Priore, G. Del

Abstract Background: Currently, the gold standard in cancer diagnosis, prognosis, and treatment is assessing cancer cell mitosis in tumor tissue biopsies, which identifies patients with more aggressive cancer subtypes that may require more tailored treatment regimens. Circulating tumor cells (CTCs), cancer cells that detach from primary tumors, enter the bloodstream, and seed metastatic sites, are a well-known non-invasive blood biomarker which can also be used to aid in stratifying metastatic breast cancer (mBC) patients with highly aggressive subtypes. Initial pilot studies have identified a particular subtype of CTCs which are undergoing mitosis that appears to correlate with very poor survival outcomes compared to non-mitotic CTCs alone. In a multi-institutional prospective study, we isolated CTCs from the blood of n=167 mBC patients to categorize mitotic CTCs and evaluate their association with progression-free survival (PFS) and overall survival (OS) over a 3 year period. Methods: Peripheral blood samples (7.5ml) were collected from n=167 mBC patients progressing on systemic therapies and prior to new systemic therapy induction (i.e. chemotherapy n=102, hormone therapy n=13, immunotherapy n=63, targeted therapy n=62). Samples were processed using CellSieve microfilters to isolate CTCs via size exclusion and fluorescently stained for CD45, Cytokeratin, and DAPI. CTCs were imaged, enumerated, and subtyped based on the presence of ≥1 mitotic event, using previously established visual indicators. PFS and OS were assessed over 3 years by censored univariate and multivariate analyses. Results: CTCs were detected in ∼47% (n=79/167) of patients, while mitotic CTCs were identified in a subset of this group, ∼48% (n=38/79). The presence of any CTC was associated with poor outcomes (PFS HR=0.49, 95%CI=0.3-0.7, p=0.0002 and OS: HR=0.42, 95%CI=0.3-0.7, p=0.0002). However, the presence of mitotic CTCs correlated with significantly poorer outcomes compared to non-mitotic CTCs (PFS: HR=0.3, 95%CI=0.2-0.5, p<0.0001 and OS: HR=0.3, 95%CI=0.2-0.6, p=0.0002). Further, patients with no CTCs had a median PFS (mPFS) of 6 months and median OS (mOS) of 22 months, patients with non-mitotic CTCs had a mPFS of 4 months and mOS of 14.5 months; patients with mitotic CTCs had a mPFS of 2.4 months and mOS of 4.4 months. Notably, 95% of patients with a mitotic CTC progressed within 6 months, regardless of systemic therapy type. Interestingly, there were 5 patients who survived longer than 1 year with a mitotic CTC all of which were receiving specific targeted therapies. Conclusion: In this study, the detection of CTCs indicated poor outcomes in patients. Additional cytological analysis of CTCs further identified CTCs undergoing mitosis which correlated to highly aggressive disease with the poorest survival rates. These results indicate that while the presence of CTCs is a prognostic indicator for survival, subtyping CTCs and identifying mitosis may correlate to therapy response. This study highlights the need for larger, more comprehensive studies to expand upon and validate these findings. Citation Format: A. B. Duffy, M. Cristofanilli, C. Reduzzi, W. V. Williams, G. Del Priore, S. Chumsri, C. Tang, T. Iwase, N. T. Ueno, D. L. Adams. Mitotic Circulating Tumor Cells Correlates with Poor Clinical Outcomes in Metastatic Breast Cancer Patients over a 3 Year Period [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS4-01-05.

