63 Olaparib in patients with biallelic BRCA1/2 inactivation Eight biopsies could not be sequenced due to a low tumor cellularity (< 30%) and one was sequenced despite a tumor cellularity below the threshold, confirming the qualifying BRCA mutation, but HRD-score and bi-allelic call could not be extracted (Supplementary Table 1). From seven of 13 patients with successful baseline biopsy WGS, pre-enrollment WGS data were also available. Additionally, from eight patients with failed baseline study WGS, pre-enrollment WGS data were available, and from three patients no WGS data were available and information on bi-allelic status and HRD-score from these patients could not be retrieved. Based on a consensus of findings from the pre-enrollment and the baseline study biopsies, 15 out of 24 patients had confirmed bi-allelic BRCA LoF and a high HRD-score (Supplementary Table 1). In two patients with prostate cancer the call for bi-allelic loss could not be made due to low tumor purity, but in one of them, the high HRD-score suggests complete LoF of BRCA2. In six other patients, baseline WGS showed a low HRD-score and only mono-allelic loss (N=4) or no BRCA variant at all (N=2, 9%) (Supplementary Table 1). CLINICAL BENEFIT Fourteen of 24 patients (58%, 95% CI 37% - 78%) had CB upon treatment with olaparib. The objective response rate was 29% (7/24 patients), median time on treatment was 5.8 months (95% CI 1.8 – 9.2 months). At data cut-off (5 November 2020), one patients was still on treatment. The median PFS in this cohort was 7 months (95% CI 2-8 months) and the median OS was 13 months (95% CI 7 – NA months) (Figure 1). CB was observed across tumor types, including non-BRCA histologies such as cholangiocarcinoma, and in patients with both germline and somatic BRCA alterations (Figure 2, Supplementary Table 1). In the group of patients with CB, the median treatment duration was 9.1 months (95% CI 8.4 – NA months). The difference in outcome between bi-allelic LoF of BRCA1 and BRCA2 was not statistically significant (Fishers’ exact value 0.2445). CB was predominantly observed in patients with tumors harboring a bi-allelic LoF alteration of BRCA1 or BRCA2, and with an HRD genomic signature, with few exceptions: one patient with prostate cancer had prolonged stable disease, while having no signs of genomic bi-allelic BRCA loss. The pre-enrollment molecular data showed a somatic BRCA2 mutation with 24% variant allele frequency (VAF), while in the baseline study biopsy WGS data, no evidence of a BRCA alteration or HRD was found. As indicated before, the most likely cause of this discordance is tumor heterogeneity. It is known that patients with BRCA-associated tumor types can benefit from PARPi even if the tumor has only mono-allelic BRCA LoF13. Another possible explanation for the clinical benefit in this patient may be that the dominant tumor clone indeed had a BRCA2 alteration, in combination with a post-translational silencing of BRCA2, resulting in functional HRD. CB was also observed in two patients whose details regarding bi-allelic LoF and HRD-score were unknown. Both patients had BRCA associated tumor types and were included based on a germline test only, with no WGS results available to confirm the target. None of the four patients with confirmed mono-allelic loss had CB. Of the fifteen patients with confirmed bi-allelic BRCA1/2 LoF, eleven had CB (73%). 3
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