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Prevalence of gains and losses according to histological features

List of genes in MLPA probe mixes

About 5% of colorectal adenomas are estimated to progress to colorectal cancer. However, it is important to identify which adenomas actually carry a high risk of progression, because these serve as intermediate endpoints, for example, in screening programs. In clinical practice, adenomas with a size of ≥10 mm, villous component and/or high-grade dysplasia, called advanced adenomas, are considered high risk, although solid evidence for this classification is lacking. Specific DNA copy number changes are associated with adenoma-to-carcinoma progression. We set out to determine the prevalence of cancer-associated events (CAE) in advanced and nonadvanced adenomas. DNA copy number analysis was performed on archival tissues from three independent series of, in total, 297 adenomas (120 nonadvanced and 177 advanced) using multiplex ligation-dependent probe amplification or low-coverage whole-genome DNA sequencing. Alterations in two or more CAEs were considered to mark adenomas as high risk. Two or more CAEs were overall present in 25% (95% CI, 19.0–31.8) of advanced adenomas; 23% (11/48), 36% (12/33), and 23% (22/96) of the advanced adenomas in series 1, 2, and 3, respectively, and 1.7% (1/58) and 4.8% (3/62) of the nonadvanced adenomas, in series 1 and 2, respectively. The majority of advanced adenomas do not show CAEs, indicating that only a subset of these lesions is to be considered high risk. Nonadvanced adenomas have very low prevalence of CAEs, although those with CAEs should be considered high risk as well. Specific DNA copy number alterations may better reflect the true progression risk than the advanced adenoma phenotype. Cancer Prev Res; 11(7); 403–12. ©2018 AACR.

PDF file, 4178K, Frequency plot of different subtypes, dysplasia, origin, gender, age and location.

Figure S3. FOXP3 evaluated on-treatment.

PDF file, 100K, Supplementary table 1; Common chromosomal regions present in more than 10% of polypoid or flat colorectal adenomas. Supplementary table 2; All known genes and microRNA located on 5q15-q31.1 Supplementary table 3; Cross tables of A; APC mutation status versus phenotype, B; APC mutation status versus presents of 5qloss in flat adenomas, C; APC mutation status versus presents of 5q loss in polypoid adenomas, D; The amount of beta-catenin binding repeats versus phenotype. Supplementary table 4; Significant different chromosomal regions (FDR

Purpose:The CROSS trial established neoadjuvant chemoradiotherapy (nCRT) for patients with resectable esophageal adenocarcinoma (rEAC). In the PERFECT trial, we investigated the feasibility and efficacy of nCRT combined with programmed-death ligand-1 (PD-L1) inhibition for rEAC.Patients and Methods:Patients with rEAC received nCRT according to the CROSS regimen combined with five cycles of atezolizumab (1,200 mg). The primary endpoint was the feasibility of administering five cycles of atezolizumab in ≥75% patients. A propensity score–matched nCRT cohort was used to compare pathologic response, overall survival, and progression-free survival. Exploratory biomarker analysis was performed on repeated tumor biopsies.Results:We enrolled 40 patients of whom 85% received all cycles of atezolizumab. Immune-related adverse events of any grade were observed in 6 patients. In total, 83% proceeded to surgery. Reasons for not undergoing surgery were progression (n = 4), patient choice (n = 2), and death (n = 1). The pathologic complete response rate was 25% (10/40). No statistically significant difference in response or survival was found between the PERFECT and the nCRT cohort. Baseline expression of an established IFNγ signature was higher in responders compared with nonresponders (P = 0.043). On-treatment nonresponders showed either a high number of cytotoxic lymphocytes (CTL) with a transcriptional signature consistent with expression of immune checkpoints, or a low number of CTLs.Conclusions:Combining nCRT with atezolizumab is feasible in patients with rEAC. On the basis of our exploratory biomarker study, future studies are necessary to elucidate the potential of neoadjuvant immunotherapy in patient subgroups.See related commentary by Catenacci, p. 3269

PDF file, 1948K, Frequency plot of different histology types.

Purpose: Flat adenomas are a subgroup of colorectal adenomas that have been associated with a more aggressive clinical behavior compared with their polypoid counterparts. Here, we aimed to compare one of the molecular changes most explicitly associated with adenoma to carcinoma progression, that is, chromosomal instability, between flat and polypoid colorectal adenomas.Experimental Design: Consecutive series of 83 flat and 35 polypoid adenomas were analyzed for DNA copy number changes using a high-resolution array comparative genomic hybridization platform, microsatellite instability (MSI) status, and for mutations in the adenomatous polyposis coli (APC) gene. Immunohistochemical stainings for CD3, CD8, and FoxP3 expression were carried out.Results: Patterns of DNA copy number changes differed between the two phenotypes, with significantly more frequent loss of 5q14.3 and 5q15-q31.1 in flat adenomas, whereas losses of 1p36.32-p35.3, 10q25.3, 17p12, and chromosome 18 were more frequent in polypoid adenomas (false discovery rate < 0.2). MSI was observed in one flat adenoma. As the 5q15-q31.1 region harbors the APC locus, APC mutation status was investigated, showing significantly less mutations in flat adenomas (P = 0.04). An initial exploration of a possible association of 5q loss with inflammation indicated that tumor-infiltrating lymphocytes were more abundant in the stroma of flat adenomas compared with that of polypoid adenomas.Conclusion: Flat and polypoid adenomas have partially distinct chromosomal profiles, consistent with differences in the biology underlying these phenotypes. Alterations more specific to flat adenomas, in particular 5q loss, may be associated with inflammation. Clin Cancer Res; 18(17); 4560–9. ©2012 AACR.

Table S1. Combined positivity score (CPS) in responders and non-responders at baseline. Table S2. Immunohistochemistry markers evaluated at baseline and on-treatment.

PDF file, 464K, Concept of colorectal carcinogenesis and histological images of adenomas.

Contains fulltext : 249895.pdf (Publisher’s version ) (Open Access) AIM: The aim of this study was to evaluate interobserver variability between individual pathologists and a panel of pathologists in the histopathological assessment of advanced colorectal neoplasms in the Dutch bowel cancer screening population. METHODS AND RESULTS: Histological slides of adenomas with high-grade dysplasia and early colorectal carcinomas (CRC) from 20 different laboratories were reviewed by the pathology panel of the Dutch bowel screening programme. Interobserver variability was reported by descriptive statistics. In addition, potential clinical consequences of discrepancies were evaluated. A total of 104 cases of adenomas with high-grade dysplasia and 83 early CRCs were reviewed. Discrepancies were observed in 41 of 104 (39.4%) adenoma cases, which potentially had clinical consequences in 16 (15.4%) cases. For CRC, discrepancies were shown in 44 of 83 cases (53.0%) and would have potentially led to alternative treatment strategies in 25 (30.1%) cases. Most frequently, discrepancies were observed in the assessment of lymphovascular invasion (23 of 73 cases, 31.5%). CONCLUSION: This study showed that considerable interobserver variability is present in the histopathological assessment of advanced colorectal neoplasia, which may impact upon treatment choices. Additional stains and education, as well as intercollegial consultation, might decrease this variability.

