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Published on: March 30, 2020
Dual-fluorescence isogenic high-content screening for MUC16/CA125 selective agents
Thapi D Rao1, Nestor Rosales, David R Spriggs
1Laboratory of Clinical Pharmacology, Department of Medicine, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA.
Abstract:
Most of the currently used cancer chemotherapies are based on compounds that inhibit general cellular mechanisms, such as DNA replication or tubulin function, and lack specificity in relation to features of the cancer cell. Recent advances in genomic studies have increased our knowledge of tumor cell biology, and a panoply of new targets have been postulated. This has provided an opportunity to develop and validate drugs that specifically target cancer cells through their unique genetic characteristics. Identification of MUC16/CA125 both as a marker and a driver of transformation led us to design a target-based high-content screen to identify and classify compounds that exhibit differential effect on MUC16-expressing cells. We developed a coculture assay in 384-well plate containing isogenic ovarian cancer cells that are positive or negative for the MUC16 protein. High-throughput screening of our small molecule pilot library led to the identification of compounds preferentially cytotoxic to MUC16(+) or MUC16(-) cells, using a Preferential Score analysis. We compared screening results in both A2780 and SK-OV-3 ovarian cancer cells in single and coculture settings. We also identified compounds that were cytotoxic for both types of ovarian cancer cells regardless of the MUC16 status. Compounds that were preferentially targeting MUC16 cells were subsequently confirmed by caspase-induction assays. The isogenic, dual-color fluorescence strategy is an innovative approach that can effectively identify novel drug candidates, selectively targeting cancer cells that have unique molecular properties.
Insights
This study developed a novel screening method to find cancer drugs targeting MUC16-expressing cells. The approach identified compounds selectively killing MUC16-positive ovarian cancer cells, offering new therapeutic strategies.
Area of Science:
- Oncology
- Drug Discovery
- Molecular Biology
Background:
- Current cancer chemotherapies lack specificity, targeting general cellular mechanisms rather than unique cancer cell features.
- Genomic advancements reveal new tumor targets, enabling the development of drugs with specific cancer cell targeting capabilities.
- MUC16/CA125 is identified as a marker and driver of transformation, presenting a target for selective drug development.
Purpose of the Study:
- To design and validate a target-based high-content screen to identify compounds differentially affecting MUC16-expressing cells.
- To discover novel drug candidates with selective cytotoxicity against MUC16-positive or MUC16-negative ovarian cancer cells.
- To evaluate an innovative isogenic, dual-color fluorescence strategy for identifying targeted cancer therapeutics.
Main Methods:
- A coculture assay using isogenic ovarian cancer cells (MUC16-positive and MUC16-negative) in a 384-well plate format.
- High-throughput screening of a small molecule library using Preferential Score analysis to identify differentially cytotoxic compounds.
- Confirmation of MUC16-selective compounds through caspase-induction assays and comparison of results in single and coculture settings.
Main Results:
- Identification of compounds preferentially cytotoxic to MUC16(+) or MUC16(-) ovarian cancer cells.
- Discovery of compounds effective against both MUC16-positive and MUC16-negative ovarian cancer cells.
- Validation of MUC16-targeting compounds using caspase-induction assays, confirming selective cytotoxicity.
Conclusions:
- The developed isogenic, dual-color fluorescence screening strategy is an effective method for identifying novel drug candidates.
- This approach enables the selective targeting of cancer cells based on unique molecular properties, such as MUC16 expression.
- The findings pave the way for developing more specific and effective cancer therapies, particularly for ovarian cancer.
