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Evaluation of the safety and effectiveness of neoadjuvant combined chemoimmunotherapy in the treatment of locally advanced esophageal squamous cell carcinoma: a retrospective single-arm cohort study

Background Transforming potentially resectable advanced esophageal squamous cell carcinoma (ESCC) into resectable ESCC through preoperative induction therapy is an important component of ESCC comprehensive treatment. Immune checkpoint inhibitor (ICI) therapy has been shown to have significant effects in the treatment of advanced ESCC, but its role in the neoadjuvant treatment of potentially resectable ESCC is unclear. This study aims to investigate the safety and effectiveness of camrelizumab combined with chemotherapy in the neoadjuvant treatment of ESCC. Methods We recruited consecutive patients with potentially resectable ESCC who received preoperative camrelizumab in combination with chemotherapy. Data including demographic data, clinicopathological characteristics, neoadjuvant treatment regimens, lesion changes observed by imaging, and surgical details were retrospectively collected through specially designed forms. Toxic effects of neoadjuvant therapy on hematology, gastrointestinal tract, liver, kidney, skin, and thyroid were also collected. Imaging assessments were performed every 1–2 treatment cycles. Follow-up is based on the patient’s regular admission to the hospital for examination and treatment, at least 3 months after surgery. Results A total of 66 patients with locally advanced ESCC were included in this study, including 8 patients with stage II, 29 patients with stage III, and 29 patients with stage IVA. The objective response rate (ORR) of the neoadjuvant immunotherapy combined with chemotherapy was 75.76% (50/66), and no one developed disease progression. A total of 60 patients underwent surgery, and the R0 resection rate was 98.3% (59/60). The pathological complete remission (pCR) rate and the major pathological response (MPR) rate was 6.7% (4/60) and 20% (12/60), respectively. There were 14 cases of treatment-related adverse reactions >3, but no perioperative deaths occurred. Conclusions Neoadjuvant immunotherapy combined with chemotherapy followed by surgical resection may be an available treatment for patients with locally advanced ESCC.


Introduction
Esophageal cancer is the 6th leading cause of cancer related mortality worldwide and accounted for 1 of every 18 cancer-related deaths in 2020 (1). Esophageal cancer is mainly divided into 2 histologic subtypes; that is, esophageal squamous cell carcinoma (ESCC), and esophageal adenocarcinoma (2). Today, ESCC represents the major burden of esophageal cancer and is most common in Eastern Asia and Central Asia (3,4).
More than 80% of patients with esophageal cancer are diagnosed with middle to advanced stage when they first present (5). Among them, nearly half of the patients with esophageal cancer are diagnosed with locally advanced disease which require varying treatment regimens, including surgical resection and/or chemoradiation therapy (6,7). The radical resection of esophageal cancer, including esophagectomy and extensive lymph node dissection, is the standard surgical treatment for nonmetastatic esophageal cancer (8). And adjuvant treatment has been demonstrated to reduce the postoperative recurrence of esophageal cancer (9). van Hagen et al. compared surgery after chemoradiotherapy to surgery alone in patients with esophageal cancer and found that preoperative chemoradiotherapy improved the survival of patients with potentially resectable esophageal cancer (10). The NEOCRTEC5010 study also showed that patients with locally advanced ESCCs who received neoadjuvant chemoradiotherapy (nCRT) plus surgery had better survival than those who received surgery alone (11). Currently, trimodality therapy involving neoadjuvant chemotherapy (nCT) or nCRT followed by esophagectomy and postoperative adjuvant therapy has become the preferred treatments for locally advanced ESCCs (12,13).
Immune checkpoint inhibitors (ICIs) have gradually become an indispensable part of the treatment of advanced esophageal cancer due to their remarkable therapeutic effects, and have also shown significant efficacy in adjuvant therapy for esophageal cancer (14). The ONO-4538-07 (15), CheckMate-032 (16), KEYNOTE-028 (17), and KEYNOTE-180 (18) studies revealed that the objective response rates (ORRs) of immunotherapy for advanced esophageal cancer were 17%, 12%, 30% and 9.9%, respectively. The KEYNOTE-181 (19) and ESCORT (20) studies have shown that immunotherapy administered as a 2nd-line treatment significantly improved the overall survival (OS) of patients with advanced esophageal cancer compared to chemotherapy. Immunotherapy combined with chemotherapy has been studied by CheckMate 649 (21), , and ESCORT-1st (23) as the 1st-line treatment for advanced esophageal cancer, and was found to significantly improved OS compared to chemotherapy alone. CheckMate 577 study results show that nivolumab can significantly prolong disease-free survival (DFS) in adjuvant treatment of esophageal cancer after surgery (24). Based on the above clinical trial results, we speculate that ICIs combined with chemotherapy will also have reliable safety and efficacy in the neoadjuvant treatment of ESCC. However, the role of immunotherapy in the neoadjuvant treatment of ESCCs has not been reported. Therefore, this study attempted to explore the role of neoadjuvant ICIs combined chemotherapy in the treatment of ESCC.
As one of the PD-1 inhibitors, camrelizumab has shown stable anti-cancer effects and is also the most commonly used immunotherapy drug in our department (25). In this study, we aimed to investigate the safety and effectiveness of camrelizumab combined with chemotherapy as a neoadjuvant regimen in the treatment of locally advanced ESCC. We present the following article in accordance with the TREND reporting checklist (available at https://atm. amegroups.com/article/view/10.21037/atm-22-4268/rc).

