Dataset Information

Establishment and characterization of a novel murine model of pancreatic cancer cachexia.

ABSTRACT:

Background

Cachexia is a complex metabolic and behavioural syndrome lacking effective therapies. Pancreatic ductal adenocarcinoma (PDAC) is one of the most important conditions associated with cachexia, with >80% of PDAC patients suffering from the condition. To establish the cardinal features of a murine model of PDAC-associated cachexia, we characterized the effects of implanting a pancreatic tumour cell line from a syngeneic C57BL/6 KRAS^G12D P53^R172H Pdx-Cre^+/+ (KPC) mouse.

Methods

Male and female C57BL/6 mice were inoculated subcutaneously, intraperitoneally, or orthotopically with KPC tumour cells. We performed rigorous phenotypic, metabolic, and behavioural analysis of animals over the course of tumour development.

Results

All routes of administration produced rapidly growing tumours histologically consistent with moderate to poorly differentiated PDAC. The phenotype of this model was dependent on route of administration, with orthotopic and intraperitoneal implantation inducing more severe cachexia than subcutaneous implantation. KPC tumour growth decreased food intake, decreased adiposity and lean body mass, and decreased locomotor activity. Muscle catabolism was observed in both skeletal and cardiac muscles, but the dominant catabolic pathway differed between these tissues. The wasting syndrome in this model was accompanied by hypothalamic inflammation, progressively decreasing brown and white adipose tissue uncoupling protein 1 (Ucp1) expression, and increased peripheral inflammation. Haematological and endocrine abnormalities included neutrophil-dominant leukocytosis and anaemia, and decreased serum testosterone.

Conclusions

Syngeneic KPC allografts are a robust model for studying cachexia, which recapitulate key features of the PDAC disease process and induce a wide array of cachexia manifestations. This model is therefore ideally suited for future studies exploring the physiological systems involved in cachexia and for preclinical studies of novel therapies.

SUBMITTER: Michaelis KA

PROVIDER: S-EPMC5659050 | biostudies-literature | 2017 Oct

REPOSITORIES: biostudies-literature

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Publications

Establishment and characterization of a novel murine model of pancreatic cancer cachexia.

Michaelis Katherine A KA Zhu Xinxia X Burfeind Kevin G KG Krasnow Stephanie M SM Levasseur Peter R PR Morgan Terry K TK Marks Daniel L DL

Journal of cachexia, sarcopenia and muscle 20170720 5

<h4>Background</h4>Cachexia is a complex metabolic and behavioural syndrome lacking effective therapies. Pancreatic ductal adenocarcinoma (PDAC) is one of the most important conditions associated with cachexia, with >80% of PDAC patients suffering from the condition. To establish the cardinal features of a murine model of PDAC-associated cachexia, we characterized the effects of implanting a pancreatic tumour cell line from a syngeneic C57BL/6 KRAS<sup>G12D</sup> P53<sup>R172H</sup> Pdx-Cre<sup> ...[more]

PMID: 28730707

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Project description:BackgroundThe majority of patients with pancreatic cancer develops cachexia. The mechanisms underlying cancer cachexia development and progression remain elusive, although tumour-derived factors are considered to play a major role. Pancreatic tumour organoids are in vitro three-dimensional organ-like structures that retain many pathophysiological characteristics of the in vivo tumour. We aimed to establish a pancreatic tumour organoid biobank from well-phenotyped cachectic and non-cachectic patients to enable identification of tumour-derived factors driving cancer cachexia.MethodsOrganoids were generated from tumour tissue of eight pancreatic cancer patients. A comprehensive pre-operative patient assessment of cachexia-related parameters including nutritional status, physical performance, body composition, and inflammation was performed. Tumour-related and cachexia-related characteristics of the organoids were analysed using histological stainings, targeted sequencing, and real-time-quantitative PCR. Cachexia-related factors present in the circulation of the patients and in the tumour organoid secretome were analysed by enzyme-linked immunosorbent assay.ResultsThe established human pancreatic tumour organoids presented typical features of malignancy corresponding to the primary tumour (i.e. nuclear enlargement, multiple nucleoli, mitosis, apoptosis, and mutated KRAS and/or TP53). These tumour organoids also expressed variable levels of many known cachexia-related genes including interleukin-6 (IL-6), TNF-α, IL-8, IL-1α, IL-1β, Mcp-1, GDF15, and LIF. mRNA expression of IL-1α and IL-1β was significantly reduced in organoids from cachectic vs. non-cachectic patients (IL-1α: -3.8-fold, P = 0.009, and IL-1β: -4.7-fold, P = 0.004). LIF, IL-8, and GDF15 mRNA expression levels were significantly higher in organoids from cachectic vs. non-cachectic patients (LIF: 1.6-fold, P = 0.003; IL-8: 1.4-fold, P = 0.01; GDF15: 2.3-fold, P < 0.001). In line with the GDF15 and IL-8 mRNA expression levels, tumour organoids from cachectic patients secreted more GDF15 and IL-8 compared with organoids from non-cachectic patients (5.4 vs. 1.5 ng/mL, P = 0.01, and 7.4 vs. 1.3 ng/mL, P = 0.07, respectively).ConclusionsThis novel human pancreatic tumour organoid biobank provides a valuable tool to increase our understanding of the mechanisms driving cancer cachexia. Our preliminary characterization of the secretome of these organoids supports their application in functional studies including conditioned medium approaches and in vivo transplantation models.

