T cell neoepitope discovery in colorectal cancer by high throughput profiling of somatic mutations in expressed genes.

OBJECTIVE: Patient-specific (unique) tumour antigens, encoded by somatically mutated cancer genes, generate neoepitopes that are implicated in the induction of tumour-controlling T cell responses. Recent advancements in massive DNA sequencing combined with robust T cell epitope predictions have allowed their systematic identification in several malignancies. DESIGN: We undertook the identification of unique neoepitopes in colorectal cancers (CRCs) by using high-throughput sequencing of cDNAs expressed by standard cancer cell cultures, and by related cancer stem/initiating cells (CSCs) cultures, coupled with a reverse immunology approach not requiring human leukocyte antigen (HLA) allele-specific epitope predictions. RESULTS: Several unique mutated antigens of CRC, shared by standard cancer and related CSC cultures, were identified by this strategy. CD8+ and CD4+ T cells, either autologous to the patient or derived from HLA-matched healthy donors, were readily expanded in vitro by peptides spanning different cancer mutations and specifically recognised differentiated cancer cells and CSC cultures, expressing the mutations. Neoepitope-specific CD8+ T cell frequency was also increased in a patient, compared with healthy donors, supporting the occurrence of clonal expansion in vivo. CONCLUSIONS: These results provide a proof-of-concept approach for the identification of unique neoepitopes that are immunogenic in patients with CRC and can also target T cells against the most aggressive CSC component.

Gastric cancer: texture analysis from multidetector computed tomography as a potential preoperative prognostic biomarker.

OBJECTIVES: To investigate the association between preoperative texture analysis from multidetector computed tomography (MDCT) and overall survival in patients with gastric cancer. METHODS: Institutional review board approval and informed consent were obtained. Fifty-six patients with biopsy-proved gastric cancer were examined by MDCT and treated with surgery. Image features from texture analysis were quantified, with and without filters for fine to coarse textures. The association with survival time was assessed using Kaplan-Meier and Cox analysis. RESULTS: The following parameters were significantly associated with a negative prognosis, according to different thresholds: energy [no filter] - Logarithm of relative risk (Log RR): 3.25; p = 0.046; entropy [no filter] (Log RR: 5.96; p = 0.002); entropy [filter 1.5] (Log RR: 3.54; p = 0.027); maximum Hounsfield unit value [filter 1.5] (Log RR: 3.44; p = 0.027); skewness [filter 2] (Log RR: 5.83; p = 0.004); root mean square [filter 1] (Log RR: - 2.66; p = 0.024) and mean absolute deviation [filter 2] (Log RR: - 4.22; p = 0.007). CONCLUSIONS: Texture analysis could increase the performance of a multivariate prognostic model for risk stratification in gastric cancer. Further evaluations are warranted to clarify the clinical role of texture analysis from MDCT. KEY POINTS: • Textural analysis from computed tomography can be applied in gastric cancer. • Preoperative non-invasive texture features are related to prognosis in gastric cancer. • Texture analysis could help to evaluate the aggressiveness of this tumour.

Risk factor analysis for involvement of resection margins in gastric and esophagogastric junction cancer: an Italian multicenter study.

BACKGROUND: Resection margin (RM) involvement is associated with negative prognosis after gastrectomy. Intraoperative frozen section (IFS) analysis allows radical resection to be achieved in a single operation but is time-consuming and resource-consuming. The aim of this study was to assess risk factors associated with RM involvement to identify patients who would benefit from IFS analysis. METHODS: We retrospectively analyzed patients who underwent gastrectomy with curative intent for gastric or esophagogastric junction (EGJ) cancer from 2000 to 2014 in six Italian hospitals. RM status was assessed by IFS analysis and/or definitive histopathology examination. A set of 21 potential risk factors were compared in a multivariate analysis between patients with positive RMs on IFS analysis or definitive histopathology examination and a control cohort of similar patients with negative RMs, with the samples stratified into three subgroups (T1, T2-T4 Lauren intestinal pattern, T2-T4 Lauren diffuse/mixed pattern). RESULTS: One hundred forty-five patients had positive RMs. Survival was significantly worse in positive RM patients than in negative RM patients (89.5 months vs 28.9 months). Multivariate analysis showed that in T1 cancers a margin distance of less than 2 cm is a risk factor for RM involvement (odds ratio 15.7), in T2-T4 intestinal pattern cancers, serosa invasion (odds ratio 6.0), EGJ location (odds ratio 4.1), and a margin distance of less than 3 cm (odds ratio 4.0) are independent risk factors, and in T2-T4 diffuse/mixed pattern cancers, lymphatic infiltration (odds ratio 4.2), tumor diameter greater than 4 cm (odds ratio 3.5), EGJ location (odds ratio 2.8), and serosa invasion (odds ratio 2.2) are independent risk factors. CONCLUSIONS: Survival after gastrectomy is negatively affected by positive RMs. IFS analysis should be routinely used in patients with a high risk of positive RMs, especially in diffuse pattern cancers.

