Animal Models Of Cancer Cachexia Research Articles

Combination therapy of corylifol A and anamorelin attenuates cachexia by decreasing pro-inflammatory cytokines, inhibiting lipolysis and increasing food intake in a murine colon cancer model

PurposeIn our previous study, we found that corylifol A (CYA), an active component derived from Psoralea corylifolia L, exhibited ameliorating effects on cancer cachexia. Anamorelin (AML), a ghrelin receptor agonist, is the only approved drug for the clinical treatment of cancer cachexia. Here, we checked the combination of CYA and AML on muscle atrophy and fat lipolysis in cancer cachexia mice to explore the possible clinic therapeutic potency of the combination on cancer cachexia.MethodsC26 colon tumor-bearing mice, a well-established animal model of cancer cachexia, were used to evaluate the effects of combined therapy involving CYA and AML on cancer cachexia and compared with those of CYA or AML alone. Histological analysis was performed to assess the changes in gastrocnemius (GAS) muscle tissue and epididymal adipose tissue (eWAT) and the possible involved molecular pathways were explored using western blotting and ELISA.ResultsTreatments of CYA, AML, or CYA+AML all significantly ameliorated the body weight decrease, muscle atrophy as well as fat loss in the tumor-bearing mice. Notably, the ameliorating effects of CYA+AML on fat loss were more significant than CYA or AML alone. The eWAT weight of mice in healthy control, C26 model, C26 +CYA, C26 +AML or C26 +CYA +AML group was 0.48 ± 0.07, 0.21 ± 0.10, 0.26 ± 0.10, 0.37 ± 0.10 and 0.48 ± 0.18 g, respectively. CYA+AML could increase food intake, significantly decrease the serum levels of inflammatory cytokines (TNF-α and IL-6) and inhibit the activation (phosphorylation) of hormone-sensitive lipase (HSL) in the eWAT tissues of the C26 tumor-bearing mice. Treatment of CYA along could significantly decrease the serum TNF-α level of the C26 tumor-bearing mice. Treatment of AML alone could increase food intake but could not decrease the serum levels of inflammatory cytokines of the C26 tumor-bearing mice.ConclusionThese results suggested that the effects of CYA+AML include both the appetite-stimulating effects of AML and the anti-inflammatory effects of CYA. CYA+AML was better than CYA or AML alone in ameliorating cancer cachexia, especially fat loss, of C26 tumor-bearing mice and might be a novel and beneficial therapeutic option for cancer cachexia.

198. MODELING CANCER CACHEXIA USING PATIENT TUMOR ORGANOID-DERIVED XENOGRAFTS IN ESOPHAGEAL SQUAMOUS CELL CARCINOMA

Abstract Background Cancer cachexia is a metabolic and functional multifactorial body wasting syndrome and is one of the major causes of death in late-stage patients. Esophageal squamous cell carcinoma (ESCC) presents one of the highest morbidity and mortality rates due to cancer cachexia. However, there is limited effective and approved targeted treatment. Cancer cachexia animal models are highly limited, with few cancer types available as a pre-clinical drug testing platform and with none related to esophageal carcinoma. Methods We have developed a panel of ESCC patient-derived organoid xenograft (PDOX) mouse models based on our latest patient-derived organoid cultures from Hong Kong patients. Results Our PDOX mouse model displays cachexic phenotypes that traditional cell line-derived xenografts lack, including body wasting and functional signs. The slow-proliferating characteristic of the ESCC PDOX on mice allows development of cachexic conditions before the humane endpoint. Our comprehensive panel of PDOX lines allowed us to identify full spectrum and differential cachexia severities across mice bearing different PDOXs, indicating a heterogenous nature and allowing faithful mimicking of various patient conditions. Molecular analysis showed that several classic cachexia-related gene expressions were up-regulated in the isolated liver and inguinal adipose tissues of cachexic mice, along with elevated circulating Il-6 levels. Conclusion Our ESCC PDOX model potentially serves as a useful pre-clinical cachexia-targeted drug testing platform, providing more treatment options for ESCC patients for better disease management. Acknowledgments We acknowledge the Research Grants Council (Theme-based Research Scheme T12–701/17-R to MLL) and the Food and Health Bureau (Health and Medical Research Fund 06171566 to VZY) of the Hong Kong Special Administrative Region for funding support. We acknowledge DSMZ (German Collection of Microorganisms and Cell Culture) for the KYSE cell lines. We acknowledge the University of Hong Kong LKS Faculty of Medicine Centre for PanorOmic Sciences for providing imaging facilities.

Platelet status in cancer cachexia progression in ApcMin/+ mice.

