Redaelli, Matteo

(1) Background: Mammographic breast density (MBD) is a well-established predictor of breast cancer risk, yet the biological mechanisms underlying this association remain incompletely understood. MBD is characterized by alterations in breast stromal architecture, including increased collagen deposition and changes in immune cell composition. Given emerging evidence that the breast harbors a resident microbiome, we investigated whether the breast tissue microbiome correlates with MBD. (2) Methods: Adjacent normal breast tissue was collected under sterile conditions from 33 women undergoing surgery for benign or malignant breast disease. DNA was extracted and subjected to 16S rRNA gene sequencing (Illumina MiSeq). (3) Results: We observed a non-significant trend toward lower α-diversity in high-MBD samples compared to low-MBD samples, p = 0.13. β-Diversity analyses identified a modest association between MBD and microbial community composition (MiRKAT p = 0.049). A random forest-based model incorporating genus-level relative abundances improved prediction of MBD over clinical characteristics alone, identifying Corynebacterium (Actinobacteria) and other genera as key predictors. (4) Conclusions: Breast tissue microbial features vary with mammographic breast density, suggesting a potential association with density-associated breast cancer risk. These exploratory findings warrant validation in larger cohorts to better elucidate biological mechanisms and clinical relevance.

ABSTRACT Background and Objectives Neoadjuvant chemoimmunotherapy (NACI) for node‐positive breast cancer may induce immune cell activation in regional lymph nodes (LN), confounding post‐NACI imaging. Here we evaluated RECIST 1.1 criteria to predict post‐NACI pathologic LN status. Methods We studied patients with biopsy‐proven cN+ non‐distant metastatic breast cancer receiving NACI operated on 03/2020–09/2024 with both pre‐ and post‐NACI cross‐sectional imaging. Per RECIST 1.1, LN short axis diameter (SAD) was measured in target (defined as SAD ≥ 15 mm) and nontarget (SAD 10–14.9 mm) LNs. Groups were compared using Fisher′s exact test. Results 75 patients, median age 53 years, were studied: 61% cN1, 12% cN2, and 27% cN3. Baseline median number of imaging‐suspicious LN was 4 (IQR 2‐5). Post‐NACI, 64% had a nodal pathologic complete response (pCR/ypN0). 55% met RECIST 1.1 LN assessment criteria. 21 of 32 (66%) with an imaging CR (iCR) were ypN0, while 8 of 9 (89%) without an iCR were ypN0. Neither target ( p = 0.24) nor combined target/nontarget LN iCR ( p = 0.76) predicted ypN status. Nodal pCR rates were higher in those with ≤ 1 versus > 1 suspicious LN post‐NACI (71% vs. 42%, p = 0.03). Conclusions RECIST 1.1 criteria did not predict nodal pCR for NACI‐treated node‐positive breast cancer patients. Other post‐NACI imaging assessment strategies are needed.

Abstract Breast cancer is the leading cause of cancer death among women worldwide. Most cases occur in those without identifiable modifiable risk factors, highlighting critical gaps in our understanding of breast cancer biology including the influence of the gut microbiome. Here we studied the gut microbiome with shallow shotgun metagenomic sequencing from preoperative stool samples in a well-characterized clinical cohort of 131 women, median age 59 years, undergoing breast-conserving surgery for benign (n=23), high-risk (n=47), or malignant (n=61) breast disease. Taxonomic profiling was performed with Sourmasth 4.2.4 with GTDBv202 as reference database; functional profiling of metagenomic data employed HUMAnN 3.6 with identified gene families mapped to MetaCyc pathways. Differences in alpha diversity in terms of species richness were observed across diagnosis groups (benign, high-risk, malignant) and was lowest for high-risk lesions (p=0.031). Decreased alpha diversity was also associated with higher body mass index (BMI) (Shannon p=0.12, Inverse Simpson p=0.025). Significant beta diversity differences across diagnosis groups also were seen (Bray-Curtis p=0.044, Jaccard p=0.013) as were differences in association with BMI (Bray-Curtis, Jaccard, both p=0.002). No significant diversity differences associated with age, mammographic breast density or menopausal status were observed. Functional pathway profiling stratified by BMI showed a correlation between increasing BMI and greater relative abundance of genes in nicotinamide adenine dinucleotide (NAD) biosynthesis, folate metabolism, and aromatic amino acid metabolism pathways (p=0.0279, 0.0667 and 0.0860, respectively) - pathways implicated in breast cancer detection, survival and prognosis. Cross-referencing our significantly (FDR <5%) differentially abundant taxonomic findings with these functional pathway data identified enrichment of several Blautia species, linked to NAD biosynthesis via tryptophan metabolism, in patients with higher BMI. We also noted enrichment of Dorea formicigenerans, a species associated with increased breast cancer risk in obese women due to its pro-inflammatory effects, in individuals with a higher BMI. Here we report significant differences in gut microbiome diversity, taxonomy and functional pathway signals in association with BMI in women with breast disease. Differentially abundant species seen with rising BMI may contribute to functional pathways that drive increased breast cancer risk and may provide an actionable target for risk reduction independent of conventional risk reduction strategies. Citation Format: Jack W. Sample, Matteo Redaelli, Stephen Johnson, Tanya L. Hoskin, Marina R. Walther-Antonio, Jun Chen, Tina J. Hieken. Obesity alters cancer-related microbial pathways in women with benign and malignant breast disease [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2213.