A study of 103 patients with VHL retinal manifestations and 108 patients without VHL retinal manifestations extracted from the French VHL database revealed that the number of hemangioblastomas appeared higher in patients who had a substitution when compared with patients with a truncated protein. Detected by PCR and SSCP, this missense mutation (p.Q132P (c.395A>C)) was found in a VHL patient without retinal hemangioblastomas. Other manifestations are not clearly stated. No family information was provided. This is possibly previously published in PMID: 15300849

A study of 103 patients with VHL retinal manifestations and 108 patients without VHL retinal manifestations extracted from the French VHL database revealed that the number of hemangioblastomas appeared higher in patients who had a substitution when compared with patients with a truncated protein. Detected by PCR and SSCP, this mutation p.V130L (c.388G>C) was found in 3 VHL patients. 2 members were said to have retinal hemangioblastomas, while the other member was said not to have this phenotype. Other manifestations are not clearly stated. No family information or relation was provided. This is possibly previously published in PMID: 15300849

A study of 103 patients with VHL retinal manifestations and 108 patients without VHL retinal manifestations extracted from the French VHL database revealed that the number of hemangioblastomas appeared higher in patients who had a substitution when compared with patients with a truncated protein. Detected by PCR and SSCP, this mutation p.C162W (c.486C>G) was found in 4 VHL patients. All 4 didn’t have retinal hemangioblastomas, while other manifestations are not clearly stated. No family information or relation was provided. This is possibly previously published in PMID: 15300849

A study of 103 patients with VHL retinal manifestations and 108 patients without VHL retinal manifestations extracted from the French VHL database revealed that the number of hemangioblastomas appeared higher in patients who had a substitution when compared with patients with a truncated protein. Detected by PCR and SSCP, this missense mutation (p.H115Q (c.345C>G)) was found in a VHL patient without retinal hemangioblastomas. Other manifestations are not clearly stated. No family information was provided. This is possibly previously published in PMID: 15300849

A study of 103 patients with VHL retinal manifestations and 108 patients without VHL retinal manifestations extracted from the French VHL database revealed that the number of hemangioblastomas appeared higher in patients who had a substitution when compared with patients with a truncated protein. Detected by PCR and SSCP, this mutation was found in a 1 patient, without retinal hemangioblastomas, but other manifestations are not clearly stated. No family information or relation was provided. Recorded in paper as c.417_418delTC, this automaps to NM_000551.3:c.419_420delTC (p.L140fs). This is possibly previously published in PMID: 15300849

A study of 103 patients with VHL retinal manifestations and 108 patients without VHL retinal manifestations extracted from the French VHL database revealed that the number of hemangioblastomas appeared higher in patients who had a substitution when compared with patients with a truncated protein. Detected by PCR and SSCP, this mutation p.R108L*52 (c.322_323insTG) was found in a 1 patient, without retinal hemangioblastomas, but other manifestations are not clearly stated. No family information or relation was provided. This is possibly previously published in PMID: 15300849

A study of 103 patients with VHL retinal manifestations and 108 patients without VHL retinal manifestations extracted from the French VHL database revealed that the number of hemangioblastomas appeared higher in patients who had a substitution when compared with patients with a truncated protein. Detected by PCR and SSCP, this synonymous mutation (p.P154= (c.462A>C)) was found in a VHL patient with retinal hemangioblastomas. Other manifestations are not clearly stated. No family information was provided.

Mutational analysis for 16 patients with clinically diagnosed VHL disease, revealed 12 germline mutations. This missense mutation was found in 4 VHL family members (proband no. 6). The 65-year-old father had renal cell carcinoma; The 37-year old sister had pancreatic cysts and the 33-year old sister had retinal angiomas. The 38-year-old proband had hemangioblastomas of the central nervous system, renal cell carcinoma, pancreatic cysts, and epididymal cystadenoma. Cosegregation with disease in multiple affected family members in a gene definitively known to cause the disease

Mutational analysis for 16 patients with clinically diagnosed VHL disease, revealed 12 germline mutations. This missense mutation was found in a 43-year-old VHL type 1 patient with renal cell carcinoma, pancreatic cysts, an endolymphatic sac tumor, and an epididymal /ovarian cystadenoma (proband no. 5) and no family history. HPO terms: renal cell carcinoma, multiple pancreatic cysts, epididymal cysts, endolymphatic sac tumors.