Cachexia, a complex wasting syndrome, significantly affects the quality of life and treatment options for cancer patients. Studies have reported a strong correlation between high platelet count and decreased survival in cachectic individuals. Therefore, this study aimed to investigate the immunopathogenesis of cancer cachexia using the ApcMin/+ mouse model of spontaneous colorectal cancer. The research focused on identifying cellular elements in the blood at different stages of cancer cachexia, assessing inflammatory markers and fibrogenic factors in the skeletal muscle, and studying the behavioral and metabolic phenotype of ApcMin/+ mice at the pre-cachectic and severely cachectic stages. Platelet measurements were also obtained from other animal models of cancer cachexia - Lewis Lung Carcinoma and Colon 26 adenocarcinoma. Our study revealed that platelet number is elevated prior to cachexia development in ApcMin/+ mice and can become activated during its progression. We also observed increased expression of TGFβ2, TGFβ3, and SMAD3 in the skeletal muscle of pre-cachectic ApcMin/+ mice. In severely cachectic mice, we observed an increase in Ly6g, CD206, and IL-10 mRNA. Meanwhile, IL-1β gene expression was elevated in the pre-cachectic stage. Our behavioral and metabolic phenotyping results indicate that pre-cachectic ApcMin/+ mice exhibit decreased physical activity. Additionally, we found an increase in anemia at pre-cachectic and severely cachectic stages. These findings highlight the altered platelet status during early and late stages of cachexia and provide a basis for further investigation of platelets in the field of cancer cachexia.

Effects of S-pindolol in mouse pancreatic and lung cancer cachexia models.

It is known that S-pindolol attenuates muscle loss in animal models of cancer cachexia and sarcopenia. In cancer cachexia, it also significantly reduced mortality and improved cardiac function, which is strongly compromised in cachectic animals. Here, we tested 3mg/kg/day of S-pindolol in two murine cancer cachexia models: pancreatic cancer cachexia (KPC) and Lewis lung carcinoma (LLC). Treatment of mice with 3mg/kg/day of S-pindolol in KPC or LLC cancer cachexia models significantly attenuated the loss of body weight, including lean mass and muscle weights, leading to improved grip strength compared with placebo-treated mice. In the KPC model, treated mice lost less than half of the total weight lost by placebo (-0.9±1.0 vs. -2.2±1.4g for S-pindolol and placebo, respectively, P<0.05) and around a third of the lean mass lost by tumour-bearing controls (-0.4±1.0 vs. -1.5±1.5g for S-pindolol and placebo, respectively, P<0.05), whereas loss of fat mass was similar. In the LLC model, the gastrocnemius weight was higher in sham (108±16mg) and S-pindolol tumour-bearing (94±15mg) mice than that in placebo (83±12mg), whereas the soleus weight was only significantly higher in the S-pindolol-treated group (7.9±1.7mg) than that in placebo (6.5±0.9). Grip strength was significantly improved by S-pindolol treatment (110.8±16.2 vs. 93.9±17.1g for S-pindolol and placebo, respectively). A higher grip strength was observed in all groups; whereas S-pindolol-treated mice improved by 32.7±18.5g, tumour-bearing mice only show minimal improvements (7.3±19.4g, P<0.01). S-pindolol is an important candidate for clinical development in the treatment of cancer cachexia that strongly attenuates loss of body weight and lean body mass. This was also seen in the weight of individual muscles and resulted in higher grip strength.

Abstract 363: Modelling cancer cachexia using a comprehensive panel of heterogeneous esophageal squamous cell carcinoma xenografts for therapeutic application

Abstract Background: Esophageal squamous cell carcinoma (ESCC) is one of deadliest esophageal malignancies in Asia. Cancer cachexia is a multifactorial body wasting syndrome and is one of the major causes of death in late-stage patients (1). It is characterized by skeletal muscle loss and atrophy, adipose and body wasting, and metabolic dysregulations and stress (1). ESCC is one of the cancer types with the highest morbidity and mortality rates due to cancer cachexia (2). However, there are few effective and approved treatments targeting cancer cachexia. Currently, cancer cachexia animal models are highly limited with few cancer types available as a pre-clinical drug testing platform, with none related to esophageal carcinoma (3). Results: Here we have developed a panel of ESCC patient-derived organoid xenograft (PDOX) mouse model based on our latest patient-derived organoid cultures originating from Hong Kong patients. Our immunodeficient mouse-based PDOX model resembles different cachexic phenotypes that cell line-derived xenograft lacks, including body (skeletal muscle and adipose tissue) wasting and functional signs (e.g. lack of body movement, reduced muscle strength, and loss of appetite). The slow-proliferating characteristic of the ESCC PDOX allows development of a cachexic condition before humane endpoint. Furthermore, our full panel of PDOX lines allowed us to identify the full spectrum, differential cachexia severity across mice bearing different PDOXs, indicating a heterogenous nature. We have comprehensively characterized the cachexia phenotypes and performed proof-of-concept preclinical drug tests using established anti-cachexic drugs; treated PDOX-bearing mice have improved body condition and function compared to the control, which were further verified by histological examinations of tissue samples. Molecular analysis showed that several classic cachexia related gene expressions were down-regulated in isolated liver (Il1b) and inguinal adipose tissues (adipose triglyceride lipase). Conclusions: Our ESCC PDOX model potentially serves as a great pre-clinical cachexia-targeted drug testing platform, providing more treatment options for ESCC patients for better disease management. Acknowledgement: We acknowledge the Research Grants Council (TRS T12-701/17-R to MLL) and the Food and Health Bureau (HMRF 06171566 to VZY) of Hong Kong Special Administrative Region for funding supports. We acknowledge DSMZ for the KYSE cell lines. We acknowledge the HKUMed-CPOS for providing imaging facilities.

