One of the major issues in clinical oncology is the ability of cancer cells to make resistance to chemotherapy drugs, that this lead to chemotherapy failure. The usage of the CRISPR-Cas9 system tosuppress drug resistance genes in different cancers is a potential therapeutic strategy to enhance chemotherapy outcome . CRISPR-Cas9 genome editing technology have many advantages and it hasformerly initiated to supplant necessary genome editing technologies, such as transcription activator-like effector nucleases (TALENs) and zinc finger nucleases (ZFNs). The goal of current study was topresent an overview of the crispr-cas9 applications in this field. The search in this study was based on the keywords crispr-cas9 and chemotherapy resistance followed by Scopus, PubMed and Web ofScience. In fast increasing number of experiments, Crispr-cas9 system has been used to remove drug resistance or to find the gene accountable for chemotherapy resistance. The development of multidrugresistance (MDR) is one of the major problems in the chemotherapy treatment of osteosarcoma. Multidrug resistance gene 1 (MDR1), which produce the membrane efflux pump P-glycoprotein (P-gp), have an significant role in the drug resistance process. The application of the CRISPR-Cas9 system to target exon 5 of MDR1 in osteosarcoma cell lines efficiently knocked out MDR1, coming back drug resistance. So, this procedure may also be used as a potential therapy for T790 M acquired drug resistant mutations in EGFR-mutant lung cancer. T790M-related drug resistance may result from change of inhibitors binding in the ATP pocket of EGFR and restored binding affinity for ATP. CRISPR-Cas9-mediated genome editing tool has revolutionized the area of gene therapy, which makeswide application potential for therapeutic manipulations of cancer. Although in CRISPR/Cas9 system we faced with problems, including off-target effects, transfer system, safety, and ethical subject, but isbeing practical in biotechnology and clinical work like the studies mentioned above. CRISPR/Cas9- mediated preclinical study and clinical trials should be encouraged and carry out in several cancers andother diseases like neurodegenerative disorders