Project description:IntroductionCancer cachexia, highly prevalent in lung cancer, is a debilitating syndrome characterized by involuntary loss of skeletal muscle mass and is associated with poor clinical outcome, decreased survival and negative impact on tumour therapy. Various lung tumour-bearing animal models have been used to explore underlying mechanisms of cancer cachexia. However, these models do not simulate anatomical and immunological features key to lung cancer and associated muscle wasting. Overcoming these shortcomings is essential to translate experimental findings into the clinic. We therefore evaluated whether a syngeneic, orthotopic lung cancer mouse model replicates systemic and muscle-specific alterations associated with human lung cancer cachexia.MethodsImmune competent, 11 weeks old male 129S2/Sv mice, were randomly allocated to either (1) sham control group or (2) tumour-bearing group. Syngeneic lung epithelium-derived adenocarcinoma cells (K-rasG12D ; p53R172HΔG ) were inoculated intrapulmonary into the left lung lobe of the mice. Body weight and food intake were measured daily. At baseline and weekly after surgery, grip strength was measured and tumour growth and muscle volume were assessed using micro cone beam CT imaging. After reaching predefined surrogate survival endpoint, animals were euthanized, and skeletal muscles of the lower hind limbs were collected for biochemical analysis.ResultsTwo-third of the tumour-bearing mice developed cachexia based on predefined criteria. Final body weight (-13.7 ± 5.7%; P < 0.01), muscle mass (-13.8 ± 8.1%; P < 0.01) and muscle strength (-25.5 ± 10.5%; P < 0.001) were reduced in cachectic mice compared with sham controls and median survival time post-surgery was 33.5 days until humane endpoint. Markers for proteolysis, both ubiquitin proteasome system (Fbxo32 and Trim63) and autophagy-lysosomal pathway (Gabarapl1 and Bnip3), were significantly upregulated, whereas markers for protein synthesis (relative phosphorylation of Akt, S6 and 4E-BP1) were significantly decreased in the skeletal muscle of cachectic mice compared with control. The cachectic mice exhibited increased pentraxin-2 (P < 0.001) and CXCL1/KC (P < 0.01) expression levels in blood plasma and increased mRNA expression of IκBα (P < 0.05) in skeletal muscle, indicative for the presence of systemic inflammation. Strikingly, RNA sequencing, pathway enrichment and miRNA expression analyses of mouse skeletal muscle strongly mirrored alterations observed in muscle biopsies of patients with lung cancer cachexia.ConclusionsWe developed an orthotopic model of lung cancer cachexia in immune competent mice. Because this model simulates key aspects specific to cachexia in lung cancer patients, it is highly suitable to further investigate the underlying mechanisms of lung cancer cachexia and to test the efficacy of novel intervention strategies.

Project description:Pancreatic cancer (PC) induced cachexia is a complex metabolic syndrome associated with significantly increased morbidity and mortality and reduced quality of life. The pathophysiology of cachexia is complex and poorly understood. Many molecular signaling pathways are involved in PC and cachexia. Though our understanding of cancer cachexia is growing, therapeutic options remain limited. Thus, further discovery and investigation of the molecular signaling pathways involved in the pathophysiology of cachexia can be applied to development of targeted therapies. This review focuses on three main pathophysiologic processes implicated in the development and progression of cachexia in PC, as well as their utility in the discovery of novel targeted therapies. Skeletal muscle wasting is the most prominent pathophysiologic anomaly in cachectic patients and driven by multiple regulatory pathways. Several known molecular pathways that mediate muscle wasting and cachexia include transforming growth factor-beta (TGF-?), myostatin and activin, IGF-1/PI3K/AKT, and JAK-STAT signaling. TGF-? antagonism in cachectic mice reduces skeletal muscle catabolism and weight loss, while improving overall survival. Myostatin/activin inhibition has a great therapeutic potential since it plays an essential role in skeletal muscle regulation. Overexpression of insulin-like growth factor binding protein-3 (IGFBP-3) leads to increased ubiquitination associated proteolysis, inhibition of myogenesis, and decreased muscle mass in PC induced cachexia. IGFBP-3 antagonism alleviates muscle cell wasting. Another component of cachexia is profound systemic inflammation driven by pro-cachectic cytokines such as interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-?), and interferon gamma (INF-?). IL-6 antagonism has been shown to reduce inflammation, reduce skeletal muscle loss, and ameliorate cachexia. While TNF-? inhibitors are clinically available, blocking TNF-? signaling is not effective in the treatment of cancer cachexia. Blocking the synthesis or action of acute phase reactants and cytokines is a feasible therapeutic strategy, but no anti-cytokine therapies are currently approved for use in PC. Metabolic alterations such as increased energy expenditure and gluconeogenesis, insulin resistance, fat tissue browning, excessive oxidative stress, and proteolysis with amino acid mobilization support tumor growth and the development of cachexia. Current innovative nutritional strategies for cachexia management include ketogenic diet, utilization of natural compounds such as silibinin, and supplementation with ?3-polyunsaturated fatty acids. Elevated ketone bodies exhibit an anticancer and anticachectic effect. Silibinin has been shown to inhibit growth of PC cells, induce metabolic alterations, and reduce myofiber degradation. Consumption of ?3-polyunsaturated fatty acids has been shown to significantly decrease resting energy expenditure and regulate metabolic dysfunction.

Dataset Information

Establishment and characterization of a novel murine model of pancreatic cancer cachexia.

Background

Methods

Results

Conclusions

Publications

Establishment and characterization of a novel murine model of pancreatic cancer cachexia.

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