Molecular mechanisms of cancer cachexia-related loss of skeletal muscle mass: data analysis from preclinical and clinical studies.

Cancer cachexia is a systemic hypoanabolic and catabolic syndrome that diminishes the quality of life of cancer patients, decreases the efficiency of therapeutic strategies and ultimately contributes to decrease their lifespan. The depletion of skeletal muscle compartment, which represents the primary site of protein loss during cancer cachexia, is of very poor prognostic in cancer patients. In this review, we provide an extensive and comparative analysis of the molecular mechanisms involved in the regulation of skeletal muscle mass in human cachectic cancer patients and in animal models of cancer cachexia. We summarize data from preclinical and clinical studies investigating how the protein turnover is regulated in cachectic skeletal muscle and question to what extent the transcriptional and translational capacities, as well as the proteolytic capacity (ubiquitin-proteasome system, autophagy-lysosome system and calpains) of skeletal muscle are involved in the cachectic syndrome in human and animals. We also wonder how regulatory mechanisms such as insulin/IGF1-AKT-mTOR pathway, endoplasmic reticulum stress and unfolded protein response, oxidative stress, inflammation (cytokines and downstream IL1ß/TNFα-NF-κB and IL6-JAK-STAT3 pathways), TGF-ß signalling pathways (myostatin/activin A-SMAD2/3 and BMP-SMAD1/5/8 pathways), as well as glucocorticoid signalling, modulate skeletal muscle proteostasis in cachectic cancer patients and animals. Finally, a brief description of the effects of various therapeutic strategies in preclinical models is also provided. Differences in the molecular and biochemical responses of skeletal muscle to cancer cachexia between human and animals (protein turnover rates, regulation of ubiquitin-proteasome system and myostatin/activin A-SMAD2/3 signalling pathways) are highlighted and discussed. Identifying the various and intertwined mechanisms that are deregulated during cancer cachexia and understanding why they are decontrolled will provide therapeutic targets for the treatment of skeletal muscle wasting in cancer patients.

Fuzheng Xiaoai Decoction 1 ameliorated cancer cachexia-induced muscle atrophy via Akt-mTOR pathway

Ethnopharmacological relevanceFuzheng Xiaoai Decoction 1 (FZXAD1) is a clinical experience prescription for the treatment of cancer patients at an advanced stage. FZXAD1 has been used for more than 10 years in the clinic and can effectively improve the deficiency syndrome of cancer patients. However, its mechanisms need further clarification. Aim of the studyTo check the effects of FZXAD1 in colon 26 (C26) cancer cachexia mice and try to clarify the mechanisms of FZXAD1 in ameliorating cancer cachexia symptoms. Materials and methodsAn animal model of cancer cachexia was constructed with male BALB/c mice bearing C26 tumor cells. Food intake, body weight and tumor size were measured daily during the animal experiment. Tissue samples in different groups including tumor and gastrocnemius muscle, were dissected and weighed at the end of the assay. Serum biochemical indicators such as total protein (TP), glucose (GLU) and alkaline phosphatase (ALP) were also detected. Network pharmacology-based analysis predicted the possible targets and signaling pathways involved in the effects of FZXAD1 on cancer cachexia therapy. Western blotting assays of the gastrocnemius muscle tissues from C26 tumor-bearing mice were then used to confirm the predicted possible targets of FZXAD1. ResultsThe results of animal experiments showed that FZXAD1 could ameliorate cancer cachexia by alleviating the muscle wasting as well as kidney atrophy and increasing the body weight of cancer cachexia mice. AKT1, MTOR, MAPK3, HIF1A and MAPK1 were predicted as the core targets of FZXAD1. Western blotting confirmed the prediction that FZXAD1 increased the expression levels of phosphorylated Akt and mTOR in the muscle tissues. In addition, FZXAD1 treatment obviously ameliorated the increased levels of HIF-1α and phosphorylated Erk1/2 in C26 tumor-bearing mice. ConclusionFZXAD1 effectively ameliorated cancer cachexia in an animal model of mice, which is consistent with its efficacy in the treatment of cancer patients. The mechanisms of FZXAD1 might be mainly based on its alleviating effects on muscle atrophy by activating the Akt-mTOR pathway and thus helping to maintain body weight.

The atypical β-blocker S-oxprenolol reduces cachexia and improves survival in a rat cancer cachexia model.

