Analysis of RAS gene mutations in cytogenetically normal de novo acute myeloid leukemia patients reveals some novel alterations

Afia Muhammad Akram; Asma Chaudhary; Humera Kausar; Fayez Althobaiti; Afshan Syed Abbas; Zawar Hussain; Naz Fatima; Erum Zafar; Wajiha Asif; Umair Afzal; Zoufishan Yousaf; Amjad Zafar; Steve M Harakeh; Samina Qamer

doi:10.1016/j.sjbs.2021.04.089

Analysis of RAS gene mutations in cytogenetically normal de novo acute myeloid leukemia patients reveals some novel alterations

Saudi J Biol Sci. 2021 Jul;28(7):3735-3740. doi: 10.1016/j.sjbs.2021.04.089. Epub 2021 May 6.

Authors

Affiliations

¹ Department of Zoology, Division of Science and Technology, University of Education, Township, Lahore, Pakistan.
² Department of Biotechnology, Kinnaird College for Women, Lahore, Pakistan.
³ Department of Biotechnology, College of Science, Taif University, P.O. Box 11099, Taif 21944, Saudi Arabia.
⁴ Department of Zoology, University of Education, Lower Mall Campus, Lahore, Pakistan.
⁵ Molecular Biology Laboratory, Department of Zoology, University of the Punjab, Lahore, Pakistan.
⁶ Department of Oncology, Mayo Hospital, Anarkali Bazar, Lahore, Pakistan.
⁷ Special Infectious Agents Unit, King Fahd Medical Research Center, King Abdulaziz University, Jeddah 21589, Saudi Arabia.
⁸ Department of Zoology, Government College University, Faisalabad, Pakistan.

Abstract

Rat sarcoma gene (RAS) holds great importance in pathogenesis of acute myeloid leukemia (AML). The activated mutations in Neuroblastoma rat sarcoma viral oncogene homolog (NRAS) and Kirsten rat sarcoma viral oncogene homolog (KRAS) confers proliferative and survival signals, deliberating numerous effects on overall survival and progression free survival in AML patients. In this study thirty one (31) blood samples of adult newly diagnosed AML patients were collected to identify possible incidence of mutations through amplification of KRAS (exon 1 and 2) and NRAS gene (exon 1 and 2) using polymerase chain reaction (PCR). Amplicons were then subjected to sequencing and were analyzed through Geneious Prime 2019. Five of thirty one (16.12%) patients had altered sites in either NRAS or KRAS. The NRAS mutations were observed in three AML patients (N = 3, 9.67%). A novel missense mutation NRAS-I36R (239 T > G) representing a substitution of single nucleotide basepair found in NRAS exon 1 while exon 2 was detected with heterozygous mutation NRAS-E63X (318G > T) and insertion (A), resulting in frameshift of the amino acid sequence and insertion of two nucleotide basepairs (TA) in two of the patients. KRAS mutations (N = 2, 6.45%) were found in exon 1 whereas no mutations in KRAS exon 2 were detected in our patient cohort. Mutation in KRAS Exon 1, KRAS-D30N (280G > A) was observed in two patients and one of them also had a novel heterozygous mutation KRAS-L16N (240G > C). In addition there was no statistically significant association of mutRAS gene of AML patients with several prognostic markers including age, gender, karyotyping, CD34 positivity, cytogenetic abnormalities, total leukocyte count, white blood cell count and French-American-British (FAB) classification. However, the presence of mutRAS gene were strongly associated (p = 0.001) with increased percentage of bone marrow blasts. The prevalence of mutations in correlation with clinical and hematological parameter is useful for risk stratification in AML patients.

Keywords: Acute myeloid leukemia (AML); Kirsten rat sarcoma viral oncogene homolog (KRAS); Neuroblastoma rat sarcoma viral oncogene homolog (NRAS); RAS gene mutations.