The Italian Research Group for Gastric Cancer (GIRCG) guidelines for gastric cancer staging and treatment: 2015.

This article reports the guidelines for gastric cancer staging and treatment developed by the GIRCG, and contains comprehensive indications for clinical management, including radiological, endoscopic, surgical, pathological, and oncological paths.

Apparent diffusion coefficient by diffusion-weighted magnetic resonance imaging as a sole biomarker for staging and prognosis of gastric cancer.

OBJECTIVE: To investigate the role of apparent diffusion coefficient (ADC) from diffusion-weighted magnetic resonance imaging (DW-MRI) when applied to the 7th TNM classification in the staging and prognosis of gastric cancer (GC). METHODS: Between October 2009 and May 2014, a total of 89 patients with non-metastatic, biopsy proven GC underwent 1.5T DW-MRI, and then treated with radical surgery. Tumor ADC was measured retrospectively and compared with final histology following the 7th TNM staging (local invasion, nodal involvement and according to the different groups - stage I, II and III). Kaplan-Meier curves were also generated. The follow-up period is updated to May 2016. RESULTS: Median follow-up period was 33 months and 45/89 (51%) deaths from GC were observed. ADC was significantly different both for local invasion and nodal involvement (P<0.001). Considering final histology as the reference standard, a preoperative ADC cut-off of 1.80×10-3 mm2/s could distinguish between stages I and II and an ADC value of ≤1.36×10-3 mm2/s was associated with stage III (P<0.001). Kaplan-Meier curves demonstrated that the survival rates for the three prognostic groups were significantly different according to final histology and ADC cut-offs (P<0.001). CONCLUSIONS: ADC is different according to local invasion, nodal involvement and the 7th TNM stage groups for GC, representing a potential, additional prognostic biomarker. The addition of DW-MRI could aid in the staging and risk stratification of GC.

Surgical management of duodenal stump fistula after elective gastrectomy for malignancy: an Italian retrospective multicenter study.

BACKGROUND: Duodenal stump fistula (DSF) is a severe complication of gastrectomy. Although nonsurgical therapy is preferred, surgery is still mandatory in one third of DSF patients. The aim of this article is to analyze the surgical management of DSF and factors related to its outcome. METHODS: We performed a retrospective multicenter study using data from January 1990 to November 2011 in 16 Italian surgery centers. We collected 8,268 elective gastrectomies for malignancies, 7,987 by the laparotomic and 281 by the laparoscopic approach. Two hundred five patients developed a DSF, 75 of whom underwent surgery for DSF. We analyzed mortality and DSF healing time as well as the impact of clinical, oncological, and surgical characteristics. RESULTS: The laparoscopic approach increased the risk of DSF development (odds ratio 5.6, 95% confidence interval 2.7-10.6, P < 0.001). The indication for first DSF surgery was intra-abdominal sepsis; the failure rate was over 30%, associated with the appearance of fistulas of neighboring organs, bleeding, and the need for reoperations. The mortality rate was 28% and was related to the presence of vascular disease (P = 0.04), more than one reoperation (P = 0.05), sepsis (P < 0.001), and renal failure (P < 0.001). Fifty-four patients recovered after a median of 39 days (interquartile range 22-68 days); the need to perform more reoperations (P < 0.01) and the presence of an abdominal abscess (P < 0.01) led to an increase in healing time. CONCLUSIONS: Surgery for DSF has a poor prognosis. Our data will help to identify patients at risk of death, but unfortunately could not establish the best surgical procedure applicable to all cases of DSF.

