Background: Patients (pts) with advanced NSCLC and sensitizing EGFR mutations who initially respond to gefitinib or erlotinib eventually develop acquired resistance to the TKIs. Anecdotal and retrospective reports suggest that EGFR-TKI resistant cancers can respond again to gefitinib or erlotinib after an interval off the TKI. This retrospective study was undertaken to investigate the impact of EGFR-TKI retreatment after a drug-free interval in EGFR mutant NSCLC with acquired resistance to gefitinib or erlotinib. Methods: Pts with stage IV or relapsed NSCLC with sensitizing EGFR mutations and acquired resistance to EGFR-TKI seen at the DFCI/MGH between 8/00 and 8/11 who were retreated with single agent gefitinib or erlotinib after an EGFR-TKI-free interval were identified from a prospective trial. The objective tumor response (CR, PR, SD, PD) was determined using RECIST 1.1. Results: 19 pts were eligible and had adequate scans for radiographic assessments after the reinstitution of an EGFR-TKI. The response rate and median PFS to the initial course of gefitinib (n=4) or erlotinib (n=15) were 16/19 (84%) and 9.8 months (95% CI, 7.8-11.3) respectively. All pts were retreated with erlotinib after 1 to 4 intervening systemic regimens. The median interval from EGFR-TKI discontinuation to erlotinib retreatment was 11 months (range, 2-46). 4 of the 19 pts (21%) had PD as the best response to erlotinib retreatment, 14 (74%) had SD for at least 1 month, and 1 (5%) had a PR. The median PFS was 4.4 months (95% CI, 3.0-6.7). 3 pts remained on erlotinib without progression for 6 months. 3 pts had their tumors rebiopsied before (n=2) or during (n=1) erlotinib retreatment; 1 of the 3 had EGFR T790M in association with the initial sensitizing EGFR mutation, and another had a secondary PIK3CA mutation. Conclusions: Our findings suggest that erlotinib retreatment is an option for EGFR mutated NSCLC with acquired resistance to EGFR-TKI after a drug-free interval and progression on intervening therapy. Additional advanced NSCLC pts without a documented EGFR mutation who fulfill the clinical definition of acquired resistance are undergoing review to expand our cohort.