Background: T-DXd is an antibody–drug conjugate of a humanized anti-HER2 antibody bound to a topoisomerase I inhibitor by a cleavable linker. The primary analysis of DESTINY-CRC01 (DS8201-A-J203; NCT03384940), a phase 2, open-label, multicenter study of T-DXd in pts with HER2-expressing mCRC showed promising antitumor activity and a manageable safety profile (cohort A median follow-up [FU], 27.1 weeks; Siena S, ASCO 2020). We present updated longer-term efficacy and safety data. Methods: Pts had centrally confirmed HER2-expressing, RAS wild-type mCRC that progressed after ≥2 prior regimens. 6.4 mg/kg of T-DXd was administered every 3 weeks (Q3W) in 3 cohorts (A: HER2 IHC3+ or IHC2+/ISH+; B: IHC2+/ISH−; C: IHC1+). The primary end point was confirmed objective response rate (ORR) by independent central review in cohort A. Secondary end points were disease control rate (DCR; CR + PR + SD), duration of response (DOR), progression-free survival (PFS), and overall survival (OS). Results: At data cutoff (Dec 28, 2020), 86 pts (A, 53; B, 15; C, 18) received T-DXd. Median age was 58.5 y (range, 27-79), 53.5% were male, and 90.7% had left colon or rectum cancer. Median prior regimens for metastatic disease was 4 (range, 2-11). All pts had prior irinotecan; 30.2% in cohort A had prior anti-HER2 therapy. Median (m) treatment duration (all pts) was 3.0 mo (95% CI, 2.1-4.1; cohort A, 5.1 mo [95% CI, 3.9-7.6]). In cohort A (median FU, 62.4 weeks), confirmed ORR was 45.3% (24/53 pts; 95% CI, 31.6-59.6), DCR was 83.0% (44/53 pts; 95% CI, 70.2-91.9), mDOR was 7.0 mo (95% CI, 5.8-9.5), mPFS was 6.9 mo (95% CI, 4.1-8.7) with 37 (69.8%) PFS events, and mOS was 15.5 mo (95% CI, 8.8-20.8) with 36 (67.9%) OS events. These results are consistent with the primary analysis. Confirmed ORR was 43.8% (7/16 pts; 95% CI, 19.8-70.1) in pts with prior anti-HER2 therapy, 57.5% (23/40 pts; 95% CI, 40.9-73.0) in pts with IHC3+ status, and 7.7% (1/13 pts; 95% CI, 0.2-36.0) in pts with IHC2+/ISH+ status. In cohorts B and C, mPFS was 2.1 mo (95% CI, 1.4-4.1) and 1.4 mo (95% CI, 1.3-2.1); mOS was 7.3 mo (95% CI, 3.0-NE) and 7.7 mo (95% CI, 2.2-13.9), respectively. Treatment-emergent adverse events (TEAEs) of grade (G) ≥3 occurred in 65.1% of pts (56/86); the most common TEAEs were hematologic and gastrointestinal. TEAEs leading to drug discontinuation occurred in 13 pts (15.1%). 8 pts (9.3%) had interstitial lung disease (ILD) adjudicated by an independent committee as related to T-DXd (4 G2; 1 G3; 3 G5). Conclusions: T-DXd at 6.4 mg/kg Q3W showed promising activity and durability with longer-term FU in this pt population. The safety profile was consistent with prior results; ILD continues to be recognized as an important identified risk that requires careful monitoring and intervention as needed. These results support continued exploration of T-DXd in pts with HER2-overexpressing mCRC. Clinical trial information: NCT03384940