Optic nerve invasion (ONI) is an important risk factor for extraocular metastasis in retinoblastoma. This study evaluates the impact of pre-enucleation chemotherapy, delayed enucleation, and adjuvant chemotherapy on survival in patients with varying degree of ONI on histopathology.A retrospective review of consecutive enucleated eyes with any degree of ONI from 29 Chinese treatment centers 2012-2017. Children with other high-risk histopathological features, extraocular disease at diagnosis or bilateral enucleation were excluded.Among 2500 enucleated eyes, 386 eyes with isolated ONI (one eye per child) met the inclusion criteria: prelaminar (n = 204), intralaminar (n = 70), retrolaminar without tumor at transected optic nerve (n = 96), or retrolaminar with tumor at transected end (n = 16). Primary enucleation was performed in 59% of cases, whereas 41% underwent secondary enucleation after eye salvage therapies. Delayed enucleation beyond six months from diagnosis significantly increased the likelihood of tumor at optic nerve transection (21% vs. 2%; P < 0.001). The five-year cause-specific survival (CSS) was 96.7% overall: prelaminar (100%), intralaminar (98%), retrolaminar without transected end involvement (94%), or tumor at transected optic nerve (58%). Secondarily enucleated eyes with retrolaminar ONI without transected end involvement had lower CSS than those primarily enucleated (85.6% vs. 100%; P = 0.022). The five-year CSS was higher but not statistically significant, for eyes treated with or without adjuvant chemotherapy: intralaminar ONI (100% vs. 95.7%; P = 0.193), retrolaminar ONI without tumor at transected end (100% vs. 93.5; P = 0.413), and tumor at the transected end (80.0% vs. 45.0%; P = 0.192).Timely enucleation reduces the risk of tumor involvement at optic nerve transection. Delay in enucleation by pre-enucleation chemotherapy may reduce the effectiveness of subsequent adjuvant chemotherapy.