Patients
This study was designed as a retrospective single-arm cohort study, which consecutively included all locally advanced ESCC patients who received preoperative camrelizumab combined with chemotherapy at the Thoracic Surgery Department of The First Affiliated Hospital, Zhejiang University School of Medicine from May 2019 to May 2021. The reported objective response rate (ORR) of immunotherapy combined with chemotherapy for the first-line treatment of advanced ESCC was 45% (22). We estimate that the ORR of immunotherapy combined with chemotherapy for ESCC neoadjuvant treatment is about 75%, taking α =0.05 (bilateral), 1−β =0.80, resulting in an estimated sample size of 20. We obtained informed consent to use each patient's medical record information upon admission, and the study was approved by the Clinical Research Ethics Committee of The First Affiliated Hospital, Zhejiang University School of Medicine (2021 IIT No. 742). The study was conducted in accordance with the Declaration of Helsinki (as revised in 2013) (26).
All diagnoses were made after pathological analysis of tissue biopsies obtained with esophagogastroduodenoscopy (EGD). Patients were excluded if they met any of the following criteria: (I) had not undergone a pretreatment imaging evaluation; (II) had undergone serial imaging evaluation <2 times; (III) had previously received surgical or endoscopic interventional treatment for ESCC; (IV) had previously received radiotherapy or systemic anti-cancer therapy; (V) suffered from active tuberculosis; (VI) had an esophageal fistula before treatment; and/or (VII) had distant metastases. The electronic medical records of the Hospital contained the medical history, laboratory examination results, imaging examination results, medication details, surgical records, and pathological results of the inpatients and outpatients.
The primary endpoint of this study was the pathological remission rate of ESCC after neoadjuvant therapy, and the secondary endpoints were ORR, surgical resection rate and adverse reactions. Follow-up data was obtained at the patients' routine hospital admission for examinations or treatments. If a patient was not admitted according to the treatment plan, the patient was contacted by telephone. Follow-up was performed for at least 3 months after surgery.

