T lymphocyte­related immune response and immunotherapy in gastric cancer (Review).

Gastric cancer (GC) remains a global healthcare challenge because of its high incidence and poor prognosis. The efficacy of current chemotherapy regimens for advanced GC is limited. T cells, which have been implicated in the progression of GC, have a significant impact in the tumor microenvironment. With a more detailed understanding of the mechanisms underlying the cancer immunoediting process, immunotherapy may become a promising treatment option for patients with GC. Several clinical trials are currently investigating different mechanisms targeting the tumor immune response. The present review summarized T cell-involved immune responses and various immunotherapy strategies for GC.

Comparison of SOX and CAPOX in patients with advanced gastric cancer after laparoscopic D2 gastrectomy: A randomized controlled trial.

BACKGROUND: Optimal adjuvant chemotherapy after laparoscopic surgery in gastric cancer (GC) patients is still undefined. We aimed to evaluate the efficacy of S-1 plus oxaliplatin (SOX) and capecitabine plus oxaliplatin (CAPOX) in patients with GC after laparoscopic gastrectomy. METHODS: A non-inferiority randomized controlled clinical trial was performed in China. Patients with advanced GC who underwent laparoscopic D2 gastrectomy were randomly assigned to receive SOX and CAPOX regimens. RESULTS: In total, 191 patients were screened between May 2018 and June 2019, and 140 (73.3%) were included in the modified intent-to-treat analysis (mITT), of whom 69 and 71 were assigned to the SOX and CAPOX groups, respectively. The SOX group had similar 3-year overall survival (OS) and disease-free survival to the CAPOX group. Subgroup analysis revealed significantly better OS in the SOX group for male patients ([HR] = 0.395; 95% [CI], 0.153-1.019; p = 0.045), age >60 (HR = 0.219; 95% [CI], 0.064-0.753; p = 0.016), tumors in the gastric antrum (HR = 0.273; 95% [CI], 0.076-0.981; p = 0.047), and moderately differentiated tumors (HR = 0.338; 95% [CI], 0.110-1.041; p = 0.041). There were no significant differences observed in terms of adverse events and recurrence patterns between the two groups. CONCLUSION: Adjuvant SOX was non-inferior to CAPOX treatments for patients with GC who underwent curative laparoscopic D2 gastrectomy. For male patients, aged >60 years, tumors in the gastric antrum, and moderately differentiated tumors, adjuvant SOX may achieve an improvement compared with CAPOX.

Clinical outcome and prognostic factors of T4N0M0 colon cancer after R0 resection: A retrospective study.

BACKGROUND: Paradoxically, patients with T4N0M0 (stage II, no lymph node metastasis) colon cancer have a worse prognosis than those with T2N1-2M0 (stage III). However, no previous report has addressed this issue. AIM: To screen prognostic risk factors for T4N0M0 colon cancer and construct a prognostic nomogram model for these patients. METHODS: Two hundred patients with T4N0M0 colon cancer were treated at Tianjin Medical University General Hospital between January 2017 and December 2021, of which 112 patients were assigned to the training cohort, and the remaining 88 patients were assigned to the validation cohort. Differences between the training and validation groups were analyzed. The training cohort was subjected to multivariate analysis to select prognostic risk factors for T4N0M0 colon cancer, followed by the construction of a nomogram model. RESULTS: The 3-year overall survival (OS) rates were 86.2% and 74.4% for the training and validation cohorts, respectively. Enterostomy (P = 0.000), T stage (P = 0.001), right hemicolon (P = 0.025), irregular review (P = 0.040), and carbohydrate antigen 199 (CA199) (P = 0.011) were independent risk factors of OS in patients with T4N0M0 colon cancer. A nomogram model with good concordance and accuracy was constructed. CONCLUSION: Enterostomy, T stage, right hemicolon, irregular review, and CA199 were independent risk factors for OS in patients with T4N0M0 colon cancer. The nomogram model exhibited good agreement and accuracy.

