Skeletal muscle analysis of cancer patients reveals a potential role for carnosine in muscle wasting.

BACKGROUND: Muscle wasting during cancer cachexia is mediated by protein degradation via autophagy and ubiquitin-linked proteolysis. These processes are sensitive to changes in intracellular pH ([pH]i ) and reactive oxygen species, which in skeletal muscle are partly regulated by histidyl dipeptides, such as carnosine. These dipeptides, synthesized by the enzyme carnosine synthase (CARNS), remove lipid peroxidation-derived aldehydes, and buffer [pH]i . Nevertheless, their role in muscle wasting has not been studied. METHODS: Histidyl dipeptides in the rectus abdominis (RA) muscle and red blood cells (RBCs) of male and female controls (n = 37), weight stable (WS: n = 35), and weight losing (WL; n = 30) upper gastrointestinal cancer (UGIC) patients, were profiled by LC-MS/MS. Expression of enzymes and amino acid transporters, involved in carnosine homeostasis, was measured by Western blotting and RT-PCR. Skeletal muscle myotubes were treated with Lewis lung carcinoma conditioned medium (LLC CM), and ß-alanine to study the effects of enhancing carnosine production on muscle wasting. RESULTS: Carnosine was the predominant dipeptide present in the RA muscle. In controls, carnosine levels were higher in men (7.87 ± 1.98 nmol/mg tissue) compared with women (4.73 ± 1.26 nmol/mg tissue; P = 0.002). In men, carnosine was significantly reduced in both the WS (5.92 ± 2.04 nmol/mg tissue, P = 0.009) and WL (6.15 ± 1.90 nmol/mg tissue; P = 0.030) UGIC patients, compared with controls. In women, carnosine was decreased in the WL UGIC (3.42 ± 1.33 nmol/mg tissue; P = 0.050), compared with WS UGIC patients (4.58 ± 1.57 nmol/mg tissue), and controls (P = 0.025). Carnosine was significantly reduced in the combined WL UGIC patients (5.12 ± 2.15 nmol/mg tissue) compared with controls (6.21 ± 2.24 nmol/mg tissue; P = 0.045). Carnosine was also significantly reduced in the RBCs of WL UGIC patients (0.32 ± 0.24 pmol/mg protein), compared with controls (0.49 ± 0.31 pmol/mg protein, P = 0.037) and WS UGIC patients (0.51 ± 0.40 pmol/mg protein, P = 0.042). Depletion of carnosine diminished the aldehyde-removing ability in the muscle of WL UGIC patients. Carnosine levels were positively associated with decreases in skeletal muscle index in the WL UGIC patients. CARNS expression was decreased in the muscle of WL UGIC patients and myotubes treated with LLC-CM. Treatment with ß-alanine, a carnosine precursor, enhanced endogenous carnosine production and decreased ubiquitin-linked protein degradation in LLC-CM treated myotubes. CONCLUSIONS: Depletion of carnosine could contribute to muscle wasting in cancer patients by lowering the aldehyde quenching abilities. Synthesis of carnosine by CARNS in myotubes is particularly affected by tumour derived factors and could contribute to carnosine depletion in WL UGIC patients. Increasing carnosine in skeletal muscle may be an effective therapeutic intervention to prevent muscle wasting in cancer patients.

The Impact of Acute Systemic Inflammation Secondary to Oesophagectomy and Anastomotic Leak on Computed Tomography Body Composition Analyses.

This study aimed to longitudinally assess CT body composition analyses in patients who experienced anastomotic leak post-oesophagectomy. Consecutive patients, between 1 January 2012 and 1 January 2022 were identified from a prospectively maintained database. Changes in computed tomography (CT) body composition at the third lumbar vertebral level (remote from the site of complication) were assessed across four time points where available: staging, pre-operative/post-neoadjuvant treatment, post-leak, and late follow-up. A total of 20 patients (median 65 years, 90% male) were included, with a total of 66 computed tomography (CT) scans analysed. Of these, 16 underwent neoadjuvant chemo(radio)therapy prior to oesophagectomy. Skeletal muscle index (SMI) was significantly reduced following neoadjuvant treatment (p < 0.001). Following the inflammatory response associated with surgery and anastomotic leak, a decrease in SMI (mean difference: -4.23 cm2/m2, p < 0.001) was noted. Estimates of intramuscular and subcutaneous adipose tissue quantity conversely increased (both p < 0.001). Skeletal muscle density fell (mean difference: -5.42 HU, p = 0.049) while visceral and subcutaneous fat density were higher following anastomotic leak. Thus, all tissues trended towards the radiodensity of water. Although tissue radiodensity and subcutaneous fat area normalised on late follow-up scans, skeletal muscle index remained below pre-treatment levels.

Adipose depot gene expression and intelectin-1 in the metabolic response to cancer and cachexia.

