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Single-Cell Signaling Network Profiling of Pooled CRISPR Screens by Mass Cytometry

Award Number: R21CA313195
ORGANIZATION: NATIONAL CANCER INSTITUTE
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)
PERIOD OF PERFORMANCE START DATE: 05/07/2026
PERIOD OF PERFORMANCE END DATE: 04/30/2028

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Single-Cell Signaling Network Profiling of Pooled CRISPR Screens by Mass Cytometry - Project Summary/Abstract Deciphering how gene perturbations reshape cellular signaling networks is essential for un- derstanding disease mechanisms and discovering therapeutic targets. However, current pooled CRISPR screening platforms lack the scalability, throughput, and protein-level resolution needed to capture complex cellular responses, especially those governed by post-translational modifications. This collaborative project between Dr. Xiaokang Lun’s lab at the University of Minnesota and Dr. Jellert Gaublomme’s lab at Columbia University seeks to overcome these limitations by developing CRISPRmap-CyTOF, a novel single-cell screening platform that integrates high-complexity pooled CRISPR perturbations with high-dimensional protein-level profiling using mass cytometry. The platform builds on the CRISPRmap combinatorial barcoding strategy (developed by the Gaublomme lab) and a photocrosslinking-based DNA stabilization method (developed by the Lun lab), combining mass cytometry-based signaling network analysis to simultaneously decode thousands of gene perturbations and quantify ∼30 protein or phospho-epitope markers in individual cells. This one-step, high-throughput readout enables proteome-resolved CRISPR screens at single-cell resolution across millions of cells. In Aim 1, we will re-engineer CRISPRmap for compatibility with mass cytometry, optimizing probe chemistry, staining conditions, and barcode decoding algorithms to enable high-fidelity linkage between gene perturbation and signaling phenotypes. In Aim 2, we will apply CRISPRmap-CyTOF to systematically profile early and late signaling responses to a kinome-scale CRISPR knockout library in chronic myeloid leukemia (CML) cells, with and without tyrosine kinase inhibitor (TKI) treatment. These experiments will identify kinases that drive CML progression, uncover novel TKI resistance mechanisms, and generate integrative signaling biomarkers predictive of disease outcome. Together, this work will deliver a transformative technology for functional genomics and a mecha- nistic framework for dissecting therapy resistance in cancer.


Issue Date FY	Funding FY	Legal Entity Name	Legal Entity Address	Legal Entity City	Legal Entity State	Legal Entity Zip Code	Legal Entity COUNTY	Legal Entity COUNTRY	Assistance Listing	Award Code	Budget Year	Action Date	Action Type	Action Amount

Issue Date FY: 2026 ( Subtotal = $406,325 )
2026	2026	REGENTS OF THE UNIVERSITY OF MINNESOTA	2221 UNIVERSITY AVE SE STE 100	MINNEAPOLIS	MN	55414	HENNEPIN	USA	Cancer Biology Research	000	1	5/7/2026	NEW	$406,325
														Subtotal = $406,325

Grand Total All Awards = $406,325

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Single-Cell Signaling Network Profiling of Pooled CRISPR Screens by Mass Cytometry

Award Number: R21CA313195

ORGANIZATION: NATIONAL CANCER INSTITUTE

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

PERIOD OF PERFORMANCE START DATE: 05/07/2026

PERIOD OF PERFORMANCE END DATE: 04/30/2028

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