VOLUME 12 NUMBER 2 2019 ISSN 2218-7979 eISSN 2409-370X International Journal of Biology and Chemistry Al-Farabi Kazakh National University International Journal of Biology and Chemistry is published twice a year by al-Farabi Kazakh National University, al-Farabi ave., 71, 050040, Almaty, Republic of Kazakhstan website: http://ijbch.kaznu.kz/ Any inquiry for subscriptions should be sent to: Prof. Mukhambetkali Burkitbayev, al-Farabi Kazakh National University al-Farabi ave., 71, 050040, Almaty, Republic of Kazakhstan e-mail: [email protected] © 2019 al-Farabi Kazakh National University Int. j. biol. chem. (Online) EDITORIAL The most significant achievements in the field of natural sciences are reached in joint collaboration, where important roles are taken by biology and chemistry. Therefore publi- cation of a Journal, displaying results of current studies in the field of biology and chemis- try, facilitates highlighting of theoretical and practical issues and distribution of scientific discoveries. One of the basic goals of the Journal is to promote the extensive exchange of informa- tion between the scientists from all over the world. We welcome publishing original papers and materials of biological and chemical conferences, held in different countries (after the process of their subsequent selection). Creation of special International Journal of Biology and Chemistry is of great impor- tance, because a great amount of scientists might publish their articles and it will help to widen the geography of future collaboration. We will be glad to publish also the papers of the scientists from the other continents. The Journal aims to publish the results of the experimental and theoretical studies in the field of biology, biotechnology, chemistry and chemical technology. Among the em- phasized subjects are: modern issues of technologies for organic synthesis; scientific basis of the production of physiologically active preparations; modern issues of technologies for processing of raw materials; production of new materials and technologies; study on chemi- cal and physical properties and structure of oil and coal; theoretical and practical issues in processing of hydrocarbons; modern achievements in the field of nanotechnology; results of studies in the fields of biology, biotechnology, genetics, nanotechnology, etc. We hope to receive papers from a number of scientific centers, which are involved in the application of the scientific principles of biological and chemical sciences on practice and carrying out research on the subject, whether it relates to the production of new materials, technologies and ecological issues. International Journal of Biology and Chemistry 12, № 2, 4 (2019) IRSTI 34.23.53 https://doi.org/10.26577/ijbch-2019-v2-1 1*O.K. Khamdiyeva , 2S. Pack , 1Z.M. Biyasheva , 2Z. Abdullaev , 3D.R. Kaidarova , 4 N.P. Kabysheva 1Al-Farabi Kazakh National University, Almaty, Kazakhstan 2The Chromosome Pathology Unit, Laboratory of Pathology, National Cancer Institute, National Institutes of Health in Bethesda, MD, USA 3Kazakh Institute of Oncology and Radiology, Almaty, Kazakhstan 4Kazakh Scientific Center of Quarantine and Zoonotic Diseases, Almaty, Kazakhstan *e-mail: [email protected] Mutation and expression of the C-KIT gene on population of Kazakhstan Abstract: Lung cancer is still a leading cause of death from malignant tumors worldwide. Due to late diagnosis, results of treatment remain unsatisfactory. In Kazakhstan about 3,669 new cases of lung cancer are detected every year, with a 5-year survival rate for 2016 equal to 48.0%. The aim of the current work is to study the expression of C-KIT protein and identify mutations in patients with lung cancer living in Almaty region. Literature data showed that anomalous expression of the corresponding gene and presence of mutations lead to a number of malignant neoplasms. Creation of drugs targeting C-KIT protein, promotes the development of clinical diagnosis and treatment of cancer. Blood samples and biopsy material obtained from patients diagnosed with lung cancer, treated at the Almaty Oncology Center and residing in Almaty region served the objects of the study. Written informed consent was obtained from all patients. Analysis of restriction fragment length polymorphism and immunohistochemical analysis followed the polymerase chain reaction. For immunohistochemical analysis tissue fixed in formalin and embedded in paraffin blocks was used. The presence of mutations in codon 557 was revealed by the genetic analysis of 11 exon of C-KIT gene. RFLP analysis and sequencing showed mutations in the codon. Immunohistochemical analysis revealed overexpression of C-KIT gene in four (9.09%) patients, 14 (31.82%) patients had moderate expression, 24 (54.54%) patients had weak expression and two (4.55%) patients had no expression. In total, 18 (40.91%) patients had a positive response to immunohistochemical analysis. This suggests that the disorders occur in the cells of tumor tissue and are not inherited. In fact, in most cases the occurrence of lung cancer can be associated with smoking. Key words: C-KIT