BACKGROUND:Sufficient high-quality protein intake is required to prevent sarcopenia in older adults. Plant-based proteins have been reported to have lower anabolic properties when compared with animal-based proteins. Whether the lower quality of plant-based protein can be improved, thereby resulting in an anabolic response noninferior to an equivalent amount of animal-based protein, remains to be established in older adults. OBJECTIVES:To compare postprandial muscle protein synthesis rates following ingestion of a single bolus of a soy- and pea-derived protein blend (PLANT), with a soy-pea protein blend fortified with free leucine (PLANT+LEU), or whey protein (WHEY) in older males. METHODS:In this randomly assigned, double-blind, parallel-group design, 45 healthy older males [aged 69 ± 5 y, BMI (in kg·m‒2) 26.2 ± 3.2] were selected to ingest a 20 g protein blend combining 12 g soy plus 8 g pea protein (PLANT), 20 g of the soy-pea protein blend fortified with 2 g leucine (PLANT+LEU), or 20 g WHEY. Primed continuous L-[ring-13C6]-phenylalanine infusions were applied, with blood and muscle sampling ≤4 h after protein ingestion to assess plasma amino acid profiles and muscle protein synthesis rates. RESULTS:WHEY increased plasma essential amino acid concentrations more than PLANT and PLANT+LEU over the 4 h postprandial period (Incremental area under the curve: 135 ± 20 compared with 99 ± 21 compared with 105 ± 21 mmol·240 min·L‒1, respectively; P < 0.001). Plasma peak leucine concentrations were higher after PLANT+LEU ingestion compared with PLANT and WHEY (567 ± 74 μmol·L‒1 compared with 310 ± 49 μmol·L‒1 compared with 471 ± 74 μmol·L‒1, respectively; P < 0.001). Postprandial muscle protein synthesis rates averaged 0.034 ± 0.010 %·h‒1, 0.035 ± 0.012 %·h‒1, and 0.034 ± 0.010 %·h‒1 following PLANT, PLANT+LEU, and WHEY ingestion, respectively (treatment P = 0.828), and were not increased when compared with postabsorptive values. CONCLUSIONS:Ingestion of 20 g of protein alone, independent of its quality, is not enough to increase muscle protein synthesis rates in older males. More work is needed to define the preferred combination of both protein quality and quantity to stimulate muscle protein synthesis in an older population. This trial was registered at clinicaltrials.gov as NCT05711095.

Several studies demonstrated curative responses of combined radio-immunotherapy, although not standard-of-care for most cancers. Increased fiber intake has been associated with improved radiotherapy and immunotherapy outcomes, but fiber compositions’ impact remains unclear. This study aimed to explore whether dietary fiber composition influences the therapeutic outcome of combined radio-immunotherapy in a preclinical cancer model. A syngeneic mouse model of colon cancer (CT26) (female BALB/cOla Hsd) was used. Mice were randomized into three groups ( n = 12) of iso-caloric diets with different fiber compositions. Five instances of local radiotherapy on tumors, combined with injections of anti-PD-L1, were administered over 5 and 10 days. Diets’ impact was assessed on progression-free survival, SCFA levels in fecal and cecal samples, gut microbiome composition, and immunological profile. Progression-free survival was different between compositions, as well as their gut microbiota community structure, at all measured time-points. Therapeutic outcome (cure) was negatively associated with the relative abundance of Bacteroides and positively with Atopobiaceae Family . There was no association with SCFA levels. Cured mice displayed smaller spleens containing increased proportions of CD8+ T-cells and decreased proportions of myeloid-derived suppressor cells. Our data suggest that fiber composition may influence therapeutic outcome of combined radio-immunotherapy treatment in vivo.