Preoperative locoregional staging of gastric cancer: is there a place for magnetic resonance imaging? Prospective comparison with EUS and multidetector computed tomography.

BACKGROUND: The aim of this study was to prospectively compare the diagnostic performance of magnetic resonance imaging (MRI), multidetector computed tomography (MDCT) and endoscopic ultrasonography (EUS) in the preoperative locoregional staging of gastric cancer. METHODS: This study had Institutional Review Board approval, and informed consent was obtained from all patients. Fifty-two patients with biopsy-proven gastric cancer underwent preoperative 1.5-T MRI, 64-channel MDCT and EUS. All images were analysed blind, and the results were compared with histopathological findings according to the seventh edition of the TNM classification. After the population had been divided on the basis of the local invasion (T1-3 vs T4a-b) and nodal involvement (N0 vs N+), sensitivity, specificity, positive and negative predictive value, and accuracy were calculated and diagnostic performance measures were assessed using the McNemar test. RESULTS: For T staging, EUS showed higher sensitivity (94%) than MDCT and MRI (65 and 76%; p = 0.02 and p = 0.08). MDCT and MRI had significantly higher specificity (91 and 89%) than EUS (60%) (p = 0.0009 and p = 0.003). Adding MRI to MDCT or EUS did not result in significant differences for sensitivity. For N staging, EUS showed higher sensitivity (92%) than MRI and MDCT (69 and 73%; p = 0.01 and p = 0.02). MDCT showed better specificity (81%) than EUS and MRI (58 and 73%; p = 0.03 and p = 0.15). CONCLUSIONS: Our prospective study confirmed the leading role of EUS and MDCT in the staging of gastric cancer and did not prove, at present, the value of the clinical use of MRI.

Cancer-initiating cells from colorectal cancer patients escape from T cell-mediated immunosurveillance in vitro through membrane-bound IL-4.

Cancer-initiating cells (CICs) that are responsible for tumor initiation, propagation, and resistance to standard therapies have been isolated from human solid tumors, including colorectal cancer (CRC). The aim of this study was to obtain an immunological profile of CRC-derived CICs and to identify CIC-associated target molecules for T cell immunotherapy. We have isolated cells with CIC properties along with their putative non-CIC autologous counterparts from human primary CRC tissues. These CICs have been shown to display "tumor-initiating/stemness" properties, including the expression of CIC-associated markers (e.g., CD44, CD24, ALDH-1, EpCAM, Lgr5), multipotency, and tumorigenicity following injection in immunodeficient mice. The immune profile of these cells was assessed by phenotype analysis and by in vitro stimulation of PBMCs with CICs as a source of Ags. CICs, compared with non-CIC counterparts, showed weak immunogenicity. This feature correlated with the expression of high levels of immunomodulatory molecules, such as IL-4, and with CIC-mediated inhibitory activity for anti-tumor T cell responses. CIC-associated IL-4 was found to be responsible for this negative function, which requires cell-to-cell contact with T lymphocytes and which is impaired by blocking IL-4 signaling. In addition, the CRC-associated Ag COA-1 was found to be expressed by CICs and to represent, in an autologous setting, a target molecule for anti-tumor T cells. Our study provides relevant information that may contribute to designing new immunotherapy protocols to target CICs in CRC patients.

Prognostic utility of diffusion-weighted MRI in oesophageal cancer: is apparent diffusion coefficient a potential marker of tumour aggressiveness?

