Deciphering the potential pathogeny of rare tracheal adenoid cystic carcinoma by single-cell RNA-sequencing

Tracheal adenoid cystic carcinoma (TACC) is an extremely rare type of cancer, characterized by slow growth, but high rates of recurrence and metastasis. The treatment of TACC is greatly hampered by the limited understanding of its molecular characteristics. We sought to conduct an in-depth analysis of this type of cancer through single-cell RNA-sequencing (RNA-seq) to provide important evidence for the formulation of treatment strategies for TACC.

The peripheral blood, tumor tissue, and adjacent tissue of the same TACC patient was obtained and tested by single-cell RNA-seq. A bioinformatics analysis was conducted to identify the cell clones with key biological functions and determine their molecular characteristics. Fluorescent multiplex immunohistochemistry (mIHC) was used to confirm the markers with abnormal expression.

RNA-seq and mIHC verification were successfully performed in one and four TACC patients, respectively. All the sequenced cells were divided into three main categories of epithelial, immune, and stromal cells. In relation to the epithelial cells in the tumor tissue, ciliated cells expressing commonly observed stem-cell markers were found at the beginning of the pseudo-time curve, indicating that they might be related to the origin of the TACC. Further analysis revealed eight genes in the ciliated cells may warrant further investigation. Additionally, the function of immune cells like natural killer (NK) cells and cytotoxic T cells were found to be impaired, which might be the reason for the expansion of TACC. Notably, a distinct subgroup of mesothelial cells disguised as stromal cells was identified. These cells possessed a malignant phenotype and probably interact with macrophages through the MIF-(CD74 + CXCR4 or CD44) pathway. The high density of the CD68⁺CD74⁺ cells and their close distance to the Vimentin⁺PANCK⁺ cells was confirmed by mIHC. These findings suggest that these markers could be used in the development of drugs targeting TACC.

Our research showed that TACC might originate from ciliated epithelial cells. Abnormal gene expression triggered by external factors, along with the dysfunction of NK and T cells, are critical factors in the genesis and progression of TACC. Developing drugs that target MIF-CD74 signaling pathways may lead to breakthroughs in the treatment of TACC.