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CRISPR-Cas9 Screen Reveals New Drug Combination for Treating Liver Cancer

Using a combinatorial CRISPR-Cas9 screening platform, researchers have demonstrated how this novel two-drug regime can be used in combination with sorafenib to curb tumour cell growth.

Accounting for more than 700,000 deaths each year, liver cancer is the sixth most common cancer and the third leading cause of cancer-related deaths worldwide. Since 1980, the incidence rates of liver cancer have more than tripled and death rates have more than doubled. While treatments such as molecularly targeted therapies have shown promising results, tumours are often susceptible to developing drug resistance which diminishes the effects of even the most effective drugs including sorafenib, the mainstay FDA-approved first-line drug for hepatocellular carcinoma (HCC) treatment. Evidently, there is a need to expand treatment options available for patients.

Exploring the potential of drug combination, a team of researchers from the LKS Faculty of Medicine at the University of Hong Kong (HKUMed) has successfully repurposed ifenprodil, an approved vasodilator, to be used in combination with sorafenib. Compared to standalone treatments, drug combinations can better reduce the risk of drug resistance and tumour relapse.

“Successful drug repurposing saves the cost and time that otherwise would be needed for developing new therapeutic agents with uncertain efficacy and safety profile. It also increases the chance of clinical translation of the findings from bench to bedside as the identified drug combination could be readily tested in future trials for treating HCC. Our work has identified a potentially useful drug combination to be further tested for the treatment of HCC patients from approved drugs, which could possibly save or prolong patients’ life,” said Dr. Stephanie Ma, Associate Professor of the School of Biomedical Sciences, HKUMed, who co-led the study.

To discover and develop drugs, scientists normally use CRISPR-Cas9 to knockout genes and identify druggable targets because the platform is not only simple but also precise. In the study, the HKUMed researchers initiated dual-genetic knockouts in a pool of cells linked with DNA barcodes that specify the types of genetic alterations they carry. They then used their CombiGEM-CRISPR v2.0 screening platform to produce multiplexable gene knockouts and characterised the survival of these cancer cells.

Unlike conventional drug screening arrays that demand scientists to handle numerous independent multi-wells, the CombiGEM-CRISPR v2.0 screening platform only requires a simple experimental setup. This is because the number of DNA barcodes carried by a large population of cells grown in the same culture dish could be counted in high volume with the help of high-throughput sequencing technologies. By screening genes and their combinations from which hits can be translated directly into drug combinations using existing drugs, non-conventional drugs and drug combinations could be discovered and repurposed for treating cancers.

Focusing on druggable targets with upregulated expressions in HCC cancer stem cells, their combinatorial screen revealed two combinations that harbour a common target – NMDAR1. Its paired targets are FLT4 and FGFR3, which are kinases that can be inhibited by sorafenib. Most notably, the scientists discovered that the genetic ablation of the identified gene combinations successfully inhibited the growth and self-renewal ability of HCC cells. To establish the clinical significance of NMDAR1 in HCC, the researchers used The Cancer Genome Atlas database and found that HCC patients with a low level of NMDAR1 expression have better survival outcomes.

In addition, the research group demonstrated the improved inhibition effect of the drug combination and uncovered its underlying mechanisms. When ifenprodil and sorafenib were administrated together, cell growth and stemness were significantly reduced in several HCC cell lines, patient-derived organoids, and tumour xenograft models. The scientists attributed these magnified effects to the upregulation of unfolded protein response, which triggers the arrest of the cell cycle, and the downregulation of genes linked to WNT-signalling and stemness.

Having successfully demonstrated the cancer-suppressing potential of combining this two-drug regimen with sorafenib, their findings and new CombiGEM-CRISPR v2.0 screening platform are expected to accelerate the discovery and development of new liver cancer treatments and broaden options for patients.

“The application of the CombiGEM-CRISPR v2.0 platform has broadened our scope in search for effective combinations of actionable targets and approved/repurposed drugs for HCC in a rapid and simple manner, and could be extended to other cancers and diseases,” added Dr. Alan Wong Siu-lun, Assistant Professor of the School of Biomedical Sciences, HKUMed, who co-led the research. [APBN]

Source: Xu et al. (2021). A Combinatorial CRISPR–Cas9 Screen Identifies Ifenprodil as an Adjunct to Sorafenib for Liver Cancer Treatment. Cancer Research, 81(24).