Relationship between laparoscopic total gastrectomy-associated postoperative complications and gastric cancer prognosis.

This study aimed to investigate the incidence and prognosis of postoperative complications after laparoscopic total gastrectomy (LTG) for gastric cancer (GC). We retrospectively enrolled 411 patients who underwent curative LTG for GC at seven institutions between January 2004 and December 2018. The patients were divided into two groups, complication group (CG) and non-complication group (non-CG), depending on the presence of serious postoperative complications (Clavien-Dindo grade III [≥ CD IIIa] or higher complications). Short-term outcomes and prognoses were compared between two groups. Serious postoperative complications occurred in 65 (15.8%) patients. No significant difference was observed between the two groups in the median operative time, intraoperative blood loss, number of lymph nodes harvested, or pathological stage; however, the 5-year overall survival (OS; CG 66.4% vs. non-CG 76.8%; p = 0.001), disease-specific survival (DSS; CG 70.1% vs. non-CG 76.2%; p = 0.011), and disease-free survival (CG 70.9% vs. non-CG 80.9%; p = 0.001) were significantly different. The Cox multivariate analysis identified the serious postoperative complications as independent risk factors for 5-year OS (HR 2.143, 95% CI 1.165-3.944, p = 0.014) and DSS (HR 2.467, 95% CI 1.223-4.975, p = 0.011). A significant difference was detected in the median days until postoperative recurrence (CG 223 days vs. non-CG 469 days; p = 0.017) between the two groups. Serious postoperative complications after LTG negatively affected the GC prognosis. Efforts to decrease incidences of serious complications should be made that may help in better prognosis in patients with GC after LTG.

Prognostic value of MAGEA4 in primary lung cancer depends on subcellular localization and p53 status.

Melanoma antigen family A4 (MAGEA4), a cancer/testis antigen, is overexpressed and is thus an immunotherapy target in various malignant tumors, including non-small cell lung cancer. However, whether MAGEA4 induces or inhibits the apoptosis of lung cancer cells remains controversial, as is its prognostic significance, particularly since there is no reliable method with which to detect MAGEA4 specifically. In this study, we optimized assay conditions to detect MAGEA4 based on cells transiently transfected with MAGEA genes, and found that MAGEA4 was expressed in four of eight non-small cell lung cancer cell lines, and in 25.4% of clinical lung cancer specimens. We also found that MAGEA4 overexpression decreased apoptosis, as measured by the levels of cleaved caspase-3 in stably transfected 293F cells. Notably, patients with nuclear MAGEA4, but not p53 expression exhibited a significantly poorer survival than those expressing both nuclear MAGEA4 and p53. Indeed, multivariate analysis identified nuclear MAGEA4 as an independent prognostic factor (P=0.0042), albeit only in the absence of p53. In this study, to the best of our knowledge, we are the first to demonstrate that the function and prognostic value of MAGEA4 depends on its subcellular localization and on the p53 status.

Circular versus linear stapling in esophagojejunostomy after laparoscopic total gastrectomy for gastric cancer: a propensity score-matched study.

PURPOSE: We used propensity score matching to compare the complication rates after laparoscopic total gastrectomy (LTG) with esophagojejunostomy (EJS) performed using a circular or a linear stapler. METHODS: We retrospectively enrolled all patients who underwent curative LTG between November 2004 and March 2016. Patients were categorized into the circular and linear groups according to the stapler type used for the subsequent EJS. Patients in the groups were matched using the following propensity score covariates: age, sex, body mass index, American Society of Anesthesiologists physical status, extent of lymph node dissection, and Japanese Classification of Gastric Carcinoma stage. Clinicopathological characteristics and surgical outcomes were compared. RESULTS: We identified 66 propensity score-matched pairs among 379 patients who underwent LTG. There was no significant between-group difference in the median operative time, extent of lymph node dissection, number of lymph nodes resected, rate of conversion to open surgery, or number of surgeries performed by a surgeon certified by the Japanese Society of Endoscopic Surgery. In the circular and linear groups, the rate of all complications (Clavien-Dindo [CD] classification ≥ I; 21 vs. 26%, respectively; p = 0.538), complications more severe than CD grade III (14 vs. 14%, respectively; p = 1.000), and occurrence of EJS leakage and stenosis more severe than CD grade III (5 vs. 2%, p = 0.301; 9 vs. 8%, p = 0.753, respectively) were comparable. CONCLUSIONS: There is no difference in the postoperative complication rate related to the type of stapler used for EJS after LTG.

Clinical Implications of CD4+CD25+Foxp3+Regulatory T Cell Frequencies After CHP-MAGE-A4 Cancer Vaccination.

