Single cell analysis reveals immune subtypes associated with CRT treatment

Cervical cancer (CC) is one of the most prevalent female malignancies globally in low- and middle-income regions, accounting for 7.5% of all female cancer deaths.Despite advancements in medical technology and treatment protocols,the combination of radiation therapy and chemotherapy, known as chemoradiotherapy (CRT), remains the standard of care for locally advanced cervical cancer. However, a substantial proportion of patients do not achieved a complete response to CRT, leading to treatment failure and poor clinical outcomes.Herein, we profiled the transcriptome of 50649 cells derived from six partial responsive(PR) tumor tissues and two complete responsive(CR) tissues of CSCC through scRNA-seq. We displayed a comprehensive cell landscape of the TME and investigated its function and lineage tracking. Overall design: Advanced CSCC patients(IVA or IVB) received cisplatin(75mg/m2) and paclitaxel(135-175mg/m2) for two cycles. Then, patients received radiotherapy (45Gy for 25 fractions) accompanied by cisplatin treatment(35-40 mg/m2, once a week). At last, patients received four cycles of cisplatin(75mg/m2) and paclitaxel(135-175mg/m2) treatment. The clinical response for each target lesion was evaluated based on RECIST v.1.1 criteria.Eight tumor tissues (two tumor tissues from CR patients and six tumor tissues from non-responsive PR patients) were obtained from post-treatment of CRT in CSCC patients for scRNA-seq.Tumor tissues were washed three times with PBS and then were sliced into 1-3 mm3 pieces. The tissue pieces were subsequently transferred to a 10-mL digestion medium containing 0.2% collagenase I/II, DNAse I (Sigma), and 25 units dispase in DMEM. The samples were placed on an incubated orbital shaker at 37C and 250 RPM for 15 min. Approximately 20 mL of ice-cold PBS containing 5% fetal bovine serum was added to the samples, and the samples were then filtered through a 40-µm cell strainer. After centrifugation at 1500 RPM for 5 min at 4C, 5 mL of red blood cell lysis buffer was added and mixed for 15 min. The samples were then centrifuged; single cells were resuspended in a sorting buffer, and then counted using an automatic cell counter.