PURPOSE: To investigate the role of the apparent diffusion coefficient (ADC) as a potential prognostic biomarker in the evaluation of the aggressiveness of oesophageal cancer. MATERIALS AND METHODS: Between November 2009 and December 2013, 43 patients with evidence of oesophageal or oesophago-gastric junction cancer were referred to our institution and prospectively entered in our database. The final study population consisted of 23 patients (18 men; 5 women; mean age, 64.62 ± 10.91 years) who underwent diffusion-weighted Magnetic Resonance before surgical intervention. Specifically, 14 were directly treated with surgery and 9 were addressed to chemo/radiotherapy beforehand. Two radiologists independently measured mean tumour ADC and inter-observer agreement (Spearman's and intraclass correlation coefficient [ICC]) was assessed. In the univariate analysis, overall survival curves related to pathological ADC, pT, pN, tumour location and histotype were fitted using the Kaplan-Meier method. Survival curves were then compared using the log-rank test. RESULTS: Inter-observer reproducibility was very good (Spearman's rho = 0.95; ICC = 0.94). At a total median follow-up of 19 months (2-49 months), 4 patients had died. The median follow-up was 18.50 months (5-49 months) for the surgery-only group (1/4 events, 25 %) and 24 months (2-34 months) for the chemo/radiotherapy group (3/4 events, 75 %). Survival time at 48 months for the overall population was 59 % (±0.11), while for the surgery-only group and the chemo/radiotherapy group was 90 % (±0.09) and 61 % (±0.34), respectively. In the univariate analysis, ADC values below or equal to 1.4 × 10(-3) mm(2)/s were associated with a negative prognosis both in the total population (P = 0.016) and in the surgery-only group (P < 0.001). CONCLUSION: Despite the biggest limitation of our study (i.e. the small study population), we were able to show that pathological ADC could be considered a prognostic factor for oesophageal cancer. DWI might be introduced into clinical practice as a promising and reliable technique in the diagnostic pathway of this tumour.

Prognostic Role of Diffusion-weighted MR Imaging for Resectable Gastric Cancer.

PURPOSE: To prospectively investigate the role of apparent diffusion coefficient (ADC) calculated from diffusion-weighted magnetic resonance (MR) imaging as a potential prognostic biomarker in the evaluation of the aggressiveness of gastric cancer. MATERIALS AND METHODS: This prospective study had institutional review board approval. Informed consent was obtained from all patients. Between October 2009 and December 2013, a total of 99 patients (65 men, 34 women; mean age, 62.02 years; age range, 32.33-85.15 years) with biopsy-proved cancer (28 esophagogastric junction and 71 gastric cancers) were examined with a 1.5-T MR imaging system, including T1-, T2-, and diffusion-weighted sequences. ADC measurements were obtained. Seventy-one patients were directly treated with surgery, while 28 underwent neoadjuvant chemotherapy beforehand. Pathologic ADC, pathologic T and N stages, tumor location, surgical approach, and histologic subtype were investigated with univariate and multivariate analyses by using the Cox regression model. RESULTS: At a total median follow-up period of 21 months, 31 patients had died. The median follow-up was 25 months for the surgery-only group (19 of 31 events [61%]) and 28 months for the chemotherapy group (12 of 31 events [39%]). In the multivariate analysis, ADC values of 1.5 × 10(-3) mm(2)/sec or lower were associated with a negative prognosis, both in the total population (log-relative risk, 1.73; standard error, 0.56; P = .002) and in the surgery-only (log-relative risk, 1.97; standard error, 0.66; P = .003) and chemotherapy (log-relative risk, 2.93; standard error, 1.41; P = .03) groups, along with other significant prognostic factors (in particular, pathologic T and N stages). CONCLUSION: Pathologic ADC represents a strong independent prognostic factor in the evaluation of the aggressiveness of gastric cancer, in addition to clinical and surgical variables.

Response to chemotherapy in gastric adenocarcinoma with diffusion-weighted MRI and (18) F-FDG-PET/CT: correlation of apparent diffusion coefficient and partial volume corrected standardized uptake value with histological tumor regression grade.