Introduction: In early rectal cancer, organ sparing treatment strategies such as local excision have gained popularity. The necessity of radical surgery is based on the histopathological evaluation of the local excision specimen. This study aimed to describe diagnostic variability between pathologists, and its impact on treatment allocation in patients with locally excised early rectal cancer. Materials and methods: Patients with locally excised pT1-2 rectal cancer were included in this prospective cohort study. Both quantitative measures and histopathological risk factors (i.e. poor differentiation, deep submucosal invasion, and lymphatic- or venous invasion) were evaluated. Interobserver variability was reported by both percentages and Fleiss’ Kappa- (ĸ) or intra-class correlation coefficients. Results: A total of 126 patients were included. Ninety-four percent of the original histopathological reports contained all required parameters. In 73 of the 126 (57.9%) patients, at least one discordant parameter was observed, which regarded histopathological risk factors for lymph node metastases in 36 patients (28.6%). Interobserver agreement among different variables varied between 74% and 95% or ĸ 0.530–0.962. The assessment of lymphovascular invasion showed discordances in 26% (ĸ = 0.530, 95% CI 0.375–0.684) of the cases. In fourteen (11%) patients, discordances led to a change in treatment strategy. Conclusion: This study demonstrated that there is substantial interobserver variability between pathologists, especially in the assessment of lymphovascular invasion. Pathologists play a key role in treatment allocation after local excision of early rectal cancer, therefore interobserver variability needs to be reduced to decrease the number of patients that are over- or undertreated.

Objective: Biological sex differences in cancer are increasingly acknowledged. Here, we examined these differences in clinicopathological characteristics and survival in microsatellite instability (MSI)-high and microsatellite stable (MSS) gastric cancer (GC).Design: We analysed MSI status by polymerase chain reaction (PCR) and/or mismatch repair (MMR) status by immunohistochemistry in a pooled analysis of individual patient data from one retrospective cohort from Cologne, and the randomised phase III clinical trials D1/D2 and CRITICS. All patients had resectable adenocarcinoma of the stomach and/or gastro-oesophageal junction. Patients were treated with either surgery only or perioperative chemo(radio) therapy.Results: MSI and/or MMR analyses on 1307 tumours resulted in 1192 (91.2%) MSS and/or MMR proficient (MMRP) [median age, 65 years; 759 males (63.7%); 619 treated with surgery only (51.9%)], and 115 (8.8%) MSI-high [median age, 69 years; 67 males (58.3%); 76 treated with surgery only (66.1%)] GC cases. Males had shorter overall survival (OS) than female MSI-high GC (5-year OS 34.7% vs. 69.7%; hazard ratio (HR) 2.68, 95%CI 1.60 to 4.49; p < 0.001). Females with MSI-high had longer OS than those with MSS/MMRP GC ( HR 0.61, 95%CI 0.41 to 0.92; p = 0.02). Males with MSI-high did not have longer OS than those with MSS/MMRP GC (HR 1.26, 95%CI 0.94 to 1.69; p = 0.12).Conclusions: MSI-high GC males had a significantly worse prognosis compared to their female counterparts in three independent cohorts. In addition, the favourable prognostic value of MSI was only seen in females and not in males. These observations emphasise the need to consider sex differences in prognosis and treatment effects in oncology. Clinical trial registration: The CRITICS trial is registered at ClinicalTrials.gov, number NCT00407186; EudraCT, number 2006-004130-32; and CKTO, 2006-02. (C) 2022 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

Purpose: The CROSS trial established neoadjuvant chemoradiotherapy (nCRT) for patients with resectable esophageal adenocarcinoma (rEAC). In the PERFECT trial, we investigated the feasibility and efficacy of nCRT combined with programmed-death ligand-1 (PD-L1) inhibition for rEAC. Patients and Methods: Patients with rEAC received nCRT according to the CROSS regimen combined with five cycles of atezolizumab (1,200 mg). The primary endpoint was the feasibility of administering five cycles of atezolizumab in ≥75% patients. A propensity score–matched nCRT cohort was used to compare pathologic response, overall survival, and progression-free survival. Exploratory biomarker analysis was performed on repeated tumor biopsies. Results: We enrolled 40 patients of whom 85% received all cycles of atezolizumab. Immune-related adverse events of any grade were observed in 6 patients. In total, 83% proceeded to surgery. Reasons for not undergoing surgery were progression (n = 4), patient choice (n = 2), and death (n = 1). The pathologic complete response rate was 25% (10/40). No statistically significant difference in response or survival was found between the PERFECT and the nCRT cohort. Baseline expression of an established IFNγ signature was higher in responders compared with nonresponders (P = 0.043). On-treatment nonresponders showed either a high number of cytotoxic lymphocytes (CTL) with a transcriptional signature consistent with expression of immune checkpoints, or a low number of CTLs. Conclusions: Combining nCRT with atezolizumab is feasible in patients with rEAC. On the basis of our exploratory biomarker study, future studies are necessary to elucidate the potential of neoadjuvant immunotherapy in patient subgroups. See related commentary by Catenacci, p. 3269

Background Improvement of radiotherapy efficacy requires better insight in the dynamic responses that occur during irradiation. Here, we aimed to identify the molecular responses that are triggered during clinically applied fractionated irradiation. Methods Gene expression analysis was performed by RNAseq or microarray analysis of cancer cells or xenograft tumors, respectively, subjected to 3–5 weeks of 5 × 2 Gy/week. Validation of altered gene expression was performed by qPCR and/or ELISA in multiple cancer cell lines as well as in pre- and on-treatment biopsies from esophageal cancer patients (NCT02072720). Targeted protein inhibition and CRISPR/Cas-induced gene knockout was used to analyze the role of type I interferons and cGAS/STING signaling pathway in the molecular and cellular response to fractionated irradiation. Results Gene expression analysis identified type I interferon signaling as the most significantly enriched biological process induced during fractionated irradiation. The commonality of this response was confirmed in all irradiated cell lines, the xenograft tumors and in biopsies from esophageal cancer patients. Time-course analyses demonstrated a peak in interferon-stimulated gene (ISG) expression within 2–3 weeks of treatment. The response was accompanied by a variable induction of predominantly interferon-beta and/or -lambda, but blocking these interferons did not affect ISG expression induction. The same was true for targeted inhibition of the upstream regulatory STING protein while knockout of STING expression only delayed the ISG expression induction. Conclusions Collectively, the presented data show that clinically applied fractionated low-dose irradiation can induce a delayed type I interferon response that occurs independently of interferon expression or STING signaling. These findings have implications for current efforts that aim to target the type I interferon response for cancer treatment.

Aim To evaluate the prognostic value of tumor markers in a European cohort of patients with resectable gastric cancer. Methods We performed a post hoc analysis of the CRITICS trial, in which 788 patients received perioperative therapy. Association between survival and pretreatment CEA, CA 19-9, alkaline phosphatase, neutrophils, hemoglobin and lactate dehydrogenase were explored in uni- and multivariable Cox regression analyses. Likelihoods to receive potentially curative surgery were investigated for patients without elevated tumor markers versus one of the tumor markers elevated versus both tumor markers elevated. The association between tumor markers and the presence of circulating tumor DNA (ctDNA) was explored in 50 patients with available ctDNA data. Results In multivariable analysis, in which we corrected for allocated treatment and other baseline characteristics, elevated pretreatment CEA (HR 1.43; 95% CI 1.11–1.85, p p p Conclusion CEA and CA 19-9 were independent prognostic factors for survival in a large cohort of European patients with resectable gastric cancer. No relationship was found between tumor markers and ctDNA. These factors could potentially guide treatment choices and should be included in future trials to determine their definitive position. Trial registration ClinicalTrial.gov identifier: NCT00407186. EudraCT number: 2006-00413032.