Beta-blockers and selected stereoisomers of beta-blockers, like bisoprolol and S-pindolol (ACM-001), have been shown to be effective in preclinical cancer cachexia models. Here, we tested the efficacy of stereoisomers of oxprenolol in two preclinical models of cancer cachexia-the Yoshida AH-130 rat model and the Lewis lung carcinoma (LLC) mouse model. In the Yoshida AH130 hepatoma rat cancer cachexia model and compared with placebo, 50mg/kg/d S-oxprenolol (HR: 0.49, 95% CI: 0.28-0.85, P=0.012) was superior to 50mg/kg/d R-oxprenolol (HR: 0.83, 95% CI 0.38-1.45, P=0.51) in reducing mortality (=reaching ethical endpoints). Combination of the three doses (12.5, 25 and 50mg/kg/d) that had a significant effect on body weight loss in the S-oxprenolol groups vs the same combination of the R-oxprenolol groups lead to a significantly improved survival of S-oxprenolol vs R-oxprenolol (HR: 1.61, 95% CI: 1.08-2.39, P=0.0185). Interestingly, there is a clear dose dependency in S-oxprenolol-treated (5, 12.5, 25 and 50mg/kg/d) groups, which was not observed in groups treated with R-oxprenolol. A dose-dependent attenuation of weight and lean mass loss by S-oxprenolol was seen in the Yoshida rat model, whereas R-oxprenolol had only had a significant effect on fat mass. S-oxprenolol also non-significantly reduced weight loss in the LLC model and also improved muscle function (grip strength 428±25 and 539±37g/100g body weight for placebo and S-oxprenolol, respectively). However, there was only a minor effect on quality of life indicators food intake and spontaneous activity in the Yoshida model (25mg/kg/S-oxprenolol: 11.9±2.5g vs placebo: 4.9±0.8g, P=0.013 and also vs 25mg/kg/d R-oxprenolol: 7.5±2.6g, P=0.025). Both enantiomers had no effects on cardiac dimensions and function at the doses used in this study. Western blotting of proteins involved in the anabolic/catabolic homoeostasis suggest that anabolic signalling is persevered (IGF-1 receptor, Akt) and catabolic signalling is inhibited (FXBO-10, TRAF-6) by S-pindolol, but not he R-enantiomer. Expression of glucose transporters Glut1 and Glut 4 was similar in all groups, as was AMPK. S-oxprenolol is superior to R-oxprenolol in cancer cachexia animal models and shows promise for a human application in cancer cachexia.

C/EBPβ promotes the expression of atrophy-inducing factors by tumours and is a central regulator of cancer cachexia.

BackgroundCCAAT/enhancer‐binding protein β (C/EBPβ) is a transcription factor whose high expression in human cancers is associated with tumour aggressiveness and poor outcomes. Most advanced cancer patients will develop cachexia, characterized by loss of skeletal muscle mass. In response to secreted factors from cachexia‐inducing tumours, C/EBPβ is stimulated in muscle, leading to both myofibre atrophy and the inhibition of muscle regeneration. Involved in the regulation of immune responses, C/EBPβ induces the expression of many secreted factors, including cytokines. Because tumour‐secreted factors drive cachexia and aggressive tumours have higher expression of C/EBPβ, we examined a potential role for C/EBPβ in the expression of tumour‐derived cachexia‐inducing factors.MethodsWe used gain‐of‐function and loss‐of‐function approaches in vitro and in vivo to evaluate the role of tumour C/EBPβ expression on the secretion of cachexia‐inducing factors.ResultsWe report that C/EBPβ overexpression up‐regulates the expression of 260 secreted protein genes, resulting in a secretome that inhibits myogenic differentiation (−31%, P < 0.05) and myotube maturation [−38% (fusion index) and −25% (myotube diameter), P < 0.05]. We find that knockdown of C/EBPβ in cachexia‐inducing Lewis lung carcinoma cells restores myogenic differentiation (+25%, P < 0.0001) and myotube diameter (+90%, P < 0.0001) in conditioned medium experiments and, in vivo, prevents muscle wasting (−51% for small myofibres vs. controls, P < 0.01; +140% for large myofibres, P < 0.01). Conversely, overexpression of C/EBPβ in non‐cachectic tumours converts their secretome into a cachexia‐inducing one, resulting in reduced myotube diameter (−41%, P < 0.0001, EL4 model) and inhibition of differentiation in culture (−26%, P < 0.01, EL4 model) and muscle wasting in vivo (+98% small fibres, P < 0.001; −76% large fibres, P < 0.001). Comparison of the differently expressed transcripts coding for secreted proteins in C/EBPβ‐overexpressing myoblasts with the secretome from 27 different types of human cancers revealed ~18% similarity between C/EBPβ‐regulated secreted proteins and those secreted by highly cachectic tumours (brain, pancreatic, and stomach cancers). At the protein level, we identified 16 novel secreted factors that are present in human cancer secretomes and are up‐regulated by C/EBPβ. Of these, we tested the effect of three factors (SERPINF1, TNFRSF11B, and CD93) on myotubes and found that all had atrophic potential (−33 to −36% for myotube diameter, P < 0.01).ConclusionsWe find that C/EBPβ is necessary and sufficient to induce the secretion of cachexia‐inducing factors by cancer cells and loss of C/EBPβ in tumours attenuates muscle atrophy in an animal model of cancer cachexia. Our findings establish C/EBPβ as a central regulator of cancer cachexia and an important therapeutic target.