PURPOSE: To assess whether changes in diffusion-weighted MRI (DW-MRI) and (18) F-fluoro-2-deoxyglucose positron emission tomography/computed tomography ((18) F-FDG PET/CT), correlate with treatment response to neoadjuvant therapy (NT), as expressed by tumor regression grade (TRG), from locally advanced gastric adenocarcinoma (GA). MATERIALS AND METHODS: Seventeen patients underwent both DW-MRI and (18) F-FDG-PET/CT scans before and after the end of NT. Apparent diffusion coefficient (ADC) and mean standardized uptake value (SUV) corrected for partial volume effect (PVC-SUVBW-mean ) were evaluated and compared with histopathological TRG. RESULTS: Pre- and post-NT and percentage changes for ADC and PVC-SUVBW-mean were assessed. Post-NT ADC and ΔADC showed a significant inverse correlation with TRG (r = -0.71; P = 0.0011 and r = -0.78; P = 0.00020, respectively) and significant differences in their mean values were found between responders (TRG 1-2-3) and nonresponders (TRG 4-5) (P = 0.0009; P = 0.000082, respectively). No correlations with TRG were found for pre-NT ADC and for all PVC-SUVBW-mean values as well as between ΔADC and Δ PVC-SUVBW-mean . CONCLUSION: DW-MRI seems more accurate than (18) F-FDG-PET/CT and ADC modifications may represent a reproducible tool to assess tumor response for GA.

Duodenal stump fistula after gastric surgery for malignancies: a retrospective analysis of risk factors in a single centre experience.

BACKGROUND: Duodenal stump fistula (DSF) is the most severe surgical complication after gastrectomy. This study was designed to assess the incidence, to observe the consequences, and to identify the risk factors associated with DSF after gastrectomy. METHODS: All procedures involving total or sub-total gastrectomy for cancer, performed between January 1987 and June 2012 in a single institution, were prospectively entered into a computerized database. Risk factors analysis was performed between DSF patients, patients with complete uneventful postoperative course and patients with other major surgical complications. RESULTS: Over this 25 years period, 1287 gastrectomies were performed. DSF was present in 32 cases (2.5 %). Mean post-operative onset was 6.6 days. 19 patients were treated conservatively and 13 surgically. Mean DSF healing time was 31.2 and 45.2 days in the two groups, respectively. Mortality was registered in 3 cases (9.37 %), due to septic shock (2 cases) and bleeding (1 case). In monovariate analysis, heart disease (p < 0.001), pre-operative lymphocytes number (p = 0.003) and absence of manual reinforcement over duodenal stump (p < 0.001) were found to be DSF-specific risk factors, whereas liver cirrhosis (p = 0.002), pre-operative albumin levels (p < 0.001) and blood losses (p = 0.002) were found to be non-DSF-specific risk factors. In multivariate analysis heart disease (OR 5.18; p < 0.001), liver cirrhosis (OR 13.2; p < 0.001), bio-humoral nutritional status impairment (OR 2.29; p = 0.05), blood losses >300 mL (OR 4.47; p = 0.001) and absence of manual reinforcement over duodenal stump (OR 30.47; p < 0.001) were found to be independent risk factors for DSF development. CONCLUSIONS: Duodenal stump fistula still remains a life-threatening complication after gastric surgery. Co-morbidity factors, nutritional status impairment and surgical technical difficulties should be considered as important risk factors in developing this awful complication.

Clinical competence in the surgery of rectal cancer: the Italian Consensus Conference.

BACKGROUND AND AIM: The literature continues to emphasize the advantages of treating patients in "high volume" units by "expert" surgeons, but there is no agreed definition of what is meant by either term. In September 2012, a Consensus Conference on Clinical Competence was organized in Rome as part of the meeting of the National Congress of Italian Surgery (I Congresso Nazionale della Chirurgia Italiana: Unità e valore della chirurgia italiana). The aims were to provide a definition of "expert surgeon" and "high-volume facility" in rectal cancer surgery and to assess their influence on patient outcome. METHOD: An Organizing Committee (OC), a Scientific Committee (SC), a Group of Experts (E) and a Panel/Jury (P) were set up for the conduct of the Consensus Conference. Review of the literature focused on three main questions including training, "measuring" of quality and to what extent hospital and surgeon volume affects sphincter-preserving procedures, local recurrence, 30-day morbidity and mortality, survival, function, choice of laparoscopic approach and the choice of transanal endoscopic microsurgery (TEM). RESULTS AND CONCLUSION: The difficulties encountered in defining competence in rectal surgery arise from the great heterogeneity of the parameters described in the literature to quantify it. Acquisition of data is difficult as many articles were published many years ago. Even with a focus on surgeon and hospital volume, it is difficult to define their role owing to the variability and the quality of the relevant studies.

