103 Matching Annotations
  1. Mar 2021
  2. www.ncbi.nlm.nih.gov www.ncbi.nlm.nih.gov
    Functional characterization of 84 PALB2 variants of uncertain significance
    10
    1. shannon_mcnulty 18 Mar 2021
      Cells reconstituted with WT-PALB2 showed substantially less sensitivity to olaparib than cells expressing p.A1025R and p.I944N (Fig. 4a). Similar results were observed for cisplatin treatment, although the difference in sensitivity was less pronounced (Fig. 4b). p.L24S, p.L1070P, and p.L35P were also associated with greater sensitivity to olaparib (Fig. 4c) and cisplatin (Fig. 4d) than WT-PALB2.

      AssayResult: 0.01 µM: 100; 0.1 µM: 65; 0.8 µM: 18; 1 µM: 15

      AssayResultAssertion: Abnormal

      Approximation: Exact cisplatin concentrations and assay result values not reported; values estimated from Figures 4b and 4d.

      CGType:FunctionalAssayResult FuncAssay:4 Variant:4 ClinVarID:657328 CAID:CA279502031
    2. shannon_mcnulty 18 Mar 2021
      Cells reconstituted with WT-PALB2 showed substantially less sensitivity to olaparib than cells expressing p.A1025R and p.I944N (Fig. 4a). Similar results were observed for cisplatin treatment, although the difference in sensitivity was less pronounced (Fig. 4b). p.L24S, p.L1070P, and p.L35P were also associated with greater sensitivity to olaparib (Fig. 4c) and cisplatin (Fig. 4d) than WT-PALB2.

      AssayResult: 0.01 µM: 80; 0.1 µM: 52; 0.8 µM: 18; 1 µM: 5

      AssayResultAssertion: Abnormal

      Approximation: Exact cisplatin concentrations and assay result values not reported; values estimated from Figures 4b and 4d.

      CGType:FunctionalAssayResult FuncAssay:4 Variant:56 ClinVarID:216752 CAID:CA339433
    3. shannon_mcnulty 18 Mar 2021
      Cells reconstituted with WT-PALB2 showed substantially less sensitivity to olaparib than cells expressing p.A1025R and p.I944N (Fig. 4a). Similar results were observed for cisplatin treatment, although the difference in sensitivity was less pronounced (Fig. 4b). p.L24S, p.L1070P, and p.L35P were also associated with greater sensitivity to olaparib (Fig. 4c) and cisplatin (Fig. 4d) than WT-PALB2.

      AssayResult: 0.01 µM: 102; 0.1 µM: 65; 0.8 µM: 18; 1 µM: 10

      AssayResultAssertion: Abnormal

      Approximation: Exact cisplatin concentrations and assay result values not reported; values estimated from Figures 4b and 4d.

      CGType:FunctionalAssayResult FuncAssay:4 Variant:85 ClinVarID:230588 CAID:CA10580054
    4. shannon_mcnulty 18 Mar 2021
      Cells reconstituted with WT-PALB2 showed substantially less sensitivity to olaparib than cells expressing p.A1025R and p.I944N (Fig. 4a). Similar results were observed for cisplatin treatment, although the difference in sensitivity was less pronounced (Fig. 4b). p.L24S, p.L1070P, and p.L35P were also associated with greater sensitivity to olaparib (Fig. 4c) and cisplatin (Fig. 4d) than WT-PALB2.

      AssayResult: 0.01 µM: 85; 0.1 µM: 40; 0.8 µM: 20; 1 µM: 13

      AssayResultAssertion: Abnormal

      Approximation: Exact cisplatin concentrations and assay result values not reported; values estimated from Figures 4b and 4d.

      CGType:FunctionalAssayResult FuncAssay:4 Variant:42 ClinVarID:187262 CAID:CA197176
    5. shannon_mcnulty 18 Mar 2021
      Cells reconstituted with WT-PALB2 showed substantially less sensitivity to olaparib than cells expressing p.A1025R and p.I944N (Fig. 4a). Similar results were observed for cisplatin treatment, although the difference in sensitivity was less pronounced (Fig. 4b). p.L24S, p.L1070P, and p.L35P were also associated with greater sensitivity to olaparib (Fig. 4c) and cisplatin (Fig. 4d) than WT-PALB2.

      AssayResult: 0.01 µM: 90; 0.1 µM: 60; 0.8 µM: 15; 1 µM: 15

      AssayResultAssertion: Abnormal

      Approximation: Exact cisplatin concentrations and assay result values not reported; values estimated from Figures 4b and 4d.

      CGType:FunctionalAssayResult FuncAssay:4 Variant:92 CAID:CA923726356
    6. shannon_mcnulty 18 Mar 2021
      Cells reconstituted with WT-PALB2 showed substantially less sensitivity to olaparib than cells expressing p.A1025R and p.I944N (Fig. 4a). Similar results were observed for cisplatin treatment, although the difference in sensitivity was less pronounced (Fig. 4b). p.L24S, p.L1070P, and p.L35P were also associated with greater sensitivity to olaparib (Fig. 4c) and cisplatin (Fig. 4d) than WT-PALB2.