Phenotypic features of cancer cachexia-related loss of skeletal muscle mass and function: lessons from human and animal studies.

Cancer cachexia is a complex multi‐organ catabolic syndrome that reduces mobility, increases fatigue, decreases the efficiency of therapeutic strategies, diminishes the quality of life, and increases the mortality of cancer patients. This review provides an exhaustive and comprehensive analysis of cancer cachexia‐related phenotypic changes in skeletal muscle at both the cellular and subcellular levels in human cancer patients, as well as in animal models of cancer cachexia. Cancer cachexia is characterized by a major decrease in skeletal muscle mass in human and animals that depends on the severity of the disease/model and the localization of the tumour. It affects both type 1 and type 2 muscle fibres, even if some animal studies suggest that type 2 muscle fibres would be more prone to atrophy. Animal studies indicate an impairment in mitochondrial oxidative metabolism resulting from a decrease in mitochondrial content, an alteration in mitochondria morphology, and a reduction in mitochondrial metabolic fluxes. Immuno‐histological analyses in human and animal models also suggest that a faulty mechanism of skeletal muscle repair would contribute to muscle mass loss. An increase in collagen deposit, an accumulation of fat depot outside and inside the muscle fibre, and a disrupted contractile machinery structure are also phenotypic features that have been consistently reported in cachectic skeletal muscle. Muscle function is also profoundly altered during cancer cachexia with a strong reduction in skeletal muscle force. Even though the loss of skeletal muscle mass largely contributes to the loss of muscle function, other factors such as muscle–nerve interaction and calcium handling are probably involved in the decrease in muscle force. Longitudinal analyses of skeletal muscle mass by imaging technics and skeletal muscle force in cancer patients, but also in animal models of cancer cachexia, are necessary to determine the respective kinetics and functional involvements of these factors. Our analysis also emphasizes that measuring skeletal muscle force through standardized tests could provide a simple and robust mean to early diagnose cachexia in cancer patients. That would be of great benefit to cancer patient's quality of life and health care systems.

Cachexia induced by Yoshida ascites hepatoma in Wistar rats is not associated with inflammatory response in the spleen or brain

Recent data indicate that peripheral, as well as hypothalamic pro-inflammatory cytokines play an important role in the development of cancer cachexia. However, there are only a few studies simultaneously investigating the expression of inflammatory molecules in both the periphery and hypothalamic structures in animal models of cancer cachexia. Therefore, using the Yoshida ascites hepatoma rat's model of cancer cachexia we investigated the gene expression of inflammatory markers in the spleen along with the paraventricular and arcuate nuclei, two hypothalamic structures that are involved in regulating energy balance. In addition, we investigated the effect of intracerebroventricular administration of PS-1145 dihydrochloride (an Ikβ inhibitor) on the expression of selected inflammatory molecules in these hypothalamic nuclei and spleen. We observed significantly reduced food intake in tumor-bearing rats. Moreover, we found significantly decreased expression of IL-6 in the spleen as well as decreased NF-κB in the paraventricular nucleus of rats with Yoshida ascites hepatoma. Similarly, expression of TNF-α, IL-1β, NF-κB, and COX-2 in the arcuate nucleus was significantly reduced in tumor-bearing rats. Administration of PS-1145 dihydrochloride reduced only the gene expression of COX-2 in the hypothalamus. Based on our findings, we suggest that the growing Yoshida ascites hepatoma decreased food intake by mechanical compression of the gut and therefore this model is not suitable for investigation of the inflammation-related mechanisms of cancer cachexia development.

The Mechanical Stimulation of Myotubes Counteracts the Effects of Tumor-Derived Factors Through the Modulation of the Activin/Follistatin Ratio.