Background: Emerging evidence suggests a positive impact of long-chain polyunsaturated fatty acids (lc-PUFAs) on radiotherapy and anti-PD-L1 efficacy, however whether this translates into better outcomes in multimodality combination treatments remains unclear. We hypothesized that dietary lc-PUFAs can improve the outcome of RT/IT combination treatment. Methods: Effects of different lc-PUFAs on CT26 surface PDL-1 expression were measured in vitro in absence or presence of radiotherapy using flow cytometry. Immunocompetent BALB/cOlaHsd mice, inoculated with CT26 cells, were randomized across different isocaloric AIN93M-based diets enriched with either eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) rich fish oil, arachidonic acid (ARA) rich ARASCO oil or combination of both. When tumors reached ±200 mm3, irradiation (2.33 Gy, QD5) was started. anti-PD-L1 (10 mg/kg, i.p., Q2Dx5) treatment started 1 day later. Study endpoint was defined as time to reach four times starting volume. Effects on splenic cytotoxic CD8+ T-cell number and activity was measured by flow cytometry. Results: All lc-PUFAs tested and irradiation, upregulated PD-L1 expression in vitro with a stronger increase when combined. n-3 lc-PUFA-rich fish oil administration may support responsiveness to combined RT/anti-PD-L1 therapy, compared to control diet based on soybean oil, although borderline significant (hazard ratio 0.35 [0.11–1.02], p = 0.053). No effects of the oil blends were observed on tumor take, body weight, food intake or activation of splenic cytotoxic CD8+ T-cells. Conclusion: A modest reduction in the risk of local tumor failure was observed upon n-3 lc-PUFA-rich fish oil administration, highlighting a need for further research.

Dietary β-glucan has the potential to selectively counteract LPS-induced immunosuppression.

To meet the global dietary protein demands, a trend towards plant-based protein (PBP) sources to replace animal-derived protein is currently ongoing. However, PBPs may not have the same anabolic capacity to stimulate muscle protein synthesis (MPS) as dairy proteins. For vulnerable populations with specific medical needs, it is especially important to validate the anabolic properties of PBPs. In this study, a blend of pea and soy protein isolate, with or without additional leucine, was compared to whey protein isolate on MPS in aged mice. 25-Months aged C57BL/6J-mice received an oral gavage with 70 mg of whey protein isolate (W), PS protein isolate (PS; ratio 51:49), PS fortified with 19

Beta-glucans and arabinoxylans are known for their immunostimulatory properties. However, in vivo these have been documented almost exclusively following parenteral administration, underemphasizing oral intake. C57BL/6 mice are fed either a control diet or a diet supplemented with yeast-derived whole β-glucan particle (yWGP) or with rice-derived arabinoxylan (rice bran-1) at a concentration of 1%, 2.5%, or 5% weight/weight (w/w) for 2 weeks. Thereafter, cells from blood, bone marrow, and spleen are collected for ex vivo stimulation with various microbial stimuli. Dietary intake of yWGP for 2 weeks at concentrations of 1% and 2.5% w/w increases ex vivo cytokine production in mouse blood and bone marrow, whereas 5% w/w yWGP shows no effect. In the spleen, cytokine production remains unaffected by yWGP. At a concentration of 1% w/w, rice bran-1 increases ex vivo cytokine production by whole blood, but 2.5% and 5% w/w cause inhibitory effects in bone marrow and spleen. This study demonstrates that dietary yWGP and rice bran-1 induce immune priming in mouse blood and bone marrow, with the strongest effects observed at 1% w/w. Future human trials should substantiate the efficacy of dietary β-glucans and arabinoxylans to bolster host immunity, focusing on dose optimization.

We demonstrate divergent incorporation and washout patterns for EPA and DHA following high and low-dose EPA+DHA incubation in C2C12 myotubes, with higher concentrations favoring n-3 PUFA incorporation. Lower n-3 PUFA concentrations increased MPS without further upregulating the mTORC1 signaling pathway. Our study provides novel insights into the temporal incorporation and washout dynamics of EPA and DHA and, specifically, their combined effect on MPS, thereby advancing knowledge regarding dietary n-3 PUFA prescription to promote skeletal muscle health in humans.