Background Palliative systemic therapy is currently standard of care for patients with extensive metastatic colorectal cancer (mCRC). A biomarker predicting chemotherapy benefit which prevents toxicity from ineffective treatment is urgently needed. Therefore, a previously developed tissue‐derived microRNA profile to predict clinical benefit from chemotherapy was evaluated in tissue biopsies and serum from patients with mCRC. Methods Samples were prospectively collected from patients (N = 132) who were treated with capecitabine or 5‐FU/LV with oxaliplatin ± bevacizumab. Response evaluation was performed according to RECIST 1.1 after three or four cycles, respectively. Baseline tissue and serum miRNAs expression levels of miR‐17‐5p, miR‐20a‐5p, miR‐30a‐5p, miR‐92a‐3p, miR‐92b‐3p, and miR‐98‐5p were quantified with RT‐qPCR and droplet digital PCR, respectively. Combined predictive performance of selected variables was tested using logistic regression analysis. Results From 132 patients, 81 fresh frozen tissue biopsies from metastases and 93 serum samples were available. Based on expression levels of miRNAs in tissue, progressive disease could be predicted with an AUC of 0.85 (95% CI:0.72‐0.91) and response could be predicted with an AUC of 0.70 (95% CI:0.56‐0.80). This did not outperform clinical parameters alone (respectively P = .14 and P = .27). Expression levels of miR‐92a‐3p and miR‐98‐5p in serum significantly improved the predictive value of clinical parameters for response to chemotherapy (AUC 0.74, 95% CI:0.64‐0.84, P = .003) in this cohort. Conclusions The additive predictive value to clinical parameters of the tissue‐derived six miRNA profile for clinical benefit could not be validated in patients with mCRC treated with first‐line systemic therapy. Although miR‐92a‐3p and miR‐98‐5p serum levels improved the predictive value of clinical parameters, it remained insufficient for clinical decision‐making., For miR‐17‐5p, miR‐20a‐5p, miR‐30a‐5p, miR‐92a‐3p, miR‐92b‐3p and miR‐98‐5p no correlation was observed between tissue and serum miRNA expression levels in patients with metastatic colorectal cancer (mCRC). The predictive value of this tissue‐derived 6 miRNA profile for clinical benefit from systemic treatment could not be validated in patients with mCRC starting first line systemic therapy.

Gastric cancer (GC) patients at high risk of developing peritoneal metastasis (PM) as a single site of metastasis after curative treatment may be candidates for adjuvant prophylactic strategies. Here we investigated risk factors for metachronous isolated PM in patients who were treated in the CRITICS trial (NCT00407186). Univariable and multivariable analyses on both metachronous isolated PM and ‘other events’, i.e., (concurrent) distant metastasis, locoregional recurrence or death, were performed using a competing risk model and summarized by cumulative incidences. Isolated PM occurred in 64 of the 606 (11%) included patients. Diffuse or mixed histological subtype, ypT4 tumor stage and LNhigh (ypN3 lymph node stage or a lymph node ratio &gt, 20%) were independent risk factors for isolated PM in both univariable and multivariable analyses. Likewise, LNhigh was an independent risk factor for ‘other events’. Patients with tumors who were positive for all three independent risk factors had the highest two-year cumulative incidence of 43% for isolated PM development. In conclusion, diffuse or mixed histological subtype, ypT4 and LNhigh were identified as independent risk factors for isolated PM in patients treated with preoperative chemotherapy followed by surgical resection. The combination of these factors may identify a subgroup that may benefit from PM-preventing treatment strategies.

Objective: We examined the association between surgical hospital volume and both overall survival (OS) and disease-free survival (DFS) using data obtained from the international CRITICS (ChemoRadiotherapy after Induction chemotherapy In Cancer of the Stomach) trial. Summary Background Data: In the CRITICS trial, patients with resectable gastric cancer were randomized to receive preoperative chemotherapy followed by adequate gastrectomy and either chemotherapy or chemoradiotherapy. Methods: Patients in the CRITICS trial who underwent a gastrectomy with curative intent in a Dutch hospital were included in the analysis. The annual number of gastric cancer surgeries performed at the participating hospitals was obtained from the Netherlands Cancer Registry; the hospitals were then classified as low-volume (1 – 20 surgeries/year) or high-volume (≥21 surger-surgeries/year) and matched with the CRITICS trial data. Univariate and multivariate analyses were then performed to evaluate the hazard ratio (HR) between hospital volume and both OS and DFS. Results: From 2007 through 2015, 788 patients were included in the CRITICS trial. Among these 788 patients, 494 were eligible for our study; the median follow-up was 5.0 years. Five-year OS was 59.2% and 46.1% in the high-volume and low-volume hospitals, respectively. Multivariate analysis revealed that undergoing surgery in a high-volume hospital was associated with higher OS [HR ¼ 0.69, 95% confidence interval (CI) ¼ 0.50 – 0.94, P ¼ 0.020] and DFS (HR ¼ 0.73, 95% CI: 0.54 – 0.99, P ¼ 0.040). Conclusions: In the CRITICS trial, hospitals with a high annual volume of gastric cancer surgery were associated with higher overall and DFS. These findings emphasize the value of centralizing gastric cancer surgeries in the Western world.

Changes in glycosylation during tumour progression are a key hallmark of cancer. One of the glycan moieties generally overexpressed in cancer are sialic acids, which can induce immunomodulatory properties via binding to Siglec receptors. We here show that Pancreatic Ductal Adenocarcinoma (PDAC) tumour cells present an increased sialylation that can be recognized by Siglec-7 and Siglec-9 on myeloid cells. We identified the expression of the α2,3 sialyltransferases ST3GAL1 and ST3GAL4 as main contributor to the synthesis of ligands for Siglec-7 and Siglec-9 in tumour cells. Analysing the myeloid composition in PDAC, using single cell and bulk transcriptomics data, we identified monocyte-derived macrophages as contributors to the poor clinical outcome. Tumour-derived sialic acids dictate monocyte to macrophage differentiation via signalling through Siglec-7 and Siglec-9. Moreover, triggering of Siglec-9 in macrophages reduce inflammatory programmes, while increasing PD-L1 and IL-10 expression, illustrating that sialic acids modulate different myeloid cells. This work highlights a critical role for sialylated glycans in controlling immune suppression and provides new potential targets for cancer immunotherapy in PDAC., Alterations in glycosylation in tumours facilitate tumour progression. Here, the authors show that pancreatic ductal adenocarcinomas present increased sialylation, which stimulates the polarisation of monocytes via Siglec receptors, resulting in the generation of immune suppressive tumour associated macrophages.

