Evaluating gut microbiome as a biomarker of pathological complete response in patients with early locally advanced triple negative breast cancer (LA TNBC): A pilot study.
Authors: Sarfraz Humaira , Czerniecki Brian J. , Stankiewicz Kaylee , Jaglal Michael Vishal , Jain Shalini , Yadav Hariom , Liu Min , Shukla Rohit , Kumar Vivek , Alhomsi Muhammad , Hathout Mahmoud , Tuz Zahra Fatima , Darwin Alicia , Joshi Utsav , Sen Amit , Muzaffar Jameel , Bari Shahla 
Published: May 2024

593

Background: It has previously been shown that microbial metabolic pathways are differentially activated in immunotherapy resistance, hence can serve as potential biomarker as well as target for therapeutic intervention. With the advent of Immune Checkpoint Blockers (ICB) in the neoadjuvant (NA) setting of locally advanced (LA) TNBC, we hypothesized that similar microbial metabolite mediated resistance may be at play. We are presenting data demonstrating differences in microbiome and metabolites in these patients. Methods: 28 patients who received NA pembrolizumab and chemotherapyfor LA TNBC were identified at Moffitt Cancer Center and Tampa General Hospital between February 2022 to October 2023. Blood and stool samples were collected at therapy initiation, at completion of neoadjuvant treatment and/or when the patients developed grade 3 or higher toxicity to therapy. We evaluated stool metagenomics and untargeted stool/plasma metabolomics as correlatives. Patient characteristics were as follows: 21 Caucasian, 3 African-American, 2 Hispanic, 2 unknown, median age 51 years, median body mass index (BMI) 28.5 kg/m2. 17 patients had stage II, while 11 patients had stage III TNBC. Results: Response was assessed in 27 patients, of which 11 had pCR (ypT0/is ypN0) while 16 had residual disease (RD). 1 patient progressed, while on NA therapy, while 8 patients developed grade 3-4 immune related adverse events leading to treatment discontinuation and death in one patient. We conducted shallow shotgun metagenomic sequencing and untargeted metabolomic profiling on pre and post treatment fecal and plasma samples respectively to identify gut microbial diversity, composition and differentially abundant metabolites associated with pCR.There wasno difference in alpha and beta diversity; however significant enrichment of microbiota with genus Akkermansia HR 0.04, p 0.03 was noted in those with pCR. We observed higher abundance of tryptophan metabolizing clostridium species HR 50541, p <0.001, along with differential activation of microbial tryptophan pathway in those with residual disease, specifically higher abundance of hydroxyindole acetic acid HR 1.54 P<0.005, indole propionic acid HR 1.75 p 0.05 and indole butyric acid HR 1.86 p 0.04. Conclusions: To our knowledge this is the first analysis of differences in the microbiome and microbial metabolism in patients with LA TNBC treated with NA ICB. Despite being a small study, it shows a more aggressive disease pattern with higher RD than noted in KEYNOTE 522 and shows significant concern for serious immune mediated adverse effects. This highlights the importance of biomarkers to stratify patients most likely to benefit from ICB. Enrichment of microbiota and immunosuppressive tryptophan metabolites were associated with pCR and should be validated in large studies as biomarker of response to NA ICB in TNBC.