Background: This study investigated the efficacy of chemoradiotherapy (CRT) followed by durvalumab as neoadjuvant therapy of locally advanced rectal cancer.Patients and methods: The PANDORA trial is a prospective, phase II, open-label, single-arm, multicenter study aimed at evaluating the efficacy and safety of preoperative treatment with durvalumab (1500 mg every 4 weeks for three administrations) following long-course radiotherapy (RT) plus concomitant capecitabine (5040 cGy RT in 25-28 fractions over 5 weeks and capecitabine administered at 825 mg/m2 twice daily). The primary endpoint was the pathological complete response (pCR) rate; secondary endpoints were the proportion of clinical complete remissions and safety. The sample size was estimated assuming a null pCR proportion of 0.15 and an alternative pCR proportion of 0.30 (a = 0.05, power = 0.80). The proposed treatment could be considered promising if >= 13 pCRs were observed in 55 patients (EudraCT: 2018-004758-39; NCT04083365).Results: Between November 2019 and August 2021, 60 patients were accrued, of which 55 were assessable for the study's objectives. Two patients experienced disease progression during treatment. Nineteen out of 55 eligible patients achieved a pCR (34.5%, 95% confidence interval 22.2% to 48.6%). Regarding toxicity related to durvalumab, grade 3 adverse events (AEs) occurred in four patients (7.3%) (diarrhea, skin toxicity, transaminase increase, lipase increase, and pancolitis). Grade 4 toxicity was not observed. In 20 patients (36.4%), grade 1-2 AEs related to durvalumab were observed. The most common were endocrine toxicity (hyper/hypothyroidism), dermatologic toxicity (skin rash), and gastrointestinal toxicity (transaminase increase, nausea, diarrhea, constipation).Conclusion: This study met its primary endpoint showing that CRT followed by durvalumab could increase pCR with a safe toxicity profile. This combination is a promising, feasible strategy worthy of further investigation.

The combination of preoperative chemo- and radiotherapy (CRT) is the standard treatment in locally advanced rectal cancer (LARC). The benefit of the addition of immunotherapy in the neoadjuvant treatment is under investigation. However, a wide genomic characterization is needed to better stratify LARC patients (pts) for the most adequate treatment. We analysed cancer tissues from 48 pts enrolled in the PANDORA trial (NCT04083365) in which a capecitabine-based concomitant CRT followed by durvalumab was administrated. DNA was extracted from bioptic chemo-naïve formalin-fixed paraffin-embedded samples. Genetic alterations of more than 500 genes, microsatellite instability (MSI) and tumor mutational burden (TMB) status were characterized using TSO500HT panel on an Illumina Novaseq 6000 instrument. PD-L1 expression was evaluated by the combined positive score (CPS). Pathological response was centrally determined on surgical specimens and was evaluable for 46 pts. A pathological complete response (pCR) was defined as ypT0N0M0. Pathogenic variants on APC, TP53, KRAS, FBXW7, PIK3CA, SMAD4, AMER1 and ARID1A genes were the most frequent mutations. Overall, 17/46 (37%) pts achieved a pCR. Interestingly, ARID1A and SMAD4 alterations occur only in patients without a pCR, being mutated in the 17% and 21% of pts, respectively. In particular, a more significant association was reached for SMAD4 when either pathogenetic variants and variants of uncertain significance (VUS) were considered (p=0.019). Only 4% and 31% of pts had MSI and high TMB, respectively, but no significant association with response was found. High TMB status was significantly related to ARID1A, FBXW7, MYC and RICTOR1 mutations. SMAD4 mutations resulted significantly associated with low levels of PD-L1 expression (p=0.017). Indeed, 75% of the mutated pts as compared to the 27% of the wild type pts reported PD-L1 CPS values less than 10. SMAD4 mutations are related to the absence of complete response and to a low PD-L1 expression. SMAD4 alterations should need further investigation in LARC pts treated by combined CRT followed by durvalumab.