Rationale: Processing and blending of purified protein sources can improve amino acid (AA) bioavailability and protein quality. A specific combination of dairy proteins (Whey and Casein) rich in essential AA and plant-based protein isolates (Pea and Soy) rich in conditionally essential AA has been shown to provide a well-balanced AA profile [1]; however less is known how this translates to muscle protein synthesis (MPS). Hence, the aim of this study was to investigate the effect of this P4 protein blend compared to single dairy-based protein sources on MPS in aged mice undergoing starvation.

P4, a specific combination of dairy proteins (whey and casein) and plant-based protein isolates (pea and soy), has been shown to provide a more balanced amino acid (AA) profile than its single constituent proteins; however, less is known about how this translates to muscle protein synthesis (MPS). The aim of this study was to investigate the effect of P4 compared to whey or casein against fasted control on MPS. C57BL/6J mice, aged 25 months, were fasted overnight, followed by oral gavage of either whey, P4, casein, or water as a fasted control. Thirty minutes after ingestion, puromycin (0.04 µmol∙g−1 bodyweight) was subcutaneously injected; 30-min thereafter, mice were sacrificed. MPS was measured by the SUnSET method, and signalling proteins were determined in the left-tibialis anterior (TA) muscle by the WES technique. AA composition was determined in plasma and right-TA muscle. Dried blood spots (DBS) were analysed for postprandial AA dynamics at 10, 20, 45, 60 min. MPS was 1.6-fold increased with whey (p = 0.006) and 1.5-fold with P4 compared to fasted (p = 0.008), while no change was seen with casein. This was confirmed by a significant increase of phosphorylated/total ratio of 4E-BP1 for both whey (p = 0.012) and P4 (p = 0.001). No changes were observed in p70S6K and mTOR phosphorylation/total ratio with whey or P4. Intramuscular leucine levels were lower for P4 (0.71 µmol∙g dry weight−1) compared to whey (0.97 µmol∙g dry weight−1) (p = 0.0007). Ten minutes postprandial, DBS showed significantly increased blood AA levels of BCAAs, histidine, lysine, threonine, arginine, and tyrosine for P4 versus fasted. In conclusion, a hybrid mix of dairy and plant-based proteins (P4) resulted in a MPS response that was similar to whey protein in aged mice after fasting. This suggests that other anabolic triggers beyond leucine or the well-balanced amino acid profile and bioavailability of the blend benefit stimulation of MPS.

This review aims to critically evaluate the efficacy of long-chain ո-3 PUFA ingestion in modulating muscle protein synthesis (MPS), with application to maintaining skeletal muscle mass, strength and function into later life. Ageing is associated with a gradual decline in muscle mass, specifically atrophy of type II fibres, that is exacerbated by periods of (in)voluntary muscle disuse. At the metabolic level, in otherwise healthy older adults, muscle atrophy is underpinned by anabolic resistance which describes the impaired MPS response to non-pharmacological anabolic stimuli, namely, physical activity/exercise and amino acid provision. Accumulating evidence implicates a mechanistic role for n-3 PUFA in upregulating MPS under stimulated conditions (post-prandial state or following exercise) via incorporation of EPA and DHA into the skeletal muscle phospholipid membrane. In some instances, these changes in MPS with chronic ո-3 PUFA ingestion have translated into clinically relevant improvements in muscle mass, strength and function; an observation evidently more prevalent in healthy older women than men. This apparent sexual dimorphism in the adaptive response of skeletal muscle metabolism to EPA and DHA ingestion may be related to a greater propensity for females to incorporate ո-3 PUFA into human tissue and/or the larger dose of ingested ո-3 PUFA when expressed relative to body mass or lean body mass. Future experimental studies are warranted to characterise the optimal dosing and duration of ո-3 PUFA ingestion to prescribe tailored recommendations regarding n-3 PUFA nutrition for healthy musculoskeletal ageing into later life.

