Abstract 1389: Single-cell sequencing to identify genetic variant accelerating shedding of CTCs into peripheral blood in gastric cancer patients

Early detection of disseminated tumor cells in the peripheral blood of patients with early stage gastric cancer could help to improve the outcome after tumor resection. The aim of this study is to evaluate the prognostic significance of tumor-related mRNA for the detection of circulating tumor cells in gastric cancer patients by a reverse-transcriptase polymerase chain reaction (RT-PCR) method. We simultaneously analyzed human telomerase reverse transcriptase (hTERT), cytokeratin-19 (CK-19), cytokeratin-20 (CK-20) and carcinoembryonic antigen (CEA) mRNA (messenger RNA) expression in the peripheral blood of 42 gastric cancer patients and 30 healthy individuals. Additionally, analyses were carried out for the correlation of these four molecular markers with patients’ clinicopathologic features, as well as the occurrence of postoperative recurrence/metastasis. Among 42 gastric cancer patients, the prevalence of mRNA for hTERT, CK-19, CK-20, and CEA was 61.9% (26/42), 69% (29/42), 61.9% (26/42), and 78.6% (33/42), respectively. All 30 healthy individuals were negative for hTERT and CEA mRNA, while two were positive for either CK-19 mRNA or CK-20 mRNA. Positive CEA mRNA was significantly correlated with tumor size (p= 0.008), vessel invasion (p= 0.001), depth of tumor invasion (p= 0.007), lymph node metastasis (p< 0.001), and TNM stage (p< 0.001). In addition, the multivariate logistic regression demonstrated that CEA mRNA expression was an independent and significant predictor for postoperative recurrence/metastasis (p= 0.032). Our findings suggest that CEA mRNA may be a more reliable marker than hTERT, CK-19 and CK-20 for the detection of circulating cancer cells in gastric cancer patients' peripheral blood. Patients with positive CEA mRNA expression in peripheral blood have a significantly higher risk of postoperative recurrence/metastasis.

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Clinical significance of changes of TLR4/MyD88/NF-κB signaling pathway in peripheral blood mononuclear cells of gastric cancer patients with Helicobacter pylori infection

World Chinese Journal of Digestology ◽

10.11569/wcjd.v29.i6.319 ◽

2021 ◽

Vol 29 (6) ◽

pp. 319-324

Author(s):

Yang Wang ◽

Xin-Mei Zou ◽

Qin-Mei Pan ◽

Li-Ping Zhong

Keyword(s):

Gastric Cancer ◽

Helicobacter Pylori ◽

Cancer Patients ◽

Clinical Significance ◽

Peripheral Blood Mononuclear Cells ◽

Peripheral Blood ◽

Mononuclear Cells ◽

Peripheral Blood Mononuclear ◽

Blood Mononuclear Cells ◽

Gastric Cancer Patients

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Single-Cell Sequencing Reveals the Transcriptome and TCR Characteristics of pTregs and in vitro Expanded iTregs

Frontiers in Immunology ◽

10.3389/fimmu.2021.619932 ◽

2021 ◽

Vol 12 ◽

Author(s):

Zhenzhen Hui ◽

Jiali Zhang ◽

Yu Zheng ◽

Lili Yang ◽

Wenwen Yu ◽

...

Keyword(s):

Colorectal Cancer ◽

Single Cell ◽

Cancer Patients ◽

Peripheral Blood ◽

Critical Role ◽

Interleukin 2 ◽

Treg Cells ◽

Single Cell Sequencing ◽

Colorectal Cancer Patients

Regulatory T cells (Tregs) play a critical role in the maintenance of immune tolerance and tumor evasion. However, the relative low proportion of these cells in peripheral blood and tissues has hindered many studies. We sought to establish a rapamycin-based in vitro Treg expansion procedure in patients diagnosed with colorectal cancer and perform single-cell sequencing to explore the characteristics of Treg cells. CD25+ cells enriched from peripheral blood mononuclear cells (PBMC) of colorectal tumor patients were cultured in X-VIVO15 medium, supplemented with 5% human AB serum, L-glutamine, rapamycin, interleukin-2 (IL-2), and Dynabeads human Treg expander for 21 days to expand Tregs. Treg cells with satisfactory phenotype and function were successfully expanded from CD4+CD25+ cells in patients with colorectal cancer. The median expansion fold was 75 (range, 20–105-fold), and >90.0% of the harvest cells were CD4+CD25+CD127dim/− cells. The ratio of CD4+CD25+Foxp3+ cells exceeded 60%. Functional assays showed that iTregs significantly inhibited CD8+T cell proliferation in vitro. Single-cell sequencing showed that the transcriptome of pTreg (CD4+CD25+CD127dim/− cells isolated from PBMC of colorectal cancer patients) and iTreg (CD4+CD25+CD127dim/− cells expanded in vitro according to the above regimen) cells were interlaced. pTregs exhibited enhanced suppressive function, whereas iTregs exhibited increased proliferative capacity. TCR repertoire analysis indicated minimal overlap between pTregs and iTregs. Pseudo-time trajectory analysis of Tregs revealed that pTregs were a continuum composed of three main branches: activated/effector, resting and proliferative Tregs. In contrast, in vitro expanded iTregs were a mixture of proliferating and activated/effector cells. The expression of trafficking receptors was also different in pTregs and iTregs. Various chemokine receptors were upregulated in pTregs. Activated effector pTregs overexpressed the chemokine receptor CCR10, which was not expressed in iTregs. The chemokine CCL28 was overexpressed in colorectal cancer and associated with poor prognosis. CCR10 interacted with CCL28 to mediate the recruitment of Treg into tumors and accelerated tumor progression. Depletion of CCR10+Treg cells from tumor microenvironment (TME) could be used as an effective treatment strategy for colorectal cancer patients. Our data distinguished the transcriptomic characteristics of different subsets of Treg cells and revealed the context-dependent functions of different populations of Treg cells, which was crucial to the development of alternative therapeutic strategies for Treg cells in autoimmune disease and cancer.

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