Diabetic foot ulcers, complicated by osteomyelitis, can be treated by surgical resection, dead space filling with gentamicin-loaded calcium sulphate-hydroxyapatite (CaS-HA) biocomposite, and closure of soft tissues and skin. To assess the feasibility of this treatment regimen, we conducted a multicenter retrospective cohort study of patients after failed conventional treatments. From 13 hospitals we included 64 patients with forefoot (n = 41 (64%)), midfoot (n = 14 (22%)), or hindfoot (n = 9 (14%)) ulcers complicated by osteomyelitis. Median follow-up was 43 (interquartile range, 20-61) weeks. We observed wound healing in 54 patients (84%) and treatment success (wound healing without ulcer recurrence) in 42 patients (66%). Treatment failures (no wound healing or ulcer recurrence) led to minor amputations in four patients (6%) and major amputations in seven patients (11%). Factors associated with treatment failures in univariable Cox regression analysis were gentamicin-resistant osteomyelitis (hazard ratio (HR), 3.847; 95%-confidence interval (CI), 1.065-13.899), hindfoot ulcers (HR, 3.624; 95%-CI, 1.187-11.060) and surgical procedures with gentamicin-loaded CaS-HA biocomposite that involved minor amputations (HR, 3.965; 95%-CI, 1.608-9.777). In this study of patients with diabetic foot ulcers, complicated by osteomyelitis, surgical treatment with gentamicin-loaded CaS-HA biocomposite was feasible and successful in 66% of patients. A prospective trial of this treatment regimen, based on a uniform treatment protocol, is required.