Mutational analysis for 16 patients with clinically diagnosed VHL disease, revealed 12 germline mutations. This mutation was found in a 30-year-old VHL patient with hemangioblastomas of the central nervous system, retinal angiomas, renal cell carcinoma, and pancreatic cysts (proband no. 16). HPO terms: hemangioblastoma, retinal hemangioblastoma, renal cell carcinoma, multiple pancreatic cysts.

Data was collected from 82 VHL mutation carriers in the Dutch VHL surveillance program. One patient was found with this missense varaint in the VHL gene. The patient was 27Y at the last follow-up and presented with: pancreatic neuroendocrine tumour and pheochromocytoma. Family history was not described. This cohort was published later in PMID 31087189.

Data was collected from 82 VHL mutation carriers in the Dutch VHL surveillance program. Seven patients were found with this missense variant in the VHL gene. The ages of the patients was represented as a range 7-88 (mean 45). Phenotypes were grouped together: pheochromocytoma and renal cysts, although it is unclear if only one of the seven patients had renal cysts. Family history was not described. This cohort was published later in PMID 31087189.

Data was collected from 82 VHL mutation carriers in the Dutch VHL surveillance program. Two patients were found with the missense mutation, c.241C>T, in the VHL gene. One patient was 34Y and the other was 59Y at last follow-up. Their phenotypes were grouped together: retinal hemangioblastoma, cerebellar hemangioblastoma, spinal cord hemangioblastoma, renal cell carcinoma, pancreatic neuroendocrine tumour, pheochromocytoma, and renal cysts. Family history was not described. The group was classified as a 2a/b which was described by the author to included de novo VHL mutation carriers or VHL mutation carriers having a family history with diagnosis of pheochromocytoma and other VHL related manifestations.

In a study of 426 unrelated VHL patients, 111 were discovered to have alterations in the VHL gene. 18 novel variants were identified in VHL patients, but none were present in 200 unaffected control individuals. DNA samples from peripheral blood leukocytes underwent polymerase chain reaction (PCR) amplification, denaturing high-performance liquid chromatography (DHPLC), and direct sequence analysis of the entire coding sequence and intron-exon boundaries. This splice site variant, c.463+1G>T, was found in an asymptomatic 8 year-old with family history of VHL disease (index case 242).

In a study of 426 unrelated VHL patients, 111 were discovered to have alterations in the VHL gene. 18 novel variants were identified in VHL patients, but none were present in 200 unaffected control individuals. DNA samples from peripheral blood leukocytes underwent polymerase chain reaction (PCR) amplification, denaturing high-performance liquid chromatography (DHPLC), and direct sequence analysis of the entire coding sequence and intron-exon boundaries. The T157I (c.470C>T) germline missense mutation of the VHL gene was found in two VHL patients (23Y and 26Y). Both had retinal hemangioblastomas and pheochromocytoma (index cases 8 and 19). The 23 yr old patient also had hemangioblastomas of the central nervous system.

In a study of 426 unrelated VHL patients, 111 were discovered to have alterations in the VHL gene. 18 novel variants were identified in VHL patients, but none were present in 200 unaffected control individuals. DNA samples from peripheral blood leukocytes underwent polymerase chain reaction (PCR) amplification, denaturing high-performance liquid chromatography (DHPLC), and direct sequence analysis of the entire coding sequence and intron-exon boundaries. This nonsense mutation, c.481C>T, was found in 3 VHL patients. The 17-year-old VHL patient only had hemangioblastomas of the central nervous system (CNS) (index case 218). The 24-year-old VHL patient had CNS hemangioblastomas and clear cell renal cell carcinoma (index case 46). The 29-year-old VHL patient had CNS and retinal capillary hemangioma and pheochromocytoma (index case 79). ACMG code as follows: There is very strong evidence for pathogenicity in the form of a nonsense variant in a gene where loss-of-function is a known mechanism of disease.

In a study of 426 unrelated VHL patients, 111 were discovered to have alterations in the VHL gene. 18 novel variants were identified in VHL patients, but none were present in 200 unaffected control individuals. DNA samples from peripheral blood leukocytes underwent polymerase chain reaction (PCR) amplification, denaturing high-performance liquid chromatography (DHPLC), and direct sequence analysis of the entire coding sequence and intron-exon boundaries. The L153P (c.458T>C) germline missense mutation of the VHL gene was found in a 36 yr old VHL patient with renal cysts as well as central nervous system and retinal hemangioblastoma (index case 191).