Different Methods of Minimally Invasive Esophagojejunostomy After Total Gastrectomy for Gastric Cancer: Outcomes from Two Experienced Centers.

BACKGROUND: Esophagojejunostomy after minimally invasive total gastrectomy (MITG) for gastric cancer (GC) is technically challenging. Failure of the esophagojejunal anastomosis can lead to significant morbidity, leading to short- and long-term quality of life (QoL) impairment or mortality. The optimal reconstruction method following MITG remains controversial. We evaluated outcomes of minimally invasive esophagojejunostomy after laparoscopic or robotic total gastrectomies. METHODS: We retrospectively reviewed MITG patients between 2015 and 2020 at two high-volume centers in China and the United States. Eligible patients were divided into groups by different reconstruction methods. We compared clinicopathologic characteristics, postoperative outcomes, including complication rates, overall survival rate (OS), disease-free survival rate (DFS), and patient-reported QoL. RESULTS: GC patients (n = 105) were divided into intracorporeal esophagojejunostomy (IEJ, n = 60) and extracorporeal esophagojejunostomy (EEJ, n = 45) groups. EEJ had higher incidence of wound infection (8.3% vs 13.3%, P = 0.044) and pneumonia (21.7% vs 40.0%, P = 0.042) than IEJ. The linear stapler (LS) group was inferior to the circular stapler (CS) group in reflux [50.0 (11.1-77.8) vs 44.4 (0.0-66.7), P = 0.041] and diarrhea [33.3 (0.0-66.7) vs 0.0 (0.0-66.7), P = 0.045] while LS was better than CS for dysphagia [22.2 (0.0-33.3) vs 11.1 (0.0-33.3), P = 0.049] and eating restrictions [33.3 (16.7-58.3) vs 41.7 (16.7-66.7), P = 0.029] at 1 year. OS and DFS did not differ significantly between LS and CS. CONCLUSIONS: IEJ anastomosis generated better results than EEJ. LS was associated with a better patient eating experience, but more diarrhea and reflux compared with CS. Clinical and patient-reported outcomes show the superiority of IEJ with the LS reconstruction method in MITG for GC.

Optimal Reconstruction After Laparoscopic Distal Gastrectomy: A Single-Center Retrospective Study.

OBJECTIVES: Although laparoscopic distal gastrectomy has been widely used for distal gastric cancer, the best functional reconstruction type has not yet been established. Based on previous experience, we propose a modified uncut Roux-en-Y anastomosis. This study aimed to compare the outcomes of different intracorporeal anastomoses after laparoscopic distal gastrectomy. METHODS: From April 2015 to August 2020, the data of 215 patients who underwent laparoscopic distal gastrectomy was collected. The patients were divided into 4 groups according to the digestive tract reconstruction method, Billroth-I, Billroth-II, Roux-en-Y, and the modified uncut Roux-en-Y. Clinicopathologic characteristics, surgery details, short-term outcomes, and postoperative nutritional status were analyzed. RESULTS: The operation time of Billroth-I anastomosis was significantly shorter (216.2 ± 25.8 min, P < .001) than that of other methods. There was no difference in postoperative complications and OS among the 4 reconstruction methods. The incidences of esophagitis, gastritis, and bile reflux were significantly lower in the Roux-en-Y and uncut Roux-en-Y group (P < .001) 1 year after surgery. And the postoperative albumin and PNI levels in uncut Roux-en-Y group were higher than those in other groups(P < .05). On multivariate analysis, age and reconstruction type were independently related to esophagitis, gastritis, and bile reflux. Serum albumin and the prognostic nutritional index were significantly higher in the uncut Roux-en-Y group than other groups (P < .05). CONCLUSIONS: All 4 reconstruction techniques are feasible and safe. The Roux-en-Y and uncut Roux-en-Y are superior to Billroth-Ⅰ and Billroth-Ⅱ+Braun in terms of reflux esophagitis, gastritis, and bile reflux. Uncut Roux-en-Y may result in better PNI than the others.

A nomogram to predict the risk of colorectal anastomotic leakage combining inflammatory-nutritional and abdominal aorta calcium index.

