Study design
This domestic multicenter, randomized controlled clinical study compares neoadjuvant immunotherapy and chemoradiotherapy (Group A) with neoadjuvant chemoradiotherapy (Group B) for the treatment of patients with locally advanced resectable ESCC (stage cII-III), as well as conduct relevant translational medicine research. Patients eligible for enrollment will be randomly assigned to Groups A and B in a 1:1 ratio in Phase 1, with pathological complete response (pCR) rate as the study endpoint, and 176 patients are planned to be enrolled. 300 patients will continue to be enrolled in Phase 2/3 after the Phase 1 target has been attained, and they will be equally divided between Groups A and B by randomization. There will be 476 patients signed up in total. The trial flow chart is presented in Fig. 1. All methods are carried out in accordance with declaration of Helsinki and relevant guidelines."
Target population
Both male and female patients with thoracic ESCC will be regarded as eligible for enrollment in the experiment. Responsible researchers assess patients' conditions and confirm that they fit the requirements for selection. Additionally, only participants who provide written informed permission are permitted to participate in the trial.
Sample size considerations
The sample size for this prospective randomized controlled clinical study was determined using the preliminary data from the Department of Thoracic Surgery, Zhongshan Hospital, Fudan University.
The primary endpoint of the first phase of the study was pCR. According to the early data results, the pCR rate was 50% in trial group A and 27% in trial group B. The randomization ratio was 1:1, the power was 0.90, the alpha was 0.05, and the calculation was based on 15% shedding. A total of 176 cases were required and randomly allocated to trial groups A (neoadjuvant immunotherapy combination with chemoradiotherapy) and B (neoadjuvant chemoradiotherapy group) using the software PASS (Power Analysis and Sample Size).
Overall survival served as the study's main goal during the second phase (OS). According to our group's CMISG1701 trial, the 3-year survival rate was approximately 55% in the neoadjuvant chemoradiotherapy group (group B), while the expected 3-year survival rate was 65% in the neoadjuvant immunotherapy combining chemoradiotherapy group (group A). A total of 476 cases were enrolled and distributed equally between the two groups using the software PASS (Power Analysis and Sample Size), with a randomization ratio of 1:1, power = 0.80, alpha = 0.05, enrollment time of 36 months, total study time of 72 months, and based on 10% shedding.
Therefore, 176 subjects (88 in each group) were enrolled in the first phase of the study. After the target was reached, 300 subjects (150 in each group) were enrolled in the second and third phase, making a total of 476 subjects enrolled in the whole study.
Inclusion criteria
Patients that qualify must fulfill each of the following conditions:
① Patients having a gastroscopic biopsy-type of pathology-confirmed diagnosis of ESCC and tissue specimens saved before therapy.
② The primary tumor is in the chest, and the primary site of EC depends on where in the esophagus the mass is located (upper thoracic esophagus: from the entrance of the thorax up to the level of the lower border of the arch of azygos vein, 20 cm to < 25 cm from the incisors on endoscopy; middle thoracic esophagus: from the lower border of the arch of azygos vein up to the level of the lower pulmonary vein, 25 cm to < 30 cm from the incisors on endoscopy; lower thoracic esophagus: upper from the level of the lower pulmonary vein, down to 30 cm to < 40 cm from the incisor on gastric endoscopy)
As determined by the aforementioned tests, patients with preoperative clinically feasible esophageal cancer for surgical resection (with or without significant tumor invasion, mediastinal lymph node enlargement, and distant organ metastasis by means of enhanced CT of the chest and abdomen, ultrasound of the cervical lymph nodes; if the primary tumor are suspected to be T4b, multiple mediastinal lymph node metastases, or the presence of remote metastases, whole-body PET-CT, ultrasound endoscopy (EUS) (optional) and other examinations to further clarify the clinical stage).
④ Physical status score: ECOG 0 to 1, age: 18 years, physical age: 75 years, and anticipated survival: 12 months.
⑤The major organs of the five subjects are all functioning normally, as are the subjects' blood counts, lung, liver, kidney, and heart functions. The following criteria must be met by laboratory indicators: White blood cells > 4. 0×109/L, absolute neutrophil count (ANC) ≥ 2.0×109/L, platelet count > 100×109/L, hemoglobin > 90g/L, and pulmonary function: FEV1 ≥ 1.2L, FEV1% ≥50%, and DLCO ≥ 50% are all blood parameters.
Description: FEV1(L): the force spirometry measurement in liters.
FEV1%: Force Spirometry Measured Value/Expected Value (%).
DLCO%: carbon monoxide breath diffusion function measured value/expected value (%);
Liver function: serum bilirubin less than 1.5 times the upper limit of normal; ALT and AST less than 1.5 times the upper limit of normal.
Blood creatinine (SCr) is 120 mol/L and creatinine clearance (CCr) is 60 ml/min, indicating proper renal function.
⑥ able to read and understand the study's consent form.