In a study of 114 unrelated VHL families, 85 germline mutations were found. VHL mutations were detected in affected family members, but not in unaffected family members or 96 normal individuals.  This mutation was found in a VHL family of 4 individuals (family ID 4409), using PCR and SSCP sequencing. The mutation of c.340G>C was referred to in the paper by c.553G>C due to a difference in transcript sequence. 4/4 retinal angioma; 2/4 CNS hemangioblastoma; 1/4 RCC; 1/4 Pheo. Walther et al., 1999 also reports a a partial update of this family, describing that Family 4409 also had pancreatic neuroendocrine tumour but not how many members were affected by PNET. (ACMG code: PP4).

In a study of 114 unrelated VHL families, 85 germline mutations were found. VHL mutations were detected in affected family members, but not in unaffected family members or 96 normal individuals. This mutation was also found in a VHL type 1 family of 27 individuals (family ID 3003), using PCR and SSCP sequencing. The mutation of c.502_503insTTGTTCGT was referred to in the paper by c.715insTTGTTCGT due to a difference in transcript sequence. Five of 11 individuals were identified with renal cell carcinoma, 11 with retinal hemangioblastomas, and 9 with CNS hemangioblastomas. Previously published in PMID 8956040 (type 1 3003NCI); and PMID 9829911. ACMG codes as follows: PVS1 a frameshift variant in a gene where loss of function is a known mechanism of disease.

A cohort of 141 patients with hemangioblastomas of the central nervous system was analyzed. 81 patients had germline mutations in the VHL gene, analyzed through peripheral blood by Southern blotting and SSCP analysis. This missense mutation (c.500G>A) was recorded in the paper as c.713G>A due to a difference in transcript sequence. It was found in one patient with a hemangioblastoma of the central nervous system. Any family information is unknown. No other phenotypes described.

A cohort of 141 patients with hemangioblastomas of the central nervous system was analyzed. 81 patients had germline mutations in the VHL gene, analyzed through peripheral blood by Southern blotting and SSCP analysis. This nonsense mutation (c.548C>A) was recorded in the paper as c.761C>A due to a difference in transcript sequence. It was found in 4 patients with hemangioblastomas of the central nervous system. Any family information or relations is unknown. No other phenotypes described.

A cohort of 141 patients with hemangioblastomas of the central nervous system was analyzed. 81 patients had germline mutations in the VHL gene, analyzed through peripheral blood by Southern blotting and SSCP analysis. This missense mutation (c.241C>T) was recorded in the paper as c.454C>T due to a difference in transcript sequence. It was found in 1 patient with a hemangioblastoma of the central nervous system. Any family information is unknown. No other phenotypes described.

A cohort of 141 patients with hemangioblastomas of the central nervous system was analyzed. 81 patients had germline mutations in the VHL gene, analyzed through peripheral blood by Southern blotting and SSCP analysis. This splice site mutation (c.463+1G>C) was recorded in the paper as c.676+1G>C due to a difference in transcript sequence. It was found in 2 patients with hemangioblastomas of the central nervous system. Any family information or relations is unknown. No other phenotypes described.

A case study on a 35 year old female of Indian (South Asian) descent was presented. She presented 6 years prior (2006) with severe abdominal pain prompting an abdominal ultrasound. Imaging revealed a left renal mass measuring 3.5 × 4.0 × 3.5cm (biopsy consistent with papillary renal cell carcinoma). Final pathology revealed a 3.2cm low-grade primary renal carcinoid (neuroendocrine neoplasm of the kidney) with two of four lymph nodes involved with metastatic disease. She currently remains disease free after 31 months. A clear cell renal cell carcinoma tissue sample removed surgically from a patient with VHL detected a germline VHL mutation: R167Q (c.500G>A), which was used as a positive control. This was described in the paper was c.713G>A due to a difference in transcript reference used.

3 unrelated patients with unexpected or non-syndromic phenotypes, accompanied by a VHL component tumor received germline gene panel testing as part of the PROACTIVE study at the Dana-Farber Cancer Institute, each detected with a low penetrant variant. At age 32, patient 3 (female) presented with posterior headaches, which a CT scan and MRI revealed to be a cerebellar hemangioblastoma. Treatment involved a gross total resection of the lesion. Somatic testing of a left cerebellar mass revealed a somatic VHL variant: p.Leu63fs*4 (c.188_188T>TG); updated to p.R64fs (c.189dupG). Single gene testing of VHL revealed a heterozygous germline VHL p.Pro81Ser (c.241C>T) variant. To this day at age 40, she continues to be monitored. Her mother (age 72) and two of her three siblings (ages 38 and 36) carry the same VHL p.Pro81Ser variant, but have been negative for hemangioblastoma or any other VHL component tumor. ACMG: PP4