Background: Complete surgical resection (R0) of the primary tumor and regional lymphadenectomy is the treatment of choice for gastric cancer. Perioperative chemotherapy is considered a standard approach for locally advanced gastric and gastroesophageal junction (GEJ) cancer. With this study we examined morbidity, mortality, and outcomes in patients after total gastrectomy plus at least D2 lymphadenectomy for operable, locally advanced GEJ and gastric cancer.

4084 Background: Studies on Asian, US, and German patients have moved some criticisms on the validity of the 7th edition of the AJCC classification to discriminate outcome of gastric cancer stages. We investigated the effect of this AJCC classification in a high-quality surgical populations of patients receiving D2 lymphadenectomy. Methods: From the prospective database at San Salvatore Hospital, Pesaro, we identified 515 patientswith gastroesophageal junction (Siewert II and III) or stomach adenocarcinoma who underwent gastrectomy with curative intent from 1998 to 2010. Lymphadenectomy extended to the 3rd level 12p/b nodes (D2/D3) was performed in all patients. Overall survival (OS) probabilities, calculated from the date of surgery to the date of death, from any cause, were estimated using the Kaplan-Meier method and compared using the log-rank test. Results: 58% of patients were male,median age was 73 years (range 36-96). Median number of examined lymph nodes was 32 (range, 1-89), and only 8.9% of patients had less than 15 examined lymph nodes; 96 patients received adjuvant chemo- or chemoradiotherapy. As shown in the table, we proposed a revised staging system (Pesaro Staging System, PSS), which performs better than the 7th edition of AJCC classification in terms of survival differences between stages. Conclusions: This study confirms once again that the 7th edition of the AJCC classification does not discriminate adequately the outcome from stage to stage. In a European population of patients undergoing gastrectomy plus at least D2 lymphadenectomy, the revised staging system, PSS, better defines patient prognosis. [Table: see text]

4038 Background: Activation of the c-MET oncogene promotes tumor growth, invasion and metastasis. It has been shown that c-MET mutations are almost lacking in GC, while there is limited information on c-MET activation by CNV in caucasians. We investigated c-MET CNV in GC patients in Italy. Methods: A single-institution surgical database was used for this retrospective analysis. High risk stage II-III consecutive patients who underwent gastrectomy (R0 with D2 lymphadenectomy) between 1998 and 2007 were selected. For each case, formalin-fixed paraffin-embedded tumor specimens were used for DNA extraction and c-METCNV detection by the TaqMan Gene Copy Number Assay (Applied Biosystem). The relative number of copies of three c-MET regions (Hs01602615 in exon 3, Hs05027935 in exon 12, Hs02884964 in exon 21) was determined. A sample was defined as c-METCNV>2 when this status was found in at least two of the three studied c-METregions. Results were analyzed for association with clinico-pathologic features and survival outcomes. Results: In 214 gastric carcinomas 98 cases (45%) showed c-MET CNV>2. No significant association with clinico-pathologic features was detected except for tumor histotype according to Lauren’s classification (p=0.01). c-MET CNV>2 occurred in 63/98 (64%) and 35/98 (36%) cases of intestinal and non-intestinal subtypes, respectively, while c-MET CNV<2 occurred in 64/116 (55%) and 52/116 (45%) cases of intestinal and non-intestinal subtypes, respectively. After a minimun follow-up time of three years, there were 87 relapses (40%). Log-rank tests showed significantly worse relapse-free survival (p=0.0008) and overall survival (p=0.0001) in patients with c-MET CNV>2. In multivariate models, these associations remained significant together with tumor stage. Conclusions: qPCR showed c-MET CNV in GC caucasian patients with impact on clinical outcomes. This finding is relevant to the current development of anti-MET therapeutics in this lethal disease. Ongoing analyses are refining the prognostic role of c-MET CNV considering CNV subgroups within the >2 copies group.