Identification of molecular predictive markers of response to neoadjuvant chemoradiation could aid clinical decision‐making in patients with localized oesophageal cancer. Therefore, we subjected pretreatment biopsies of 75 adenocarcinoma (OAC) and 16 squamous cell carcinoma (OSCC) patients to targeted next‐generation DNA sequencing, as well as biopsies of 85 OAC and 20 OSCC patients to promoter methylation analysis of eight GI‐specific genes, and subsequently searched for associations with histopathological response and disease‐free (DFS) and overall survival (OS). Thereby, we found that in OAC, CSMD1 deletion (8%) and ETV4 amplification (5%) were associated with a favourable histopathological response, whereas SMURF1 amplification (5%) and SMARCA4 mutation (7%) were associated with an unfavourable histopathological response. KRAS (15%) and GATA4 (7%) amplification were associated with shorter OS. In OSCC, TP63 amplification (25%) and TFPI2 (10%) gene promoter methylation were associated with an unfavourable histopathological response and shorter DFS (TP63) and OS (TFPI2), whereas CDKN2A deletion (38%) was associated with prolonged OS. In conclusion, this study identified candidate genetic biomarkers associated with response to neoadjuvant chemoradiotherapy in patients with localized oesophageal cancer., Using DNA sequencing and methylation panels on pretreatment biopsies, we identified candidate molecular alterations associated with response [tumour regression grade (TRG) or survival] to neoadjuvant chemoradiotherapy in localized oesophageal adenocarcinoma (OAC) and squamous cell carcinoma (OSCC). These findings may assist approaches to further individualize oesophageal cancer treatment.

Immunotherapy is a new anti-cancer treatment option, showing promising results in clinical trials. To investigate potential immune biomarkers in esophageal adenocarcinoma (EAC), we explored immune landscape patterns in the tumor microenvironment before and after neoadjuvant chemoradiation (nCRT). Sections from matched pretreatment biopsies and post-nCRT resection specimens (n = 188) were stained for (1) programmed death-ligand 1 (PD-L1, CD274); (2) programmed cell death protein 1 (PD-1, CD279), forkhead box P3 (FOXP3), CD8, pan-cytokeratin multiplex; and (3) an MHC class I, II duplex. The densities of tumor-associated immune cells (TAICs) were calculated using digital image analyses and correlated to histopathological nCRT response [tumor regression grade (TRG)], survival, and post-nCRT immune patterns. PD-L1 positivity defined by a combined positive score of >1 was associated with a better response post-nCRT (TRG 1–3 versus 4, 5, p = 0.010). In addition, high combined mean densities of CD8 +, FOXP3 +, and PD-1 + TAICs in the tumor epithelium and stroma of biopsies were associated with a better response (TRG 1–3 versus 4, 5, p = 0.025 and p = 0.044, respectively). Heterogeneous TAIC density patterns were observed post-nCRT, with significantly higher CD8 + and PD-1 + TAIC mean densities compared with biopsies (both p = 0.000). Three immune landscape patterns were defined post-nCRT: ‘inflamed’, ‘invasive margin’, and ‘desert’, of which ‘inflamed’ was the most frequent (57%). Compared with matched biopsies, resection specimens with ‘inflamed’ tumors showed a significantly higher increase in CD8 + density compared with non-inflamed tumors post-nCRT (p = 0.000). In this cohort of EAC patients, higher TAIC densities in pretreatment biopsies were associated with response to nCRT. This warrants future research into the potential of the tumor-immune landscape for patient stratification and novel (immune) therapeutic strategies.

Importance: The optimal staging for gastric cancer remains a matter of debate. Objective: To evaluate the value of 18F-fludeoxyglucose-positron emission tomography with computed tomography (FDG-PET/CT) and staging laparoscopy (SL) in addition to initial staging by means of gastroscopy and CT in patients with locally advanced gastric cancer. Design, Setting, and Participants: This multicenter prospective, observational cohort study included 394 patients with locally advanced, clinically curable gastric adenocarcinoma (=cT3 and/or N+, M0 category based on CT) between August 1, 2017, and February 1, 2020. Exposures: All patients underwent an FDG-PET/CT and/or SL in addition to initial staging. Main Outcomes and Measures: The primary outcome was the number of patients in whom the intent of treatment changed based on the results of these 2 investigations. Secondary outcomes included diagnostic performance, number of incidental findings on FDG-PET/CT, morbidity and mortality after SL, and diagnostic delay. Results: Of the 394 patients included, 256 (65%) were men and mean (SD) age was 67.6 (10.7) years. A total of 382 patients underwent FDG-PET/CT and 357 underwent SL. Treatment intent changed from curative to palliative in 65 patients (16%) based on the additional FDG-PET/CT and SL findings. FDG-PET/CT detected distant metastases in 12 patients (3%), and SL detected peritoneal or locally nonresectable disease in 73 patients (19%), with an overlap of 7 patients (2%). FDG-PET/CT had a sensitivity of 33% (95% CI, 17%-53%) and specificity of 97% (95% CI, 94%-99%) in detecting distant metastases. Secondary findings on FDG/PET were found in 83 of 382 patients (22%), which led to additional examinations in 65 of 394 patients (16%). Staging laparoscopy resulted in a complication requiring reintervention in 3 patients (0.8%) without postoperative mortality. The mean (SD) diagnostic delay was 19 (14) days. Conclusions and Relevance: This study's findings suggest an apparently limited additional value of FDG-PET/CT; however, SL added considerably to the staging process of locally advanced gastric cancer by detection of peritoneal and nonresectable disease. Therefore, it may be useful to include SL in guidelines for staging advanced gastric cancer, but not FDG-PET/CT..

Liquid biopsies are providing new opportunities for detection of residual disease in cell-free DNA (cfDNA) after surgery but may be confounded through identification of alterations arising from clonal hematopoiesis. Here, we identify circulating tumor-derived DNA (ctDNA) alterations through ultrasensitive targeted sequencing analyses of matched cfDNA and white blood cells from the same patient. We apply this approach to analyze samples from patients in the CRITICS trial, a phase III randomized controlled study of perioperative treatment in patients with operable gastric cancer. After filtering alterations from matched white blood cells, the presence of ctDNA predicts recurrence when analyzed within nine weeks after preoperative treatment and after surgery in patients eligible for multimodal treatment. These analyses provide a facile method for distinguishing ctDNA from other cfDNA alterations and highlight the utility of ctDNA as a predictive biomarker of patient outcome to perioperative cancer therapy and surgical resection in patients with gastric cancer., Identifying patients that will respond to a particular therapy remains a key challenge in precision oncology. Here, in gastric cancer, the authors show that circulating tumour DNA can predict recurrence, provided that the signal from white blood cells is filtered out.

Background The occurrence of a venous thromboembolism (VTE) is common in patients with cancer. Gastric cancer has been associated with one of the highest risks for VTE. Chemotherapy, especially cisplatin has been associated with a high VTE risk. In this study, risk factors for VTE occurrence and their potential impact on subsequent therapeutic interventions were investigated in patients who underwent preoperative chemotherapy, in the CRITICS gastric cancer trial. Patients and methods Patients with resectable gastric cancer were preoperatively treated with three cycles of 3‐weekly epirubicin, cisplatin or oxaliplatin, and capecitabine (ECC/EOC). VTE was defined as any thrombus in the venous system, excluding superficial and/or device related VTEs. Potential risk factors were analyzed in a multivariable regression model with age, gender, Body Mass Index (BMI), tumor localization, Lauren classification, type of chemotherapy (ECC/EOC), (cardiovascular) comorbidity, and previous VTE as independent risk factors. The impact of VTE on completion rate of preoperative chemotherapy, surgical resection rate, postoperative complications, and start of postoperative therapy were investigated. Results Of 781 patients, 78 (10%) of 781 patients developed a VTE during preoperative chemotherapy. On multivariable analysis, BMI ≥ 30 kg/m2 and previous VTE were associated with VTE occurrence (reference BMI, High BMI and previous VTE were independent risk factors for VTE occurrence during preoperative chemotherapy in patients with resectable gastric cancer. Comparable proportions of patients proceeded to surgery and started postoperative therapy, suggesting that VTE occurrence during preoperative treatment did not affect receipt of further treatment.