Fourty-four of 389 patients with VHL (between 1988-1999) screened positive for PNETs either by pathologic analysis of tissue specimens or by characteristic radiographic appearance on CT and MRI. VHL gene germline mutations were determined by quantitative Southern blotting to detect deletions of the gene and gene rearrangements as well as complete gene sequencing. Patient 14, a 42 year old male, was found to harbour the c.490 C>T mutation in the VHL gene (stated as 703 C to T in the paper which used the Latif sequence for reference) leading to a premature stop codon at protein position 164 in exon 3. He presented with a single pathologic pancreatic neuroendocrine tumor. ACMG evidence codes: 'PP4' since the patient presents phenotypes highly specific for a disease with a single genetic etiology.

Fourty-four of 389 patients with VHL (between 1988-1999) screened positive for PNETs either by pathologic analysis of tissue specimens or by characteristic radiographic appearance on CT and MRI. VHL gene germline mutations were determined by quantitative Southern blotting to detect deletions of the gene and gene rearrangements as well as complete gene sequencing. One patient (15), a 26 year old female, was found to harbour the p.Val181Glyfs*20 (c.540_543delCGTC) VHL gene mutation, stated in the paper as c.753_756delCGTC according to an older transcript reference. She presented at 21Y with a single radiologic pancreatic neuroendocrine tumor. No family information was present.

Fourty-four of 389 patients with VHL (between 1988-1999) screened positive for PNETs either by pathologic analysis of tissue specimens or by characteristic radiographic appearance on CT and MRI. VHL gene germline mutations were determined by quantitative Southern blotting to detect deletions of the gene and gene rearrangements as well as complete gene sequencing. One patient (20), a 68-year-old male, was found to harbour the c.278G>A mutation in the VHL gene (stated as 491 G to A in the paper which used the Latif sequence as reference) leading to an amino acid change of Gly to Asp at protein position 93. He presented with a single radiologic pancreatic neuroendocrine tumor. ACMG evidence codes: 'PP4' since the patient presents phenotypes highly specific for a disease with a single genetic etiology.

Of 31 Brazilian patients, five were found to harbour the c.499C>T (p.Arg167Trp) mutation of the VHL gene. Genomic DNA was extracted and all VHL coding regions were analyzed by Sanger sequencing. MLPA was performed to investigate large deletions using SALSA® MLPA® P016 VHL probe mix. Data analysis was performed with Genescan 3.7. Two of these patients were familials. Individual 12 of family 6, age 36, presented with paraganglioma, pheochromocytoma, pancreatic endocrine tumor (9.6cm), renal cyst, pancreatic cyst, retinal hemangioblastoma as well as a local duodenal invasion. Individual 13 of family 6, a 16 year old male, presented with paraganglioma and a right-side pheochromocytoma (1.7cm). Individual 5 of family 3, age 30, presented with paraganglioma, pheochromocytoma, pancreatic endocrine tumor, renal cell carcinoma pancreatic cyst, CNS hemangioblastoma and retinal hemangioblastoma. Individual 11 of family 5, age 26, presented with paraganglioma, pheochromocytoma, and pancreatic endocrine tumor. Finally, individual 25 of family 18, age 35Y, presented with paraganglioma, pheochromocytoma at age 26Y, pancreatic endocrine tumor at 35Y, and CNS hemangioblastoma.

Of 31 Brazilian patients, one was found to harbour the c.309_322del14(p.G104Hfs*23) mutation of the VHL gene. Genomic DNA was extracted and all VHL coding regions were analyzed by Sanger sequencing. MLPA was performed to investigate large deletions using SALSA® MLPA® P016 VHL probe mix. Data analysis was performed with Genescan 3.7. This patient, age 15, presented with renal and pancreatic cysts, CNS and retinal hemangioblastoma. It is not clear if this mutation is familial.

Of 31 Brazilian patients, one was found to harbour the c.540delC (p.I180Mfs*22) mutation of the VHL gene. Genomic DNA was extracted and all VHL coding regions were analyzed by Sanger sequencing. MLPA was performed to investigate large deletions using SALSA® MLPA® P016 VHL probe mix. Data analysis was performed with Genescan 3.7. This patient, age 14, presented with CNS and retinal hemangioblastoma. It is not clear if this mutation is familial.