Background: Anastomotic leakage is a serious complication after colorectal cancer surgery, which affects the quality of life and the prognosis. This study aims to create a novel nomogram to predict the risk of anastomotic leakage for patients with colorectal cancer based on the preoperative inflammatory-nutritional index and abdominal aorta calcium index. Methods: 292 patients at Tianjin Medical University General Hospital (Tianjin, China) from January 2018 to October 2021 who underwent colorectal cancer surgery with a primary anastomosis were retrospectively reviewed. A nomogram was constructed based on the results of multivariate logistic regression model. The calibration curves and receiver operating characteristic curves were used to verify the efficacy of the nomogram. Results: Univariate and multivariate analyses showed that tumor location (P = 0.002), preoperative albumin (P = 0.006), preoperative lymphocyte (P = 0.035), preoperative neutrophil to lymphocyte ratio (P = 0.024), and superior mesenteric artery calcium volumes score (P = 0.004) were identified as the independent risk factors for postoperative anastomotic leakage in patients with colorectal carcinoma. A nomogram was constructed based on the results of the multivariate analysis, and the C-index of the calibration curves was 0.913 (95%CI: 0.870-0.957) in the training cohort and 0.840 (95%CI: 0.753-0.927) in the validation cohort. Conclusion: The nomogram, combining basic variables, inflammatory-nutritional index and abdominal aorta calcium index, could effectively predict the possibility of postoperative anastomotic leakage for patients with colorectal cancer, which could guide surgeons to carry out the appropriate treatment for the prevention of anastomotic leakage.

Impact of Neoadjuvant Therapy on Minimally Invasive Surgical Outcomes in Advanced Gastric Cancer: An International Propensity Score-Matched Study.

BACKGROUND: No international consensus on the treatment of advanced gastric cancer (AGC) exists. In the absence of well-designed, comparative studies between neoadjuvant versus adjuvant strategies, concerns about increased risk of postoperative complications remain barriers to neoadjuvant chemotherapy (NAC) for AGC. We evaluated surgical outcomes of AGC patients who received minimally invasive radical gastrectomy with D2 lymphadenectomy after NAC. METHODS: We collected data from two high-volume gastric cancer programs in the United States and China between January 2015 and December 2019 with the last follow-up in February 2020. AGC patients undergoing minimally invasive radical surgery were included. After propensity score-matching, surgical outcomes were analyzed. Risk-factor of complications was analyzed in the whole cohort. RESULTS: After 1:1 propensity score-matching, 97 patients were included in each cohort. NAC + surgery cohort was younger (58.2 ± 10.3 vs. 61.3 ± 9.6, P = 0.036) with lower preoperative WBC count (5.7 ± 2.8 vs. 6.9 ± 2.1 × 109/ml) than the surgery upfront cohort. NAC was not a risk-factor for postoperative complications (odds ratio [OR], 0.859; 95% confidence interval [CI], 0.46-1.60; P = 0.633). Overall risk-factors of postoperative complications included age ≥ 60 years (OR, 21.338; 95% CI, 5.00-91.05; P < 0.001), tumor size ≥ 5 cm (OR, 1.24; 95% CI, 1.08-1.83; P < 0.001), operation time ≥ 240 min (OR, 5.53; 95% CI, 1.26-24.26; P = 0.012), and ASA classification ≥ II (OR, 13.14; 95% CI, 4.12-24.73; P < 0.001). CONCLUSIONS: NAC before minimally invasive radical gastrectomy with D2 lymphadenectomy does not increase postoperative complications, and these findings support broader application of NAC and MIS for AGC. Additional studies are required to determine the effect of NAC on long-term survival.

A tetravalent single chain diabody (CD40/HER2) efficiently inhibits tumor proliferation through recruitment of T cells and anti-HER2 functions.