Exclusion criteria
Patients meeting any of the following criteria are not eligible for this trial:
① Clinical stage of (AJCC/UICC 8th Edition) T4b inoperable resection (as determined by two senior thoracic surgeons), multiple lymph node enlargement (estimated ≥ 6 lymph node metastases), multi-station lymph node enlargement (estimated ≥ 3 lymph node stations), or distant metastases (M1) as determined by imaging such as thoracic and abdominal enhancement CT, cervical lymph node ultrasound, whole body PET-CT (optional) or EBUS (optional) or patients with distant metastases (M1).
② Patients who have received or are receiving other chemotherapy, radiation therapy or targeted therapy.
③ Gastroscopy pathology of non-squamous carcinoma.
④ Previous other tumors (except for those who had cervical carcinoma in situ or localized basal cell carcinoma of the skin and were cured).
Other exclusion criteria.
⑤ History of autoimmune disease.
⑥ Recent or current use of hormones or immunosuppressive drugs.
(7) Prior immunotherapy.
⑧ Prior history of severe hypersensitivity to antibody drugs.
⑨ Previous or ongoing chronic or recurrent autoimmune disease.
⑩ Interstitial lung disease, pulmonary fibrosis, diverticulitis, or systemic ulcerative gastrointestinal inflammation.
⑪ Confirmed history of congestive heart failure; poorly controlled angina with medication; transmural myocardial infarction confirmed by electrocardiogram (ECG); poorly controlled hypertension; clinically significant heart valve disease; or high-risk uncontrolled arrhythmia.
⑫ Severe uncontrolled intercurrent systemic disease, such as active infection or poorly controlled diabetes mellitus; abnormal coagulation, bleeding tendency, or ongoing thrombolytic or anticoagulant therapy.
⑬ Women with a positive serum pregnancy test or who are breastfeeding, and men and women of childbearing age who are unwilling to use adequate contraception during study drug therapy.
⑭ Known active infection with immunodeficiency virus (HIV), hepatitis B virus (HBV) or hepatitis C virus (HCV) or known HIV seropositivity; including HBV or HCV surface antigen positivity (RNA).
⑮ Known allergy to any investigational drug.
⑯History of organ transplantation (including bone marrow auto-transplantation and peripheral stem cell transplantation).
⑰ Presence of peripheral neurological disorders or history of significant psychiatric disorders and central nervous system disorders.
⑱ Patients who are co-administering antitumor drugs outside the study protocol.
⑲ Presence of other factors that may result in forced termination of the study mid-stream, such as the presence of other serious medical conditions (including psychiatric disorders) requiring comorbid treatment, alcoholism, substance abuse, family or social factors in the judgment of the investigator
Study interventions
The detailed treatment regimens of two arms are presented in Fig. 2.
Neoadjuvant immunotherapy combined with chemoradiotherapy (group A)
Paclitaxel at 50 mg/m2 and carboplatin at AUC: 2 mg/(ml.min) at 30-minute intervals on day 1; radiotherapy at 1.8 Gy/day on days 2 to 6 for a total of 5 weeks. This includes a total of 41.4 Gy for a total of 23 doses, with only 3 doses in cycle 5; and 200 mg/dose of Tirelizumab on days 1 and 22 administered intravenously over 30 minutes (no less than 20 minutes and no more than 60 minutes).
Neoadjuvant chemoradiotherapy (group B)
On day 1, paclitaxel at 50 mg/m2 and AUC: 2 mg/(ml.min) of carboplatin at 30-minute intervals, and on days 2–6, 1.8 Gy/day of radiation therapy for 5 days, weekly once a week for a total of 5 sessions, with a total of 41.4 Gy of radiotherapy for a total of 23 sessions, and only 3 sessions of radiotherapy in session 5.
Esophagectomy (both groups)
Four weeks after the conclusion of neoadjuvant therapy, the patient's physical condition and the results of the enhanced CT of the chest and abdomen, ultrasound of the cervical lymph nodes, blood routine, liver function, kidney function, blood biochemistry, tumor markers, ECG, lung function, and PET-CT were reviewed to determine the impact of neoadjuvant therapy. The surgery would be performed 4 ~ 8 weeks after neoadjuvant therapy if the criteria for surgery are met.
The recommended Mckeown procedure is combined with an esophagectomy and full thoracoabdominal lymph node dissection as the surgical treatment. Lymph node dissection is recommended to include the left cardia, right cardia, left gastric artery parietal and diaphragmatic, inferior esophageal parietal, middle esophageal parietal, inferior ramus, superior esophageal parietal, left and right laryngeal nerve parietal lymph nodes. The site of the lymph node dissection is marked, and it is advised to dissect at least 15 lymph nodes.
Outcome measures
Primary outcomes
Phase I: comparison of the pathological complete response (pCR) rate between the two groups.
Phase II: comparison of overall survival (OS) between the two groups.
Secondary outcomes
Phase I: Comparison of the incidence and severity of adverse events, as well as safety indicators such as the rate of intermediate chest opening, surgical complications, 30-day and 90-day postoperative mortality, postoperative hospital stay, and reoperation rate; comparison of the effectiveness indicators such as R0 resection rate, positive lymph node rate and response, number of lymph nodes cleared, and tumor regression grade (TRG) of the primary tumor between the two groups.