Cytoreductive Surgery and Hyperthermic Intraperitoneal Chemotherapy (CRS-HIPEC) may be curative for colorectal cancer patients with peritoneal metastases (PMs) but it has a high rate of morbidity. Accurate preoperative patient selection is therefore imperative, but is constrained by the limitations of current imaging techniques. In this pilot study, we explored the feasibility of circulating tumor (ct) DNA analysis to select patients for CRS-HIPEC. Thirty patients eligible for CRS-HIPEC provided blood samples preoperatively and during follow-up if the procedure was completed. Targeted Next-Generation Sequencing (NGS) of DNA from PMs was used to identify bespoke mutations that were subsequently tested in corresponding plasma cell-free (cf) DNA samples using droplet digital (dd) PCR. CtDNA was detected preoperatively in cfDNA samples from 33% of patients and was associated with a reduced disease-free survival (DFS) after CRS-HIPEC (median 6.0 months vs median not reached, p = 0.016). This association could indicate the presence of undiagnosed systemic metastases or an increased metastatic potential of the tumors. We demonstrate the feasibility of ctDNA to serve as a preoperative marker of recurrence in patients with PMs of colorectal cancer using a highly sensitive technique. A more appropriate treatment for patients with preoperative ctDNA detection may be systemic chemotherapy in addition to, or instead of, CRS-HIPEC.

Aim: To evaluate treatment-related toxicity, treatment compliance, surgical complications and event-free survival (EFS) in older (≥70 years) versus younger (

Background Minimally invasive diagnostic biomarkers for patients with pancreatic ductal adenocarcinoma (PDAC) and distal cholangiocarcinoma (dCCA) are warranted to facilitate accurate diagnosis. This study identified diagnostic plasma proteins based on proteomics of tumor secretome. Materials and Methods Secretome of tumor and normal tissue was collected after resection of PDAC and dCCA. Differentially expressed proteins were measured by mass spectrometry. Selected candidate biomarkers and carbohydrate antigen 19‐9 (CA19‐9) were validated by enzyme‐linked immunosorbent assay in plasma from patients with PDAC (n = 82), dCCA (n = 29), benign disease (BD; n = 30), and healthy donors (HDs; n = 50). Areas under the curve (AUCs) of receiver operator characteristic curves were calculated to determine the discriminative power. Results In tumor secretome, 696 discriminatory proteins were identified, including 21 candidate biomarkers. Thrombospondin‐2 (THBS2) emerged as promising biomarker. Abundance of THBS2 in plasma from patients with cancer was significantly higher compared to HDs (p < .001, AUC = 0.844). Combined expression of THBS2 and CA19‐9 yielded the optimal discriminatory capacity (AUC = 0.952), similarly for early‐ and late‐stage disease (AUC = 0.971 and AUC = 0.911). Remarkably, this combination demonstrated a power similar to CA19‐9 to discriminate cancer from BD (AUC = 0.764), and THBS2 provided an additive value in patients with high expression levels of bilirubin. Conclusion Our proteome approach identified a promising set of candidate biomarkers. The combined plasma expression of THBS2/CA19‐9 is able to accurately distinguish patients with PDAC or dCCA from HD and BD. Implications for Practice The combined plasma expression of thrombospondin‐2 and carbohydrate antigen 19‐9 is able to accurately diagnose patients with pancreatic cancer and distal cholangiocarcinoma. This will facilitate minimally invasive diagnosis for these patients by distinguishing them from healthy individuals and benign diseases., This article identifies diagnostic plasma proteins to distinguish patients with pancreatic ductal adenocarcinoma and distal cholangiocarcinoma from benign disease and health donors and evaluates these new markers for additive value with CA19‐9 at different disease stages.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by a relative paucity of cancer cells that are surrounded by an abundance of nontumor cells and extracellular matrix, known as stroma. The interaction between stroma and cancer cells contributes to poor outcome, but how proteins from these individual compartments drive aggressive tumor behavior is not known. Here, we report the proteomic analysis of laser-capture microdissected (LCM) PDAC samples. We isolated stroma, tumor, and bulk samples from a cohort with long- and short-term survivors. Compartment-specific proteins were measured by mass spectrometry, yielding what we believe to be the largest PDAC proteome landscape to date. These analyses revealed that, in bulk analysis, tumor-derived proteins were typically masked and that LCM was required to reveal biology and prognostic markers. We validated tumor CALB2 and stromal COL11A1 expression as compartment-specific prognostic markers. We identified and functionally addressed the contributions of the tumor cell receptor EPHA2 to tumor cell viability and motility, underscoring the value of compartment-specific protein analysis in PDAC.

Identification of predictive biomarkers for targeted therapies requires information on drug exposure at the target site as well as its effect on the signaling context of a tumor. To obtain more insight in the clinical mechanism of action of protein kinase inhibitors (PKIs), we studied tumor drug concentrations of protein kinase inhibitors (PKIs) and their effect on the tyrosine-(pTyr)-phosphoproteome in patients with advanced cancer. Tumor biopsies were obtained from 31 patients with advanced cancer before and after 2 weeks of treatment with sorafenib (SOR), erlotinib (ERL), dasatinib (DAS), vemurafenib (VEM), sunitinib (SUN) or everolimus (EVE). Tumor concentrations were determined by LC-MS/MS. pTyr-phosphoproteomics was performed by pTyr-immunoprecipitation followed by LC-MS/MS. Median tumor concentrations were 2&ndash, 10 &micro, M for SOR, ERL, DAS, SUN, EVE and &gt, 1 mM for VEM. These were 2&ndash, 178 &times, higher than median plasma concentrations. Unsupervised hierarchical clustering of pTyr-phosphopeptide intensities revealed patient-specific clustering of pre- and on-treatment profiles. Drug-specific alterations of peptide phosphorylation was demonstrated by marginal overlap of robustly up- and downregulated phosphopeptides. These findings demonstrate that tumor drug concentrations are higher than anticipated and result in drug specific alterations of the phosphoproteome. Further development of phosphoproteomics-based personalized medicine is warranted.