Neoadjuvant therapy
The neoadjuvant treatment strategy comprised 2-4 cycles of camrelizumab combined with platinum-containing dualdrug chemotherapy (platinum + paclitaxel), which was administered intravenously by ward nurses, with a cycle every 3 weeks. The dose of camrelizumab was 200 mg each time. The platinum-based chemotherapy regimen was 75 mg/m 2 of cisplatin, area under the curve (AUC) of the plasma concentration-time curve after a single dose =5 of carboplatin, or 80 mg/m 2 of nedaplatin. The paclitaxel regimen was 260 mg/m 2 of albumin-bound paclitaxel.
After 2 cycles of treatment, the patients were evaluated by computed tomography (CT) or positron emission tomography (PET)-CT to determine resectability of the ESCC. If the tumor regression was not significant, neoadjuvant therapy was continued, and the possibility of surgery was evaluated again after 1-2 cycles.

Tumor response evaluation
Within 1 week of the initial treatment, we performed systematic imaging evaluations of the patients using the baseline data, including PET-CT, CT of the esophagus, endoscopic ultrasound, cranial magnetic resonance imaging, and abdominal ultrasound. Next, we performed imaging evaluations by CT of the esophagus every 2 treatment cycles until the patient underwent surgery or changed treatment. Tumor location, degree of differentiation, clinical TNM (cTNM), cTNM staging after neoadjuvant therapy (ycTNM), and pathological TNM staging after neoadjuvant therapy (ypTNM) were determined according to the TNM staging system of the American Joint Committee on Cancer (8th edition) (27). Tumor treatment response was evaluated according to the Response Evaluation Criteria in Solid Tumor version 1.1 (RECST 1.1) (28). Complete response (CR) was defined as the disappearance of all target lesions; partial remission (PR) was defined as a reduction in the total diameter of target lesions by at least 30%; progressive disease (PD) was defined as an increase in the total diameter of target lesions by at least 20% or the appearance of new lesions; stable disease (SD) was defined as neither CR, PR, nor PD. Patients who achieved CR or PR were defined as responders, while those who had SD and PD were nonresponders.

Neoadjuvant therapy-related adverse events
During neoadjuvant therapy, routine hematological and serum biochemical tests were performed every week and included myocardial enzyme spectrum, thyroid function, and coagulation function tests performed every 3 weeks. Gastrointestinal reactions and skin reactions were evaluated during the examinations.

Surgical treatment
The surgical treatment of inferior and medialis ESCC after neoadjuvant therapy included open or video-assisted thoracoscopic Ivor-Lewis esophagectomy with at least a two-field lymphadenectomy. If a tumor was located in the upper esophagus, McKeown esophagectomy and neck lymph node dissection were performed. These operations are performed by the same medical team, usually four surgeons per operation. The length of the surgery, estimated blood loss, and postoperative complications were recorded.

Pathological examination
The original pathology report described the macroscopic and microscopic appearance of the tumor, including the pathological type, degree of differentiation, depth of invasion, resection margins, lymph nodes, and tumor regression grade (TRG). According to the College of American Pathologists (CAP) and the National Comprehensive Cancer Network (NCCN) guidelines, the TRG was determined by the estimated percentage of residual viable tumor cells relative to the original tumor area and was then be classified into the following 4 categories: TRG 0 (no surviving cancer cells), TRG 1 (residual cancer cells ≤10%), TRG 2 (10%< residual tumor cells ≤50%), and TRG 3 (residual cancer cells >50%). The major pathological response (MPR) was defined as the residual tumor cells not exceeding 10%, which includes TRG 0 and TRG 1.

Statistical analysis
The categorical variables are presented as frequencies and percentages and the continuous variables are presented as the median and interquartile range (IQR). In this study, most of the analyses are descriptive. To explore clinicopathological factors associated with treatment response, we divided patients into PR and SD groups according to treatment response. The differences between PR and SD groups were compared using the chi-square test or the paired chi-square test. For continuous variables, the differences between PR and SD groups were compared using the t-test or the Wilcoxon test. All the analyses were performed using R software version 4.1.2 (R Project for Statistical Computing, Vienna, Austria). A two-tailed P value <0.05 was considered significant.