Cytotoxicity (i.e. cell death) is the core mechanism by which chemotherapy induces its anti-cancer effects. Unfortunately, this same mechanism underpins the collateral damage it causes to healthy tissues. The gastrointestinal tract is highly susceptible to chemotherapy’s cytotoxicity, resulting in ulcerative lesions (termed gastrointestinal mucositis, GI-M) that impair the functional capacity of the gut leading to diarrhea, anorexia, malnutrition and weight loss, which negatively impact physical/psychological wellbeing and treatment adherence. Preventing these side effects has proven challenging given the overlapping mechanisms that dictate chemotherapy efficacy and toxicity. Here, we report on a novel dietary intervention that, due to its localized gastrointestinal effects, is able to protect the intestinal mucosal from unwanted toxicity without impairing the anti-tumor effects of chemotherapy. The test diet (containing extensively hydrolyzed whey protein and medium chain triglycerides (MCTs)), was investigated in both tumor-naïve and tumor-bearing models to evaluate its effect on GI-M and chemo-efficacy, respectively. In both models, methotrexate was used as the representative chemotherapeutic agent and the diet was provided ad libitum for 14 days prior to treatment. GI-M was measured using the validated biomarker plasma citrulline, and chemo-efficacy defined by tumor burden (cm3/g body weight). The test diet significantly attenuated GI-M (P = 0.03), with associated reductions in diarrhea (P < 0.0001), weight loss (P < 0.05), daily activity (P < 0.02) and maintenance of body composition (P < 0.02). Moreover, the test diet showed significant impact on gut microbiota by increasing diversity and resilience, whilst also altering microbial composition and function (indicated by cecal short and brained chain fatty acids). The test diet did not impair the efficacy of methotrexate against mammary adenocarcinoma (tumor) cells. In line with the first model, the test diet minimized intestinal injury (P = 0.001) and diarrhea (P < 0.0001). These data support translational initiatives to determine the clinical feasibility, utility and efficacy of this diet to improve chemotherapy treatment outcomes.

AbstractBackgroundPhysical activity is associated with a lower risk of colorectal cancer (CRC) and CRC‐specific mortality. However, evidence for a causal relationship between physical activity and disease progression is lacking. Here, we have used CRC organoids to create a novel mouse model for spontaneous metastasis formation to multiple organs. We have used this model to assess the influence of voluntary exercise on disease progression.MethodsCollagen‐embedded murine colorectal tumour organoids were transplanted into the livers of immunocompetent C57Bl/6 mice using microsurgery. Voluntary exercise in tumour‐bearing mice was modelled by offering running wheels continuously (n = 12) or 3 h/day (n = 12) versus no wheel access (n = 12). Running wheel revolutions were cumulatively measured every 30 min and physical activity was continuously monitored by infrared cameras. Food intake was monitored throughout the experiment and body composition was assessed with echoMRI. Animals were sacrificed 14 weeks after tumour initiation. Tumour load was quantified by EpCAM immunohistochemistry staining. Systemic inflammation parameters were assessed in blood plasma by a multiplex immunoassay.ResultsTumour growth was initiated by implantation of CRC organoids into the livers of immunocompetent mice. The resulting tumours spontaneously formed distant metastases to non‐implanted liver lobes and to the lungs. Mice with access to the running wheels for 3 h/day ran relatively short distances (2.3 ± 0.3 km/night; 221 ± 29 km total distance) with relatively high intensity (wheel revolutions/h). Mice with continuous access to the running wheels ran significantly longer distances (6.6 ± 3.0 km/night; 600 ± 290 km total distance) with a significantly lower intensity. Both exercise groups showed increased lean body mass, and decreased fat mass and body weight compared with tumour‐bearing control mice. Food intake was unaffected by exercise or tumour growth. Primary tumour growth was not significantly affected by exercise. However, mice with continuous wheel access (long distance‐lower intensity group) displayed increased lung metastasis and decreased liver metastasis formation, when compared with the sedentary control group. Short distance‐higher intensity exercise did not affect metastasis formation. Analysis of blood cytokine levels revealed that mice with continuous wheel access displayed signs of systemic inflammation.ConclusionsThese results suggest that exercise has the potential to influence the patterns and extent of metastasis in CRC, and that the degree and intensity of exercise are likely to be important variables. Confirmation of these results in additional preclinical models with or without systemic treatment is warranted.