Clinical significance of the pT4 category in colon cancer is increasing with several therapeutic implications. The aim of this study was to evaluate variability in diagnosing pT4a colon cancer. Twelve pathologists classified 66 preselected scanned Hematoxylin/Eosin-stained slides with tumor cells at a distance of 25–1500 μm (n = 22), 0–25 μm (n = 22), or on (n = 22) the peritoneal surface. Inter- and intraobserver variability were calculated using Kappa statistics. For interlaboratory variability, pathology reports of pT3 and pT4a colon cancer were extracted from the Dutch Pathology Registry between 2012 and 2015. The proportion of pT4a (pT4a/(pT3+pT4a)) was compared between 33 laboratories. Potential risk of understaging was assessed by determining the average number of blocks taken from pT3 and pT4a N0-2M0 tumors with metachronous peritoneal metastasis. Interobserver variability among 12 pathologists was 0.50 (95%CI 0.41–0.60; moderate agreement). Intraobserver variability (8 pathologists) was 0.71 (substantial agreement). A total of 7745 reports with pT3 or pT4aN0-2M0 colon cancer from 33 laboratories were included for interlaboratory analysis. Median percentage of pT4a was 15.5% (range 3.2–24.6%). After adjustment for case mix, 8 labs diagnosed pT4a significantly less or more frequently than the median lab. Metachronous peritoneal metastases were histologically verified in 170 of 6629 pT3 and in 129 of 1116 pT4a tumors, with a mean number of blocks of 4.03(SD 1.51) and 4.78 (SD 1.76) taken from the primary tumors, respectively (p

Background: Perioperative chemotherapy plus surgery is one recommended standard treatment for patients with resectable gastric and esophageal cancer. Even with a multimodality treatment more than half of patients will relapse following surgical resection. Patients who have a poor response to neoadjuvant chemotherapy and have an incomplete (R1) resection or have metastatic lymph nodes in the resection specimen (N+) are especially at risk of recurrence. Current clinical practice is to continue with the same chemotherapy in the adjuvant setting as before surgery. In the phase II randomized EORTC VESTIGE trial (NCT03443856), patients with high risk resected gastric or esophageal adenocarcinoma will be randomized to either adjuvant chemotherapy (as before surgery) or to immunotherapy with nivolumab and low dose ipilimumab (nivolumab 3 mg/kg IV Q2W plus Ipilimumab 1 mg/kg IV Q6W for 1 year). The primary endpoint of the study is disease free survival, with secondary endpoints of overall survival, safety and toxicity, and quality of life. This is an open label randomized controlled multi-center phase-2 superiority trial. Patients will be randomized in a 1:1 ratio to study arms. The trial will recruit 240 patients; recruitment commenced July 2019 and is anticipated to take 30 months. Detailed inclusion/exclusion criteria, toxicity management guidelines, and statistical plans for EORTC VESTIGE are described in the manuscript. Clinical Trial Registration: The trial is registered with www.ClinicalTrials.gov, identifier: NCT03443856.

BACKGROUND: Patients with longstanding inflammatory bowel disease (IBD; ie, ulcerative colitis and Crohn's disease) have an increased risk of colorectal cancer (CRC). Due to ongoing inflammation, IBD-associated dysplastic lesions can develop. These lesions have an increased risk to progress to cancer compared with sporadic adenomas, which are also found in these patients. Differentiating between these 2 types of dysplasia remains challenging, both clinically and histologically, while treatment strategies may differ. Therefore, the aim of this study was to investigate molecular alterations associated with colorectal dysplasia to cancer progression in IBD and evaluate to what extent these alterations differ from sporadic adenomas. METHODS: DNA copy number aberrations and mutation analyses of 48 genes were performed by next-generation sequencing in 43 IBD-associated dysplastic lesions, 30 of which were dysplastic and 13 of which were cancers. Results were compared with existing DNA copy number and mutation data from 118 sporadic adenomas and 24 sporadic cancers. RESULTS: Inflammatory bowel disease-associated dysplastic lesions harbor patterns of DNA copy number aberrations comparable to carcinomas, which are rare in sporadic adenomas. TP53 mutation was the most frequent mutation observed in IBD-associated dysplastic lesions and in cancers. FBXW7 was mutated significantly more often in IBD-associated dysplastic lesions than in sporadic adenomas. CONCLUSIONS: Inflammatory bowel disease-associated dysplastic lesions show more DNA copy number aberrations than sporadic adenomas. TP53 and FBXW7 mutations appear to be involved in the development of IBD-associated dysplastic lesions and cancer. These findings indicate that IBD-associated dysplastic lesions are more genomically unstable, possibly reflecting a faster progression toward cancer.

Chemotherapy combined with the angiogenesis inhibitor bevacizumab (BVZ) is approved as a first-line treatment in metastatic colorectal cancer (mCRC). Limited clinical benefit underpins the need for improved understanding of resistance mechanisms and the elucidation of novel predictive biomarkers. We assessed germline single-nucleotide polymorphisms (SNPs) in 180 mCRC patients (Angiopredict [APD] cohort) treated with combined BVZ + chemotherapy and investigated previously reported predictive SNPs. We further employed a machine learning approach to identify novel associations. In the APD cohort IL8 rs4073 any A carriers, compared to TT carriers, were associated with worse progression-free survival (PFS) (HR = 1.51, 95% CI:1.03–2.22, p-value = 0.037) and TBK1 rs7486100 TT carriers, compared to any A carriers, were associated with worse PFS in KRAS wild-type (wt) patients (HR = 1.94, 95% CI:1.04–3.61, p-value = 0.037), replicating previous findings. Machine learning identified novel associations in genes encoding the inflammasome protein NLRP1 and the ER protein Sarcalumenin (SRL). A negative association between PFS and carriers of any A at NLRP1 rs12150220 and AA for SRL rs13334970 in APD KRAS wild-type patients (HR = 4.44, 95% CI:1.23–16.13, p-value = 0.005), which validated in two independent clinical cohorts involving BVZ, MAVERICC and TRIBE. Our findings highlight a key role for inflammation and ER signalling underpinning BVZ + chemotherapy responsiveness.

Contains fulltext : 229811.pdf (Publisher’s version ) (Open Access) Gastric cancer is the fifth most common cancer worldwide and has a high mortality rate. In the last decades, treatment strategy has shifted from an exclusive surgical approach to a multidisciplinary strategy. Treatment options for patients with resectable gastric cancer as recommended by different worldwide guidelines, include perioperative chemotherapy, pre- or postoperative chemoradiotherapy and postoperative chemotherapy. Although gastric cancer is a heterogeneous disease with respect to patient-, tumor-, and molecular characteristics, the current standard of care is still according to a one-size-fits-all approach. In this review, we discuss the background of the different treatment strategies in resectable gastric cancer including the current standard, the specific role of radiotherapy, and describe the current areas of research and potential strategies for personalization of therapy.

Consensus molecular subtyping is an RNA expression-based classification system for colorectal cancer (CRC). Genomic alterations accumulate during CRC pathogenesis, including the premalignant adenoma stage, leading to changes in RNA expression. Only a minority of adenomas progress to malignancies, a transition that is associated with specific DNA copy number aberrations or microsatellite instability (MSI). We aimed to investigate whether colorectal adenomas can already be stratified into consensus molecular subtype (CMS) classes, and whether specific CMS classes are related to the presence of specific DNA copy number aberrations associated with progression to malignancy. RNA sequencing was performed on 62 adenomas and 59 CRCs. MSI status was determined with polymerase chain reaction-based methodology. DNA copy number was assessed by low-coverage DNA sequencing (n = 30) or array-comparative genomic hybridisation (n = 32). Adenomas were classified into CMS classes together with CRCs from the study cohort and from The Cancer Genome Atlas (n = 556), by use of the established CMS classifier. As a result, 54 of 62 (87%) adenomas were classified according to the CMS. The CMS3 ‘metabolic subtype’, which was least common among CRCs, was most prevalent among adenomas (n = 45; 73%). One of the two adenomas showing MSI was classified as CMS1 (2%), the ‘MSI immune’ subtype. Eight adenomas (13%) were classified as the ‘canonical’ CMS2. No adenomas were classified as the ‘mesenchymal’ CMS4, consistent with the fact that adenomas lack invasion-associated stroma. The distribution of the CMS classes among adenomas was confirmed in an independent series. CMS3 was enriched with adenomas at low risk of progressing to CRC, whereas relatively more high-risk adenomas were observed in CMS2. We conclude that adenomas can be stratified into the CMS classes. Considering that CMS1 and CMS2 expression signatures may mark adenomas at increased risk of progression, the distribution of the CMS classes among adenomas is consistent with the proportion of adenomas expected to progress to CRC.