Self-expanding metal stents (SEMS) can alleviate malignant colorectal obstruction (MCRO) and avoid emergency decompressive surgery, usually reducing morbidity and mortality compared with elective situations. Combining SEMS placement with elective surgery also appeared safer and more effective than emergency surgery, with higher rates of primary anastomosis, lower rates of colostomy, shorter hospital stays and lower overall complications rate. The purpose of this study was to review the use of SEMS as the initial interventional approach in the management of acute MCRO, evaluating the effectiveness both as a bridge to surgery and definitive palliative treatment. 92 patients (57 males, 35 females, age range 48-89 years, mean age 74.5) underwent the insertion of a nitinol SEMS presenting with acute MCRO were retrospectively evaluated during the period of time from January 2002 through March 2010. The SEMS was inserted into the obstructive site by using trough-the-scope method. Technical success was defined as accurate SEMS deployment across the stricture on the first attempt; clinical success was defined as decompression and relief of obstructive colonic symptoms maintained without intervention or serious device-related complications. Gastroenterology and Digestive Endoscopy Unit of San Salvatore Hospital, Pesaro - Italy. Technical success was achieved in 92.3% (85/92) of patients with a 95% CI of 87% to 98% and initial clinical success in 97.6% (83/85) with a 95% CI of 94% to 100%. The stent was placed in the rectum (18/92 - 19.5%), sigmoid (46/92 - 50%), left colon (23/92 - 25%), transverse colon (4/92 - 4.3%), right colon (1/92 - 1%). In 29 patients (31.5%) the SEMS were attempted as primary palliative definitive treatment, whereas in 63 (68.5%) were placed as a bridge to surgery. All patients with operable tumours were successfully bridge to elective surgery within a median of 14 days from SEMS placement. SEMS-related perforation occurred in 4.3% of patients (4/92), obstruction in 7.6% (7/92) and migration was reported in 6.5% (6/92) of cases. In general endoscopic practice SEMS insertion is an effective and safe treatment for patients with acute MCRO, providing time for a complete preoperative evaluation and avoiding emergency surgery with considerably lower morbidity and mortality rates. This procedure provides an effective outcome in patients with advanced cancer as a palliative therapy to surgery with satisfactory results. Moreover after palliative stent insertion the more rapid and less complicated recovery may facilitate the early administration of chemotherapy.

PURPOSE To investigate whether prognosis of patients with high-risk gastric cancer may depend on MET copy number gain (CNG) or an activating truncation within a deoxyadenosine tract element (DATE) in the promoter region of the MET ligand HGF. PATIENTS AND METHODS A single-institution cohort of 230 patients with stage II/III gastric cancer was studied. Formalin-fixed paraffin-embedded tumor specimens were used for DNA extraction. Quantitative polymerase chain reaction (qPCR) for MET CNG and sequencing for HGF DATE truncation (< 25 deoxyadenosines instead of 30) were used. Results were analyzed for association with disease-free survival (DFS) and overall survival (OS). To assess the reliability of the qPCR measurement, a random sample of cases was reanalyzed using an alternative assay (fluorescent in situ hybridization [FISH]) with calculation of the intracorrelation coefficient (ICC). RESULTS In 216 assessable patients, MET CNG five or more copies and homozygous HGF-truncated DATE occurred in 21 patients (10%) and 30 patients (13%), respectively. Patients with MET CNG five or more copies (MET-positive) showed significantly worse prognosis with multivariate hazard ratio (HR) of 3.02 (95% CI, 1.71 to 5.33; P < .001) for DFS and multivariate HR of 2.91 (95% CI, 1.65 to 5.11; P < .001) for OS. The agreement between qPCR and FISH was high, with ICC = 0.9% (95% CI, 0.81% to 0.95%; the closer the ICC is to 1, the greater is the agreement). HGF-truncated DATE did not show relevant prognostic effect. CONCLUSION In this study, qPCR revealed approximately 10% of white patients with gastric cancer harboring MET CNG of five or more copies. This marker was significantly associated with unfavorable prognosis. This information is relevant to the current clinical development of anti-MET compounds.

