Mosaicism in von Hippel-Lindau disease: lessons from kindreds with germline mutations identified in offspring with mosaic parents

Abstract

von Hippel-Lindau disease (VHL [MIM 193300]) is a heritable autosomal dominant multiple-neoplastic disorder with high penetrance. It is characterized by brain and spinal-cord hemangioblastomas, retinal angiomas, clear-cell renal carcinoma, neuroendocrine tumors and cysts of the pancreas, pheochromocytomas, endolymphatic-sac tumors, and papillary cystadenomas of the epididymis and broad ligament. Although most index cases have a positive family history of VHL, some do not and may represent de novo cases. Cases without a family history of VHL may or may not have a germline mutation in their VHL tumor-suppressor gene. We present two cases of VHL mosaicism. In each of two families, standard testing methods (Southern blot analysis and direct sequencing) identified the germline mutation in the VHL gene of the offspring, but not in their clinically affected parent. Additional methods of analysis of the affected parents' blood detected the VHL-gene mutation in a portion of their peripheral blood lymphocytes. In one case, detection of the deleted allele was by FISH, and, in the second case, the 3-bp deletion was detected by conformational sensitive gel electrophoresis and DNA sequencing of cloned genomic DNA. Mosaicism in VHL is important to search for and recognize when an individual without a family history of VHL has VHL. Patients diagnosed without family histories of the disease have been reported in as many as 23% of kindreds with VHL. Identification of individuals potentially mosaic for VHL will affect counseling of families, and these individuals should themselves be included in clinical screening programs for occult disease.

Figure 1

Pedigrees of families 1 (top) and 2 (bottom), which have VHL with parental mosaicism. Arrows indicate probands. Blackened circles and squares indicate individuals with VHL disease. For keys to disease codes, see table 1.

Figure 2

Radiographic images for mosaic parents and affected offspring. A, 27-year-old woman with VHL, from family 1 (individual III:1). Computed tomography of the abdomen demonstrates multiple cysts in the pancreas (arrows). B, 49-year-old woman, with mosaicism for VHL, from family 1 (individual II:2). Computed tomography of the abdomen demonstrates milder cystic changes within the pancreas. C, 23-year-old woman, with VHL, from family 2 (individual IV:6). Magnetic resonance imaging of the spine with gadolinium enhancement demonstrates hemangioblastoma (arrow) in the low brainstem and a tiny additional hemangioblastoma at C2-3 (curved arrow). D, 48-year-old man, with VHL mosaicism, from family 2 (individual III:4) demonstrates hemangioblastomas (arrows) at the same site (as IV:6) in the low brainstem and an additional lesion in the cerebellum. Additional sections (not shown) showed multiple cerebellar hemangioblastomas.

Figure 3

Quantitative Southern blot analysis, family 1. Southern blots of EcoRI- and AseI-digested genomic DNA were hybridized to probes specific for the VHL gene (g7) (Latif et al. 1993) and the human beta-globin gene (Stolle et al. 1987). With equal loading of DNA, as assessed by the intensity of the beta-globin gene band, complete deletion of the VHL gene is apparent from the decreased intensity of the VHL gene band in patient samples, relative to that in unaffected controls. Samples with a partial deletion of the VHL gene exhibit an abnormally migrating band. Lanes 1, 2, and 7, Control DNA. Lane 3, DNA from III:1. Lane 4, DNA from II:2. Lanes 5 and 6, DNA from patients with complete or partial deletions of the VHL gene, respectively.

Figure 4

DNA-sequence analysis, family 2. A portion of the VHL gene containing exon 1 sequence was amplified as described elsewhere (Stolle et al. 1998) from either genomic DNA or plasmid DNA containing a cloned DNA insert and was subjected to DNA-sequence analysis. A, Sequence of genomic DNA from III:4. B, Sequence of genomic DNA from IV:6. C, Sequence of cloned DNA from III:4. Note that the sequence in B indicates that this patient is heterozygous for a frameshift mutation. The sequence of the cloned DNA in C reveals that the frameshift is due to a 3-bp (TTC) deletion in this region.

Figure 5

CSGE of samples from family 2. Genomic DNA from control (lane 1) and patient (lanes 2–5) samples was amplified with primers for exon 1. PCR products were denatured, reannealed, and analyzed by CSGE (Ganguly et al. 1993). Genomic DNA was extracted from the peripheral blood of a normal control (lane 1), peripheral blood from IV:6 (lane 2), peripheral blood from III:4 (lane 3), buccal cells from III:4 (lane 4), cultured skin fibroblasts from III:4 (lane 5). Arrows indicate position of the shifted bands.