Abstract Background Management of metastatic breast cancer (MBC) continues to present obstacles, as many pts develop resistance to or cannot tolerate standard of care treatments. Bria-IMT combines two modalities to both overcome immune exhaustion and minimize the toxic safety profiles commonly observed in contemporary late stage therapies: an allogeneic whole-cell vaccine (SV-BR-1-GM) and checkpoint inhibition (CPI). The SV-BR-1-GM cell line originates from breast cancer cells and are engineered to secrete GM-CSF and irradiated before administration. These cells upregulate surface presentation of tumor associated antigens on both HLA class I and II, enhancing tumor recognition by T cells. Coadministration w/ an anti-PD-1 CPI, the vaccine response is reinforced, counteracting the immuno-suppressive tumor microenvironment. Methods This phase I/II study evaluated the Bria-IMT regimen: low-dose cyclophosphamide (Day –2) followed by SV-BR-1-GM and CPI (pembrolizumab or retifanlimab), then low-dose local peg- interferon α. Cycles q3w. In the phase II study, pts were randomized 1:1 to start CPI immediately at cycle 1 (C1) or delayed start at cycle 2 (C2). There were 2 formulations of SV-BR-1-GM w/ IFNγ stimulation (IFNγ) or unstimulated (PH3). Cancer-associated macrophage-like cells (CAMLs), circulating tumor cells (CTCs), and PD-L1 expression scores on these cells were monitored as biomarkers. Candida skin test assessed anergy status before cycle 1, and a delayed type hypersensitivity (DTH) skin test to SV-BR-1-GM was assessed at each cycle w/ a small SV-BR-1-GM dose before the full dose. Results Results of the randomized phase II study cohort are reported (n = 32). Retifanlimab was co-administered w/ SV-BR-1-GM at C1 or C2. Median age 61 (41-80) years; median number of prior regimens was 6 (2-13). 20(63%) pts were ER+/PR+/HER2-; 3(9%) HER2+, and 9 (28%) triple-negative (TNBC). The Bria-IMT regimen was well tolerated, w/ no drop-outs attributable to Bria-IMT. The majority of adverse events (93.8%) grade 1-2, w/ (40.6%) grades 3 and 4. Common AEs included injection site reactions (53%), nausea (43.8%), and fatigue (34%). The median progression free survival (PFS) was 3.5 (range: 1.8-24.5) and overall survival (OS) 9.5 (2.4-31.6) months. One patient remains on therapy, approaching 25 months of PFS. PFS and OS among pts who received immediate vs. delayed start of CPI (PFS 3.7 vs. 3.3 mo, HR: 0.59; 95% CI 0.3-1.2, p=0.16 | OS 13.3 vs. 7.4 months, HR: 0.78; 95% CI 0.34-1.8, p = 0.56). Pts who received the SV-BR-1-GM Ph3 formulation (n=21) had an increase in both PFS (4.1 vs 2.6 months, p = 0.005) and OS (16.6 vs. 9.1 months, HR: 0.6; 95% CI 0.2 - 1.4, p = 0.24); best objective response rate was 12% and the clinical benefit rate was 53%. In ER+/PR+/HER2-, overall OS was 11.7 (3.5-31.6) mo, 17.3 mo (3.5-27.9) in Ph3; HER2+ 7.2 (4.7-24.5) mo, all received Ph3; TNBC 9.9 (2.4-23.0) mo, 16.6 mo (2.4-23.0) in Ph3. In pts w/ available data, pts w/ post-dose CTC count < 5 significantly improved OS compared w/ a CTC count > 5 (16.6 vs. 5.5 months, p = 0.0003). Pts w/ positive DTH response had significantly improved PFS (3.6 vs. 2.4 months, p = 0.01) and OS (11.9 vs. 4.7 months, p < 0.0001). Conclusions In heavily pretreated MBC pts, the Bria-IMT regimen demonstrated promising results and a favorable safety profile across all subtypes of breast cancer. A clinically meaningful, though not statistically significant, difference in OS was observed between pts receiving CPI at C1 versus C2. Superior outcomes were observed in pts receiving SV-BR-1-GM w/o IFNγ incubation, guiding selection of Ph3 formulation for ongoing and subsequent clinical trials. Pts w/ post-dose CTC count < 5, and positive DTH response had significantly improved outcomes. On basis of these results, a randomized phase III trial comparing the Bria-IMT regimen to physician's choice is ongoing (NCT06072612). Trial information: NCT03328026. Citation Format: C. Nangia, S. Chumsri, K. Rowland, L. Negret, J. Knecth, B. Bayer, T. Aghajanian, W. Williams, G. Del Priore, C. Calfa. Survival Results of Phase II Bria-IMT Allogenic Whole Cell-Based Cancer Vaccine [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS1-13-23.