gene, mutation, expression, immunohistochemical analysis. Introduction caused by aggressiveness, invasive and metastatic disease potential, as well as difficulties in detect- Cancer remains the leading cause of death world- ing it in the early stages. Therefore, the search for wide, despite impressive advances in the early diag- pathogenetic approaches in the treatment of cancer nosis and treatment of patients with malignant neo- remains an urgent problem today. The lack of sig- plasms. Incidence rates in Kazakhstan are highest nificant achievements in the fight against cancer is among CIS countries [1]; with 36 813 cases detected largely due to an insufficient level of diagnosis. For in 2016 [2]. Lung cancer is one of the most common the vast majority of people, the cancer is still diag- types of cancer and the leading cause of death from nosed in the late stages of development (73.5%), in cancer worldwide [3]. This type of cancer is the most which the possibilities of modern treatment methods common in the world, and in 2016, it amounted to cannot be fully realized, as patients may have resis- 9.9% of the total number of new cases of cancer in tance to chemotherapy and targeted drugs. Ten years Kazakhstan. In 2016, about 3,635 new cases of lung ago, therapeutic strategies for treating lung cancer cancer were detected in Kazakhstan, with five-year were based on a histological type of classification. survival equal to 48.0%, and 2,498 death cases [2]. However, in recent years, one of the most interesting High mortality from lung cancer is believed to be advances in the treatment of lung cancer has been the © 2019 al-Farabi Kazakh National University Int. j. biol. chem. (Online) O.K. Khamdiyeva et al. 5 understanding of genetic changes in cells. Achieve- The PCR products were analyzed using electro- ments in genomic sequencing and identification of phoresis on 1% agarose gel. The correspondence of molecular markers over the past decade have clearly the obtained fragments was evaluated using the DNA demonstrated that cancer is a heterogeneous disease Ladder Gene Ruler 100 bp marker (Thermo Fisher [4; 5]. Scientific, USA). Resrtriction endonucleases were The studies using some modern methods of mo- selected using WatCut online software for SNP- lecular biology have allowed us to fundamentally RFLP analysis. The PCR products were digested change our ideas about the characteristics of the vari- with restriction endonucleases SsiI and FokI. 5 μL of ous forms of the disease that arise and, consequently, the PCR product and 10U of endonuclease were used the treatment tactics and its predicted course. De- in the reaction carried out at 3˚C for 3 h. Restricted velopments in genotyping have changed the clinical DNA was analyzed on 8% polyacrylamide gel. The practice of treating lung cancer and have shown that polyacrylamide gel was stained with ethidium bro- genetic changes in the EGFR, ALK, ROS1, HER2, mide. DNA Ladder Gene Ruler 100 bp (Thermo KIT and BRAF genes are powerful predictive bio- Fisher Scientific, USA) was used for detection of the markers in the treatment of lung cancer [6-10]. restriction fragments length. In this regard, the purpose of the current research Tissue microarrays. Before creating a Tissue Mi- was to study the expression of C-KIT protein and croarrays (ТМАs) slide, all tumor tissues were clas- identify mutations in patients with lung cancer living sified according to the international classification of in Almaty region. stages of malignant neoplasms – TNM [11]. As a material for the creation of ТМАs slides and Materials and methods immunohistochemical analysis, 44 histological ma- terials obtained from patients with lung cancer fixed Patients. Surgical specimens were obtained dur- in formalin and embedded in paraffin blocks were ing the surgery from patients diagnosed with a lung used. Each selected area of lung tissue was analyzed cancer. All patients underwent a survey, which con- by a qualified pathologist to determine histological tained information on clinical data, gender, age and types of tumors. From selected sites, selected tissue bad habits. Patients signed a voluntary informed was carefully taken with a hollow cylinder (2 mm consent before participating in the study. The study in diameter) and transferred into one common paraf- protocol was approved by the local ethics committee fin block. Then, a 3-4 micron thick section was cut of the medical faculty of the Higher School of Public from this block using a microtome (RM2255, Leica, Health at Al-Farabi Kazakh National University (Al- Wetzlar, Germany) and placed on a glass slide [12] maty, Kazakhstan) – protocol No. IRB – A024 from (Figure 1). September 22, 2017. DNA preparation. Genomic DNA was isolated and purified from formalin-fixed paraffin-embedded tissues using the GeneJet Genomic DNA Purifica- tion Kit (Thermo Fisher Scientific, USA). Purified DNA samples were aliquoted and stored in a freezer at -20˚C. PCR and RLFP analysis. Specific primers (for- ward 5’-GATCTATTTTTCCCTTTCTC-3’ and re- verse 5’-AGCCCCTGTTTCATACTGAC-3’) were designed using the PrimerQuest Tool.