Gastroenteric neuroendocrine neoplasms classification: comparison of prognostic models.

BACKGROUND: Gastroenteric neuroendocrine neoplasms (GE-NENs) display highly variable clinical behavior. In an attempt to assess a better prognostic description, in 2010, the World Health Organization (WHO) updated its previous classification, and the European Neuroendocrine Tumors Society (ENETS) proposed a new grading and TNM-based staging system. In the current study, the authors evaluated the prognostic significance of these models and compared their efficacy in describing patients' long-term survival to assess the best prognostic model currently available for clinicians. METHODS: The study cohort was composed of 145 patients with extrapancreatic GE-NEN who were observed from 1986 to 2008 at a single center and were classified according to the WHO and ENETS classifications. Survival evaluations were performed using Kaplan-Meyer analyses on 131 patients. Only deaths from neoplasia were considered. A P value < .05 was considered significant. Prognostic efficacy was assessed by determining the Harrell concordance index (c-index). RESULTS: Both the 2010 WHO and the ENETS classification were able to efficiently divide patients into classes with different prognoses. According to the model comparison, the ENETS TNM-based staging system appeared to be the strongest. All combined models were effective prognostic predictors, but the model that included the 2010 WHO classification plus ENETS staging had a higher c-index. CONCLUSIONS: Both the 2010 WHO classification and the ENETS staging system are valid instruments for GE-NENs prognostic assessment, with TNM-based stage appearing to be the best available choice for clinicians, both alone and in association with other classifications.

Apparent diffusion coefficient modifications in assessing gastro-oesophageal cancer response to neoadjuvant treatment: comparison with tumour regression grade at histology.

OBJECTIVES: To assess changes in apparent diffusion coefficient (ΔADC) and volume (ΔV) after neoadjuvant treatment (NT), and tumour regression grade (TRG) in gastro-oesophageal cancers (GEC), and to discriminate responders from non-responders. METHODS: Thirty-two patients with biopsy-proven locally-advanced GEC underwent diffusion weighted magnetic resonance imaging (DWI) pre- and post-NT. Lesion ADC, volume, ΔADC and ΔV were calculated. TRG 1-2-3 patients were classified as R; TRG 4-5 as non-responders. ΔADC-TRG and ΔV-TRG correlations, pre-NT and post-NT ADC, ΔADC and ΔV cut-off values for responders and non-responders were calculated. Two readers measured mean tumour ADCs and interobserver variability was calculated. (Spearman's and intraclass correlation coefficient [ICC]). RESULTS: The interobserver reproducibility was very good both for pre-NT (Spearman's rho = 0.8160; ICC = 0.8993) and post-NT (Spearman's rho = 0.8357; ICC = 0.8663). Responders showed lower pre-NT ADC (1.32 versus 1.63 × 10(-3) mm(2)/s; P = 0.002) and higher post-NT ADC (2.22 versus 1.51 × 10(-3) mm(2)/s; P = 0.001) than non-responders and ADC increased in responders (ΔADC, 85.45 versus -8.21 %; P = 0.00005). ΔADC inversely correlated with TRG (r = -0.71, P = 0.000004); no difference in ΔV between responders and non-responders (-50.92 % versus -14.12 %; P = 0.068) and no correlation ΔV-TRG (r = 0.02 P = 0.883) were observed. CONCLUSIONS: The ADC can be used to assess gastro-oesophageal tumour response to neoadjuvant treatment as a reliable expression of tumour regression. KEY POINTS: • DWI is now being used to assess many cancers. • Change in ADC measurements offer new information about oesophageal tumours. • ADC changes are more reliable than dimensional criteria in assessing neoadjuvant treatment. • Such ADC assessment could optimise management of locally advanced gastro-oesophageal cancers.

Ex vivo enrichment of circulating anti-tumor T cells from both cutaneous and ocular melanoma patients: clinical implications for adoptive cell transfer therapy.

