Background It remains unclear whether extended lymphadenectomy provides oncological advantages in colorectal cancer. This multicentre RCT aimed to address this issue. Methods Patients with resectable primary colonic cancer were enrolled in four hospitals registered in the COLD trial, and randomized to D2 or D3 dissection in a 1 : 1 ratio. Data were analysed to assess the safety of D3 dissection. Results The study included the first 100 patients randomized in this ongoing trial. Ninety‐nine patients were included in the intention‐to‐treat (ITT) analysis (43 D2, 56 D3). Ninety‐two patients received the allocated treatment and were included in the per‐protocol (PP) analysis: 39 of 43 in the D2 group and 53 of 56 in the D3 group. There were no deaths. The 30‐day postoperative morbidity rate was 47 per cent in the D2 group and 48 per cent in the D3 group, with a risk ratio of 1·04 (95 per cent c.i. 0·68 to 1·58) (P = 0·867). There were two anastomotic leaks (5 per cent) in the D2 group and none in the D3 group. Postoperative recovery, complication and readmission rates did not differ between the groups in ITT and PP analyses. Mean lymph node yield was 26·6 and 27·8 in D2 and D3 procedures respectively. Good quality of complete mesocolic excision was more frequently noted in the D3 group (P = 0·048). Three patients in the D3 group (5 per cent) had metastases in D3 lymph nodes. D3 was never the only affected level of lymph nodes. N‐positive status was more common in the D3 group (46 per cent versus 26 per cent in D2), with a risk ratio of 1·81 (95 per cent c.i. 1·01 to 3·24) (P = 0·044). Conclusion D3 lymph node dissection is feasible and may be associated with better N staging. Registration number: NCT03009227 ( http://www.clinicaltrials.gov).

e15081 Background: Colorectal cancer (CRC) is a common pathology in the world; the annual incidence rate reaches 1 million cases, and the annual mortality rate exceeds 500,000. The target therapy of EGFR with monoclonal drugs improves the survival of patients with this pathology. However, the effectiveness of therapy depends on the presence of mutations in genes involved in the EGFR signaling cascade, in particular KRAS. The purpose of this study was to determine the frequency of somatic mutations in the KRAS gene in tumor samples of patients from the South Russia diagnosed with colorectal cancer, as well as to analyze the effect of point mutations G12A, G12C, G12D, G12R, G12S, G12V and G13D on tumor metastasis. Methods: 744 patients diagnosed with colorectal adenocarcinoma were examined for mutations in 12th and 13th codons of KRAS. Somatic mutations in the KRAS gene were detected with reagent kit Biolink (Russia) using FFPE DNA samples of tumor tissues. Statistical analysis was performed using the method of binary logistic multiple regression and χ2-test. Results: Mutations in the KRAS gene were found in 32% of patients in the pooled sample; the most common mutation was the replacement of G12D in the KRAS gene (36%). Less common detectable mutations in the KRAS gene were G13D (18%), G12V (16%), G12A (10%), G12C (8%), G12S (10%), G12R (2%). Metastatic CRC was found in 86% of carriers of the wild-type allele and in 88% of carriers of the mutant allele. The presence of a G12V substitution almost 6 times increased the risk of metastasis in patients with CRC (OR = 5.89; 95% CI: 0.8-43.45; p = 0.049). Conclusions: Mutations in the KRAS gene were found in 32% of patients with CRC in the South of Russia. A significant association of G12V replacement with a high risk of metastasis was established. Further study of somatic mutations in the KRAS gene will allow to choose the most effective therapeutic strategies in the presence of one or another of the studied mutations.