5 cases treated for thoracic paraganglioma in Freiburg, Germany aged 6–70 at the time of diagnosis were identified and had germline VHL mutations identified using constitutional genomic DNA from blood samples. The mutation p.Gly93Ser (legacy notation c.490G>A converted to c.277G>A) was found in case 3, a 6Y boy with thoracic paraganglioma. His parents were negative for the mutation, supporting de novo germline mutatation. ACMG: PM6

Patients with ELSTs were identified and included in the International Endolymphatic Sac Tumor Registry. The registry contained 93 patients as of July 2014, and of those, 25 were sporadic, and 68 associated with VHL. Patients with ELSTs and clinically confirmed diagnoses of VHL were included in this study. Statistics of this VHL subgroup are as follows: mean age 30, 62% female, 91% symptomatic, 54% with retinal angioma, 72% with CNS hemangioblastoma, 46% with renal cell carcinoma, 19% with pheochromocytoma, and 12% with pancreatic neuroendocrine tumour. In the data from this study, only endolymphatic sac tumours can be confirmed as a phenotype for this mutation (c.337C>T.), which was found in 1 individual from France.

Patients with ELSTs were identified and included in the International Endolymphatic Sac Tumor Registry. The registry contained 93 patients as of July 2014, and of those, 25 were sporadic, and 68 associated with VHL. Patients with ELSTs and clinically confirmed diagnoses of VHL were included in this study. Statistics of this VHL subgroup are as follows: mean age 30, 62% female, 91% symptomatic, 54% with retinal angioma, 72% with CNS hemangioblastoma, 46% with renal cell carcinoma, 19% with pheochromocytoma, and 12% with pancreatic neuroendocrine tumour. In the data from this study, only endolymphatic sac tumours can be confirmed as a phenotype for this mutation (c.339delA), which was found in 1 individual from Germany.


GroupID: 15

Case: Sex unknown, Age unknown, Italian

DiseaseAssertion: VHL

FamilyInfo: Unknown. 1 registrant was found with this variant; no family info was mentioned

CasePresentingHPOs: HP:0030393 (Endolymphatic sac tumor)

CaseHPOFreeText: N/A

CaseNotHPOs: N/A

CaseNotHPOFreeText: N/A

CasePreviousTesting: N/A

PreviouslyPublished: N/A

SupplementalData: N/A

Variant: p.R120G (c.358A>G)

CaseProblemVariantFreeText: N/A

LegacyVariant: N/A

ClinVarID: 223199 https://www.ncbi.nlm.nih.gov/clinvar/variation/223199/

CAID: N/A

gnomAD: N/A

VariantEvidence: N/A

MutationType: missense_variant; transition

CivicName: R120G (c.358A>G)

MultipleGeneVariants: N/A

Two cases of VHL patients presenting with an endolympathic sac tumor are reported. Genetic testing revealed the mutation: H115Q (c.345C>G) in the second patient. The second patient had a familial history of VHL disease but had never been screened for central nervous system manifestations when she experienced a sudden left hearing loss at 18 years old. Additional manifestations included: CNS hemangioblastoma and pancreatic cysts. During 18 years of surveillance, the endolympathic sac tumor increased in size following a two-phase growth pattern alternating phases of quiescence and phases of continuous growth.

Two cases of VHL patients presenting with an endolympathic sac tumor are reported. The first patient had no familial history of VHL disease and was 21 years old when he experienced a sudden right hearing loss. Genetic testing revealed the mutation: R113* (c.337C>T). Additional manifestations included: retinal hemangioblastoma, CNS hemangioblastoma, pheochromocytoma, renal cell carcinoma, and pancreatic cysts. After 20 years of radiological surveillance, the size of his endolymphatic sac tumor remained stable.

Eight children with a history of polycythemia and elevated serum erythropoietin level were evaluated. Three germline mutations were identified in 4 patients while, mutations were not detected for other 4 patients. Peripheral blood samples were taken and PCR products of either each VHL exon or the cDNA were purified and sequenced using a DNA sequencing kit (BigDye Terminator Cycle Sequencing Ready Reaction; Applied Biosystems, Foster City, CA). The sequencing reaction was performed in the Peltier Model P200 thermocycler. The products were then analyzed on the ABI Prism 377 DNA Sequencer (Applied Biosystems) according to the manufacturer's protocol. Clonality was determined by a transcriptional assay of exonic polymorphisms in 5 X-linked genes. An in vitro assay of erythroid progenitors' sensitivity to Epo was also performed. This missense mutation was found in a family of three patients. Patient 1, was a 16 year old Ukrainian girl who was heterozygous for the c. 376G>C mutation. She was diagnosed with polycythemia at age 10 and since then had presented with pulmonary capillary hemangiomatosis, abnormality of the kidney, hemangioma and her polycythemia had maintained. Her brother, patient 2, was 10 years old and also heterozygous for the mutation but only presented with polycythemia. He had an abnormally high erythropoietin level (271 mIU/mL). The children were found to have inherited the mutation from their father (no patient ID) who was also heterozygous for the mutation but did not have polycythemia. All three patients had hypersensitivity of erythroid progenitors to Epo.

