CRISPR-enabled point-of-care genotyping for APOL1 genetic risk assessment
Detecting genetic variants enables risk factor identification, disease screening, and initiation of preventative therapeutics. However, current methods, relying on hybridization or sequencing, are unsuitable for point-of-care settings. In contrast, CRISPR-based-diagnostics offer high sensitivity and specificity for point-of-care applications. While these methods have predominantly been used for pathogen sensing, their utilization for genotyping is limited. Here, we report a multiplexed CRISPR-based genotyping assay using LwaCas13a, PsmCas13b, and LbaCas12a, enabling the simultaneous detection of six genotypes. We applied this assay to identify genetic variants in the APOL1 gene prevalent among African Americans, which are associated with an 8-30-fold increase in the risk of developing kidney disease. Machine learning facilitated robust analysis across a multicenter clinical cohort of more than 100 patients, accurately identifying their genotypes. In addition, we optimized the readout using a multi-analyte lateral-flow assay demonstrating the ability for simplified genotype determination of clinical samples. Our CRISPR-based genotyping assay enables cost-effective point-of-care genetic variant detection due to its simplicity, versatility, and fast readout.
Authors
- Robert Greensmith (ORCID: https://orcid.org/0000-0001-8142-0127)
- Anand S. Dighe (ORCID: https://orcid.org/0000-0003-4130-0758)
- Anja Schütz (ORCID: https://orcid.org/0000-0002-0606-2574)
- Jakob J. Metzger (ORCID: https://orcid.org/0000-0001-6813-4425)
- Bernhard Hemmer (ORCID: https://orcid.org/0000-0001-5985-6784)
- Alexander J. Schubert (ORCID: https://orcid.org/0000-0002-6907-5276)
- Michael M. Kaminski (ORCID: https://orcid.org/0000-0003-0429-7027)
- Isadora T. Lape
- Cristian V. Riella (ORCID: https://orcid.org/0000-0003-3753-9459)
- Leonardo V. Riella (ORCID: https://orcid.org/0000-0002-7636-3196)
- Xiao Tan (ORCID: https://orcid.org/0000-0002-3874-9557)
- Sarah Wendlinger
- Josefine Rau
- Nora Diederich
Institutions
- Broad Institute (US)
- Beth Israel Deaconess Medical Center (US)
- Harvard University (US)
- Max Delbrück Center (DE)
- TUM Klinikum (DE)
- Massachusetts General Hospital (US)
- Munich Cluster for Systems Neurology (DE)
- Berlin Institute of Health at Charité - Universitätsmedizin Berlin (DE)
- Center for Systems Biology (US)
- Technical University of Munich (DE)
- Massachusetts Institute of Technology (US)
- Charité - Universitätsmedizin Berlin (DE)
Publication Details
- Journal
- EMBO Molecular Medicine
- Published
- 2024-09-13
- DOI
- https://doi.org/10.1038/s44321-024-00126-x
- Citations
- 7
- Primary Topic
- Renal Diseases and Glomerulopathies
- Type
- article
- Field-Weighted Citation Impact
- 2.37
Funders
- Deutsche Forschungsgemeinschaft
- Berlin Institute of Health