Tumor-infiltrating lymphocytes (TILs) have been successfully used for adoptive cell transfer (ACT) immunotherapy; however, due to their scarce availability, this therapy is possible for a limited fraction of cutaneous melanoma patients. We assessed whether an effective protocol for ex vivo T-cell expansion from peripheral blood mononuclear cells (PBMCs), suitable for ACT of both cutaneous and ocular melanoma patients, could be identified. PBMCs from both cutaneous and ocular melanoma patients were stimulated in vitro with autologous, irradiated melanoma cells (mixed lymphocyte tumor cell culture; MLTCs) in the presence of IL-2 and IL-15 followed by the rapid expansion protocol (REP). The functional activity of these T lymphocytes was characterized and compared with that of TILs. In addition, the immune infiltration in vivo of ocular melanoma lesions was analyzed. An efficient in vitro MLTC expansion of melanoma reactive T cells was achieved from all PBMC's samples obtained in 7 cutaneous and ocular metastatic melanoma patients. Large numbers of melanoma-specific T cells could be obtained when the REP protocol was applied to these MLTCs. Most MLTCs were enriched in non-terminally differentiated T(EM) cells homogeneously expressing co-stimulatory molecules (e.g., NKG2D, CD28, CD134, CD137). A similar pattern of anti-tumor activity, in association with a more variable expression of co-stimulatory molecules, was detected on short-term in vitro cultured TILs isolated from the same patients. In these ocular melanoma patients, we observed an immune infiltrate with suppressive characteristics and a low rate of ex vivo growing TILs (28.5% of our cases). Our MLTC protocol overcomes this limitation, allowing the isolation of T lymphocytes with effector functions even in these patients. Thus, anti-tumor circulating PBMC-derived T cells could be efficiently isolated from melanoma patients by our novel ex vivo enrichment protocol. This protocol appears suitable for ACT studies of cutaneous and ocular melanoma patients.

Laparoscopy-assisted gastrectomy versus open gastrectomy for gastric cancer: a monoinstitutional Western center experience.

BACKGROUND: Laparoscopic gastrectomy (LAG) is safe for benign lesions; however, such surgery for cancer remains controversial. The aim of this study was to compare technical feasibility and oncologic efficacy of laparoscopic versus open gastrectomy for gastric carcinoma. METHODS: Between January 2002 and November 2008, 109 gastric cancer patients underwent LAG (92 distal gastrectomy and 17 total gastrectomy) at our hospital. These patients were compared with 269 gastric cancer patients who underwent conventional open gastrectomy (OG; 171 distal gastrectomy and 98 total gastrectomy) during the same period. RESULTS: Operation time was significantly longer in the LAG group than in the OG group. Estimated blood loss in the LAG group was significantly less than in the OG group. The morbidity rate was higher than in the OG group (p < 0.0001). The distance of the proximal resection margin was significantly lower in the OG group (2.8 ± 1.9 vs. 3.8 ± 2.5; p = 0.014). The mean number of nodes resected with LAG was 31 ± 14 and that with OG was 27 ± 13 (p = 0.002). The mean survival time was 53 months in both groups. There were no differences regarding overall patient survival at a mean time of follow-up of 33 months. CONCLUSIONS: LAG with extended lymphadenectomy for gastric cancer is a feasible and safe procedure and has several advantages despite a higher rate of morbidity. Moreover, this method can achieve a radical oncologic equivalent resection and it does not have a deleterious effect on cancer-related outcome.

Predictors of morbidity and mortality in patients submitted to cytoreductive surgery plus hyperthermic intraperitoneal chemotherapy for ovarian carcinomatosis: a multicenter study.