BACKGROUND: Cancer cachexia is a poorly understood metabolic consequence of cancer. During cachexia, different adipose depots demonstrate differential wasting rates. Animal models suggest adipose tissue may be a key driver of muscle wasting through fat-muscle crosstalk, but human studies in this area are lacking. We performed global gene expression profiling of visceral (VAT) and subcutaneous (SAT) adipose from weight stable and cachectic cancer patients and healthy controls. METHODS: Cachexia was defined as >2% weight loss plus low computed tomography-muscularity. Biopsies of SAT and VAT were taken from patients undergoing resection for oesophago-gastric cancer, and healthy controls (n = 16 and 8 respectively). RNA was isolated and reverse transcribed. cDNA was hybridised to the Affymetrix Clariom S microarray and data analysed using R/Bioconductor. Differential expression of genes was assessed using empirical Bayes and moderated-t-statistic approaches. Category enrichment analysis was used with a tissue-specific background to examine the biological context of differentially expressed genes. Selected differentially regulated genes were validated by qPCR. Enzyme-linked immunosorbent assay (ELISA) for intelectin-1 was performed on all VAT samples. The previously-described cohort plus 12 additional patients from each group also had plasma I = intelectin-1 ELISA carried out. RESULTS: In VAT vs. SAT comparisons, there were 2101, 1722, and 1659 significantly regulated genes in the cachectic, weight stable, and control groups, respectively. There were 2200 significantly regulated genes from VAT in cachectic patients compared with controls. Genes involving inflammation were enriched in cancer and control VAT vs. SAT, although different genes contributed to enrichment in each group. Energy metabolism, fat browning (e.g. uncoupling protein 1), and adipogenesis genes were down-regulated in cancer VAT (P = 0.043, P = 5.4 × 10-6 and P = 1 × 10-6 respectively). The gene showing the largest difference in expression was ITLN1, the gene that encodes for intelectin-1 (false discovery rate-corrected P = 0.0001), a novel adipocytokine associated with weight loss in other contexts. CONCLUSIONS: SAT and VAT have unique gene expression signatures in cancer and cachexia. VAT is metabolically active in cancer, and intelectin-1 may be a target for therapeutic manipulation. VAT may play a fundamental role in cachexia, but the down-regulation of energy metabolism genes implies a limited role for fat browning in cachectic patients, in contrast to pre-clinical models.

Plasma Metabolomics Identifies Lipid and Amino Acid Markers of Weight Loss in Patients with Upper Gastrointestinal Cancer.

Cachexia is a multifactorial wasting syndrome associated with high morbidity and mortality in patients with cancer. Diagnosis can be difficult and, in the clinical situation, usually relies upon reported weight loss. The 'omics' technologies allow us the opportunity to study the end points of many biological processes. Among these, blood-based metabolomics is a promising method to investigate the pathophysiology of human cancer cachexia and identify candidate biomarkers. In this study, we performed liquid chromatography mass spectrometry (LC/MS)-based metabolomics to investigate the metabolic profile of cancer-associated weight loss. Non-selected patients undergoing surgery with curative intent for upper gastrointestinal cancer were recruited. Fasting plasma samples were taken at induction of anaesthesia. LC/MS analysis showed that 6 metabolites were highly discriminative of weight loss. Specifically, a combination profile of LysoPC 18.2, L-Proline, Hexadecanoic acid, Octadecanoic acid, Phenylalanine and LysoPC 16:1 showed close correlation for eight weight-losing samples (≥5% weight loss) and nine weight-stable samples (<5%weight loss) between predicted and actual weight change (r = 0.976, p = 0.0014). Overall, 40 metabolites were associated with ≥5% weight loss. This study provides biological validation of the consensus definition of cancer cachexia (Fearon et al.) and provides feasible candidate markers for further investigation in early diagnosis and the assessment of therapeutic intervention.

The relationship between muscle mass and function in cancer cachexia: smoke and mirrors?

PURPOSE OF REVIEW: Randomized clinical trials of cancer cachexia interventions are based on the premise that an increase in the muscle mass of patients is associated with consequent improvements in muscle function, and ultimately, quality of life. However, recent trials that have succeeded in demonstrating increases in lean body mass have been unable to show associated increases in patient physical function. In this review, we examine the potential causes for this lack of association between muscle mass and function in cancer cachexia, paying particular attention to those factors that may be at play when using body composition analysis techniques involving cross-sectional imaging. Moreover, we propose a new population-specific model for the relationship between muscle mass and physical function in patients with cancer cachexia. RECENT FINDINGS: The ROMANA 1 and 2 trials of anamorelin (a novel ghrelin agonist) and the POWER 1 and 2 trials of enobosarm (a selective androgen receptor modulator) were able to demonstrate improvements in patient lean body mass, but not the functional co-primary endpoints of handgrip strength and stair climb power, respectively. We report similar confirmatory findings in other studies, and describe potential reasons for these observations. SUMMARY: The relationship between muscle mass and muscle function is complex and unlikely to be linear. Furthermore, the relationship is influenced by the techniques used to assess nutritional endpoints [e.g. computed tomography (CT)]; the nature of the chosen physical function outcome measures; and the sex and severity of the recruited cachectic patients. Such factors need to be considered when designing intervention trials for cancer cachexia with functional endpoints.

The relationship between muscle protein content and CT-derived muscle radio-density in patients with upper GI cancer.

INTRODUCTION: Cancer cachexia is a multifactorial syndrome characterized by skeletal muscle loss. Cross-sectional analysis of CT scans is a recognized research method for assessing skeletal muscle volume. However, little is known about the relationship between CT-derived estimates of muscle radio-density (SMD) and muscle protein content. We assessed the relationship between CT-derived body composition variables and the protein content of muscle biopsies from cancer patients. METHODS: Rectus abdominis biopsies from cancer patients (n = 32) were analysed for protein content and correlated with phenotypic data gathered using CT body composition software. RESULTS: Skeletal muscle protein content varied widely between patients (median µg/mg wet weight = 89.3, range 70-141). There was a weak positive correlation between muscle protein content and SMD (r = 0.406, p = 0.021), and a weak positive correlation between protein content and percentage weight change (r = 0.416, p = 0.018). CONCLUSION: The protein content of skeletal muscle varies widely in cancer patients and cannot be accurately predicted by CT-derived muscle radio-density.