Our aim was to construct a CD40×HER2 single chain diabody (ScDb) and determine its tumor-specific immune activation and anti-HER2 function. Overlap extension-polymerase chain reaction was applied in the construction of ScDb, and the protein was expressed with the pET28a (+)-Rosetta prokaryotic expression system. Soluble ScDb was purified by a nickel-nitrilotriacetic acid column. Dendritic cells (DC) was stimulated by ScDb and inhibited 4T1 cells proliferation in vitro. In 4T1 tumor mice model, lymphocyte infiltration was prominently detected in ScDb group, Caspase-3 expression was significantly upregulated. ScDb was labeled using quantum dots. Immunofluorescence assay indicated ScDb exhibited high affinity to HER2. T6-17 cells were inhibited by ScDb in vitro. The phosphorylation and expression levels of AKT, ERK were markedly decreased. In T6-17 tumor mice model. Compared to CD40 ScFv, HER2 ScFv and normal saline groups, tumor volume diminished significantly in ScDb group, and tumor cells showed extensive deformation, and pervasive karyopyknosis and karyorrhexis were found. In the present study, we successfully constructed a ScDb fragment and expressed it using a prokaryotic expression system. The in vivo and in vitro experimental results indicated that ScDb could inhibit the proliferation of tumor cells by stimulating the tumor-specific immunoreaction and blocking the HER2-related signaling pathway.

Selection and expression of CD40 single chain variable fragment by phage display and evaluation of tumor specific immune activation.

Immune escape is the final phase of the cancer immunoediting process. Researchers have found that cancer induces immune escape by inhibiting the expression of CD40L. The purpose of the present study was to select a high affinity CD40 single chain variable fragment (ScFv) and to evaluate its effect on tumor-specific immune activation. One Wistar rat was immunized with mouse CD40 antigen. CD40 ScFv with high affinity was constructed by overlap extension-polymerase chain reaction (SOE-PCR) and screened by three rounds of phage display. CD40 ScFv protein was expressed by the pET28a (+)-Rosetta prokaryotic expression system and purified using a nickel-nitrilotriacetic acid (Ni-NTA) column. CD40 ScFv significantly upregulated CD80, CD86, and MHC-II in vitro expression in dendritic cells (DCs) and upregulated the expression of IL-12 (p70) based on ELISA results. Cell counting kit-8 (CCK-8) results indicated that T lymphocytes were stimulated by DCs in an Ag + CD40 ScFv group, which also inhibited the proliferation of immortalized T6-17 cells. In an in vivo assay, 1 × 106 T6-17 cells were subcutaneously injected into BALB/c mice in the hind flank. Tumor volume curves showed that CD40 ScFv exhibited a remarkable inhibition of tumor proliferation after 15 days of treatment. Hematoxylin-eosin (H&E) staining of tumor tissues indicated that CD40 ScFv enhanced lymphocyte infiltration, which remarkably inhibited the proliferation of T6-17 cells. Furthermore, immunohistochemistry (IHC) staining revealed that caspase-3 was abundantly expressed in the T6-17 cytoplasm after CD40 ScFv treatment. In conclusion, this study revealed that high affinity CD40 ScFv could be screened by phage display and had a significant stimulating effect on DCs and inhibited the proliferation of T6-17 cells in vivo and in vitro.

Inhibition of cardiac allograft rejection in mice using interleukin-35-modified mesenchymal stem cells.

Interleukin-35 (IL-35) is a cytokine recently discovered to play a potent immunosuppressive role by intensifying the functions of regulatory T cells and inhibiting the proliferation and functions of T helper 1 and T helper 17 cells. Mesenchymal stem cells (MSCs) have recently emerged as promising candidates for cell-based immune therapy, and our previous study showed that IL-35 gene modification can effectively enhance the therapeutic effect of MSCs in vitro. In this study, we isolated adipose tissue-derived MSCs in vitro and infected them with lentiviral vectors overexpressing the IL-35 gene, thereby creating IL-35-MSCs. Subsequently, IL-35-MSCs were then injected into mice of the allogeneic heterotopic abdominal heart transplant model to determine their effect on allograft rejection. The results showed that IL-35-MSCs could continuously secrete IL-35 in vivo and in vitro, successfully alleviate allograft rejection and prolong graft survival. In addition, compared to MSCs, IL-35-MSCs showed a stronger immunosuppressive ability and further reduced the percentage of Th17 cells, increased the proportion of CD4+ Foxp3+ T cells, and regulated Th1/Th2 balance in heart transplant mice. These findings suggest that IL-35-MSCs have more advantages than MSCs in inhibiting graft rejection and may thus provide a new approach for inducing immune tolerance during transplantation.