PURPOSE:Muscle-wasting and treatment-related toxicities negatively impact prognosis of colorectal cancer (CRC) patients. Specific nutritional composition might support skeletal muscle and enhance treatment support. In this in vitro study we assess the effect of nutrients EPA, DHA, L-leucine and vitamin D3, as single nutrients or in combination on chemotherapy-treated C2C12-myotubes, and specific CRC-tumor cells.MATERIALS AND METHODS:Using C2C12-myotubes, the effects of chemotherapy (oxaliplatin, 5-fluorouracil, oxaliplatin+5-fluorouracil and irinotecan) on protein synthesis, cell-viability, caspase-3/7-activity and LDH-activity were assessed. Addition of EPA, DHA, L-leucine and vitamin D3 and their combination (SNCi) were studied in presence of above chemotherapies. Tumor cell-viability was assessed in oxaliplatin-treated C26 and MC38 CRC cells, and in murine and patient-derived CRC-organoids.RESULTS:While chemotherapy treatment of C2C12-myotubes decreased protein synthesis, cell-viability and increased caspase-3/7 and LDH-activity, SNCi showed improved protein synthesis and cell viability and lowered LDH activity. The nutrient combination SNCi showed a better overall performance compared to the single nutrients. Treatment response of tumor models was not significantly affected by addition of nutrients.CONCLUSIONS:This in vitro study shows protective effect with specific nutrition composition of C2C12-myotubes against chemotherapy toxicity, which is superior to the single nutrients, while treatment response of tumor cells remained.

Skeletal muscle mitochondrial function is the biggest component of whole-body energy output. Mitochondrial energy production during exercise is impaired in vitamin D-deficient subjects. In cultured myotubes, loss of vitamin D receptor (VDR) function decreases mitochondrial respiration rate and ATP production from oxidative phosphorylation. We aimed to examine the effects of vitamin D deficiency and supplementation on whole-body energy expenditure and muscle mitochondrial function in old rats, old mice, and human subjects. To gain further insight into the mechanisms involved, we used C2C12 and human muscle cells and transgenic mice with muscle-specific VDR tamoxifen-inducible deficiency. We observed that in vivo and in vitro vitamin D fluctuations changed mitochondrial biogenesis and oxidative activity in skeletal muscle. Vitamin D supplementation initiated in older people improved muscle mass and strength. We hypothesize that vitamin D supplementation is likely to help prevent not only sarcopenia but also sarcopenic obesity in vitamin D-deficient subjects.

Abstract Background Skeletal muscle wasting and fatigue are commonly observed in cancer patients receiving chemotherapy and associated with reduced treatment outcome and quality of life. Nutritional support may mitigate these side effects, but potential interference with chemotherapy efficacy could be of concern. Here, we investigated the effects of an ω‐3 polyunsaturated fatty acid (eicosapentaenoic acid and docosahexaenoic acid), leucine‐enriched, high‐protein (100% whey), additional vitamin D, and prebiotic fibres ‘specific nutritional composition’ (SNC) and chemotherapy on state‐of‐the‐art tumour organoids and muscle cells and studied muscle function, physical activity, systemic inflammation, and chemotherapy efficacy in a mouse model of aggressive colorectal cancer (CRC). Methods Tumour‐bearing mice received a diet with or without SNC. Chemotherapy treatment consisted of oxaliplatin and 5‐fluorouracil. Tumour formation was monitored by calliper measurements. Physical activity was continuously monitored by infrared imaging. Ex vivo muscle performance was determined by myography, muscle fatty acid composition by gas chromatography, and plasma cytokine levels by Luminex xMAP technology. Patient‐derived CRC organoids and C2C12 myotubes were used to determine whether SNC affects chemotherapy sensitivity in vitro. Results Specific nutritional composition increased muscle contraction capacity of chemotherapy‐treated tumour‐bearing mice (P < 0.05) and enriched ω‐3 fatty acid composition in muscle without affecting treatment efficacy (P < 0.0001). Mice receiving SNC maintained physical activity after chemotherapy and showed decreased systemic inflammation. Therapeutic response of CRC organoids was unaffected by SNC nutrients, while cell viability and protein synthesis of muscle cells significantly improved. Conclusions The results show that specialized nutritional support can be used to maintain muscle function and physical activity levels during chemotherapy without increasing tumour viability. Therefore, nutritional strategies have potential value in promoting cancer and chemotherapy tolerance.