A 6 year-old boy with von Hippel-Lindau (VHL) disease presented with hypertension due to bilateral pheochromocytomas. At age 13 he developed a renal carcinoma and bilateral paragangliomas, as well as clear cell RCC. His mother's age of first presentation was 39Y old and her last known age was 41Y old. She was found with retinal angiomas, bilateral pheochromocytomas and a cerebellar hemangioblastoma. Both had the same germline VHL missense mutation. This mutation was reported as c.704A>G and converted to NM_000551.3:c.491A>G. ACMG: PP4

Tissue specimens were retrieved from the archives of the Department of Pathology of the Erasmus MC (Rotterdam, The Netherlands), The University Medical Center St Radboud (Nijmegen, The Netherlands), and the University Hospital Zürich (Zürich, Switzerland). The samples were sequenced using Bigdye Terminator V3.1 cycle sequencing kit (Applied Biosystems, Foster City, CA, USA). Products were analyzed on the ABI Prism 3100 genetic analyzer (Applied Biosystems).The mutation analysis of pheochromocytoma tissue samples showed two patients, 31 year-old (F6) and 24 year-old (F87) males, with R64P germline mutations in exon 1 of the VHL tumor suppressor gene and both had no metastasis. Patient F87 was followed up 11 years later and patient F6 was followed up 21 years later. These patients belonged to the same family (family A). Besides bilateral pheochromocytoma, the paper does not provide any other phenotypic information.

301 were enrolled in a prospective clinical protocol to evaluate the natural history of vHL-associated pancreatic lesions (NCT00062166). Of these, 69 patients with pancreatic manifestations of vHL disease and missense VHL mutations were included in the current study. Individual patient phenotype was not described, but overall of the 60 patients, 57 had RCC, 41 had CNS hemangioblastomas, 28 had retinal hemangioblastoma, 23 had pheochromocytoma. 36/69 patients were female and the mean age was 50.5 years old. All patients underwent genetic testing using Sanger sequencing and Southern Blotting. Three patients were listed with this missense mutation, c.407T>C.

301 were enrolled in a prospective clinical protocol to evaluate the natural history of vHL-associated pancreatic lesions (NCT00062166). Of these, 69 patients with pancreatic manifestations of vHL disease and missense VHL mutations were included in the current study. Individual patient phenotype was not described, but overall of the 60 patients, 57 had RCC, 41 had CNS hemangioblastomas, 28 had retinal hemangioblastoma, 23 had pheochromocytoma. 36/69 patients were female and the mean age was 50.5 years old. All patients underwent genetic testing using Sanger sequencing and Southern Blotting. One patient was listed with this missense mutation, c.361G>T.

Screening of 92 unrelated patients with VHL disease revealed 61 DNA variants. No variants were found in 96 control cases. The R167W (c.499C>T) germline missense mutation was found in a VHL type 2A patient, a VHL type 2B patient and a family with 5 VHL type 2B patients (patient no. V30, V23, V1). All of these families presented with pheochromocytoma. The first two individuals had no family screened but 14 unaffected family members showed no evidence of this mutation in the family with 5 mutation-positive affected members. All of these families are stated as being previously reported in PMID 15300849.

Screening of 92 unrelated patients with VHL disease revealed 61 DNA variants. No variants were found in 96 control cases. The V166F (c.496G>T) germline missense mutation was found in a VHL type 2A (with pheochromocytoma) patient (patient no. V49). This variant was not found in 8 unaffected family members and previously reported in patients with VHL. This patient was stated to be previously published in PMID 15300849.

Screening of 92 unrelated patients with VHL disease revealed 61 DNA variants. Blood samples were isolated and sequenced by direct sequencing using prism-ready reaction dye primer cycle sequencing kits (Applied Biosystem Roissy CDG France) and an ABI 373A automated sequencer (AppliedBiosystem Roissy CDG France).No variants were found in 96 control cases. This stop gain mutation, c.481C>T, was found in 2 unrelated VHL type 1 patients (patient V3, V46). No patient information was provided and no additional family members were screened. Patient V3 was previously published in 8956040.

