Publications

PURPOSE

Posterior polymorphous corneal dystrophy (PPCD) is an autosomal-dominant disorder of the corneal endothelium associated with visually significant corneal edema and glaucoma. Statistical genetic analysis of 4 families with PPCD has demonstrated linkage to a 2.4 cM common support interval on chromosome 20 bordered by the markers D20S182 and D20S139. We sought to identify the genetic basis of PPCD linked to chromosome 20 (PPCD1) by screening the 26 positional candidate genes between these markers in a family previously mapped to the PPCD1 region.

METHODS

The coding regions of the 26 positional candidate genes mapped to the common PPCD1 support interval were amplified and sequenced in affected and unaffected individuals from a family previously linked to the PPCD1 locus. Nine other genes positioned just outside of the common PPCD1 support interval but within the autosomal-dominant congenital hereditary endothelial dystrophy interval were also screened.

RESULTS

Four DNA sequence variants in 3 of the positional candidate genes demonstrated complete segregation with the affected phenotype: p.Thr109Thr (rs6111803) in OVOL2, p.Arg56Gln (novel variant-RPSnovel) in RPS19P1, and p.Thr85Thr (rs1053834) and p.Pro99Ser (rs1053839) in C20orf79. Each of these 4 sequence variants demonstrated significant linkage with the affected phenotype in this family (P = 2.5 x 10 for RPSnovel, rs1053834 and rs1053839; P = 8.6 x 10 for rs6111803). However, we also identified each of these 4 sequence variants in > or = affected control individuals. The haplotype on which the disease-causing mutation is segregating was found to have a population frequency of 4.2% in the CEPH HapMap trios. Although a number of other previously described and novel single nucleotide polymorphisms were identified in the 35 positional candidate genes located within the PPCD1 and congenital hereditary endothelial dystrophy intervals, none segregated with the affected phenotype.

CONCLUSIONS

We report the absence of a presumed pathogenic coding region mutation in the common PPCD1 support interval. Although minor alleles of 4 single nucleotide polymorphisms were identified that segregated with the affected phenotype, the relatively high frequency of each minor allele in the general population indicates that none is a candidate for the causal variant for PPCD. Instead, the causal variant is most likely a coding region deletion or a variant in a noncoding region of the PPCD1 common support interval.

Although the clinical and pathologic diagnosis of some melanomas is clear-cut, there are many histopathologic simulators of melanoma that pose problems. Over-diagnosis of melanoma can lead to inappropriate therapy and psychologic burdens, whereas under-diagnosis can lead to inadequate treatment of a deadly cancer. We used existing data on DNA copy number alterations in melanoma to assemble panels of fluorescence in situ hybridization (FISH) probes suitable for the analysis of paraffin-embedded tissue. Using FISH data from a training set of 301 tumors, we established a discriminatory algorithm and validated it on an independent set of 169 unequivocal nevi and melanomas as well as 27 cases with ambiguous pathology, for which we had long-term follow-up data. An algorithm-using signal counts from a combination of 4 probes targeting chromosome 6p25, 6 centromere, 6q23, and 11q13 provided the highest diagnostic discrimination. This algorithm correctly classified melanoma with 86.7% sensitivity and 95.4% specificity in the validation cohort. The test also correctly identified as melanoma all 6 of 6 cases with ambiguous pathology that later metastasized. There was a significant difference in the metastasis free survival between test-positive and negative cases with ambiguous pathology (P=0.003). Sufficient chromosomal alterations are present in melanoma that a limited panel of FISH probes can distinguish most melanomas from most nevi, providing useful diagnostic information in cases that cannot be classified reliably by current methods. As a diagnostic aid to traditional histologic evaluation, this assay can have significant clinical impact and improve classification of melanocytic neoplasms with conflicting morphologic criteria.