MTX is an effective and widely used immunomodulatory drug for rheumatoid diseases. MTX osteopathy is a very rare and specific side effect, characterized by stress fractures at multiple locations in the lower extremity, hampering the patient’s mobility by pain and loss of function. In clinical practice, osteoporosis and MTX osteopathy are repeatedly confused and a comparative workup is needed to clarity it’s specifics. Furthermore, specific treatment options for MTX osteopathy need to be established. We compared patients suffering from MTX osteopathy to patients with osteoporosis (OPO). Patients underwent an extensive clinical workup including blood sampling, bone mineral density measurements, high-resolution peripheral quantitative computed tomography and muscular performance testing. Furthermore, treatment regimes in MTX osteopathy were compared with respect to regain of mobility and pain reduction. 83 patients with MTX osteopathy and 89 with OPO were included. Patients with MTX osteopathy did exhibit fractures predominantly at the lower extremity and pain scores were significantly higher (MTX: 6.75 ± 1.86 vs. OPO: 3.62 ± 2.95, p < 0.0001). MTX-caused mobility restriction was successfully reduced by treatment only if MTX was discontinued (pre-treatment: 2.16 ± 1.19 vs. post-treatment: 1.04 ± 0.87, p < 0.0001). Most mobility gain was achieved by involving anabolic treatment (anabolic: 2.1 ± 1.02 vs. antiresorptive: 1.09 ± 0.94, p < 0.05). In summary, MTX osteopathy is characterized by distinct lower extremity stress fractures leading to severe pain and immobility. Discontinuation of MTX is essential to enable treatment success and involving anabolic treatment seems to be more effectively in mobility regain as antiresorptive treatment alone.