Research Spotlight: Identification of Small Molecule Inhibitors of Neutral Ceramidase (nCDase) Via Target Based High Throughput Screening

Posted on August 18, 2021

Colon Image

Cancer is a leading cause of death worldwide. According to the CDC, cancer is the second leading cause of death in the United States, claiming the lives of nearly 600,000 Americans in 2019. The types of cancer that lead to death are numerous but the three that cause more deaths than any others are breast cancer, lung cancer, and colorectal cancer. Each of these cancers claimed close to 2 million lives worldwide in 2020. So, it is needless to say that these cancers are the subjects of intense research studies across the globe. Our Lipid Leader for this month, Dr. Yusuf Hannun, and his group of medical researchers at Stony Brook University are one of the groups providing insight into therapeutics for colorectal cancer.

Recently, Dr. Hannun published a research article identifying small molecules as inhibitors of the human neutral ceramidase (nCDase) enzyme. This enzyme is highly expressed in the colonic epithelium and converts ceramide to sphingosine. Sphingosine can then be converted to Sphingosine-1-phosphate (S1P) by sphingosine kinases (SKs). Ceramide and S1P are key molecules in colon cancer development due to their ability to promote cell proliferation, cell survival, and angiogenesis. There is substantial evidence that increased SK1 and S1P levels contribute to colon carcinogenesis. This evidence includes studies that show that 89% of human colon cancer samples demonstrated upregulation of SK1. Also, in human colon cancer tissue, there is a 50% reduction in ceramide levels compared to normal mucosa. Ceramide functions to stimulate apoptosis, induce senescence, and inhibit cell proliferation. Combining this information, sphingolipid metabolism could be a novel target for colon cancer therapy.

Dr. Hannun and his team set out to develop pharmacological inhibitors of nCDase. They used a 1536-well assay format based on the reaction between nCDase, substrate, and sodium periodate. The assay was tested against over 650,000 compounds from the Scripps Drug Discovery Library. The ceramidase inhibitor, C6-urea-ceramide, was used as a pharmacological control for the assay. The initial screening yielded almost 2500 possible hits. Of these, 125 made it through the second round of screening using a 5.64% inhibition cutoff point. Further screenings were used to identify 4 analogs that showed high activity with an IC50 < 10µM. These compounds were expanded via medicinal chemistry approaches to modify the compounds while retaining favorable drug properties. This approach revealed several chemical classes with potency ranges between 1-10 µM and maximal percent inhibitions of 70-90%.

This study could lead to a novel colon cancer therapeutic which could save thousands of lives every year if proven effective. More optimization studies, pharmacokinetic evaluations, and characterization efforts are being undertaken by Dr. Hannun and his research team. We are proud to play a small role in this crucial research effort by providing the C6 urea-ceramide compound used as the inhibition standard during the assays. We also look forward to seeing the results of further studies to identify nCDase inhibitors!

Thank you, Dr. Hannun, for your invaluable contributions to the lipid community! Don’t forget to check out Our Interview with Dr. Hannun, you won’t want to miss it! Read the full research article by clicking HERE!