Patients and treatment process
A total of 66 patients were included in this study with an overview of the neoadjuvant treatment process demonstrated in Figure 1. All cases completed follow-up. The operation rate was approximately 90.91% (60/66); 3 patients with a PR response did not wish to undergo surgery and chose radiotherapy instead. At the end of the 4th cycle, 1 patient had a PR response, but as the thoracoscopy showed that the tumor was in close contact with the adjacent trachea and could not be completely removed, radiotherapy was performed. In another 3 patients, the downstaging was not obvious, and radiotherapy was selected. With the baseline tumor size as a reference, the maximum tumor diameter change was presented using "waterfall plot" (Figure 2). To evaluate the relationship between the number of treatment cycles and the average change in the diameter of the lesions, we compared the diameter of the lesions in each treatment cycle. As Figure 3 shows, the average tumor diameter at the end of cycles 2 ( Figure 3A), 3 ( Figure 3B), and 4 ( Figure 3C) was significantly smaller than that at baseline. The average tumor diameter at the end of cycles 3 ( Figure 3D) and 4 ( Figure 3E) was also significantly reduced compared to that at the end of the 2nd cycle.
The changes in the cTNM stage before (cTNM stage) and after (ycTNM stage) neoadjuvant treatment are summarized in Table 2. There was a significant difference in T stage before and after treatment (P<0.001). After treatment, the proportion of patients with T4a (10.6% before treatment vs. 4.5% after treatment) and T4b (33.3% before treatment vs. 7.6% after treatment) decreased, the proportion of patients with T3 (43.9% before treatment vs. 40.9% after treatment) remained approximately the same, and the proportion of patients with T1 (0.0% before treatment vs. 3.0% after treatment) and T2 (12.1% before treatment vs. 43.9% after treatment) increased. In relation to the N stage, we observed a decrease in the proportion of patients with N2 (53.0% before treatment vs. 50.0% after treatment) and N3 (3.0% before treatment vs. 0.0% after treatment), and an increase in the proportion of patients with N0 (13.6% before treatment vs. 15.2% after treatment) and N1 (30.3% before treatment vs. 34.8% after treatment). The changes in N stage before and after treatment differed significantly (P<0.001). The changes in TNM stage manifested as a decrease in the proportion of patients with stage IVA (43.9% before treatment vs. 12.1% after treatment) ESCC, and an increase in the proportion of patients with stages I (0% before treatment vs. 1.5% after treatment), II (12.1% before treatment vs. 28.8% after treatment), or III (43.9% before treatment vs. 12.1% after treatment). There was a significant difference in the TNM stage between the two groups before and after treatment (P<0.001).

Surgery and pathological response
Of the 66 patients, 60 patients ultimately underwent surgery. The surgical and pathological results are summarized in Table 3. Postoperative complications included anastomotic leaks in 6 cases, of which 1 case underwent secondary surgery. Aspiration pneumonia occurred in 5 cases, all of which were treated with mechanical ventilation. One case of anastomotic stenosis underwent endoscopic treatment. One case of gastroparesis was improved after symptomatic treatment. One case of chylous leakage was treated with thoracic duct ligation. One case of intestinal obstruction was improved after gastrointestinal decompression. No perioperative deaths occurred. The pathological evaluation revealed that the incidences of TRG 0, TRG 1, TRG 2, and TRG 3 were 6.7%, 13.3%, 60.0%, and 20.0%, respectively. Of the 60 patients, 12 (20.0%) achieved an MPR.

Toxicity
The toxicities of neoadjuvant immunotherapy combined with chemotherapy are summarized in Table 4    There were 2 cases of grade 4 adverse reactions, including 1 case of agranulocytosis and 1 case of anemia. Grade 3 adverse reactions occurred in 12 patients, including anemia in 7 patients, liver damage in 2 patients, agranulocytosis in 1 patient, skin reaction in 1 patient, and anemia and skin reaction in 1 patient. These adverse reactions improved after symptomatic treatment, and no patient had to discontinue treatment due to adverse reactions.

