Abstract 3435: Notch signaling and inflammation in a murine model of sarcoma-associated cachexia

Abstract

Abstract Introduction Cachexia is characterized by progressive muscle loss. Cancer-associated cachexia (CAC) affects over 50% of oncology patients, correlates with advanced disease stage, and shorter progression-free survival. TNF-α is implicated in CAC. TNF-α and klotho are antagonistic: decreased klotho causes susceptibility to inflammation. Notch receptors regulate muscle stem cell (MSC) function. Active Notch signaling inhibits differentiation and maintains MSC quiescence. Notch upregulates Pax7 expression, inhibiting MSC differentiation. Hes1 expression indicates active Notch signaling. Sarcomas are malignancies of the connective tissues. Sarcoma patients experience musculoskeletal impairment and are uniquely susceptible to cachexia. The mechanisms of sarcoma-associated cachexia (SAC) are unknown. Here we describe SAC in a mouse model of osteosarcoma (OS). Methods 1. Mouse Model of OS: 2 × 105 K7M2 OS cells were injected into right hindlimb tibias of 4-week old SCID/Beige mice. After 4 weeks, the right hindlimb was amputated to render mice free of primary but not metastatic disease. Mice were sacrificed 10 weeks post-injection. 2. Histology: Muscle tissues from the contralateral hindlimbs of tumor-bearing animals and the hindlimbs of control animals were harvested for histology. 3. Immunohistochemistry (IHC): Immunostains for Notch3 and Pax7 were performed on tumors, contralateral hindlimbs from tumor-bearing animals, and control hindlimbs. 4. RT-PCR: The expressions of TNF-α, klotho, and Hes1 were compared in tumor tissue, muscle from tumor-bearing animals, and control muscle. 5. Co-Culture:Muscle stem cells (MSCs) were co-cultured with other MSCs or OS cells in a transwell system. MSC differentiation was assessed with IHC for fast-type myosin heavy chain. Results 1. Atrophy was appreciated in uninjected hindlimbs of sarcoma-bearing animals compared with controls. 2. Muscle from sarcoma-bearing animals displayed smaller myofibers and greater fibrosis than controls, indicating atrophy and systemic inflammation. 3. IHC for Notch3 showed intratumoral positivity. Greater Notch3 positivity, Pax7 positivity, and colocalization were appreciated in atrophic muscle from experimental animals compared with control muscle. 4. There was greater TNF-α and less klotho expression in both tumors and skeletal muscle of sarcoma-bearing animals compared with normal muscle. Hes1 expression was greater in tumor and atrophic muscle than normal muscle. 5. Co-culture with OS cells resulted in decreased MSC differentiation. Co-culture in the presence of a Notch inhibitor rescued myogenic differentiation. Conclusion These data suggest that sarcoma may cause SAC via two mechanisms: (1)-TNF-α-mediated inflammation, and (2)-the dysregulation of MSC homeostatsis by aberrant Notch signaling. The inhibition of systemic inflammation and Notch signaling may represent novel strategies to combat SAC. Citation Format: Kurt R. Weiss, Xiaodong Mu, Rashmi Agarwal. Notch signaling and inflammation in a murine model of sarcoma-associated cachexia. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3435. doi:10.1158/1538-7445.AM2015-3435