93P - Transcriptome changes of immune cells across chemotherapy of triple-negative breast cancer

Background

Immune system plays an important role in the development, treatment response, and progression of triple-negative breast cancer. Here, we aimed to assess the dynamics of cell populations and their transcriptome changes during neoadjuvant chemotherapy (NACT) in triple-negative breast cancer patients.

Methods

Mononuclear cells were purified from peripheral venous blood before NACT, after 3rd and 21st days of NACT using a ficoll density gradient. Total cell concentration and viability (Calcein/DRAQ7) were assessed by flow cytometry (Cytoflex, Beckman Coulter). Single cells were sequenced on a NextSeq 2000 (Illumina) using targeted panel Immune Response from Rhapsody Express (Becton Dickinson). Data were analyzed using Seurat and SingleR.

Results

According to single cell sequencing data we revealed 10 main populations of mononuclear leukocytes: В cells (CD79+), NK cells (KIR2DL+), classical (CD14+) and non-classical (CD16+) monocytes, Т-cytotoxic cells (CD8+), Т-helpers (CD4+), T-regs (FOXP3+), CD4 memory cells (CD4+, IL32+, KLRB+, RGS1+), CD8 naïve (CD8+, LEF1+, CCR7+, PIK3IP1+), CD4 naïve (CD4+, LEF1+, CCR7+, PIK3IP1+). 3 days after NACT overexpression of BAX, FOSb, JUN, CXCR4, CD69 genes were observed, that mediating apoptosis of CD4 memory/naïve and CD8 naïve lymphocytes, T regs and monocytes; chemotaxis and proliferation of NK and Т-cytotoxic lymphocytes. The population of blood monocytes was selectively depleted (up to 3%). 21 days after NACT overexpression of CCR2, S100A9, ITGAM, CXCL8, CCL4, CCL5 genes were observed, mediating migration of monocytes to tissue, chemotaxis of immune cells and inflammation. The population of monocytes increased by up to 20%.

Conclusions

Chemotherapy affects the quantitative and qualitative composition of mononuclear leukocytes and changes their transcriptomic profile towards proinflammatory activation. This study was funded by the Russian Science Foundation (grant #22-75-10128).

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

The authors.

Funding

Russian Science Foundation, no.22-75-10128.

Disclosure

All authors have declared no conflicts of interest.