Gene expression in late cachexia in orthotopic PDAC suggests de-differentiation but not browning of WAT. Visceral WAT represents a site of tumor micrometastasis, whether as an artifact of the experimental system or as a genuine pathophysiological mechanism. Citation Format: Sephora Jean, Brittany R. Counts, Anne E. Gowan, Samuel A. Philleo, Sha Cao, Leonidas Koniaris, Teresa A. Zimmers. Changes in adipose tissue microenvironment occurs early in murine models of pancreatic cancer cachexia [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 360.

Cachexia is the involuntary wasting of skeletal muscle and adipose tissue, affecting over 80% of patients with pancreatic ductal adenocarcinoma (PDAC). Gemcitabine and nab-paclitaxel (GemNP) combination are commonly given as first-line treatments for PDAC. Data from our lab showed GemNP reduced tumor growth and prevented muscle and fat wasting. Dimethylaminoparthenolide (DMAPT), a small molecule inhibitor, prevented muscle wasting and prolonged survival in a genetic murine model of breast cancer. It has yet to determine if GemNP in combination with DMAPT can improve indices of cachexia and overall survival. Therefore, I hypothesize that GemNP in combination with DMAPT will improve indices of cachexia and overall survival in a murine model of pancreatic cancer. Male 10-week-old C57BL/6J mice underwent orthotopic injection of 5x104 KPC cells or SHAM surgery. Mice were randomly assigned to 1 of 4 groups (N=10/group): SHAM+Vehicle, KPC+Vehicle, KPC+GemNP, KPC+GemNP+DMAPT. Gemcitabine (120mg/kg) and Nab-paclitaxel (10mg/kg) were injected intraperitoneally starting on day 4 and continued every 6 days. DMAPT (100mg/kg) was administered by gavage Monday-Friday. To determine the role of GemNP and DMAPT in vitro, KPC cells were treated with GemNP and/or DMAPT and cell viability evaluated. Additionally, we treated myotubes with GemNP with/without DMAPT to assess myotube diameter in an established KPC conditioned media. In KPC mice, GemNP (24.4days) increased survival compared to Vehicle (16.8days, p=.0007). However, the combination GemNP+DMAPT did not extend survival over GemNP alone (25.4days, p=.693). Tumor mass was similar between all groups (p=0.411). Interestingly, at the time of sacrifice, all KPC treated mice independent of treatment had similar reduction in adipose tissue and muscle mass compared to SHAM. In conclusion, the addition of DMAPT to GemNP did not extend survival over GemNP alone in an aggressive pancreatic tumor cell line. Future studies should determine if less aggressive tumor cell lines might benefit from GemNP and DMAPT.

Background: Cachexia is the progressive, unintentional wasting of muscle and adipose tissue caused by tumors. Cancer cachexia affects >80% of patients with pancreatic ductal adenocarcinoma (PDAC). Certain chemotherapy regimens cause debilitation in patients and muscle and fat wasting in mouse models. Gemcitabine with nab-paclitaxel (GemNP) is a first line treatment for PDAC. Whether GemNP promotes cachexia is unknown. Muscle wasting in cancer cachexia has been functionally linked to NF-κB, which can be induced by chemotherapies and is active in PDAC tumors. Among other functions, NF-κB induces expression of Interleukin-6 (IL-6), a major mediator of cancer cachexia. DMAPT is a small molecule NF-κB inhibitor. In prior work, DMAPT plus Gem reduced tumor burden in a genetic model of pancreatic cancer and separately, prevented muscle wasting in a genetic model of mammary cancer. Therefore, the purpose of our study was to determine whether GemNP promotes cachexia and whether DMAPT could reduce tumor burden and/or cachexia in PDAC, potentially through reducing IL-6 expression. Methods: In our in vitro model, conditioned media (CM) from KPC pancreatic cancer cells derived from the KrasG12D;Trp53R172H;Pdx1-Cre mouse, induces wasting of C2C12 myotubes. Thus, myotubes were treated with 50% KPC-CM or control CM and supplemented with 0, 5, or 10uM DMAPT for 48hrs. For in vivo modeling of PDAC cachexia, 12-week-old male C57BL/6J mice were orthotopically implanted with 5x104 KPC cells or KPC cells deleted for IL6 (KPC-IL6KO) cells; controls underwent sham surgery. Starting on day 4, every 6 days tumor-bearing mice were given 120 mg/kg gemcitabine and 10 mg/kg nab-paclitaxel (GemNP) for four rounds, then gemcitabine only for subsequent rounds, with or without 100mg/kg DMAPT (by gavage 5 days/week). Controls received vehicle only. Results: DMAPT prevented atrophy in KPC-CM treated myotubes. In vivo, GemNP increased median survival, to 23.5d in KPC + GemNP mice versus 17d in untreated KPC mice (P<0.001). GemNP + DMAPT increased survival (25d) compared to untreated KPC (P<0.001), but was similar to KPC + GemNP alone (P=0.693). There were no differences in tumor mass at euthanasia. All KPC groups lost significant body weight and fat mass compared to sham controls. In mice with KPC IL-6KO tumors, GemNP increased survival, with median survival of 39d in KPC-IL6KO + GemNP mice compared with 30d in untreated KPC-IL6KO (P<0.001). GemNP + DMAPT increased survival (35 d in KPC IL-6KO) compared to untreated KPC-IL6KO (P<0.001), but was similar to KPC IL-6KO + GemNP (P=0.276). There was a trend for GemNP + DMAPT to have smaller KPC-IL6KO tumors compared to GemNP (p=0.055). Conclusion: Regardless of IL-6 expression from tumor cells, treatment with GemNP improved survival in orthotopic PDAC without exacerbating cachexia. DMAPT did not extend survival above GemNP alone. Despite promising in vitro data and prior published work, DMAPT was not efficacious in reducing tumor growth or treating PDAC cachexia in this model. Citation Format: Brittany Counts, Sephora Jean, Reggie Wang, Teresa Zimmers. Gemcitabine nab-paclitaxel combination with DMAPT in PDAC cachexia [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 369.