CRISPR and KRAS: a match yet to be made

Abstract

CRISPR (clustered regularly interspaced short palindromic repeats) systems are one of the most fascinating tools of the current era in molecular biotechnology. With the ease that they provide in genome editing, CRISPR systems generate broad opportunities for targeting mutations. Specifically in recent years, disease-causing mutations targeted by the CRISPR systems have been of main research interest; particularly for those diseases where there is no current cure, including cancer. KRAS mutations remain untargetable in cancer. Mutations in this oncogene are main drivers in common cancers, including lung, colorectal and pancreatic cancers, which are severe causes of public health burden and mortality worldwide, with no cure at hand. CRISPR systems provide an opportunity for targeting cancer causing mutations. In this review, we highlight the work published on CRISPR applications targeting KRAS mutations directly, as well as CRISPR applications targeting mutations in KRAS-related molecules. In specific, we focus on lung, colorectal and pancreatic cancers. To date, the limited literature on CRISPR applications targeting KRAS, reflect promising results. Namely, direct targeting of mutant KRAS variants using various CRISPR systems resulted in significant decrease in cell viability and proliferation in vitro, as well as tumor growth inhibition in vivo. In addition, the effect of mutant KRAS knockdown, via CRISPR, has been observed to exert regulatory effects on the downstream molecules including PI3K, ERK, Akt, Stat3, and c-myc. Molecules in the KRAS pathway have been subjected to CRISPR applications more often than KRAS itself. The aim of using CRISPR systems in these studies was mainly to analyze the therapeutic potential of possible downstream and upstream effectors of KRAS, as well as to discover further potential molecules. Although there have been molecules identified to have such potential in treatment of KRAS-driven cancers, a substantial amount of effort is still needed to establish treatment strategies based on these discoveries. We conclude that, at this point in time, despite being such a powerful directed genome editing tool, CRISPR remains to be underutilized for targeting KRAS mutations in cancer. Efforts channelled in this direction, might pave the way in solving the long-standing challenge of targeting the KRAS mutations in cancers.

CasRx-Based Kras G12D Silencing for In Vivo Tumor Inhibition in Spontaneous Lung Adenocarcinoma

“…These versatile toolkits have facilitated the management of oncogenic mutations associated with cancer. Three distinct CRISPRdependent approaches have emerged for inhibiting these variants: 1) precisely targeting and disrupting the oncogenic mutation genomes and transcripts using DNAediting and RNA-editing tools, such as the CRISPR/Cas9 and CRISPR/Cas13a 10,13,22 .…”

“…CRISPR/Cas technology has witnessed significant advancements in recent years as a powerful gene editing tool for targeted therapies in cancer treatment [10][11][12] . In lung cancer, CRISPR/Cas9 was widely and thoroughly documented as an editing tool to directly target Kras mutations and indirectly inhibit Kras-related molecules, such as KEAP-1, NF1, and EGFR et.al, leading to tumor regression 10 . Additionally, CRISPR/Cas13a has demonstrated the ability to suppress the progression of NSCLC cells by depredating the mutated Kras transcript 13 .…”

“…Despite significant strides in understanding the anti-tumor effects of CRISPR-based gene therapy, its translation into clinical settings remains challenging due to the intrinsic limitations and our superficial cognition. The obstacles include the unavailable of sophisticated in vivo delivery vectors for large biologically active proteins like Cas9 and Cas13a, the potential safety concerns with DNA editing, and most importantly, the current proof of concept studies are heavy dependent on cell lines and patient-derived xenograft (PDX) models that inadequately represent human tumorigenesis and the natural in vivo environment 10 . To address these issues, we employed the CRISPR/CasRx systems 16 , which could be delivered by the clinical approved AAV virus, to selectively cleave oncogenic KRAS G12D transcript in a spontaneous lung adenocarcinoma model, allowing for comprehensively and faithfully understanding the therapeutic outcome, the underlying mechanism, and the potential off-targets of the CasRx-mediated in vivo therapy against the NSCLC.…”

Nucleic Acid-Based Approaches to Tackle KRAS Mutant Cancers

“…CRISPR-Cas-mediated dsDNA breaks often trigger apoptosis and can result in the emergence of inactivating mutations in TP53. Unintentional editing of a genome can lead to profound long-term complications [67][68][69][70]. Pre-existing anti-Cas antibodies in humans may further diminish editing efficiency [71].…”

Mechanism and Potential Therapy in Ameloblastoma: Akt Signaling Pathway

“…(64) Studies using CRISPR has been shown to regulate downstream molecules such as PI3K, ERK, Akt, Stat3, and c-myc, and some studies have been conducted using CRISPR to target PI3K in pancreatic cancer. (65) Although CRISPR has been shown a great potential and used to edit genes associated with a variety of diseases, its clinical application in cancer is still in infancy, and there are currently no studies using CRISPR to treat ameloblastoma cases.…”