Screening of 92 unrelated patients with VHL disease revealed 61 DNA variants. No variants were found in 96 control cases. Direct sequencing on peripheral blood was systematically performed on exon 1 PCR products. Southern blotting was used to search for larger alterations to the gene. The F136S (c.407T>C) missense mutation was found in two unrelated VHL type 1 (no pheochromocytoma) patients (patient no. V53, V94). No unaffected individuals from either family were tested for this mutation.

Medical records of 26 patients with germline mutations of the VHL gene who had been diagnosed with VHL disease in Severance Hospital (Seoul, Republic of Korea) and Samsung Medical Center (Seoul, Republic of Korea) between 2003 and 2012 were respectively reviewed. Peripheral blood leukocytes were isolated and directly sequenced. One patient presented with the c.488T>C p.Leu162Pro germline variant of the VHL gene. Patient 41, 39Y male, presented with multiple and/or bilateral renal cell carcinoma and CNS hemangioblastomas as well as a family history negative for VHL disease. Their phenotypes were revised from a previous report (PMID 19270817). He was diagnosed with VHL Type 1.

Medical records of 26 patients with germline mutations of the VHL gene who had been diagnosed with VHL disease in Severance Hospital (Seoul, Republic of Korea) and Samsung Medical Center (Seoul, Republic of Korea) between 2003 and 2012 were respectively reviewed. Peripheral blood leukocytes were isolated and directly sequenced. Patient 27, 42Y male, with VHL Type 1 presented with bilateral and/or multiple CNS hemangioblastomas and renal cell carcinoma. He had a germline missense mutation in the VHL gene, c.345C>A (p.His115Gln) (incorrectly reported in the current paper as “c.346C>A” and corrected by the authors to “c.345C>A” via email correspondence). The patient had a negative family history of VHL disease.

Medical records of 26 patients with germline mutations of the VHL gene who had been diagnosed with VHL disease in Severance Hospital (Seoul, Republic of Korea) and Samsung Medical Center (Seoul, Republic of Korea) between 2003 and 2012 were respectively reviewed. Peripheral blood leukocytes were isolated and directly sequenced. Patient 29, 47Y female, and her daughter presented with multiple and/or bilateral pheochromocytoma. The proband also presented with CNS and retinal hemangioblastomas. Both family members had the c.361G>A (p.D121N) germline VHL variant. The probands phenotypes were revised from a previous report in PMID 16142346 and was previously reported by Yang et al., in 2005 (reference 20 in the current paper). The proband was diagnosed with VHL Type 2A and her daughter with VHL Type 2C.

Medical records of 26 patients with germline mutations of the VHL gene who had been diagnosed with VHL disease in Severance Hospital (Seoul, Republic of Korea) and Samsung Medical Center (Seoul, Republic of Korea) between 2003 and 2012 were respectively reviewed. Peripheral blood leukocytes were isolated and directly sequenced using the BigDye Terminator Cycle Sequencing Ready Reaction kit on an ABI Prism 3130 genetic analyzer (Applied Biosystems). One patient presented with the c.173insC germline variant of the VHL gene. Patient 1, 17Y female previously reported in PMID 19270817, presented with CNS hemangioblastoma and a family history positive for VHL.

20 vHL families were retrospectively identified in SickKids Genetics and Cancer Genetics Program SHIRE genetics database, with data entered between 30 August 1984 and 8 August 2016. Family 5 was found with this frameshift mutation, c.408delT in the VHL gene . The female proband (ID 1) was found with hemangioblastoma, abnormality of the pancreas and had a VHL age of onset of 10Y and her last known age was 13Y. Her mother (ID 4) was found with hemangioblastoma at 37Y and was 39Y at the time of the study. Her aunt (ID 5) was found with hemangioblastoma, retinal capillary hemangioma, abnormality of the kidney, abnormality of the pancreas, had a VHL age of onset of 29Y and her last known age was 37Y. Her male cousin (ID 8) was 5Y and had no manifestations. Her grandfather (ID 13) was not genetically confirmed with the mutation, but was found with hemangioblastoma at 57Y and was 63Y at the time of the study. This family was later published in 31368132.

