Laura N. Kellman, Poornima H. Neela, Suhas Srinivasan, Zurab Siprashvili, Ronald L. Shanderson et al.
Single-nucleotide variants (SNVs) in regulatory DNA are linked to inherited cancer risk. Massively parallel reporter assays of 4,041 SNVs linked to 13 neoplasms comprising >90% of human malignancies were performed in pertinent primary human cell types and then integrated with matching chromatin accessibility, DNA looping and expression quantitative trait loci data to nominate 380 potentially regulatory SNVs and their putative target genes. The latter highlighted specific protein networks in lifetime cancer risk, including mitochondrial translation, DNA damage repair and Rho GTPase activity. A CRISPR knockout screen demonstrated that a subset of germline putative risk genes also enables the growth of established cancers. Editing one SNV, rs10411210, showed that its risk allele increases rhophilin RHPN2 expression and stimulus-responsive RhoA activation, indicating that individual SNVs may upregulate cancer-linked pathways. These functional data are a resource for variant prioritization efforts and further interrogation of the mechanisms underlying inherited risk for cancer.
Laura N. Kellman, Poornima H. Neela, Suhas Srinivasan, Zurab Siprashvili, Ronald L. Shanderson, Audrey W. Hong, Deepti Rao, Douglas F. Porter, David L. Reynolds, Robin M. Meyers, Margaret G. Guo, Xue Yang, Yang Zhao, Glenn G. Wozniak, Laura K. H. Donohue, Rajani Shenoy, Lisa A. Ko, Duy T. Nguyen, Smarajit Mondal, Omar S. Garcia, Lara E. Elcavage, Ibtihal Elfaki, Nathan S. Abell, Shiying Tao, Christopher M. Lopez, Stephen B. Montgomery, Paul A. Khavari. Functional analysis of cancer-associated germline risk variants. 2025-02-17. https://doi.org/10.1038/s41588-024-02070-5
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Primary prostate epithelial cells (ATCC). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Primary colonic epithelial cells (Cell Biologics). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Primary bronchial/tracheal epithelial cells (ATCC). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in 293T Lenti-X (Takara). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Human primary thyroid epithelial cells (Cell Biologics). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Human primary pancreatic epithelial cells (Cell Biologics). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Normal human endometrial epithelial cells (Lifeline). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Neonatal foreskin derived keratinocytes. Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Normal human astrocytes (Lonza). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Neonatal foreskin derived melanocytes. Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Human primary esophageal epithelial cells (Cell Biologics). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Human primary ovarian epithelial cells (Cell Biologics). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Human primary proximal tubular epithelial cells (Cell Biologics). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Integrated lentiMPRA of 140-bp hg19 SNV-centered genomic fragments with matched reference and alternate alleles, ten 20-bp barcodes per allele, and a minimal-promoter luciferase reporter in Human mammary epithelial cells (Lonza). Three biological replicates were sequenced as RNA barcode counts and normalized to two plasmid DNA library replicates.
Original supplemental and deposited inputs retained for this study. Download files individually or together as a ZIP; nested folders are preserved. Source reuse terms apply, and sequencing reads may be omitted.
Download all 6 files (ZIP)41588_2024_2070_MOESM1_ESM.pdf41588_2024_2070_MOESM3_ESM.xlsxGSE183301_cancer_mpra_barcode_reference.fa.gzGSE183301_cancer_mpra_counts.tsv.gzGSE183301_family.soft.gzGSE183301_oligo_name_to_snp_id_key.tsv.gz