Background: Chemotherapy may lead to systemic muscle damage. Up-regulation of developmental myosin light chain 5 (MYL5) and myosin heavy chain 8 (MYH8) genes is required for normal muscle regeneration in response to damage. However, secretion of MYL5 and MYH8 proteins into the serum suggest degradation of muscle, which, in turn, may lead to cancer-related-fatigue (CRF). In this study, we investigated (1) the effect of an exercise intervention, Exercise for Cancer Patients (EXCAP) on mRNA gene expression and serum protein levels of MYL5 and MYH8 and (2) the association of these novel biomarkers with CRF. Methods: Chemotherapy naïve cancer patients (N = 350; mean age = 55.7) from 39 community oncology practices throughout the U.S. affiliated with the URCC NCORP Research Base participated in this nationwide, multicenter, phase III RCT. Patients were randomized into 2 groups: (1) chemotherapy and (2) chemotherapy plus a 6-week aerobic and resistance exercise prescription-EXCAP. Gene expression and protein levels of MYL5 and MYH8, as well as CRF were assessed pre- and post-intervention from whole blood by qPCR, from serum by Luminex assays, and from patient-report by the Multidimensional Fatigue Symptom Inventory, respectively. Results: T-tests show MYL5, but not MYH8, mRNA levels were significantly up-regulated from pre- to post-intervention in exercisers and controls (all p < 0.01) with no significant group difference. Additionally, MYL5 and MYH8 serum protein levels significantly increased from pre to post in controls (all p < 0.05), but remained stable in exercisers. Significant group differences in these serum proteins (p < 0.01) suggest greater muscle degradation in non-exercisers. Pearson correlations revealed trends suggesting increases in MYL5 and MYH8 serum proteins are associated with increases in CRF (r = 0.09 and r = 0.11, respectively, all p < 0.10). Conclusions: Results suggest EXCAP exercise is protective from chemotherapy-induced muscle damage via its effects on MYL5 and MYH8, and changes in these novel biomarkers may mediate changes in CRF. Further research is needed to confirm these findings. NCI UGCA189961, R25 CA102618. Clinical trial information: NCT00924651