Molecular characterisation of benign and malignant thyroid dysfunction

Nodular thyroid disease is common (prevalence 2-6%) and is a significant risk factor for the development of thyroid cancer. My aim was to apply genome-wide- linkage-analysis to identify the defect in a large kindred with multi-nodular goitre (MNG) of adolescent onset progressing to papillary thyroid cancer (PTC). Genomic DNA from 18 individuals (8 affected) was hybridized to Affymetrix GeneChip® Human Mapping 10K 2.0 Arrays. Results were analysed with Affymetrix GTYPE software to produce a call rate of ~92%. Extensive quality control steps were performed (PLINK, GRR) prior to linkage analysis using Merlin software in multipoint non-parametric and parametric (dominant) model. A non-parametric LOD score of 3.01 was obtained on chromosome 20 across 20cM, the same region produced a dominant LOD score of 2.03. Haplotype analysis reduced the region of interest to 3.7 Mbp, (encodes 10 genes). Analysis of copy number variation in an affected individual (Illumina Human 660W-Quad) revealed an intronic deletion of ~1000 bp in one copy of Phospholipase-C β1 (PLCβ1), (the first in the 10 gene list), which is present in all affected family members and carriers. The deletion contained ‘ATAA’ at the junction site and this InDel was found in 1 of 105 healthy unrelated people, a similar variant was reported in the database of genomic variants in ~1% of Europeans . The deletion was not present in 70 unrelated PTC patients but was found in 4/81 with MNG (all European); the deletion frequency in the general population vs. MNG gives a X2 value of 5.076 (p=0.024). PLCβ1 expression was measured in thyroid tissues from affected family members and subjects free of the InDel and were significantly higher in the former (p< 0.02). In conclusion, the InDel identified in familial MNG occurs in a proportion of sporadic MNG. It predisposes to goitre formation, possibly by increasing PLCβ1 transcription and activating the diacyl-glycerol, PKC and MAPK pathways