Detection of Kras Gene in Colorectal Cancer Patients through Liquid Biopsy: A Cost-effective Method
Hafiz Syed, Shamim Mushtaq, Saeeda Baig
Abstract
OBJECTIVE: To detect the Kras gene through liquid biopsy, a less invasive technique in diagnosed colorectal cancer patients. STUDY DESIGN: Cross-sectional study. PLACE AND DURATION OF STUDY: Department of Oncology, Dr. Ziauddin Hospital and Bait-us-Sukoon Cancer Hospital, Karachi, from 2019 to 2020. METHODOLOGY: Circulating tumor DNA (ctDNA) in colorectal cancer patients was extracted through magnetic bead technique using MagMAX cell free DNA kit (Thermofisher, Uk). The frequency of Kras gene was quantified using a real-time polymerase chain reaction (RT-PCR) assay (qPCR). ANOVA and Chi-square tests were utilised for statistical analysis. RESULTS: Mean threshold cycle (CT) of Kras gene showed significantly higher expression 15.6 ± 1.82 (p=0.001) in stage IV CRC cases compared to early stages (19.53 ± 18.223.7 ± 2.9 and 19.8 ± 2.69 of stage 1, 2 and 3, respectively. Similarly, ΔCT mean of Kras gene at stage IV showed significantly higher expression of 2.48 ± 1.40 (0.048), compared to 2.39 ± 0.6, 3.12 ± 0.68 and 3.15 ± 0.41 of stage 1, 2 and 3, respectively. Males (n=40, 55%) showed significant association (p=0.001) with CRC compared to females (n=33, 45%). Categorisation of tumor types within different age groups revealed that colon cancer was more frequent (n=11, 15.1%) in the 41-50 age group, while rectal cancer was more frequent (n= 11, 15.1%) in the 41-50 age group, while rectal cancer was more in the 51-60 age group (n=11, 15.1%). CONCLUSION: Kras gene was detected with significantly increased levels in plasma of CRC patients at advanced stages. This confirms that liquid biopsy can be used to detect Kras gene in ctDNA of CRC patients through a magnetic bead based technique. Key Words: Liquid biopsy, Circulating tumor DNA, KRAS, Colorectal cancer, Real-time polymerase chain reaction.