OBJECTIVES: The aim of this retrospective study is to assess the incidence of morbidity and mortality related to cytoreductive surgery (CRS) plus hyperthermic intraperitoneal chemotherapy (HIPEC) and to evaluate their predictors, in patients with peritoneal metastasis of ovarian origin. METHODS: A retrospective multicenter study was carried out investigating results from eight Italian institutions. A total of 276 patients met inclusion criteria. Predictors of morbidity and mortality were evaluated with univariate and multivariate analysis. RESULTS: Overall morbidity was 71.4%, and severe complications occurred in 23.9% of the sample; 60-day mortality was 4.3%. According to univariate logistic regression models, grade 3-4 morbidity was related to Peritoneal Cancer Index (PCI) (OR 1.06; 95% CI 1.02-1.09; p<0.001), number of intraoperative blood transfusions (OR 1.21; 95% CI 1.10-1.34; p<0.001), Completeness of Cytoreduction (CC) score (OR 1.68; 95% CI 1.16-2.44; p=0.006) and number of anastomoses (OR 1.32; 95% CI 1.00-1.73; p=0.046). However, at the multivariate logistic regression analysis, only the number of intraoperative blood transfusions (OR 1.17; 95% CI 1.5-1.30; p=0.004) and PCI (OR 1.04; 95% CI 1.01-1.08; p=0.010) resulted as key predictors of severe morbidity. Furthermore, using multivariate logistic regression model, ECOG score (OR 2.45; 95% CI 1.21-4.93; p=0.012) and the number of severe complications (OR 2.16; 95% CI 1.03-4.52; p=0.042) were recorded as predictors of exitus within 60 days. CONCLUSIONS: The combination of CRS and HIPEC for treating peritoneal metastasis of ovarian origin has acceptable morbidity and mortality and, therefore, it can be considered as an option in selected patients.

Breakthrough Invasive Fungal Infections in Allogeneic Hematopoietic Stem Cell Transplantation.

Despite the recent introduction of mold-active antifungal prophylaxis (MAP), breakthrough invasive fungal infections (b-IFI) still represent a possible complication and a cause of morbidity and mortality in hematological patients and allogeneic hematopoietic stem-cell transplantation recipients (HSCT). Data on incidence and type of b-IFI are limited, although they are mainly caused by non-fumigatus Aspergillus and non-Aspergillus molds and seem to depend on specific antifungal prophylaxis and patients' characteristics. Herein, we described the clinical presentation and management of two cases of rare b-IFI which recently occurred at our institution in patients undergoing HSCT and receiving MAP. The management of b-IFI is challenging due to the lack of data from prospective trials and high mortality rates. A thorough analysis of risk factors, ongoing antifungal prophylaxis, predisposing conditions and local epidemiology should drive the choice of antifungal treatments. Early broad-spectrum preemptive therapy with a lipid formulation of amphotericin-B, in combination with a different mold-active azole plus/minus terbinafine, is advisable. The therapy would cover against rare azole-susceptible and -resistant fungal strains, as well as atypical sites of infections. An aggressive diagnostic work-up is recommended for species identification and subsequent targeted therapy.

Consensus Statement of the Italian Polispecialistic Society of Young Surgeons (SPIGC): Diagnosis and Treatment of Acute Appendicitis.

BACKGROUND: Acute appendicitis (AA) is one of the most common causes of abdominal pain requiring surgical intervention. Approximately 20% of AA cases are characterized by complications such as gangrene, abscesses, perforation, or diffuse peritonitis, which increase patients' morbidity and mortality. Diagnosis of AA can be difficult, and evaluation of clinical signs, laboratory index and imaging should be part of the management of patients with suspicion of AA. METHODS: This consensus statement was written in relation to the most recent evidence for diagnosis and treatment of AA, performing a literature review on the most largely adopted scientific sources. The members of the SPIGC (Italian Polispecialistic Society of Young Surgeons) worked jointly to draft it. The recommendations were defined and graded based on the current levels of evidence and in accordance with the criteria adopted by the American College of Chest Physicians (CHEST) for the strength of the recommendations. RESULTS: Fever and migratory pain tend to be present in patients with suspicion of AA. Laboratory and radiological examinations are commonly employed in the clinical practice, but today also scoring systems based on clinical signs and laboratory data have slowly been adopted for diagnostic purpose. The clinical presentation of AA in children, pregnant and elderly patients can be unusual, leading to more difficult and delayed diagnosis. Surgery is the best option in case of complicated AA, whereas it is not mandatory in case of uncomplicated AA. Laparoscopic surgical treatment is feasible and recommended. Postoperative antibiotic treatment is recommended only in patients with complicated AA.