Activin negatively affects muscle fibers and progenitor cells in aging (sarcopenia) and in chronic diseases characterized by severe muscle wasting (cachexia). High circulating activin levels predict poor survival in cancer patients. However, the relative impact of activin in mediating muscle atrophy and hampered homeostasis is still unknown. To directly assess the involvement of activin, and its physiological inhibitor follistatin, in cancer-induced muscle atrophy, we cultured C2C12 myotubes in the absence or in the presence of a mechanical stretching stimulus and in the absence or presence of C26 tumor-derived factors (CM), so as to mimic the mechanical stimulation of exercise and cancer cachexia, respectively. We found that CM induces activin release by myotubes, further exacerbating the negative effects of tumor-derived factors. In addition, mechanical stimulation is sufficient to counteract the adverse tumor-induced effects on muscle cells, in association with an increased follistatin/activin ratio in the cell culture medium, indicating that myotubes actively release follistatin upon stretching. Recombinant follistatin counteracts tumor effects on myotubes exclusively by rescuing fusion index, suggesting that it is only partially responsible for the stretch-mediated rescue. Therefore, besides activin, other tumor-derived factors may play a significant role in mediating muscle atrophy. In addition to increasing follistatin secretion mechanical stimulation induces additional beneficial responses in myotubes. We propose that in animal models of cancer cachexia and in cancer patients purely mechanical stimuli play an important role in mediating the rescue of the muscle homeostasis reported upon exercise.

Abstract A017: Optimization of a cachexia animal model for efficacy evaluation of candidates

Abstract Cachexia-anorexia syndrome, which is called cancer cachexia, is a common and important indicator of cancer and occurs in 30% to 80% of cancer patients. Cancer cachexia is characterized by a severe progressive body wasting, accompanied by loss of adipose tissue and skeletal muscle mass. The presence of cachexia is associated with a worsened prognosis and contributes to up to 20% of cancer deaths. In most cases, cancer cachexia patients clinically show anorexia caused through a complex system of hormones and neuropeptides. To investigate efficacy in development of new medicine, an optimized animal model is essentially required. Although several cancer cachexia animal models have been established, we focused on developing a better animal model particularly for efficacy evaluation with advanced bioimaging evaluation methods. First of all, we confirmed that an appropriate mouse strain is CDF1 in comparison with DBA1 and Balb/c after xenografting of Colon26. As the second step, we checked cell numbers with pattern of tumor growth. In human clinics, cure of cancer cachexia is usually performed with surgical removal of tumor. To make the animal model similar to clinical circumstances, we tested recovery time and pattern after removal of tumor. With optimized mouse model we tested efficacy of anticancer drug Megace 200 mg/kg as a positive control. In addition, we applied advanced bioimaging equipment such as MRI and CT to visualize actual tumor and fat with body composition. We expect that optimized animal model of cancer cachexia will prepare the way for more accurate efficacy test in preclinical state to go to a clinical trial. Citation Format: Jiyeong Bae, Ig Jun Cho, Jung Hyu Shin, Hyung Soo Yuh, Hyung Ho Moon, Hye Lim Lee, Yeong Nang Jo, Yeon Ju Park, Woo Sung Choi, Byung Hwa Hyun. Optimization of a cachexia animal model for efficacy evaluation of candidates [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A017.

Early suppression of adipocyte lipid turnover induces immunometabolic modulation in cancer cachexia syndrome.

Cancer cachexia is a multifactorial syndrome characterized by body weight loss, atrophy of adipose tissue (AT) and systemic inflammation. However, there is limited information regarding the mechanisms of immunometabolic response in AT from cancer cachexia. Male Wistar rats were inoculated with 2 × 107 of Walker 256 tumor cells [tumor bearing (TB) rats]. The mesenteric AT (MeAT) was collected on d 0, 4, 7 (early stage), and 14 (cachexia stage) after tumor cell injection. Surgical biopsies for MeAT were obtained from patients who had gastrointestinal cancer with cachexia. Lipolysis showed an early decrease in glycerol release in TB d 4 (TB4) rats in relation to the control, followed by a 6-fold increase in TB14 rats, whereas de novo lipogenesis was markedly lower in the incorporation of glucose into fatty acids in TB14 rats during the development of cachexia. CD11b and CD68 were positive in TB7 and TB14 rats, respectively. In addition, we found cachexia stage results similar to those of animals in MeAT from patients: an increased presence of CD68+, iNOS2+, TNFα+, and HSL+ cells. In summary, translational analysis of MeAT from patients and an animal model of cancer cachexia enabled us to identify early disruption in Adl turnover and subsequent inflammatory response during the development of cancer cachexia.-Henriques, F. S., Sertié, R. A. L., Franco, F. O., Knobl, P., Neves, R. X., Andreotti, S., Lima, F. B., Guilherme, A., Seelaender, M., Batista, M. L., Jr. Early suppression of adipocyte lipid turnover induces immunometabolic modulation in cancer cachexia syndrome.

The rationale for preventing cancer cachexia: targeting excessive fatty acid oxidation.

Cachexia commonly occurs at the terminal stage of cancer and has largely unclear molecular mechanisms. A recent study published in Nature Medicine, entitled “Excessive fatty acid oxidation induces muscle atrophy in cancer cachexia,” reveals that cachectic cancer cells can secrete multiple cytokines that induce excessive fatty acid oxidation, which is responsible for muscle loss in cancer cachexia. Inhibition of fatty acid oxidation using etomoxir can increase muscle mass and body weight in cancer cachexia animal models. The usage of stable cachexia animal models is also discussed in this research highlight.