Skeletal muscle wasting and fatigue are commonly observed in cancer patients receiving chemotherapy and are associated with reduced treatment outcome and quality of life. Nutritional support may mitigate these side effects, but potential interference with chemotherapy efficacy could be of concern. Here, we investigated the effects of a ω-3-PUFA (EPA and DHA), leucine-enriched, high protein (100% whey), additional vitamin D and prebiotic fibers “Specific Nutritional Composition” (SNC) and chemotherapy on state-of-the-art tumor organoid models and muscle cells and studied muscle function, physical activity, systemic inflammation, and chemotherapy efficacy in a mouse model of aggressive colorectal cancer (CRC). Chemotherapy treatment consisted of oxaliplatin and 5-fluorouracil. Tumor formation (caliper) and physical activity (infrared camera's) were assessed over time, while tumor-bearing mice received a diet with or without SNC. Ex vivo muscle performance was determined by myography, muscle fatty acid composition by gas-chromatography, and plasma cytokine levels by Luminex xMAP technology. Patient-derived CRC-organoids and C2C12-myotubes were used to determine whether SNC affects chemotherapy-sensitivity in vitro. SNC increased the muscle contraction capacity of chemotherapy-treated tumor-bearing mice (P < 0.05), and enriched ω-3 fatty acid composition in muscle without affecting treatment efficacy (P < 0.0001) compared to control tumor-bearing mice. Mice receiving SNC maintained physical activity after chemotherapy and showed decreased systemic inflammation and splenomegaly. Therapeutic response of CRC-organoids was unaffected by SNC nutrients, while C2C12 cell viability and protein synthesis significantly improved. The results show that specialized nutritional support can be used to maintain muscle function and levels of physical activity during chemotherapy without increasing tumor viability. Therefore, nutritional strategies have potential value in promoting cancer and chemotherapy tolerance. This research is part of the SCOPE project (Specialized nutrition to improve outcomes of COlorectal cancer PatiEnts) and is supported by the Province of Utrecht, The Netherlands.

Targeted exercise combined with nutritional and pharmacological strategies is commonly considered to be the most optimal strategy to reduce the development and progression of cachexia. For COPD patients, this multi-targeted treatment has shown beneficial effects. However, in many, physical activity is seriously hampered by frailty and fatigue. In the present study, effects of whole-body-vibration-training (WBV) were investigated, as potential alternative to active exercise, on body mass, muscle mass and function in tumour bearing mice. Twenty-four male CD2F1-mice (6–8 weeks, 21.5 ± 0.2 g) were stratified into four groups: control, control + WBV, C26 tumour-bearing, and C26 tumour-bearing + WBV. From day 1, whole-body-vibration was daily performed for 19 days (15 min, 45 Hz, 1.0 g acceleration). General outcome measures included body mass and composition, daily activity, blood analysis, assessments of muscle histology, function, and whole genome gene expression in m. soleus (SOL), m. extensor digitorum longus (EDL), and heart. Body mass, lean and fat mass and EDL mass were all lower in tumour bearing mice compared to controls. Except from improved contractility in SOL, no effects of vibration training were found on cachexia related general outcomes in control or tumour groups, as PCA analysis did not result in a distinction between corresponding groups. However, analysis of transcriptome data clearly revealed a distinction between tumour and trained tumour groups. WBV reduced the tumour-related effects on muscle gene expression in EDL, SOL and heart. Gene Set Enrichment Analysis showed that these effects were associated with attenuation of the upregulation of the proteasome pathway in SOL. These data suggest that WBV had minor effects on cachexia related general outcomes in the present experimental set-up, while muscle transcriptome showed changes associated with positive effects. This calls for follow-up studies applying longer treatment periods of WBV as component of a multiple-target intervention.