Discussion
The treatment options for locally advanced ESCCs usually include surgical resection, radiotherapy, chemotherapy, or a combined chemoradiotherapy (12). nCT has a significant advantage in improving the survival of ESCC patients compared to surgery alone (14). More recently,  the tolerability and efficacy of ICIs has been verified in the treatment of advanced ESCCs (15)(16)(17)(18)(19)(20)(21)(22)(23). Preoperative chemotherapy combined with immunotherapy can theoretically achieve satisfactory anti-tumor effects with tolerable side effects. In this study, we found that the ORR of neoadjuvant immunotherapy combined with chemotherapy was 75.76%, which was significantly higher than that of immunotherapy alone as the 2nd-line treatment for esophageal cancer (9.9-30%) (15)(16)(17)(18). Our neoadjuvant treatment process was set to 2-4 cycles. If the tumor regression at the end of the 2nd cycle was not satisfactory, the patient would be treated for a 3rd or 4th cycle. The imaging results show that the tumor continued to shrink after the 3rd-4th cycle of treatment. Thus, our findings suggest that for patients who have no obvious remission after 2nd cycle, the treatment can be continued up to 3rd-4th cycle of treatment. Klevebro et al. (29) used nCT for patients with esophageal cancer and found that for those with ESCC, the pathological complete remission (pCR) rate after nCT was 9%, and the MPR rate was approximately 15%. In our study, the pCR rate was approximately 6.7%, which was lower than the 9% rate of chemotherapy alone, but the MPR rate was approximately 20.0%, which was higher than the 15% rate of chemotherapy alone (29). The postoperative complication rate of patient undergoing treatment for ESCC in our study was approximately 25.0%, while the postoperative complication rate in the neoadjuvant immunotherapy combined with radiotherapy and chemotherapy group reported by Sihag et al. was approximately 88% (22/25). The discrepancy in the results may be explained by Sihag et al. reported use of immunotherapy combined with radiotherapy and chemotherapy, while we used immunotherapy combined with chemotherapy alone (30).
In terms of drug safety, the ESCORT-1st study used a treatment regimen similar to our study; that is, camrelizumab combined with paclitaxel plus platinum chemotherapy for patients with advanced ESCC (23). The ESCORT-1st study observed that during the administration of immunotherapy combined with chemotherapy to treat advanced ESCCs, the incidence of adverse reactions > grade 3 was 63.4% (189/298), and the treatment-related mortality rate was 3% (9/298) (23). In our study, we found that the incidence of grade 3-4 adverse reactions caused by neoadjuvant immunotherapy combined with chemotherapy was 21.2% (14/66), and there were no treatment-related deaths. A reason for the inconsistent results may be that the ESCC patients included in the ESCORT-1st study were all advanced or metastatic. The treatment period of the ESCORT-1st study was 6 cycles, which is longer than the 2-4 cycles of our study, and may also be a reason for the difference in safety. Overall, the toxicity of neoadjuvant immunotherapy combined with chemotherapy is acceptable. Our study had notable limitations which included its retrospective nature, small sample size, short follow-up time, and the heterogeneity of the patients. To reduce the selection bias, this study included all patients diagnosed with ESCC at our center who met inclusion criteria. Neoadjuvant concurrent chemoradiotherapy represented by the CROSS study (31) has shown a clear curative effect, but in the context of the immune era, neoadjuvant concurrent chemoradiotherapy is increasingly challenged by immunotherapy (32). Due to the lack of randomized controlled trials specifically targeting neoadjuvant immunotherapy combined with chemotherapy in patients with ESCC, we believe that our findings provide valuable insights that can be used to assist in the choice of clinical treatments.
In conclusion, our results from these preliminary analyses suggest that neoadjuvant immunotherapy combined with chemotherapy followed by surgical resection may be a safe, feasible, and effective treatment option for patients with locally-advanced ESCC. However, randomized controlled trials on larger scales are required to confirm our results. The question of whether this neoadjuvant treatment regimen can produce a survival benefit needs to be confirmed by future follow-up studies with larger sample sizes.