Cancer cachexia-when proteasomal inhibition is not enough.

Cachexia is a life threatening syndrome associated with several diseases, such as end‐stage heart failure, end‐stage renal disease, chronic obstructive pulmonary disease, chronic inflammation (i.e. rheumatoid arthritis), acquired immune deficiency syndrome, and cancer.1, 2 Cachexia is found in 31–87% of cancer patients especially in advanced disease stages.3 It is characterized by progressive weight loss, metabolic alterations, fatigue, and persistent reduction of body cell mass in response to a malignant tumour.2, 4, 5, 6 The incidence of cachexia in cancer patients is dependent on the type and site of the tumour. While patients with non‐Hodgkin's lymphoma, breast cancer, and sarcomas show low incidences, rates up to 83% in pancreatic cancer patients, and over 85% in patients with gastric cancer have been found. Additionally, around 60% of small‐cell and non‐small‐cell lung cancer patients develop cachexia.7, 8, 9 Cancer cachexia affects the function of several organs such as muscle, adipose tissue, liver, brain, immune system, and heart, collectively decreasing patients' quality of life and worsening their prognosis. Therefore, cachexia must be considered as a true multi‐organ syndrome.10 Because cancer cachexia leads to a decrease in physical performance and quality of life,11 and is associated with poor survival (accounting for more than 20% of cancer deaths,7, 12, 13, 14, 15) it is of major clinical relevance. Even more so since cachectic patients show lower response rates to chemotherapy7 and a reduced tolerance to anticancer treatment.16 Despite its importance, weight loss in cancer patients is rarely recognized, assessed,17 or treated actively.18, 19 Thus, cancer cachexia represents an important underappreciated clinical syndrome.

Diet composition as a source of variation in experimental animal models of cancer cachexia.

BackgroundA variety of experimental animal models are used extensively to study mechanisms underlying cancer cachexia, and to identify potential treatments. The important potential confounding effect of dietary composition and intake used in many preclinical studies of cancer cachexia is frequently overlooked. Dietary designs applied in experimental studies should maximize the applicability to human cancer cachexia, meeting the essential requirements of the species used in the study, matched between treatment and control groups as well as also being generally similar to human consumption.MethodsA literature review of scientific studies using animal models of cancer and cancer cachexia with dietary interventions was performed. Studies that investigated interventions using lipid sources were selected as the focus of discussion.ResultsThe search revealed a number of nutrient intervention studies (n = 44), with the majority including n‐3 fatty acids (n = 16), mainly eicosapentaenoic acid and/or docosahexaenoic acid. A review of the literature revealed that the majority of studies do not provide information about dietary design; food intake or pair‐feeding is rarely reported. Further, there is a lack of standardization in dietary design, content, source, and overall composition in animal models of cancer cachexia. A model is proposed with the intent of guiding dietary design in preclinical studies to enable comparisons of dietary treatments within the same study, translation across different study designs, as well as application to human nutrient intakes.ConclusionThe potential for experimental endpoints to be affected by variations in food intake, macronutrient content, and diet composition is likely. Diet content and composition should be reported, and food intake assessed. Minimum standards for diet definition in cachexia studies would improve reproducibility of pre‐clinical studies and aid the interpretation and translation of results to humans with cancer.

Abstract 1153: Effective treatment of cancer associated cachexia by AV-380, a GDF15 inhibitory antibody

Abstract Background: One of the most lethal and debilitating effects of cancer is the development of cancer-related anorexia-cachexia syndrome (CACS). It affects the majority of advanced cancer patients and is thought to be responsible for up to ∼30% of all cancer deaths. CACS is a complex metabolic syndrome associated with malnutrition and severe involuntary weight loss due to the loss of muscle and fat tissue, as well as the clinical manifestation of anemia, inflammation and suppression of immune functions. The precise molecular mechanisms responsible for cancer cachexia are poorly understood, thus limiting the development of effective therapeutics. Current evidence suggests that a pro-inflammatory state may be responsible for many of the symptoms associated with CACS. Growth Differentiation Factor 15 (GDF15) is a pro-inflammatory cytokine whose circulating levels are significantly increased in cachectic cancer patients and several animal models of cancer cachexia. Here we demonstrate that the inhibition of GDF15 function results in the compete reversion of the phenotypic and metabolic changes associated with CASC. Methods: Mice bearing HT-1080 human fibrosarcoma xenografts have increased plasma GDF15 levels and develop cachexia were used in this study. Tumor bearing animals were treated either with AV-380, a GDF15 neutralizing antibody or IgG1 control. The effect on body weight, muscle/fat mass and organ sizes were assessed. Metabolic changes induced by the treatment were measured by a comprehensive laboratory animal monitoring system (CLAMS). Results: Treatment with AV-380 restored body weight, muscle and fat mass as well as normal organ sizes. Analyses of the CLAMS data demonstrated that mice receiving AV-380 reversed the phenotypic and metabolic changes associated with the cachexia induced by this tumor model. Moreover, AV-380 treated mice showed an increase in locomotor activity and energy expenditure, achieving levels similar to non tumor bearing control animals. Conclusions: Inhibition of GDF15 function completely reverted body weight loss, restored normal body composition and triggered a catabolic to anabolic metabolic switch in this cancer cachexia model. The data highlights the therapeutic potential of the GDF15 inhibitory antibody AV-380 for the treatment for CACS. Citation Format: Lorena Lerner, Nianjun Tao, Brian Krieger, Richard Nicoletti, Bin Feng, Nesreen Ismail, William Winston, Yanyu Zhang, Jinwei Jiang, Solly Weiler, Jeno Gyuris. Effective treatment of cancer associated cachexia by AV-380, a GDF15 inhibitory antibody. [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 1153. doi:10.1158/1538-7445.AM2015-1153