Abstract Background: Physical activity (PA) is associated with a lower risk of colorectal cancer (CRC) initiation and death from CRC. However, evidence for a causal link between PA and disease progression is lacking. This study assesses the effect of voluntary exercise on metastases formation in a novel pre-clinical mouse model for late-stage metastatic CRC. Methods: Murine colorectal tumor organoids were transplanted into the livers of immunocompetent C57Bl/6 mice using microsurgery. Animals were fed ad libitum. Voluntary exercise in tumor-bearing (TB) mice was modeled by offering running wheels for 0h/day (n=12), 3h/day (n=12) or 24h/day (n=12). Running wheel revolutions were cumulatively measured every 30min. PA was monitored by infrared cameras. Tumor progression was monitored using in vivo Bioluminescent Imaging. Food intake was measured by weighing. Body composition was monitored by echoMRI. All animals were sacrificed 12 weeks after tumor initiation. Results: We have developed a novel model for exploring the effects of exercise on metastatic tumor progression in CRC. The model combines micro-surgical transplantation of tumor organoids, yielding spontaneously metastasizing CRC, with voluntary exercise. Voluntary exercise in both exercise groups led to an on average running distances of 2,3±0,3 km/night (total 221±29 km) in TB-3h animals and 6,6±3,0km/night (total 600±290km) in TB-24h animals. TB-3h and TB-24h animals developed more lean mass and lower fat mass compared to sedentary animals. Intermittent exercise had no effect on primary tumor growth and reduced metastasis. Vigorous exercise increased primary tumor growth (229606±186012) compared to TB-3h (152821±125902) and sedentary animals (165171±250993). Moreover, vigorous exercise non-significantly increased lung metastasis formation (2382±3539) compared to intermittent exercise (753±852) and to sedentary (1333±2902), while reducing metastatic spread to the liver and the peritoneal cavity was found. Conclusions: These results suggest that exercise has the potential to influence the patterns and extent of metastasis in CRC, and that the degree of exercise is likely to be an important variable. Confirmation of these results in additional models, for instance using patient-derived organoids, is now warranted. Citation Format: Liza A. Wijler, Daniëlle A. Raats, Miriam van Dijk, Anne M. May, Onno Kranenburg. Potential impact of voluntary exercise on patterns of metastasis in a colorectal cancer model [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3480.

Background: Dietary supplementation with leucine and fish oil rich in omega-3 fatty acids docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) has previously been shown to reduce cachexia-related outcomes in C26 tumour-bearing mice. To further explore associated processes and mechanisms we investigated changes in plasma Ca2+ levels, the involvement of parathyroid hormone related protein (PTHrP), and its possible interactions with cyclooxygenase 2 (COX-2). Methods: CD2F1 mice were subcutaneously inoculated with C26 adenocarcinoma cells or sham treated and divided in: (1) controls, (2) tumour-bearing controls, and (3) tumour-bearing receiving experimental diets. After 20 days, body and organ masses and total plasma Ca2+ levels were determined. Furthermore, effects of DHA, EPA and leucine on production of PTHrP were studied in cultured C26 cells. Results: The combination of leucine and fish oil reduced tumour-associated hypercalcemia. Plasma Ca2+ levels negatively correlated with carcass mass and multiple organ masses. DHA was able to reduce PTHrP production by C26 cells in vitro. Results indicate that this effect occurred independently of COX-2 inhibition. Conclusion: Our results suggest that cancer-related hypercalcemia may be ameliorated by a nutritional intervention rich in leucine and fish oil. The effect of fish oil possibly relates to a DHA-induced reduction of PTHrP excretion by the tumour.