      AssayResult: 0.01 µM: 65; 0.08 µM: 50; 0.8 µM: 30; 8 µM: 20

      AssayResultAssertion: Abnormal

      Approximation: Exact Olaparib concentrations and assay result values not reported; values estimated from Figures 4a and 4c.

      CGType:FunctionalAssayResult FuncAssay:3 Variant:4 ClinVarID:657328 CAID:CA279502031
    7. shannon_mcnulty 18 Mar 2021
      Cells reconstituted with WT-PALB2 showed substantially less sensitivity to olaparib than cells expressing p.A1025R and p.I944N (Fig. 4a). Similar results were observed for cisplatin treatment, although the difference in sensitivity was less pronounced (Fig. 4b). p.L24S, p.L1070P, and p.L35P were also associated with greater sensitivity to olaparib (Fig. 4c) and cisplatin (Fig. 4d) than WT-PALB2.

      AssayResult: 0.01 µM: 102; 0.1 µM: 85; 0.8 µM: 55; 1 µM: 25

      AssayResultAssertion: Normal

      ControlType: Normal; wild type PALB2 cDNA

      Approximation: Exact cisplatin concentrations and assay result values not reported; values estimated from Figures 4b and 4d.

      CGType:FunctionalAssayResult FuncAssay:4 ValidationControl:WildType AssayControl:Normal
    8. shannon_mcnulty 18 Mar 2021
      Viability assayPALB2 variants were introduced into B400 cells using mCherry-pOZC expression vector and flow cytometry for Cherry-red was performed to select for cells expressing PALB2. Sorted cells were plated in 96-well plates and exposed to increasing amounts of Olaparib or cisplatin and incubated for a period of 5 days. Presto Blue (Invitrogen) was added and incubated for 1–2 hours before measuring fluorescence intensity on a Cytation 3 microplate reader (BioTek).

      AssayGeneralClass: BAO:0003009 cell viability assay

      AssayMaterialUsed: CLO:0036938 tumor-derived cell line

      AssayDescription: Transient expression of wild type and variant mCherry-tagged PALB2 cDNA constructs in Trp53 and Palb2-null mouse cell line; exposure to increasing concentrations of cisplatin for 5 days induces interstrand-crosslink DNA damage; cell survival is determined by measuring fluorescence intensity after staining with a cell viability reagent.

      AssayReadOutDescription: Percent cell survival after treatment with cisplatin

      AssayRange: %

      AssayNormalRange: Cisplatin resistance levels comparable to that of cells expressing wild type PALB2; no numeric threshold given

      AssayAbnormalRange: Not reported

      AssayIndeterminateRange: Not reported

      ValidationControlPathogenic: 0

      ValidationControlBenign: 0

      Replication: Not reported

      StatisticalAnalysisDescription: Not reported

      CGType:FunctionalAssayResult FuncAssay:4 BAO:0003009 CLO:0036938 MONDO:0016419 HGNC:26144 AssayRangeType:Quantitative UO:0000187
    10. shannon_mcnulty 18 Mar 2021
      Results for individual PALB2 variants were normalized relative to WT-PALB2 and the p.Tyr551ter (p.Y551X) truncating variant on a 1:5 scale with the fold change in GFP-positive cells for WT set at 5.0 and fold change GFP-positive cells for p.Y551X set at 1.0. The p.L24S (c.71T>C), p.L35P (c.104T>C), p.I944N (c.2831T>A), and p.L1070P (c.3209T>C) variants and all protein-truncating frame-shift and deletion variants tested were deficient in HDR activity, with normalized fold change <2.0 (approximately 40% activity) (Fig. 1a).

      AssayResult: 0.8

      AssayResultAssertion: Abnormal

      StandardErrorMean: 0.14

      CGType:FunctionalAssayResult CG:BulkAnnotation FuncAssay:1 Variant:4 CAID:CA279502031 ClinVarID:657328
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    ncbi.nlm.nih.gov/pmc/articles/PMC7056643/
  3. www.ncbi.nlm.nih.gov www.ncbi.nlm.nih.gov
    A global functional analysis of missense mutations reveals two major hotspots in the PALB2 tumor suppressor
    3
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    ncbi.nlm.nih.gov/pmc/articles/PMC6847799/
  4. www.nature.com www.nature.com
    Functional analysis of genetic variants in the high-risk breast cancer susceptibility gene PALB2
    10
    7. shannon_mcnulty 18 Mar 2021
      To further assess the impact of the 5 selected VUS on PALB2, we examined whether they affected the accumulation of RAD51 at IR-induced DSBs by measuring the formation RAD51 foci.