BACKGROUND: First line chemotherapy is effective in 75 to 80% of patients with metastatic colorectal cancer (mCRC). We studied whether microRNA (miR) expression profiles can predict treatment outcome for first line fluoropyrimidine containing systemic therapy in patients with mCRC.METHODS: MiR expression levels were determined by next generation sequencing from snap frozen tumor samples of 88 patients with mCRC. Predictive miRs were selected with penalized logistic regression and posterior forward selection. The prediction co-efficients of the miRs were re-estimated and validated by real-time quantitative PCR in an independent cohort of 81 patients with mCRC.RESULTS: Expression levels of miR-17-5p, miR-20a-5p, miR-30a-5p, miR-92a-3p, miR-92b-3p and miR-98-5p in combination with age, tumor differentiation, adjuvant therapy and type of systemic treatment, were predictive for clinical benefit in the training cohort with an AUC of 0.78. In the validation cohort the addition of the six miR signature to the four clinicopathological factors demonstrated a significant increased AUC for predicting treatment response versus those with stable disease (SD) from 0.79 to 0.90. The increase for predicting treatment response versus progressive disease (PD) and for patients with SD versus those with PD was not significant. in the validation cohort. MiR-17-5p, miR-20a-5p and miR-92a-3p were significantly upregulated in patients with treatment response in both the training and validation cohorts.CONCLUSION: A six miR expression signature was identified that predicted treatment response to fluoropyrimidine containing first line systemic treatment in patients with mCRC when combined with four clinicopathological factors. Independent validation demonstrated added predictive value of this miR-signature for predicting treatment response versus SD. However, added predicted value for separating patients with PD could not be validated. The clinical relevance of the identified miRs for predicting treatment response has to be further explored.

BACKGROUND: Duodenal adenocarcinoma (DA) is a rare tumor for which survival data per treatment modality and disease stage are unclear. This systematic review and meta-analysis aims to summarize the current literature on patient outcome after surgical, (neo)adjuvant, and palliative treatment in patients with DA.METHODS: A systematic search was performed according to the preferred reporting items for systematic reviews and meta-analyses guidelines, to 25 April 2017. Primary outcome was overall survival (OS), specified for treatment strategy or disease stage. Random-effects models were used for the calculation of pooled odds ratios per treatment modality. Included papers were also screened for prognostic factors.RESULTS: A total of 26 observational studies, comprising 6438 patients with DA, were included. Of these, resection with curative intent was performed in 71% (range 53-100%) of patients, and 29% received palliative treatment (range 0-61%). The pooled 5-year OS rate was 46% after curative resection, compared with 1% in palliative-treated patients (OR 0.04, 95% confidence interval [CI] 0.02-0.09, p CONCLUSION: Resection with curative intent, either pancreaticoduodenectomy or segmental resection, and lack of nodal metastases, favors survival for DA. Further studies exploring multimodality (neo)adjuvant therapy are warranted to investigate their benefit.

4033 Background: ctDNA is becoming an established marker to assess tumor burden, relapse after surgery, and to identify responders in immunotherapy studies. In the phase II PERFECT trial rEAC patients were treated with neoadjuvant chemoradiotherapy (nCRT) and a PD-L1 inhibitor (van den Ende et al. CCR. 2021). Here we evaluated the potential of cell-free DNA (cfDNA) to predict pathological complete response (pCR) and recurrence. Methods: The cohort consisted of 40 patients and 145 plasma samples. EDTA blood samples were drawn at baseline (B, N = 40), in week 5 of nCRT (W5, N = 40), before surgery (OR, N = 33) and 3 months after surgery (FU, N = 32). cfDNA was isolated by affinity columns (CNAkit, QIAgen) quantified by spectrofluorometer (BioAnalyzer, Agilent), sequencing libraries were prepared for lcWGS ( < 5-fold coverage, Tag-seq, Takara) and sequenced on a NovaSeq (S4, PE150). Sequencing data were processed with an in-house pipeline. Copy number aberrations (CNA) and the tumor fraction were estimated using the ichorCNA tool. Insert sizes were recovered and we determined a Tumor Enriched Fragment Fraction (TEFF), calculated by doing the ratio of fragments between 90-150 bp and 250-320 bp (enriched in tumor signal) and fragments between 150-250 bp and 320-360 bp (poor in tumor signal). ichorCNA and TEFF were used to quantify the ctDNA fraction in plasma samples. pCR was defined as ypT0N0. Residual tumor, progression or death before surgery were considered non-pCR. Relapse-free survival (RFS) was defined as the time after surgery until recurrence. Results: The pCR rate was 25% (10/40). The median fold change TEFF between B and W5 was -0.15 (range -0.67 to 0.44) in the pCR group and 0.16 (range -1.40 to 0.76) in the non-pCR group (Mann–Whitney U; p = 0.047). Of the 17 patients in whom ctDNA was detected (TEFF≥0.3 and/or ichorCNA≥0.03) in the FU sample, 13 (76%) showed a recurrence. Of the 15 patients with no ctDNA detected 5 (33%) showed a recurrence. Patients with ctDNA detected at FU had worse RFS, HR = 2.72 (95%CI 0.96-7.71; p = 0.050). Recurrences were detected earlier by FU ctDNA than by imaging due to physical complaints with a median of 312 days (163-798 days). Conclusions: lcWGS appears to be a useful tool to predict pCR and recurrence in resectable esophageal cancer. These lcWGS results will be further combined with fragmentomics analysis and targeted mutational data (Ion Torrent next-generation sequencing) in order to assess response to immunotherapy. Clinical trial information: NCT03087864.

TPS4156 Background: Gastroesophageal adenocarcinoma (GEA) patients with metastatic lymph nodes (ypN+) or a microscopically incomplete surgical resection (R1) following neoadjuvant chemotherapy are at high risk of disease recurrence. Current practice is to continue with the same perioperative chemotherapy used prior to surgery, despite these suboptimal outcomes. Adjuvant immunotherapy with nivolumab has shown efficacy in poor risk GEA patients following chemoradiotherapy and surgery in the CheckMate 577 trial, and nivolumab and ipilimumab have demonstrated activity in advanced GEA. We hypothesise that high risk (ypN+ and/or R1) post resection GEA patients who are treated with nivolumab and ipilimumab will have better disease free survival than patients who continue with standard post-operative chemotherapy. Methods: VESTIGE is an ongoing, international, open label randomized phase II study designed to evaluate the efficacy of adjuvant nivolumab plus ipilimumab versus standard post-operative chemotherapy in high risk (ypN+ and/or R1) post resection GEA patients. Eligible patients (n=240) will be randomised 1:1 to receive post-operative adjuvant chemotherapy (identical regimen as pre-operatively) or nivolumab 3mg/kg IV q2w plus ipilimumab 1mg/kg IV q6w x 1 year. Key inclusion criteria include ypN+ and/or R1 status following neoadjuvant chemotherapy plus surgery and an adequate pre-specified surgical resection. The primary endpoint of the study is disease free survival, with secondary endpoints of overall survival, safety, toxicity and quality of life. Since start of recruitment in August 2019, the trial has recruited 85 of 240 patients at 18 sites in the Czech Republic, France, Germany, Israel, Italy, Norway, Poland, Spain, and United Kingdom. The VESTIGE translational research programme includes collection of pre-treatment biopsies, post-chemotherapy resection specimens and serial liquid biopsy on treatment to explore biomarkers predictive of immune checkpoint blockade efficacy. Clinical trial information: NCT03443856.

