Topoisomerase 1 and DNA damage: Pharmacodynamic responses and mechanism of trastuzumab deruxtecan in HER2- expressing advanced solid tumors.

3092 Background: Trastuzumab deruxtecan (T-DXd) consists of deruxtecan, a novel topoisomerase 1 (TOP1) inhibitor, covalently bound via a lysosomal protease-cleavable linker to the human epidermal growth factor receptor 2 (HER2)-targeting antibody, trastuzumab. The mechanism(s) underlying the clinical activity seen in multiple tumor types across various HER2 levels is of ongoing, active interest. To investigate the T-DXd mechanism of action, we developed a pilot clinical trial of T-DXd with a detailed pharmacodynamic analysis from TOP1 target engagement to downstream effects of DNA damage in patients with HER2-expressing (IHC 1-3+, HER2 amplified, or HER2 mutated) advanced solid tumors (NCT04294628). Methods: Based on preclinical studies modeling the trial, research tumor biopsies from consenting patients were collected at three time points: pre-treatment, post-dose Cycle 1 (48-96 hours) and pre-dose Cycle 3. The biopsies were evaluated for TOP1 inhibition, induction of stabilized TOP1covalent complexes (TOP1cc) and induction of downstream DNA damage repair (DDR) markers (RAD51, pNBS1, RPA32) using validated, quantitative multiplex immunofluorescence assays on fixed tumor sections with image analysis methodology. In addition, a retrospective analysis of Schlafen 11 expression in baseline biopsies was performed to determine its predictive value of tumor responses to T-DXd. Results: Twenty-one biopsy pairs (pre-treatment and C1, 48-96h post-dose 1) were evaluable for TOP1 molecular response and downstream DDR marker induction. TOP1 target modulation was detected in 15 (71%) of the on-treatment biopsies and markers of DDR were induced in 18 (86%) biopsy pairs. TOP1 target inhibition, robust induction of downstream DNA damage response including stalled replication fork progression, and DNA breaks were observed in HER2-amplified/2+/3+ tumors, and in HER2 1+ tumors. Conclusions: Our results confirm the intended TOP1 molecular mechanism of action of T-DXd in HER2-amplified/2+/3+ and importantly extend that finding to HER2 1+ tumors, resulting in DNA damage in nearly all cases. Pharmacodynamic biomarker studies are poised to yield important insights into the molecular effects of T-DXd in solid tumors.