      AssayGeneralClass: BAO:0000450 fluorescence microscopy

      AssayMaterialUsed: CLO:0003684 HeLa cell

      AssayDescription: Transient expression of wild type and variant PALB2 cDNA constructs in HeLa cells following PALB2 siRNA knockdown; exposure ionizing radiation induces DNA damage; RAD51 foci formation is measured by immunofluorescence microscopy 4 h after irradiation

      AssayReadOutDescription: Number of RAD51 foci per S-phase cell (determined by cyclin A detection)

      AssayRange: foci/cell

      AssayNormalRange: RAD51 foci numbers comparable to that of cells expressing wild type PALB2; no numeric threshold given

      AssayAbnormalRange: RAD51 foci numbers comparable to that of cells expressing empty vector; no numeric threshold given

      AssayIndeterminateRange: Not reported

      ValidationControlPathogenic: 0

      ValidationControlBenign: 0

      Replication: 3 independent experiments

      StatisticalAnalysisDescription: Not reported

      CGType:FunctionalAssay FuncAssay:4 BAO:0000450 CLO:0003684 MONDO:0016419 HGNC:26144 AssayRangeType:Quantitative
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    nature.com/articles/s41467-019-13194-2
  5. www.cell.com www.cell.com
    High-Throughput Reclassification of SCN5A Variants
    1
    1. shannon_mcnulty 18 Mar 2021
      Most Suspected Brugada Syndrome Variants Had (Partial) Loss of Function

      AssayResult: 0.1

      AssayResultAssertion: Abnormal

      ReplicateCount: 19

      StandardErrorMean: 0.1

      Comment: This variant had loss of function of peak current (<10% of wildtype), therefore it was considered abnormal (in vitro features consistent with Brugada Syndrome Type 1. (Personal communication: A. Glazer)

      CGType:FunctionalAssayResult CG:BulkAnnotation FuncAssay:1 Variant:4 CAID:CA014257 ClinVarID:67631
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    cell.com/ajhg/fulltext/S0002-9297(20)30162-2
  6. jmg.bmj.com jmg.bmj.com
    Blood functional assay for rapid clinical interpretation of germline TP53 variants
    1
    1. dkanavy 18 Mar 2021
      We then applied the p53 functional assay on blood samples sent to our laboratory for TP53 molecular analysis (NGS screening of the 11 exons complemented by QMPSF). Molecular and functional analyses were performed in parallel, in double blind conditions.

      AssayGeneralClass: BAOCL:20:0010044 targeted transcriptional assay

      AssayMaterialUsed: CL:2000001 peripheral blood mononuclear cell from patients

      AssayDescription: Comparative transcriptomic analysis using reverse transcription to compare peripheral blood mononuclear cells of patients with wild type or pathogenic TP53 variants in the context of genotoxic stress induced by doxorubicin treatment. p53 RNA levels were evaluated and expressed as a percentage of the mean levels obtained for the three wild-type TP53 individuals.

      AdditionalDocument: PMID: 23172776

      AssayReadOutDescription: The p53 mRNA levels were expressed as a ratio of the normal values obtained for 3 TP53 wild-type control individuals.

      AssayRange: UO:0000187 the p53 RNA levels were evaluated and expressed as a percentage of the mean levels obtained for three wild-type TP53 individuals.

      AssayNormalRange: >65%

      AssayAbnormalRange: <65%

      AssayIndeterminateRange: N/A

      AssayNormalControl: wild type TP53

      AssayAbnormalControl: LFS patient cells

      ValidationControlPathogenic: 8 individuals had seven distinct TP53 variants which could be considered as likely pathogenic or pathogenic based on their ClinVar classification or their truncating nature.

      ValidationControlBenign: 51 individuals had no detectable germline TP53 variant

      Replication: at least two wells were seeded per patient (treated and untreated) and duplicates or triplicates were performed whenever possible.

      StatisticalAnalysisDescription: Differentially expressed genes between doxorubicin-treated and untreated cells were arbitrarily defined using, as filters, a P<0.01 and fold-change cutoffs >2 or <2, for up and down regulation, respectively. The resultant signal information was analyzed using one-way analysis of variance (ANOVA, P= 0.001), assuming normality but not equal variances with a Benjamani–Hochberg correction for multiple comparisons using three groups: controls, null, and missense mutations.

      SignificanceThreshold: P=0.001

      Comment: statistical analysis and P value from previous publication.

      FuncAssay:4 BAO:0010044 CL:2000001 MONDO:0018875 HGNC:11998 UO:0000187 AssayReadOutType:Quantitative CGType:FunctionalAssay
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    jmg.bmj.com/content/early/2020/10/13/jmedgenet-2020-107059
  7. Feb 2021
  8. jmg.bmj.com jmg.bmj.com
    Blood functional assay for rapid clinical interpretation of germline TP53 variants
    78
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    jmg.bmj.com/content/early/2020/10/13/jmedgenet-2020-107059