Integrated single cell and bulk multi-omics reveals heterogeneity and early changes in pathways associated with cetuximab resistance in HNSCC sensitive cell lines

Abstract

Identifying potential mechanisms of resistance while tumor cells still respond to therapy is critical to delay acquired resistance. We generated the first comprehensive multi-omics, bulk and single cell data in sensitive head and neck squamous cell carcinoma (HNSCC) cells to identify immediate responses to cetuximab. Two pathways potentially associated with resistance were focus of the study: regulation of receptor tyrosine kinases through the transcription factor TFAP2A, and epithelial-to-mesenchymal transition (EMT) process. Single cell RNA-seq demonstrates heterogeneity, with cell specific TFAP2A and VIM expression profiles in response to treatment. RNA-seq and ATAC-seq reveal global changes within five days of cetuximab therapy, suggesting early onset of mechanisms of resistance; and corroborates cell line heterogeneity, with different TFAP2A targets or EMT markers affected by therapy. Lack of TFAP2A reduces HNSCC growth and is enhanced by cetuximab and JQ1. Regarding the EMT process, short term cetuximab therapy has the strongest effect on inhibiting migration. TFAP2A silencing does not affect cell migration, supporting an independent role for both mechanisms in resistance. Overall, we show that immediate adaptive transcriptional and epigenetic changes induced by cetuximab are heterogeneous and cell type dependent; and independent mechanisms of resistance arise while tumor cells are still sensitive to therapy.

Integrated single-cell and bulk gene expression and ATAC-seq reveals heterogeneity and early changes in pathways associated with resistance to cetuximab in HNSCC-sensitive cell lines

“…We declare that preliminary data of this study was published as an abstract 57 and presented as a poster at the 2019 AACR Annual Meeting, and the original paper before submission to peer-review was deposited at BioRxiv. 58…”

Prognostic/predictive markers in systemic therapy resistance and metastasis in breast cancer

“…38 Recently, advanced technology and assays such as single-cell genomics and spatial transcriptomics, have been applied to improve the quality of detailed assessment of the TILs, by dissecting into functional TILs, regulatory T cells, and exhausted T cells that can explain the difference response to the ICI therapy in breast cancer treatments. 39,40 Application of complex biomarker assessment is inevitable, given the wide adaptation of the ICI in the field of breast oncology; therefore, more data on the use of functional TILs is expected to emerge in the near future. 41…”

Precision Medicine Approaches to Overcome Resistance to Therapy in Head and Neck Cancers

“…Indeed, several authors report that HNSCC cells exhibiting a mesenchymal-like phenotype are resistant to CTX treatment in vitro and in vivo (xenografts) ( 174 , 176 , 177 ). Several potential mechanisms implicated in this EMT-induced CTX resistance are observed, such as (i) expression of lymphotoxin-b; (ii) methylation of EGFR that promotes the EGFR ligand-binding ability and dimerization (EGFR persistent activity) ( 169 ); (iii) secretion of CTX-containing extracellular vesicles, which lead to cancer cell protection ( 179 ); (iv) upregulation of EMT-related genes ( 133 ), especially by epigenetic regulation ( 170 , 180 ); and (v) loss of the tumor suppressor gene SMAD4, which induces JNK and MAPK pathway activation ( 172 , 173 ). Indeed, Ozawa et al.…”