Phase II: Comparison of the incidence and severity of adverse events, as well as safety indicators such as the rate of intermediate chest opening, surgical complications, mortality rate at 30 days and 90 days after surgery, postoperative hospital stay, and reoperation rate; comparison of the effectiveness indicators such as pCR rate, R0 resection rate, lymph node positive rate and response, number of lymph nodes cleared, tumor regression grade (TRG) of primary tumor, and disease-free survival (DFS) between the two groups.
♦ Pathological complete response (pCR) rate: After neoadjuvant therapy, the pathology department should evaluate the resection specimen according to established procedures and grade the tumor regression using standardized histological criteria. The level of histomorphologic regression is divided into the following four categories: grade1, no sign of vital residual tumor cells (pCR), grade2, fewer than 10% of vital residual tumor cells, grade3, between 10% and 50%, and grade4, greater than 50% according to a prior report[12].
♦ R0 resection rate: If there is no sign of a vital tumor at the proximal, distal, or circumferential resection margins, the resection is deemed to have been successful (R0). A vital tumor is microscopically positive if it can be seen at 1 mm or less from the proximal, distal, or circumferential resection margin (R1).
♦ Positive lymph nodes’ number: Record the number of lymph nodes that are positive in accordance with pathology reports.
Exploratory endpoint outcomes
Create an immune scoring system and examine its association with efficacy and prognosis by identifying immune markers in tumor tissues. Create a model based on the tumor immune scoring system to predict the pathological response and survival prognosis of neoadjuvant immunotherapy combined with chemoradiotherapy and neoadjuvant chemoradiotherapy for ESCC. Detect ctDNA levels and dynamic changes, analyzing their relationship to efficacy and prognosis as well as the monitoring and early warning value for MRD and recurrence.
Assessments, data collections and follow-up
Pre-therapeutic assessments
Transthoracic and abdominal enhancement CT, ultrasound of the cervical lymph nodes and other means to assess whether there is significant tumor invasion, enlargement of the mediastinal lymph nodes and distant organ metastases. If the primary tumor is suspected to be T4b or multiple metastases in mediastinal lymph nodes or remote metastases, whole-body PET-CT and ultrasound endoscopy (EUS) (optional) will be performed to further clarify the clinical stage.
Assessments during the treatment phase
Four weeks after the completion of neoadjuvant therapy, thoracic and abdominal enhancement CT, cervical lymph node ultrasound, blood routine, liver function, kidney function, blood biochemistry, tumor markers, electrocardiogram, lung function, and PET-CT will be reviewed to assess the effect of neoadjuvant therapy and patients’ physical condition. Surgery will be performed 4–8 weeks after neoadjuvant therapy if the patient meets the surgery criteria. If the disease progresses, radiotherapy treatment or salvage surgery is recommended.
Assessments during the follow-up phase
Safety follow-up: Follow-up every 30 days for adverse reactions related to the treatment drug within 90 days after the last dose, including survival status, adverse reactions, concomitant medication and concomitant treatment information.
Survival follow-up: The first follow-up visit was conducted 30 days after surgery and every 3 months thereafter to collect survival information and information on anti-tumor therapy and time to disease progression after the study until the subject died, was lost to follow-up, or the study was terminated. Active monitoring will be performed every 3 months by DSMC to improve study progress, ensure data integrity and patient safety.
Translational research
To determine the composition, number, and distribution of tumor-associated immune cells in the immune microenvironment of ESCC, multi-label immunofluorescence staining was applied to tumor tissue samples from preoperative biopsies. These cells included mature T cells and their subsets (helper T cells, regulatory T cells, cytotoxic T cells), NK cells, TAMs, and MDSCs. The expression of immune checkpoints PD-1/PD-L1, CTLA-4, Tim-3, LAG-3 and IDO-1 proteins were also detected to construct an immune scoring system and analyze their relationship with pathological remission and prognosis. To analyze the ctDNA levels and the correlation between its dynamics and the effectiveness of neoadjuvant therapy, patient prognosis, and disease recurrence before, after neoadjuvant therapy, 1 month after radical surgery and during follow-up, tumor tissue samples from preoperative biopsies were subjected to whole-exome sequencing. The ctDNA panel was then customized based on the tumor tissue sequencing results.
Statistical analysis
All statistical analyses were carried out with SPSS, and one-sided tests were consistently used for hypothesis testing, including the test statistic and the associated P value. A 95% confidence range was chosen, and p ≤ 0.05 will be regarded as statistically significant for the difference under test.
Quantitative indicators will be described using mean, standard deviation (normal distribution) or median, minimum, maximum (non-normal distribution). Qualitative indicators will be described using rates and composition ratios. The results are expressed mainly using statistical tables with groupings as vertical coordinates or statistical charts to give statistical results if necessary. Chi-square test was used to compare qualitative data, whereas parametric and nonparametric approaches were employed to compare quantitative data. A threshold for the statistical significance of the difference under test is p ≤ 0.05.
Statistical inference: t-test or non-parametric test was used for measurement data; chi-square test was used for comparison between groups for count data, and Fisher's exact probability method was used when necessary. The Kaplan-Meier method was used to determine survival rates, and log-rank tests were employed to compare survival rates between groups.