Mesenchymal stem cell expression of interleukin-35 protects against ulcerative colitis by suppressing mucosal immune responses.

BACKGROUND: Interleukin-35 (IL-35) has recently been identified as an immunosuppressive cytokine that has been used as a potential therapy for chronic inflammatory and autoimmune diseases. However, there remains a paucity of data regarding its potential benefits after integration into mesenchymal stem cells (MSCs). METHODS: We used a dextran sulfate sodium (DSS)-induced colitis mice model and treated them with IL-35-MSCs, MSCs or saline. The body weight was recorded daily and inflammatory processes were determined. Cytokine secretion by lamina propria lymphocytes (LPLs) and percentage of regulatory T cells (Tregs) were also measured. RESULTS: The data showed that mice in the two treated groups recovered their body weight more rapidly than mice treated with saline in the later stage of colitis. The colon lengths of IL-35-MSC-treated mice were markedly longer than those in the other two groups and the inflammation reduced significantly. Furthermore, the percentage of Foxp3 + Tregs increased significantly and the level of proinflammatory cytokines produced by LPLs decreased significantly in the IL-35-MSC-treated group. DISCUSSION: The results demonstrate that IL-35-MSCs could ameliorate ulcerative colitis by down-regulating the expression of pro-inflammatory cytokines.

Interleukin-35 Gene-Modified Mesenchymal Stem Cells Protect Concanavalin A-Induced Fulminant Hepatitis by Decreasing the Interferon Gamma Level.

Interleukin 35 (IL-35) is a relatively newly identified cytokine required for the regulatory and suppressive functions of regulatory T cells (Treg), playing an important role in the prevention of autoimmune diseases. This study used mesenchymal stem cells (MSCs) as the gene-delivery vehicles for IL-35 gene therapy and investigated their protective effects in Concanavalin A (Con A)-induced autoimmune hepatitis. Results showed that IL-35 gene modified MSCs (IL-35-MSCs) can specifically migrate to the injured liver tissues and significantly narrow the necrosis areas of injured livers. IL-35-MSCs prevented hepatocyte apoptosis by reducing the FASL expression by mononuclear cells. Although MSC treatment can alleviate liver injury to some extent, IL-35-MSCs showed a stronger protective effect, which means some novel mechanisms exist. The results show that IL-35-MSCs could decrease the level of interferon gamma secreted by liver mononuclear cells through the JAK1-STAT1/STAT4 signal pathway. In summary, this study thus demonstrates a novel and efficient treatment for Con A-induced fulminant hepatitis through negatively regulating the secretion of interferon gamma, thus providing a novel therapeutic approach for this devastating liver disease.

Mesenchymal stem cells overexpressing IL-35 effectively inhibit CD4+ T cell function.

Mesenchymal stem cells (MSCs) have recently emerged as promising candidates for cell-based immune tolerance therapy. Interleukin 35 (IL-35) is a relatively newly identified cytokine required for the regulatory and suppressive functions of regulatory T cells (Treg), playing an important role in the prevention of autoimmune diseases. In this study, we isolated adipose tissue-derived MSCs, a good vehicle for cell therapy, which were transfected with a lentivirus vector for the overexpression of the therapeutic murine IL-35 gene. IL-35 levels in transfected MSCs (IL-35-MSCs) were quantified by ELISA. Co-culture of CD4+ T cells and IL-35-MSCs resulted in the inhibition of CD4+ T cell proliferation and IL-17A secretion. In addition, IL-35-MSCs induced IL-10 production by CD4+ T cells, but did not affect IFN-γ. These findings suggested that MSCs over-expressing IL-35 had higher immunosuppressive capacity compared with non-transfected MSCs, and may provide a useful approach for basic research on gene therapy for autoimmune disorders.

