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A Dual Color Immunohistochemistry Assay for Measurement of Cereblon in Multiple Myeloma Patient Samples.

ABSTRACT: Clinical interest in the measurement of Cereblon (CRBN), the primary target of the IMiDs immunomodulatory drugs lenalidomide and pomalidomide, has been fueled by its essential requirement for antitumor or immunomodulatory activity of both drugs in multiple myeloma (MM). However, limited analyses of clinical samples for CRBN gene expression or protein levels have utilized unvalidated reagents and assays, raising uncertainty about the interpretation of these results. We previously described a highly specific rabbit monoclonal antibody CRBN65 against 65-76 AA of human Cereblon. Here we describe a validated dual color bright-field Cereblon/CD138 immunohistochemical (IHC) assay utilizing CRBN65 and a commercial mouse monoclonal CD138 antibody. Sensitivity and specificity of the assay was determined and assay precision was shown for both cytoplasmic and nuclear Cereblon in MM bone marrow samples with coefficient of variation values of 5% and 2%, respectively. The dual IHC assay was effective for detecting a continuous range of Cereblon levels in 22 MM patient bone marrow core biopsies and aspirate clots, as shown by average cytoplasmic H-scores ranging from 63 to 267 and nuclear H-scores ranging from 17 to 250. Interpathologist comparison of MM sample H-scores by 3 pathologists demonstrated good concordance (R=0.73). This dual assay demonstrated superior Cereblon IHC measurement in MM samples compared with the single IHC assay using a published commercial rabbit polyclonal Cereblon antibody and could be used to explore the potential utility of Cereblon as a biomarker in the clinic.

SUBMITTER: Ren Y

PROVIDER: S-EPMC5106091 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Cereblon and IRF4 Variants Affect Risk and Response to Treatment in Multiple Myeloma.

Project description:Multiple myeloma (MM) is a plasma-cell malignancy derived from an early precursor of the B-cell lineage characterised by bone-marrow infiltration, lytic bone lesions, and the presence of a monoclonal protein in serum and/or urine. Interferon regulatory factor 4 (IRF4) is a critical transcriptional regulator in B-cell development and function that is required during immune response for lymphocyte activation and the generation of immunoglobulin-secreting plasma cells. Immunomodulatory drugs, derivatives of thalidomide, are commonly used in therapy against MM. They are known to target a protein called cereblon (CRBN); however, the exact mechanism remains unknown. The present study aimed to assess the association of two (rs12203592 and rs872071) polymorphisms within the IRF4 gene and two (rs711613 and rs1045433) in the CRBN gene with MM susceptibility, progression, and response to treatment. For this purpose, 144 MM patients and 126 healthy individuals were genotyped for the IRF4 and CRBN alleles. The presence of the IRF4 (rs872071) G allele was more frequently detected in patients than healthy individuals (OR 1.78; P = 0.034), and this relationship was especially pronounced in women (OR 2.83; P = 0.012). The CRBN (rs711613) A allele-carriers were better responders to the treatment (P = 0.012), in particular to thalidomide including therapy (P = 0.023). These results underline the prognostic significance of the IRF4 and CRBN polymorphisms in patients with MM.

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