4053 Background: Peritoneal carcinomatosis (PC) is accountable for over 50% of metastatic spread in gastric cancer (GC). Little is known about factors contributing to the risk of metachronous PC as a single site of metastases after preoperative chemotherapy and potentially curative resection. An accurate prediction model of risk factors identifying high risk populations may pave the way for new treatment strategies, such as intraperitoneal chemotherapy. In this analysis, risk factors for the development of isolated metachronous PC after preoperative chemotherapy and surgical resection were investigated. Methods: In the CRITICS trial, 788 patients with resectable GC were randomized for preoperative chemotherapy and gastrectomy followed by either chemotherapy or chemoradiotherapy. Patients who underwent a potentially curative resection without peritoneal metastases at time of surgery were included in this analysis. Univariate and multivariate analyses were performed using a competing risk model. Results: In total, 617 patients met the inclusion criteria. Overall, 97 of 617 (16%) patients developed metachronous PC. The peritoneum was the first site of recurrence in 64 of 617 (10%) patients. Diffuse or mixed type GC, ypT4 stage and ypN3 stage or a lymph node ratio (LNR) with >20% tumor positivity were independent predictors of isolated PC in both univariate and multivariate analyses with hazard ratios (HR) of 2.90, 2.63 and 2.31, respectively (Table). ypT4 and ypN3 stage or a LNR >20% were also independent predictors of distant recurrence or death with HR of 1.46 and 2.34, respectively. Patients in whom all predictors were present had the highest 2 year cumulative incidences of both isolated PC development and other events of 39.6% and 48.8%, respectively. Conclusions: Diffuse or mixed tumor type, ypT4 and ypN3 or a LNR >20% were identified as independent risk factors for metachronous PC as a single site of metastases in a large cohort of patients treated with preoperative chemotherapy followed by surgical resection. The combination of these factors might identify a subgroup that could benefit from preventive treatment strategies.[Table: see text]

4024 Background: Carcinoembryonic antigen (CEA) and carbohydrate antigen (CA) 19-9 are well-known tumor markers. Most studies on CEA and CA 19-9 in pts with GC were performed in Asia, and/or in the metastatic setting. The aim of this study was to investigate the prognostic value of blood derived laboratory parameters in a cohort of European pts with resectable non-metastatic GC. Methods: In the CRITICS trial, 788 pts with resectable GC underwent perioperative therapy (preoperative chemotherapy plus either postoperative chemotherapy or postoperative chemoradiotherapy). Blood levels of CEA, CA 19-9, alkaline phosphatase (AP), creatinine, neutrophils, hemoglobin (Hb) and lactate dehydrogenase (LDH) were determined prior to treatment. Data for these variables were available for at least 89% of the pts. Factors significant on univariable cox regression analysis were further explored in multivariable analysis. Probabilities to undergo potentially curative surgery was investigated for factors significant on multivariable analysis. The relationship between tumor markers and the presence of circulating tumor DNA (ctDNA) was explored in 50 pts with available ctDNA data. Results: CEA and CA 19-9 were identified as independent prognostic factors for survival (Table). Probabilities to undergo potentially curative surgery were 86%, 77% and 60% for pts with no elevated tumor makers versus those with elevated CEA or elevated CA 19-9 versus those with both tumor markers elevated, respectively (p

Background: Patients with pathologic limited or no response (pNR) to neoadjuvant chemoradiation (nCRT) are subjected to curative intended esophagectomy with subsequent perioperative morbidity and mortality, but potentially only harm from nCRT. The primary aim of this study was to compare the overall survival (OS) of patients with pNR and patients who underwent primary esophagectomy to evaluate potentially benefits of nCRT in these patients. The secondary aim was to identify predictive clinicopathologic factors for pNR and pathologic complete response (pCR) to nCRT with the goal to preselect these patients before the start of treatment.Methods: From the period 2005 to 2016, 206 esophageal cancer (EC) patients treated with Carboplatin/Paclitaxel and radiotherapy with complementary esophagectomy were included in this cohort. OS of patients with pNR was compared with a historical cohort of primary surgically treated patients (n=218) after a propensity score matching resulting in a group of 68 patients with pNR after nCRT versus a group of 68 primary esophagectomy patients.Results: The OS in the pNR group and the primary esophagectomy group was comparable (P=0.986). No predictive factors were found in this cohort for pNR. Female gender (OR 2.5, 95% CI 1.2-5.3) and squamous cell carcinoma (SCC) (OR 2.6, 95% CI 1.3-5.3) were identified as independent predictive factors for pCR.Conclusions: Patients with a pNR do not benefit from nCRT followed by resection. These patients had a similar OS as those who had a primary esophagectomy alone. Although this indicates that nCRT does not negatively impact the OS of patients with pNR, patients still have an increased morbidity because of nCRT. Hence, it is important to identify factors that predict pNR. The ability to predict pNR (and pCR) will enable tailored and personalized care preventing unnecessary nCRT with increased morbidity.

Mass spectrometry-based phosphoproteomics of cancer cell and tissue lysates provides insight in aberrantly activated signaling pathways and potential drug targets. For improved understanding of individual patient's tumor biology and to allow selection of tyrosine kinase inhibitors in individual patients, phosphoproteomics of small clinical samples should be feasible and reproducible. We aimed to scale down a pTyr-phosphopeptide enrichment protocol to biopsy-level protein input and assess reproducibility and applicability to tumor needle biopsies. To this end, phosphopeptide immunoprecipitation using anti-phosphotyrosine beads was performed using 10, 5 and 1 mg protein input from lysates of colorectal cancer (CRC) cell line HCT116. Multiple needle biopsies from 7 human CRC resection specimens were analyzed at the 1 mg-level. The total number of phosphopeptides captured and detected by LC-MS/MS ranged from 681 at 10 mg input to 471 at 1 mg HCT116 protein. ID-reproducibility ranged from 60.5% at 10 mg to 43.9% at 1 mg. Per 1 mg-level biopsy sample, > 200 phosphopeptides were identified with 57% ID-reproducibility between paired tumor biopsies. Unsupervised analysis clustered biopsies from individual patients together and revealed known and potential therapeutic targets. Significance This study demonstrates the feasibility of label-free pTyr-phosphoproteomics at the tumor biopsy level based on reproducible analyses using 1 mg of protein input. The considerable number of identified phosphopeptides at this level is attributed to an effective down-scaled immuno-affinity protocol as well as to the application of ID propagation in the data processing and analysis steps. Unsupervised cluster analysis reveals patient-specific profiles. Together, these findings pave the way for clinical trials in which pTyr-phosphoproteomics will be performed on pre- and on-treatment biopsies. Such studies will improve our understanding of individual tumor biology and may enable future pTyr-phosphoproteomics-based personalized medicine.