24P - Single cell transcriptomics of the immune cells during chemotherapy in triple-negative breast cancer patients

Background

Chemotherapy is the mainstay of systemic treatment for patients with triple-negative breast cancer (TNBC) due to the lack of targets for targeted therapy (HER2/neu; ER; PR). Components of the immune system are known to be involved in the response to chemotherapeutic treatment. Here we aimed to study the dynamics of changes in immune cell composition during neoadjuvant chemotherapy (NACT).

Methods

The immune cells were purified from peripheral blood and biopsy samples of 5 TNBC patients before NACT and 21st days of first course of NACT (AC regimen). Total cell concentration and viability (Calcein/DRAQ7) were assessed by flow cytometry (Cytoflex, Beckman Coulter). Single cells were sequenced on a Genolab M platform (GeneMind Biosciences) using 10x Genomics technology for fixed multiplexed samples. Data were analyzed using Seurat and SingleR.

Results

Sequencing revealed a variety of immune cell populations including: B cells, DC cells, NK cells, CD4+, CD8+ and T regulatory lymphocytes, classical and non-classical monocytes, granulocytes and others. In the blood, the pool of classical monocytes and NK cells were depleted up to day 21 after the first NACT cycle, while naive CD4+, CD8+ T cells and end effector CD8+ T cells, were increased by day 21. In the populations of monocytes and NK cells overexpression of RGS2, ANXA1, FGL2, MX1, IFI6 genes were observed, involved in induction apoptosis. CD8+T-cells, as well as Th-1 and Th-2 CD4+ T-cells were characterized by increased expression of NFKBIA, JUN, FOS, involved in T cell differentiation. In the tumor microenvironment, depletion of CD4+ T cells, CD8+ T cells and memory B cells and an increase in T-regulatory cells and plasma cells were observed. DUSP4, LCK, CXCR4, LTB, TNFRSF18, IL2RB and PTPN7 genes were overexpressed and involved in the regulation of cytokine secretion and inhibition of TCR signaling in CD4+ and CD8+ T cells; chemotaxis-mediated inflammation in memory B cells; T cells differentiation, and induction of invasion/migration of tumor cells in T-regulatory cells and plasma cells.

Conclusions

The first cycle of NACT induced renewal of the immune cells’ composition in the blood and tumor microenvironment skewing towards to immune disfunction.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

Cancer Research Institute, Tomsk National Research Medical Center, Russian Academy of Sciences, Tomsk, Russian Federation.

Funding

Russian Science Foundation (grant #22-75-10128).

Disclosure

All authors have declared no conflicts of interest.