BACKGROUND/AIM: The aim of this study was to explore whether the treatment effect or immune response to a cancer vaccine can be predicted by the percentage of CD4+CD25+Foxp3+ regulatory T cells (Tregs) in peripheral blood mononuclear cells (PBMCs) after vaccination. PATIENTS AND METHODS: Sixteen patients (9 men, 7 women; median age 61.5 years) enrolled in the CHP-MAGE-A4 cancer vaccine clinical trial who had a fixed dose (300 µg of CHP-MAGE-A4 cancer vaccine and 0.5 Klinische Einheit (KE) of OK432 and received at least four vaccinations were investigated. Safety, immune response, and clinical effects were assessed before and after the cancer vaccination. RESULTS: Treg ratios that remained low both before and after vaccination were associated with a good prognosis, and a low Treg/CD4 lymphocyte ratio 7-weeks after the initial vaccination was correlated with a better prognosis. CONCLUSION: The Treg ratio following vaccination appears to have some utility for predicting patient prognosis.

Serum immunoglobulin E response as a marker for unfavorable prognosis following cholesteryl pullulan-MAGE A4 vaccination.

Since 2009, a cancer vaccine clinical trial was conducted with melanoma antigen gene-A4 as an immunogenic agent. The levels of IgG1, IgG2 and IgG3, which are known to be Type 1 T helper cell-associated antibodies, and the levels of IgG4 and IgE, which are known to be Type 2 T helper cell-associated antibodies, were measured and used as biomarkers for predicting therapeutic effect. The results of the present study indicated a strong positive correlation between IgG2 and IgG4, with a correlation coefficient of R=0.808 (P<0.0001). The survival time of patients in which IgE responses were induced was significantly shorter compared with the survival time of patients with no IgE induction. The results of the present study suggest that caution is required when antigen-specific IgE responses are induced during cancer vaccination therapy.

Differential detection of cytoplasmic Wilms tumor 1 expression by immunohistochemistry, western blotting and mRNA quantification.

Wilms tumor 1 (WT1) is considered to be a promising target of cancer treatment because it has been reported to be frequently expressed at high levels in various malignancies. Although WT1-targeted cancer treatment has been initiated, conclusive detection methods for WT1 are not established. The present study aimed to consolidate immunohistochemistry for WT1 with statistical basis. Transfected cells with forced WT1 expression yielded specific western blot bands and nuclear immunostaining; cytoplasmic immunostaining was not specifically recognized. Immunohistochemistry, western blotting, and quantitative reverse transcriptase-polymerase chain reaction were performed in 35 human cell lines using multiple WT1 antibodies and their results were quantified. Relationships among the quantified results were statistically analyzed; the nuclear immunostaining positively correlated with western blot bands and mRNA expression levels, whereas cytoplasmic immunostaining did not. These results indicate that nuclear immunostaining reflects WT1 expression but cytoplasmic immunostaining does not. The nuclear immunostaining was barely (3/541) observed in primary cancer of esophagus, bile duct, pancreas and lung. Although the present study has some limitations, the results indicate that the cytoplasmic immunostaining does not correlate with actual WT1 expression and prompts researchers to carefully evaluate target molecule expression in treatment of cancer.

Impact of elemental diet on early recovery after laparoscopic colectomy: findings of a randomized controlled trial.

PURPOSE: An amino acid-containing elemental diet (ED) does not require digestion for nutritional absorption, making it a good option for patients with gastrointestinal malabsorption. We conducted a randomized trial to confirm that perioperative ED enhanced the recovery of patients undergoing laparoscopic colectomy. METHODS: Patients in the intervention arm received commercially available ED from the day prior to surgery until postoperative day (POD) 3, whereas patients in the control group received a conventional perioperative diet program. To verify the endpoints, "estimated minimum length of stay in hospital after surgery" (emLOS) was defined as the number of days necessary to reach all the five criteria; namely, "sufficient oral intake", "sufficient pain control", "withdrawal of intravenous alimentation", "no abnormal findings in routine examinations", and "no rise in fever". RESULTS: A total of 102 patients were randomized, 94 of whom were analyzed (ED 45, control 49). There was no morbidity or mortality. Shorter emLOS (POD 4 vs. POD 7; p = 0.018), earlier resumption of sufficient oral intake (POD 3 vs. POD 4; p = 0.034) and faster recovery to defecation (2.2 vs. 3.1 days; p = 0.005) were observed in the ED group vs. the control group. CONCLUSIONS: The perioperative ingestion of ED by patients undergoing laparoscopic colectomy is safe and can reduce the postoperative hospital stay by supporting the acceleration of oral intake.

Clinical relevance of antigen spreading pattern induced by CHP-MAGE-A4 cancer vaccination.

AIM: To investigate the antigen spreading pattern in the CHP-MAGE-A4-vaccinated patients and analyze the clinical relevance of antigen spreading pattern as a surrogate marker of patient survival. MATERIALS & METHODS: 12 patients who had been injected with 300 µg of CHP-MAGE-A4 and 0.5 Klinische Einheit of OK-432 in more than five vaccinations were analyzed. RESULTS: Increases in the anti-MAGE-A4-specific antibody response were observed in eight patients (66.7%), compared with six patients (50%) for anti-NY-ESO-1 and five patients (41.7%) for anti-MAGE-A3 after five vaccinations. We identified frequent antigen spreading following MAGE-A4 vaccinations without associations with the clinical response or patient prognosis. CONCLUSION: Antigen spreading pattern might reflect tumor shrinkage as a response to treatment and treatment history (clinical trial registration number: UMIN000001999).

