Magbanua, MJM

Abstract Background: In MBC, CTC enumeration is highly prognostic for both progression free survival (PFS) and overall survival (OS). Multiparametric technologies enable for expanded phenotypic profiling of CTCs, including ER and HER2 assessment. Limited information is available on the concordance of CTC ER and HER2 levels compared to synchronously obtained tissue. We assessed the prognostic performance of CTC enumeration and phenotypic profiling of ER and HER2 on CTCs. Methods: The prospective, multi-institutional Aurora study enrolled pts with MBC of any subtype either at diagnosis of metastatic disease or at time of progression on any line of systemic therapy. Pts underwent a metastatic site biopsy, consented to archival primary tissue collection and donated blood samples serially every 3-4 months until progression. CTCs, from blood collected at study entry, were identified as nucleated, EpCAM+/ cytokeratin+/ CD45- cells using an automated imaging system (RareCyte, Seattle). ER and HER2 expression levels were measured by mean fluorescence intensity (MFI), with thresholds above 80 (ER) and 160 (HER2) considered positive. Comparisons were made to total RNAseq from paired metastatic tissue biopsies. PFS and OS were calculated from the time of study entry to last follow up or death. Results: Baseline CTC enumeration was performed in 154 pts. The most common clinical subtype was HR+/HER2- (59.7%), followed by HER2-positive (16.8%) and TNBC (15.5%). The median lines of prior therapy were 1 (range 0-10). At least 1 CTC was identified in 68/154 pts (44%), 30 (19%) had ≥ 5 CTCs, and 15 (9.7%) had CTC clusters. The median CTC value was 0 (0-877). The presence of CTCs (≥1 or ≥5) and CTC clusters was adversely associated with PFS and OS (Table 1 Table 1. CTC survival analyses Number Median PFS PFS HR (95% CI) Median OS OS HR (95% CI) Presence of CTC 0 CTC 86 7.37 (0.03 - 43.1) reference 20.72 (0.03 - 50.6) reference ≥ 1 CTC 68 4.7 (0.43 - 19.5) 1.65 (1.16-2.36) 12.04. (0.43-44.1) 2.13 (1.44-3.16) Presence of ≥ 5 CTC <5 CTC 124 7.7 (0.03 - 43.1) reference 19.31 (0.03 - 50.6) reference ≥ 5 CTC 30 2.88 (0.43 - 18.7) 1.77 (1.15 - 2.71) 5.74 (0.43 - 29.3) 3.59 (2.26-5.70) Presence of clusters 0 clusters 139 7.37 (0.03 - 43.1) reference 18.06 (0.03 - 50.6) reference ≥ 1 cluster 15 1.91 (0.43 - 18.7) 2.06 (1.16 - 3.68) 3.42 (0.43 - 18.7) 5.44 (2.94-10.09) ). Across all subtypes, the median CTC ER-MFI across was 36.2 (range 4.2-10,016.6) and the median HER2-MFI was 139.6 (range 4.2-2480.5). We observed high intra-patient heterogeneity of ER-MFI (CV: 0.65 [0.34-1.25]) and HER2-MFI (CV: 0.79 [0.42-1.54]). Clinical metastatic subtype from metastasis at study entry was available in 54/68 pts with CTCs. Pts with clinical ER+ MBC had a higher median ER-MFI compared to clinical TNBC (111.9 vs 68.6, p = 0.02). However, 11 pts (26.8%) with clinical ER+ MBC had all CTCs with an ER-MFI <80. Lower ER expression was observed in CTCs collected after prior exposure to a SERD (median ER-MFI: post-SERD [n=8] 27 vs post-aromatase inhibitor [n=4] 252, p<2.22e-16). No correlations were observed between the median CTC ER-MFI or HER2-MFI and paired tissue ESR1 or ERBB2 RNA expression (n=66). Conclusions: CTCs are strongly predictive of PFS and OS. CTC ER- and HER2-MFI were highly heterogeneous without clear correlation with bulk tissue RNA expression. The clinical impact of discordant clinical receptor status and CTC receptor status warrants further exploration. Citation Format: K. V. Giridhar, W. C. Nenad, H. Ye, G. L. Wheeler, T. Hinoue, A. R. Michmerhuizen, B. M. Felsheim, U. R. Chandran, B. J. Kelly, E. R. Mardis, P. W. Laird, J. M. Balko, A. C. Garrido-Castro, S. Coppens, J. Bowen, R. Nanda, N. U. Lin, C. K. Anders, C. Isaacs, J. J. Tao, J. D. Anampa, N. Jahan, L. Huppert, A. DeMichele, B. H. Park, S. Vinayak, A. L. Delson, M. M. Magbanua, M. Balic, H. A. Parsons, A. V. Lee, L. A. Carey, L. Norton, A. C. Wolff, N. E. Davidson, I. Krop, T. A. King, C. M. Perou, K. A. Hoadley, Aurora US Metastases Network. Characterizing circulating tumor cell (CTC) enumeration and phenotypic profiling in blood with synchronously obtained tissue in metastatic breast cancer (MBC) in TBCRC /AURORA US [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 PS2-07-17.

Abstract Background Invasive lobular carcinoma (ILC) differs from invasive ductal carcinoma (IDC) in its estrogen receptor (ER) signaling pathways, response to systemic therapy, and imaging features. We evaluated differences in early change in the proliferation marker Ki67 and features on magnetic resonance imaging (MRI) after novel endocrine based neoadjuvant therapy in lobular versus non-lobular cases on the I-SPY2 EOP Trial. Methods EOP is a prospective, randomized NET trial enriched for patients with ILC. Patients with stage II-III ER positive HER2 negative breast cancer with low molecular risk (Mammaprint low or Mammaprint Hi1 plus high Sensitivity to Endocrine therapy Index) were randomized to one of several endocrine therapy-based arms. Treatment arms include novel oral SERDS with or without CDK4/6 inhibitor, aromatase inhibitors, and SERMs with or without ovarian function suppression in pre-menopausal patients. Patients undergo repeat core needle biopsy with central Ki67 assessment after 3 weeks of treatment and serial breast MRI. Here, we compare endocrine therapy responsiveness as measured by change in Ki67 at 3 weeks and functional tumor volume (FTV) on pre-surgical MRI in lobular versus non-lobular cases. Results We evaluated 253 patients of whom 148 (58.5%) had non-lobular cancer (93.9% IDC), and 105 (41.5%) had lobular cancer (17.1% mixed ILC/IDC). Overall, mean age was 53.3 years, 63.8% of tumors were grade 2, and 86.5% were Mammaprint (MP) Low with the remaining being Hi1, with higher rates of MP Low in the lobular versus non-lobular cohorts (92.4% versus 83.4%, p=0.0358). At baseline, tumors in the lobular cohort had significantly lower mean Ki67 than those in the non-lobular cohort (11.2% [SD 12.8] versus 16.4% [SD 14.4%], p=0.0038). Consistent with this, the proportion with baseline tumor Ki67 <10% was significantly higher in the lobular cohort than the ductal cohort (60.4% versus 39.3%, p=0.001). At 3-week biopsy, mean Ki67 remained lower in the lobular versus non-lobular group (3.4% versus 5.4%, respectively, p=0.0415). The proportion of patients with Ki67 <10% at 3 weeks did not differ significantly between lobular and non-lobular cases (88.6% versus 79.7%, respectively, p=0.08). Among the 40 lobular cases and 88 non-lobular cases with baseline Ki67 ≥10%, week 3 Ki67 decreased to <10% in 90% of lobular cases and 72.7% of non-lobular cases (p=0.07). On MRI, mean FTV at baseline was similar among lobular and non-lobular cases (21 cc3 versus 16.3 cc3). On pre-surgical MRI, patients in the lobular cohort had significantly higher mean residual FTV than those in the non-lobular cohort (5.7 cc3 versus 3.8 cc3, p=0.034). Interestingly, when splitting the lobular cohort into mixed lobular and pure lobular, mean residual MRI FTV was 7.5 cc3 in mixed cases and 3.8 cc3 in pure lobular cases, reflecting that FTV decreased the least in mixed lobular cases (decrease of 51.1% in mixed cases, 69.7% in pure lobular cases, and 73.9% in non-lobular cases, p=0.0255). Conclusions These results highlight differences in lobular versus non-lobular breast cancer. Lobular cases had lower baseline Ki67, and similar rates of decrease in Ki67 after 3 weeks of NET than non-lobular cases. Despite this, mean FTV after completion of 6 months of NET was higher in lobular tumors. These findings may reflect differences in early versus late indicators of response to NET, or differential response patterns by histology. Alternatively, standard response measures such as Ki67 and FTV may need lobular-specific modifications. Ongoing work includes central assessment of Ki67 from surgical specimens, analysis of changes in gene expression signatures, and evaluation of surgical outcomes such as breast conservation rates by histologic subtype. Citation Format: R. Mukhtar, P. Norwood, A. Borowsky, S. Alkhafaji, K. Giridhar, C. Vaklavas, A. Elias, M. Wei, M. Goetz, N. Onishi, O. Olopade, L. Van 't Veer, L. Huppert, W. F. Symmans, L. Brown Swigart, C. Wu, C. Yau, D. Yee, M. Magbanua, E. Price, N. Hylton, L. Esserman, J. Chien. Differential early response to neoadjuvant endocrine therapy in lobular versus non-lobular breast cancer on the Endocrine Optimization Pilot (EOP) of the I-SPY2 Trial [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 PS4-02-01.

Abstract 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; and (2) 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=52), 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=32). HER2+ patients also received trastuzumab (H). Serial plasma was collected before NAT (T0), early treatment (3 weeks, T1), between regimens (12 weeks, T2), and after NAT prior to surgery (T3). Mutational profiles derived from pretreatment tumor biopsy and normal matched DNA whole exome sequencing were used to design personalized assays targeting 16 patient-specific somatic variants to detect ctDNA in serial plasma. Results: Of the 84 patients in this study, 43% were HR-/HER2- (TNBC), 35% HR+/HER2-, and 23% HER2+. In total, 74% (61 of 83), 35% (28 of 79), 14% (9 of 65), and 8% (5 of 61) were positive for ctDNA at timepoints T0, T1, T2, and T3, respectively. At T0, ctDNA positivity and levels (average number of mutant molecules detected per mL) were significantly associated with increased tumor burden (by clinical and MRI examination), more aggressive tumor biology (as reflected in higher Mammaprint scores and grade) and subtype (HER2+ and TNBC). Twenty-seven percent (27%) of the 84 patients achieved a pCR and all patients who were ctDNA-positive at T3 (n=5) did not achieve a pCR. Currently, data are being collected to: (1) assess the relationship of ctDNA and MRI imaging in predicting tumor response to therapy; (2) examine the relationship of ctDNA levels before and after NAT with 3-year DRFS and event-free survival (EFS). The results of these analyses will be 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. Citation Format: Magbanua MJM, Brown-Swigart L, Hirst GL, Yau C, Wolf D, Ma AA, Bergin E, Venters S, Sethi H, Wu H-T, Salari R, Tin T, Sawyer S, Louie M, Zimmermann B, Lin C-HJ, Keats J, Liang WS, Cuyugan L, Enriquez D, Tripathy D, Chien AJ, Forero A, DeMichele A, Liu M, Delson AL, Asare S, Esserman L, van't Veer L, I-SPY 2 Consortium. Personalized serial circulating tumor DNA (ctDNA) analysis in high-risk early stage breast cancer patients to monitor and predict response to neoadjuvant therapy and outcome in the I-SPY 2 TRIAL [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr PD2-01.