Background: Fulvestrant is a selective estrogen receptor modulator (SERM) approved as a single agent for estrogen receptor positive, HER2 negative breast cancer patients at various stages of disease and in combination with CDK4/6 inhibitors following endocrine therapy failure. Although this agent requires intramuscular injection, it has demonstrated superior activity and fewer side effects versus some oral endocrine treatments however, resistance to fulvestrant often develops. To better understand fulvestrant resistance and its utility in patients who have failed other endocrine therapies, we evaluated the agent in a panel of ER+ breast models established from patients at various stages of disease representing endocrine-sensitive and -resistant disease. Methods: Sixty-five previously developed ER+ breast XPDX models were evaluated in this study. Models were grown subcutaneously in female athymic nude mice supplemented with estradiol in drinking water when necessary. All models were characterized at early and late passages for estrogen receptor expression by immunohistochemistry and profiled using WES and RNAseq. For in vivo studies, fulvestrant was administered by subcutaneous injection at 2.5 or 5 mg per dose once weekly until study completion. In vivo study endpoints included tumor volume and time from treatment initiation with %T/C values and tumor regression reported at study completion; a %T/C of ≤ 20 versus control was considered sensitive. Tumor regression (%T/C<0) versus Day 0 tumor volume was also reported. Models were grouped by patient clinical treatment prior to sample collection and model establishment including chemo-naïve, endocrine therapies other than fulvestrant, non-endocrine therapies, and fulvestrant. Results: All models retained ER expression in evaluated passages with similar histology compared with archival clinical samples. DNA/RNA sequencing identified several model-specific variants including ESR1 point mutations and fusions. In this study, 22% of models represented chemo-naïve patients, 24% were from patients treated with non-endocrine therapies, 20% were from patients treated with fulvestrant and the remaining 34% from patients treated with endocrine therapies other than fulvestrant. In vivo, 40% of the chemo-naïve group reported sensitivity to fulvestrant including several models with tumor regressions. Activity of fulvestrant was reported in 25% of models in the non-endocrine therapies group and in 20% of models from patients treated with either fulvestrant or endocrine therapies other than fulvestrant. Several models resistant to fulvestrant from chemo-naïve or patients treated with non-endocrine therapies harbored driver mutations including ESR1, PIK3CA, and AKT1 variants or were HER2+. In models from patients pretreated with fulvestrant, those who had also received a CDK4/6 inhibitor were particularly resistant. Conclusion: We evaluated fulvestrant in a panel of ER+ breast XPDX models representing endocrine-sensitive and -resistant disease from both chemo-naïve and pretreated patients, and identified models responsive and resistant to fulvestrant therapy. This panel can be utilized as a valuable tool in better understanding fulvestrant and endocrine therapy resistance and in developing novel therapies for patients resistant to currently available hormonal therapies. Citation Format: April Cabang, Crystal Moreno, Johnnie R Flores, Jenna Boedeker, Alyssa Simonson, Jun Ma, Amy Lang, Gladys Rodriguez, Arthur Rosenthal, Kyriakos Papadopoulos, Amita Patnaik, Drew Rasco, Lon Smith, Murali Beeram, Ronald Drengler, Luis Rodriguez, Steven Abbate, Scott Ulmer, Michael J Wick. Nonclinical activity of fulvestrant in a panel of ER+ breast XPDX models representing clinically acquired and innate resistance to endocrine therapies [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-01-11.

Background: Trastuzumab deruxtecan (DS-8201a) is an antibody-drug conjugate (ADC) consisting of an anti-HER2 (human epidermal growth factor receptor 2) antibody linked to a topoisomerase I inhibitor payload using a cleavable tetrapeptide-based linker and was recently approved for unresectable or T-DM1-refractory HER2+ breast cancer. While some mechanisms for clinical T-DM1 resistance have been identified, less is known about acquired or innate resistance to DS-8201a. We established two XPDX models of ER+/HER2+ breast cancer from tissue and fluid samples collected simultaneously from the same patient. These models designated ST4480B and ST4480C were developed and characterized for receptor expression, genomic alterations, and in vivo drug sensitivities toward multiple chemotherapies and targeted agents including DS-8201a and T-DM1. Methods: ST4480B and ST4480C were established from a 70-year-old Caucasian female with ER+/HER2+ metastatic breast cancer pretreated with chemotherapy and targeted agents including T-DM1 for nine months followed by capecitabine/trastuzumab/tucatinib combination for one year prior to sample collections. ST4480B was established from a lymph node biopsy and ST4480C from a fluid sample collected the same day; both were grown subcutaneously in female athymic nude mice supplemented with estradiol. The resulting models were passaged and receptor expression confirmed immunohistochemically; genomic analysis, including WES and RNAseq, was performed to further characterize models. For in vivo studies, both models were evaluated with several chemotherapy and targeted agents alone and in combination including: trastuzumab, pertuzumab, T-DM1, DS-8201a, neratinib, tucatinib, alpelisib, everolimus, and irinotecan. In vivo study endpoints included tumor volume and time from treatment initiation with %T/C values and tumor regression reported at study completion; a %T/C of ≤ 20 versus control was considered sensitive. Tumor regression (%T/C=—<0) versus Day 0 tumor volume was also reported. Results: ST4480B and ST4480C retained comparable receptor expression (ER=2+/HER2=2+) over tested passages with similar histology compared with archival clinical samples. DNA/RNA sequencing identified several conserved variants including PIK3CAE545K and TP53Q192* mutations and an ESR1-CCDC170 fusion. However, two variants identified only in ST4480C sequences included PIK3CAG1007R and PTENS287L. In vivo, ST4480B and ST4480C were found resistant to T-DM1 up to 10 mg/kg weekly with an average %T/C of 46 and 100, respectively. However, DS-8201a treatment at 3 mg/kg weekly resulted in partial tumor regressions in ST4480B (%T/C=-68) while ST4480C was found resistant to the therapy up to 10 mg/kg weekly (%T/C=84%). Both models were found resistant to trastuzumab, pertuzumab, neratinib, tucatinib, and irinotecan but sensitive to both alpelisib and everolimus. Conclusion: We established two XPDX models representing T-DM1-resistant, ER+/HER2+ breast cancer from both tissue and fluid samples collected simultaneously from the same patient which were found differentially responsive to DS-8201a. These models can be utilized as a valuable tool in better understanding innate resistance to DS-8201a. Citation Format: Johnnie R Flores, Anna Stackpole, Abimael Garza, Alexandra Ulmer, Alyssa Simonson, Kyriakos Papadopoulos, April Cabang, Jun Ma, Amita Patnaik, Drew Rasco, Amy Lang, Gladys Rodriguez, Murali Beeram, Michael J Wick. Establishment and characterization of two simultaneously developed T-DM1-resistant, ER+/HER2+ XPDX models from the same patient with differential in vivo sensitivity to trastuzumab deruxtecan (DS-8201a) [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-01-09.