Genome-wide identification of FoxO-dependent gene networks in skeletal muscle during C26 cancer cachexia.

BackgroundEvidence from cachectic cancer patients and animal models of cancer cachexia supports the involvement of Forkhead box O (FoxO) transcription factors in driving cancer-induced skeletal muscle wasting. However, the genome-wide gene networks and associated biological processes regulated by FoxO during cancer cachexia are unknown. We hypothesize that FoxO is a central upstream regulator of diverse gene networks in skeletal muscle during cancer that may act coordinately to promote the wasting phenotype.MethodsTo inhibit endogenous FoxO DNA-binding, we transduced limb and diaphragm muscles of mice with AAV9 containing the cDNA for a dominant negative (d.n.) FoxO protein (or GFP control). The d.n.FoxO construct consists of only the FoxO3a DNA-binding domain that is highly homologous to that of FoxO1 and FoxO4, and which outcompetes and blocks endogenous FoxO DNA binding. Mice were subsequently inoculated with Colon-26 (C26) cells and muscles harvested 26 days later.ResultsBlocking FoxO prevented C26-induced muscle fiber atrophy of both locomotor muscles and the diaphragm and significantly spared force deficits. This sparing of muscle size and function was associated with the differential regulation of 543 transcripts (out of 2,093) which changed in response to C26. Bioinformatics analysis of upregulated gene transcripts that required FoxO revealed enrichment of the proteasome, AP-1 and IL-6 pathways, and included several atrophy-related transcription factors, including Stat3, Fos, and Cebpb. FoxO was also necessary for the cancer-induced downregulation of several gene transcripts that were enriched for extracellular matrix and sarcomere protein-encoding genes. We validated these findings in limb muscles and the diaphragm through qRT-PCR, and further demonstrate that FoxO1 and/or FoxO3a are sufficient to increase Stat3, Fos, Cebpb, and the C/EBPβ target gene, Ubr2. Analysis of the Cebpb proximal promoter revealed two bona fide FoxO binding elements, which we further establish are necessary for Cebpb promoter activation in response to IL-6, a predominant cytokine in the C26 cancer model.ConclusionsThese findings provide new evidence that FoxO-dependent transcription is a central node controlling diverse gene networks in skeletal muscle during cancer cachexia, and identifies novel candidate genes and networks for further investigation as causative factors in cancer-induced wasting.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-997) contains supplementary material, which is available to authorized users.

New cancer cachexia rat model generated by implantation of a peritoneal dissemination-derived human stomach cancer cell line

Cancer cachexia (CC), a syndrome characterized by anorexia and body weight loss due to low fat-free mass levels, including reduced musculature, markedly worsens patient quality of life. Although stomach cancer patients have the highest incidence of cachexia, few experimental models for the study of stomach CC have been established. Herein, we developed stomach CC animal models using nude rats subcutaneously implanted with two novel cell lines, i.e., MKN45c185, established from the human stomach cancer cell line MKN-45, and 85As2, derived from peritoneal dissemination of orthotopically implanted MKN45c185 cells in mice. Both CC models showed marked weight loss, anorexia, reduced musculature and muscle strength, increased inflammatory markers, and low plasma albumin levels; however, CC developed earlier and was more severe in rats implanted with 85As2 than in those implanted with MKN45cl85. Moreover, human leukemia inhibitory factor (LIF), a known cachectic factor, and hypothalamic orexigenic peptide mRNA levels increased in the models, whereas hypothalamic anorexigenic peptide mRNA levels decreased. Surgical removal of the tumor not only abolished cachexia symptoms but also reduced plasma LIF levels to below detectable limits. Importantly, oral administration of rikkunshito, a traditional Japanese medicine, substantially ameliorated CC-related anorexia and body composition changes. In summary, our novel peritoneal dissemination-derived 85As2 rat model developed severe cachexia, possibly caused by LIF from cancer cells, that was ameliorated by rikkunshito. This model should provide a useful tool for further study into the mechanisms and treatment of stomach CC.