Лучевая терапия является одним из методов лечения аденокарциномы толстой кишки. Большое влияние на эффективность подобной терапии оказывает радиорезистентность опухолевых клеток, зависящая от их молекулярно-генетических особенностей, к которым относится показатель копийности генов (CNV). Целью работы явилось исследование влияния копийности генов, ответственных за репарации ДНК, регуляцию клеточного цикла и апоптоза, на радиорезистентность клеток линии HT-29 в условиях воздействия стандартных доз лучевой терапии 5 и 7 Гр на протяжении 5 дней. Культивирование клеток HT-29 проходило в стерильных флаконах в среде RPMI-1640 c 10%-ной фетальной телячьей сывороткой и 50 мкг/мл гентамицина. Облучение проводили на линейном ускорителе Novalis TX. Определение относительной копийности 32 генетических локусов (AKT, ATM, BRIP, BRCA1, BRCA2, CDK1, CDKN1B, CCND1 и др.) проводили методом Real-Time qPCR. На разных уровнях кластеризации выявлено сходство по уровню копийности некоторых генов в интактных и облученных образцах. Получены данные о дифференциальном показателе копийности 5 генов в интактных и облученных при 5 и 7 Гр клетках HT-29. В клетках НТ-29, подвергнутых облучению 7 Гр, статистически значимо была повышена копийность генов BRCA2, H2AX, CASP9 и RBBP8: в 2,5 раза; 3,2 раза; 1,6 раза и 1,6 раза соответственно (р<0,05), и снижена копийность гена BCL2 в 3,6 раза (р<0,05) относительно интактных клеток. В клетках НТ-29, подвергнутых облучению 5 Гр, статистически значимо была повышена копийность CASP9 и RBBP8-в 1,5 и 1,4 раза соответственно (р<0,05). Таким образом, проведенное исследование позволило установить, что линия клеток НТ-29 исходно гетерогенна по копийности некоторых генов и лучевая терапия приводит к селективному выживанию клеток, обладающих повышенной копийностью генов BRCA2, H2AX, RBBP8 CASP9 и сниженной копийностью гена BCL2. Ключевые слова: лучевая терапия, аденокацинома толстой кишки, культура клеток, апоптоз, репарация ДНК, радиорезистентность, копийность генов.

e15508 Background: The most important stages of metastatic cascade are extravasation and invasion of malignant cells and their surviving in blood. The vast majority of circulating tumor cells (CTC) is destroyed by immune cells. The role of immune system which is able to play not only antitumor but also prooncogenic role is manifested both on local and systemic levels. We studied correlations between lymphocyte subsets` content in blood and in tumor of colorectal cancer patients (II, III, IV stages) with and without CTC. Methods: 60 colorectal cancer patients with II (n = 20), III (n = 20) and IV (n = 20) stages underwent surgery without previous chemotherapy. CTC were measured in blood by CellSearchSystem™ (JanssenDiagnostics, LLC), cell-mediated immunity was assessed in blood by flow cytometry (BD Canto II) and in tissue after surgery by immunohistochemistry; some markers of proliferation and epithelial-mesenchimal transition (EMT) in tumor cells (Ki-67, E- and N-cadherins) were also studied. Criteria of CTC-positive and CTC-negative samples were > 3 and ≤3 respectively. Correlative analysis was performed between the data of the patients with and without CTC in each stage, r was counted. Results: In CTC-positive patients with all the stages the number of strong and moderate correlations between system immunologic factors involving CD8+ appeared to be fewer than in CTC-negative ones (7 vs 19). On the contrary, the number of correlations with T regs in CTC-positive was increased: 5 vs 3 in CTC-negative patients. In patients with CTC > 3 fewer correlations were noted between factors of local and systemic immunity than in CTC-negative ones (9 vs 4 in II stage) and total disruption of all the correlations between system immunologic factors and proliferating tumor cells in III and IV stages was established. Some pathologic correlations appeared in CTC-positive patients like moderate direct one between activated T-lymphocytes` amount and Ki-67+ tumor cells. The number of correlations between intratumoral lymphocytes and tumor cells expressing proliferation and EMT markers in CTC-positive patients was decreased in comparison with CTC-negative ones (4 vs 10 in II stage, 1 vs 9 in III, 2 vs 9 in IV stage). Conclusions: The presence of CTC in colorectal cancer patients rather than tumor stage is associated with imbalance of their systemic and local immunologic factors. This provides some evidence that disruption of interactions in the immune system is at least partly due to CTC.

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