Background: Avelumab 1L maintenance + best supportive care (BSC) significantly prolonged overall survival (OS) vs BSC alone in patients (pts) with advanced UC that had not progressed with 1L platinum-based chemotherapy (median OS, 23.8 vs 15.0 months; HR, 0.76 [95% CI, 0.631-0.915]; 2-sided p = 0.0036). Avelumab 1L maintenance is now considered standard of care per international guidelines. However, data on outcomes in pts who receive 2L treatment after avelumab 1L maintenance are limited. We report a descriptive analysis of outcomes in pts enrolled in the avelumab + BSC arm of the JAVELIN Bladder 100 trial based on receipt of 2L treatment. Methods: In the phase 3 JAVELIN Bladder 100 trial (NCT02603432), eligible pts had unresectable locally advanced or metastatic UC without progression with 4-6 cycles of 1L gemcitabine + cisplatin or carboplatin. Pts were randomized 1:1 to receive avelumab + BSC or BSC alone. Exploratory analyses of time from randomization to end of 2L treatment and OS were performed in the avelumab + BSC arm in subgroups defined by 2L treatment administered by investigators after discontinuation of study treatment. Results: In the avelumab + BSC arm (n = 350), median follow-up at data cutoff (June 4, 2021) was 38.0 months. 185 pts (52.9%) had discontinued avelumab 1L maintenance treatment for any reason and received 2L treatment, whereas 122 (34.9%) had discontinued avelumab and did not receive 2L treatment. Median OS (95% CI) in subgroups is shown in the Table. In 43 pts (12.3%) who were still receiving avelumab, median treatment duration was 154.6 weeks (range, 106.7-216.0). Among pts who received 2L treatment, median time from end of avelumab 1L maintenance to start of 2L treatment was 1.35 months (range, 0.3-30.9) and median time from randomization to end of 2L treatment was 11.7 months (95% CI, 9.7-13.8). 2L treatment comprised rechallenge with platinum-based chemotherapy in 75 pts (21.4%) or other 2L treatment in 110 pts (31.4%), including 2L anti–PD-(L)1 therapy in 11 pts (3.1%). Additional data based on different types of 2L treatment will be presented. Conclusions: In this exploratory analysis from the JAVELIN Bladder 100 trial with extended follow-up, approximately 60% of pts (185/307 pts) who had discontinued avelumab received 2L treatment. Long-term OS was observed in pts who received avelumab 1L maintenance with or without 2L treatment. Clinical trial information: NCT02603432.