- Mar 2021
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www.ncbi.nlm.nih.gov www.ncbi.nlm.nih.gov
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analyzed several PALB2 variants in their response to the ICL-inducing agent cisplatin
AssayGeneralClass: BAO:0002805 cell proliferation assay
AssayMaterialUsed: CLO:0037317 mouse embryonic stem cell line
AssayDescription: Stable expression of wild type and variant PALB2 cDNA constructs in Trp53 and Palb2-null mouse cell line containing DR-GFP reporter; exposure to cisplatin for 48 h induces interstrand-crosslink DNA damage; cell survival is measured by FACS 24 h after cisplatin washout
AssayReadOutDescription: Relative resistance to cisplatin represented as cell survival relative to wild type, which was set to 100%
AssayRange: %
AssayNormalRange: Cisplatin resistance levels comparable to that of cells expressing wild type PALB2; no numeric threshold given
AssayAbnormalRange: Cisplatin resistance levels comparable to that of cells expressing empty vector; no numeric threshold given
AssayIndeterminateRange: Not reported
ValidationControlPathogenic: 2
ValidationControlBenign: 2
Replication: 2 independent experiments
StatisticalAnalysisDescription: Not reported
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sensitivity to PARPi treatment using a cellular proliferation assay
AssayGeneralClass: BAO:0002805 cell proliferation assay
AssayMaterialUsed: CLO:0037317 mouse embryonic stem cell line
AssayDescription: Stable expression of wild type and variant PALB2 cDNA constructs in Trp53 and Palb2-null mouse cell line containing DR-GFP reporter; exposure to PARP inhibitor Olaparib for 48 h inhibits end-joining mediated by PARP and sensitizes cells to DNA damage; cell survival is measured by FACS 24 h after Olaparib washout
AssayReadOutDescription: Relative resistance to PARPi represented as cell survival relative to wild type, which was set to 100%
AssayRange: %
AssayNormalRange: PARPi resistance levels comparable to that of cells expressing wild type PALB2; no numeric threshold given
AssayAbnormalRange: PARPi resistance levels ≤30% of wild type
AssayIndeterminateRange: Not reported
ValidationControlPathogenic: 12
ValidationControlBenign: 9
Replication: 2 independent experiments
StatisticalAnalysisDescription: Not reported
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A cell-based functional assay for PALB2 variants
AssayGeneralClass: BAO:0003061 reporter protein
AssayMaterialUsed: CLO:0037317 mouse embryonic stem cell line
AssayDescription: Stable expression of wild type and variant PALB2 cDNA constructs in Trp53 and Palb2-null mouse cell line containing DR-GFP reporter; I-SceI endonuclease introduces a double-stranded break in the reporter construct and efficient repair results in GFP expression, which is detected by flow cytometry
AssayReadOutDescription: Relative homologous recombination (HR) efficiency represented as mean percentages of GFP-positive cells among the mCherry-positive cells relative to wild type, which was set to 100%
AssayRange: %
AssayNormalRange: HR levels comparable to that of cells expressing wild type PALB2; no numeric threshold given
AssayAbnormalRange: HR levels ≤40% of wild type
AssayIndeterminateRange: Not reported
ValidationControlPathogenic: 12
ValidationControlBenign: 9
Replication: 2 independent experiments
StatisticalAnalysisDescription: Not reported
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