Background To investigate the erythropoietic activity and safety aspects of testosterone undecanoate (TU) injections in transgender men, assigned female at birth.Methods Twenty-three men (13 hypogonadal cisgender men and 10 transgender men) who initiated TU at the study start (naive) and 15 men (10 hypogonadal cisgender men and 5 transgender men) on steady-state treatment with TU (non-naive) were included in this prospective 1-year observational study. A control group of 32 eugonadal cisgender men was investigated once at baseline. Complete blood count, testosterone in serum and saliva, and plasma lipids, and liver enzymes were assessed.Results For naive transgender men, a significant increase in hemoglobin concentration was noted (mean (SD)), 141 (8) g/L to 151 (13) g/L, while no increase was seen in naive hypogonadal cisgender men. At the end of the study, naive transgender men exhibited comparable levels of hemoglobin, hematocrit, and testosterone levels in serum and saliva to hypogonadal cisgender men, as well as to the eugonadal cisgender men. During the study, HDL-cholesterol decreased significantly in naive transgender men, 1.4 (0.4) mmol/L to 1.2 (0.4) mmol/L, P = 0.03, whereas no significant change was noted in naive hypogonadal cisgender men. Liver enzymes remained unchanged in all groups.Conclusions After 12 months of treatment with TU in naive transgender men, hemoglobin and hematocrit increased to levels within the cisgender male reference range. A slight decrease in HDL-cholesterol was seen in naive transgender men but liver enzymes remained unchanged.