Time-dependent transition of the immunoglobulin G subclass and immunoglobulin E response in cancer patients vaccinated with cholesteryl pullulan-melanoma antigen gene-A4 nanogel.

A phase I+II clinical trial of vaccination with MAGE-A4 protein complexed with cholesteryl pullulan melanoma antigen gene-A4 nanogel (CHP-MAGE-A4) is currently underway in patients with MAGE-A4-expressing cancer. In the present study, the primary phase I endpoint was to test the safety of the administration of 300 µg CHP-MAGE-A4 with and without OK-432. Another aim of the study was to clarify the details of the specific humoral immune response to vaccination. The 9 patients enrolled for phase I were vaccinated 6 times, once every 2 weeks: 3 patients with 100 µg and 3 patients with 300 µg CHP-MAGE-A4, and 3 patients with 300 µg CHP-MAGE-A4 plus 0.5 clinical units of OK-432. Toxicities were assessed using Common Terminology Criteria for Adverse Events v3.0. Clinical response was evaluated by modified Response Evaluation Criteria in Solid Tumours. Immunological monitoring of anti-MAGE-A4-specific antibodies was performed by ELISA of pre- and post-vaccination patient sera. The 6 vaccinations produced no severe adverse events. Stable disease was assessed in 4/9 patients. Anti-MAGE-A4 total immunoglobulin (Ig)G titers increased in 7/9 patients. Efficacious anti-MAGE-A4 IgG1, 2 and 3 antibody responses were observed in 7/9 patients. Among them, positive conversions to T helper 2 (Th2)-type antibody responses (IgG4 and IgE) were observed after frequent vaccination in 4/7 patients. The Th2 conversion was possibly associated with undesirable clinical observations, including progressive disease and the appearance of a new relapse lesion. The present study suggested that frequent vaccinations activated a Th2-dominant status in the cancer patients. The identification of a time-dependent IgG subclass and IgE antibody production during vaccination protocols may be a useful surrogate marker indicating a potentially undesirable change of the immunological environment for an effective antitumor immune response in cancer patients.

The key role of calreticulin in immunomodulation induced by chemotherapeutic agents.

BACKGROUND: It has recently been shown that certain chemotherapeutic agents can improve host immune responses. The present study aimed to demonstrate the mechanism by which chemotherapeutic agents modify the tumor microenvironment and induce tumor-specific immune responses. METHODS: Three mouse cancer cell lines [CT26 mouse colon cancer cells, B16 melanoma cells and Lewis lung carcinoma (LLC)], 5 human carcinoma cell lines (human esophageal squamous cell carcinoma cell lines TE8 and HEC46 and the human pancreatic carcinoma cell lines PK-9, AsPC-1 and SUIT-2) and 5 chemotherapeutic agents [mitoxantrone (MIT), mitomycin C(MMC), 5-fluorouracil (5FU), camptothecin (CPT-11) and cisplatin (CDDP)] that are frequently used in a clinical setting for cancer treatment were utilized to investigate the surface expression level of calreticulin and HLA class I after exposure to chemotherapeutic agents. RESULTS: Increased calreticulin (CRT) expression on the surface of mouse cell lines and, moreover, increased surface expression levels of both CRT and HLA class I in all human cell lines were observed in cells treated by the chemotherapeutic agents as compared with non-treated cells. The surface expression level of CRT was significantly correlated with the HLA class I expression level in all human cell lines. CONCLUSIONS: In conclusion, chemotherapeutic drugs can improve the immunogenicity of cancer cells in a cell-specific manner through the mechanism of translocation of CRT.

Sequence confirmation and characterization of the mouse Ssxa gene: Ssxa protein is cleaved and the N-terminal cleaved fragment translocates into the nucleus.

This is the first demonstration of a stop codon in the sequence of mouse Ssxa and characterization of the biological behavior of Ssxa protein. Cancer testis antigen (CTA) is known as a target of immunotherapy against cancer, and Ssxa is one of the CTAs. Although a CTA would be useful to establish a mouse cancer vaccine model using endogenous antigen, the stop codon was not identified in the sequence of Ssxa cDNA that was previously reported. In this study, the gene sequence of Ssxa was different from the previous report in which several mouse CTAs were analyzed. Initially, we identified the correct cDNA sequence of mouse Ssxa by 3'-rapid amplification of cDNA ends and found a new exon containing the stop codon (Exon X). Ssxa mRNA expression was determined by reverse transcription-PCR (RT-PCR) in four mouse cancer cell lines and the testis but not in other normal organs. We found that the molecular weight of recombinant Ssxa protein is 12 kDa, and we generated an anti-Ssxa antibody which recognizes the C-terminus of Ssxa. Two vectors expressing fusion proteins (pSsxa-GFP and pGFP-Ssxa) were generated and fluorescence in the nucleus was observed only in the pGFP-Ssxa transfected cells. Therefore, we conclude that the N-terminal cleaved fragment of Ssxa, which has a KRAB domain (nuclear localization signal), translocates into the nucleus after cleavage of the C-terminus.