Evidence of cell fusion in carcinogen-induced mice gastric carcinoma.

The role of bone marrow-derived cells in gastric cancer formation was not fully understood. In this study, bone marrow from female green fluorescent protein transgenic mice was transplanted into male wild-type mice to generate sex-mismatched chimeric mice. The chimeric mice were treated with carcinogen to induce gastric cancer. At time of sacrifice, 18.2 % (2/11) of mice showed severe dysplasia and 25 % (3/12) of mice successfully induced with cancer. Fluorescence in situ hybridization results showed that bone marrow-derived cells participated in renewal of gastric mucosa and cell fusion was observed in both precancerous lesions and adenocarcinoma, but no sign of fusion was observed in squamous cell carcinoma. Our findings suggest that bone marrow-derived cells participate in renewal of gastric mucosa during chronic damage and might have acquired the phenotype of gastric epithelial cells through cell fusion. Fusion between gastric epithelial cells and bone marrow-derived cells was involved in increased carcinogenesis.

HER2/neu over-expression predicts poor outcome in early gastric cancer without lymph node metastasis.

PURPOSE: Human epidermal growth factor receptor 2 (HER2/neu) is involved in the pathogenesis of several types of cancer, including gastric cancer. However, there remains a paucity of data regarding the prognostic relevance of HER2/neu in early gastric cancer without lymph node metastasis (pN0 EGC). The aim of our study was to analyze whether the over-expression of HER2/neu significantly predicts poor outcomes of pN0 EGC. PATIENTS AND METHODS: Sixty-seven patients who underwent operative resection for pN0 EGC was enrolled. The HER2/neu status was examined by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). RESULTS: The HER2/neu-positive rate was 16.4%. HER2/neu over-expression showed a significant correlation with histological type (P ≤ 0.001), tumor location (P=0.022) and Lauren grade (P=0.012). Multivariate analysis showed HER2/neu serves as a good prognostic marker to predict the risk of poor outcome for pN0 EGC. (HR=1.384, 95.0% CI: 1.142-1.897 P=0.005) CONCLUSION: Considering HER2/neu over-expression significantly predicts poor outcome in pN0 EGC, accurate HER2/neu assessment would be done before endoscopic therapy. For HER2/neu-positive patients, radical surgery should be performed.

Evolution of surgery for Klatskin tumor demonstrates improved outcome: a single center analysis.

BACKGROUND AND OBJECTIVES: The rapid development of the Chinese economy has enabled advances to be made in the surgical treatment of Klatskin tumors. We retrospectively reviewed the surgical outcomes of hilar cholangiocarcinoma at a single Chinese center, focusing on the surgical procedures, radicality of the operative procedure (R0, R1 or R2), survival rates, and independent prognostic factors. This objective of the analysis was to evaluate improvements in perioperative and long-term outcomes of surgical resection of Klatskin tumors. METHODS: Between January 1998 and December 2007, 131 consecutive patients underwent operative resection for Klatskin tumor at Tianjin Medical University General Hospital, Tianjin, China. Clinicopathological data were analyzed to assess their impact on survival. RESULTS: The overall 3- and 5-year survival rates were 46% and 22%, respectively. Median overall survival for the entire cohort was 35 months. Survival increased progressively between 1998 and 2007. Preoperative biliary drainage represents a safe strategy before hepatectomy for hilar cholangiocarcinoma in patients with jaundice. Patients who underwent liver resection achieved significantly higher R0 rates than those undergoing biliary resection only. Preoperative total bilirubin level, lymph node metastasis and degree of radicality (R1 and 2 vs R0) were found to be independent factors that negatively affected overall survival. CONCLUSIONS: Surgical resection remains the mainstay of treatment for hilar cholangiocarcinoma. Development in surgical equipment, techniques, and perioperative management have increased long-term survival after curative liver resection, despite the more aggressive surgical procedures that are being performed.