4527 Background: To assess the clinical significance of tumor-infiltrating FOXP3-positive Tregs in patients with R0, stage II-III, GC. Methods: From a single-institution database, tumors of 110 consecutive GC patients who underwent R0 resection for stage II- III disease were studied for FOXP3-positive Tregs by immunohistochemistry. A minimum 3-year follow-up time and the Reporting Recommendations for Tumor Marker Prognostic Studies (REMARK) criteria were used for planning the study. The recurrences of GC had to be confirmed by cytology biopsy or surgery. None of the patients received chemotherapy or radiation therapy before or after surgery as a part of an adjuvant program. The study was conducted in a blinded fashion so that patients’ outcomes were unknown to investigators performing immunohistochemistry. The observed median number of FOXP3-positive Tregs was used as cut-point in analyses (<6 vs. >6 count). To explore the association between overall survival (OS) and relapse-free survival (RFS) with FOXP3 count stratified log-rank tests were used. To further define these relationships, Cox proportional hazards models for multivariate analyses with Hazard Ratio (HR) and 95% confidence interval (95%CI) were employed. Results: Tregs numbers were significantly higher in GC than in normal tissue (p=0.0001) and Tregs count >6 was significantly associated with vascular/lymphatic/perineural invasion (VELIPI) in the tumor (p=.03). Forty-eight patients (44%) relapsed and died after receiving cisplatin/fluorouracil-based palliative chemotherapy. In multivariate analysis, adverse RFS was associated with grading 3 (HR=2.15; 95%CI 1.19–3.89, p=0.01), stage III (HR=2.34; 95%CI 1.20–4.57, p=0.01), VELIPI (HR=1.96; 95%CI 1.08–3.55, p=0.02) and Tregs count >6 (HR=2.00; 95%CI 1.10–3.65, p=0.02), while adverse OS was associated with grading 3 (HR=2.21; 95%CI 1.22–3.98, p=0.008), stage III (HR=2.40; 95%CI 1.23–4.67, p=0.01) and Tregs count >6 (HR=2.34; 95%CI 1.27–4.28, p=0.006). Conclusions: Quantification of FOXP3-positive Tregs may be a novel marker for identifying high-risk GC patients. Present findings deserve additional investigation as Tregs may also represent an innovative therapeutic target. No significant financial relationships to disclose.

BACKGROUND:We investigated whether an endogastric capsule (EC) may be a valuable tool for collecting DNA from exfoliated cells from the gastric mucosa and for carrying out an analysis of promoter methylation status of the E-cadherin (CDH1) gene in poorly differentiated, diffuse gastric cancer (DGC).MATERIAL AND METHODS:Consecutive patients with a confirmed diagnosis of poorly differentiated DGC underwent collection of gastric juice by EC. Subjects without cancer and premalignant lesions were also accrued as controls. The samples of gastric juice were processed for DNA isolation and amplification. Then they were used for analysis of CDH1 promoter hypermethylation.RESULTS:The procedure successfully allowed the analysis of CDH1 promoter hypermethylation in 20 patients and 14 controls. This pilot study showed feasibility of the procedure and a significantly different CDH1 promoter hypermethylation status between DGC patients and controls was detected.CONCLUSIONS:The EC may represent an innovative and noninvasive tool for the analysis of a specific epigenetic change in DGC patients. Our findings deserve additional studies as this method may represent a cost-effective tool for early detection of sporadic as well as hereditary DGC in CDH1 germline mutations carriers.

We investigated the clinical significance of tumour-infiltrating FOXP3-positive regulatory T cells (Tregs) in radically resected (R0) gastric cancer. From a single-institution database, tumors of 110 patients who underwent R0 resection for stage II-III disease were studied for FOXP3-positive Tregs by immunohistochemistry. The observed median number of FOXP3-positive Tregs was used as the cut-point in analyses (<6 versus >or=6 count). Tregs were significantly higher in gastric carcinomas than in normal tissue (P = 0.0001). Tregs count >or=6 was significantly associated with vascular/lymphatic/perineural invasion (VELIPI) in the tumour (P = 0.03). Multivariate analysis showed association between adverse relapse-free survival and grading 3, stage III, VELIPI and Tregs count >or=6 (P = 0.02). Adverse overall survival was associated with grading 3, stage III, VELIPI and Tregs count >or=6 (P = 0.006). FOXP3-positive Tregs may be a novel marker for identifying high-risk gastric cancer patients. Present findings deserve additional investigation as Tregs may also represent an innovative therapeutic target.