The efficacy and safety of osimertinib were demonstrated in clinical trials; however, real-world clinical data, particularly the resistance profile, are limited. Here, we investigated the efficacy, safety, and resistance profile of osimertinib in real-world practice. We reviewed medical records of T790M mutation-positive lung cancer patients who started osimertinib between February 2016 and June 2017. Molecular pathologic data of biopsy samples obtained after acquisition of resistance to osimertinib were also analyzed. The study included 23 patients with a median age of 59 years. The median follow-up duration was 11.9 months (IQR, 4.7-15.8). Objective response was achieved in 17 (73.9%) patients, and the disease was controlled in 22 (95.7%) patients. Median progression-free survival (PFS) was 7.4 months (95% CI, 3.6-11.0). Adverse events were minimal except for one case of pneumonitis. Of 14 patients experiencing disease progression, 10 underwent re-biopsy. The T790M mutation disappeared in seven patients (70%), and one showed wild-type conversion. PFS was shorter in the T790M-loss group than in the T790M-persistent group (4.4 vs. 7.7 months). Two patients with small cell transformation responded well to subsequent chemotherapy. One patient developed a C797S mutation that became undetectable after two cycles of gemcitabine and cisplatin followed by six cycles of pembrolizumab, after which the patient responded well to osimertinib. In conclusion, osimertinib showed favorable efficacy and safety in real-world practice comparable to those observed in clinical trials. Repeat biopsy after the acquisition of resistance to osimertinib is helpful to direct further treatment strategies.