Background: ctDNA analysis offers a non-invasive approach for monitoring response and resistance to treatment. Serial ctDNA testing during neoadjuvant therapy (NAT) may provide early indicators of emerging resistance and disease progression. In this study, we analyzed ctDNA from high-risk early breast cancer patients who received NAT and definitive surgery in the I-SPY 2 TRIAL (NCT01042379). We hypothesize that (1) assessment of ctDNA levels early in treatment will improve the performance of molecular and imaging-based predictors of pathologic complete response (pCR) to NAT; (2) mutational spectrum in residual tumors are manifested early in plasma; and (3) levels of ctDNA after NAT are associated with residual cancer burden and recurrence [distant recurrence free survival (DRFS)].

Methods: ctDNA analysis was performed in 84 high-risk stage II and III breast cancer patients randomized to neoadjuvant investigational agent (n=57), AKT inhibitor MK-2206 (M) in combination with paclitaxel (T) followed by doxorubicin and cyclophosphamide (AC) (M+T->AC), or standard-of-care (T->AC) (n=27). HER2+ patients also received trastuzumab (H).

Serial plasma was collected before NAT, early treatment (3 weeks), between regimens (12 weeks), and after NAT prior to surgery. Mutational profiles derived from pretreatment tumor biopsy and germline DNA whole exome sequencing were used to design personalized assays targeting 16 variants specific to a patients' tumor to detect ctDNA in plasma. In a subset of patients who did not achieve a pCR (n=61), mutations in residual cancers were compared to those found in pretreatment tumor.

Analysis: Of the 84 patients in this analysis, 43% were HR-HER2-, 35% HR+HER2-, and 23% HER2+. 16% and 35% achieved a pCR in the control and treatment arms, respectively. Currently, data are being collected to: (1) determine the relationship between ctDNA levels during early treatment and pCR/residual cancer burden; (2) assess the relationship of ctDNA and MRI imaging in predicting tumor response to therapy; (3) examine the relationship of ctDNA levels before and after NAT with 3-year DRFS and event-free survival (EFS). The results of the analyses will be updated by August 31 and presented at the SABCS 2018 meeting.

Conclusions: Our study provides a platform to evaluate the clinical significance of ctDNA for serial monitoring of response to NAT. Accurate and early response prediction by highly sensitive ctDNA analysis can facilitate a timely and judicious change in treatment to improve patients' chances of achieving a pCR. Finally, personalized ctDNA testing may complement imaging and pathologic evaluation of tumor response to fine-tune pCR as a surrogate endpoint for improved DRFS and EFS.

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