45 individuals from 5 unrelated Chinese families were evaluated, of which 21 patients were diagnosed with VHL or showed strong evidence of having this disease. In family 5, the proband was a 10 year old male who presented with bilateral adrenal pheochromocytoma. His mother had left adrenal pheochromocytomas, and his grandmother had bilateral adrenal pheochromocytomas. Based on his clinical presentation and family history, he was diagnosed with VHL. DNA was isolated from a peripheral blood sample from the proband, and screened for a variety of pheochomocytoma and renal cell carcinoma mutations using Target Capture-Based Deep Sequencing (BGI Health.) This revealed the a heterozygous c.293A>G (p.Try98Cys) mutation in their VHL gene. ACMG evidence codes: PP4- patient’s phenotype or family history is highly specific for a disease with a single genetic etiology.

A case report of a 49Yo woman presenting with a spinal hemangioblastoma was reported. Molecular analysis confirmed the presence of the above mutation, 393C>A (N131K). Upon further workup, retinal hemangioblastoma, renal cysts, and a carotid body paraganglioma were identified. Familial screening showed 3 family members with a clinical diagnosis of VHL, who had all passed away before testing could be conducted. Clinical manifestations in these members included: renal cell carcinoma, retinal hemangioblastoma, and CNS hemangioblastoma.

A series of 28 pheochromocytomas from 28 patients from the Freiburg International Pheochromocytoma Registry were used to study somatic point mutation variants and loss of heterozygosity. Whole blood leukocytes and tumor samples underwent direct sequencing of all three exons of the VHL gene, all eight exons of the SDHB gene, all six exons of the SDHC gene, all four exons of the SDHD gene, and exons 10, 11, and 13 of the RET gene. 9 out of 28 samples were from the VHL germline mutation carriers. VHL-associated pheochromocytoma samples were collected from 3 patients (tumour ID 10,11,12) with germline missense (c.292T>C) VHL mutation. Patient 10, was found to have multiple benign tumors 4 cm in size in her adrenal gland. Patient 11, was found to have a single benign tumor 9 cm in size in her adrenal gland. Patient 12, was found to have a single benign tumor 4 cm in size in her adrenal gland. Both patients 10 and 12 had a somatic mutation in the RET gene (c. 2307T>G) and they both showed LOH in the VHL and SDHD gene. Patient 11 had LOH in the SDHB gene.

A series of 28 pheochromocytomas from 28 patients from the Freiburg International Pheochromocytoma Registry were used to study somatic point mutation variants and loss of heterozygosity. Whole blood leukocytes and tumor samples underwent direct sequencing of all three exons of the VHL gene, all eight exons of the SDHB gene, all six exons of the SDHC gene, all four exons of the SDHD gene, and exons 10, 11, and 13 of the RET gene. 9 out of 28 samples were from the VHL germline mutation carriers.VHL-associated pheochromocytoma samples were collected from 1 patient (patient 9) with germline missense (c.193T>A) VHL mutation. The patient was a 14 year old male and presented with multiple benign adrenal pheochromocytomas. The patient was found to have LOH in the SDHC gene.

A previous study of 26 Japanese, VHL families was extended to 41 additional families. Germline mutations were detected in 55 of 77 Japanese families using SSCP, direct sequencing, and Southern Blot analysis of peripheral blood samples. The Y112N (c.334T>A) germline missense mutation of the VHL gene was found in 2 VHL family members with hemangioblastomas of the central nervous system, retinal angiomas, pancreatic cysts or tumors (unspecified) (family ID 74).

A case report of a 51Y male patient presenting with headaches and a family history of VHL disease revealed multiple CNS hemangioblastomas, bilateral clear cell renal cell carcinomas (ccRCC), pheochromocytoma, and a pancreatic neuroendocrine tumor. A germline mutation of VHL p.Arg167Trp was confirmed. The patient underwent resection of the largest cerebellar lesion, total right and partial left sided nephrectomy, and adrenalectomy. Pazopanib monotherapy was initiated for treatment of metastatic RCC. The patient experienced shrinkage of some but not all spinal hemangiolastomas. After 21 months on pazopanib, the patient was placed on a drug holiday, and many of the CNS hemangioblastomas grew slightly.

37 Danish patients diagnosed with VHL before 18 years from a prospective cohort study from Denmark's National vHL Research Database (Binderup et al., 2013) were assessed. One family with 4 patients (Family ID 3) were found with a germline missense mutation in the VHL gene (G93D; c.278G>A). Patient 1 was diagnosed at 7Y and had no manifestations by the end of the study at 18Y old. Patient 2 and 3 were both diagnosed at 9Y and and both developed pheochromocytoma at age 15Y. They were both 20Y at the end of the study. Patient 4 was diagnosed at 10Y and had 2 pheochromocytomas at age 12Y, and 1 retinal hemangioblastoma at age 10Y. They were 21Y at the end of the study.

37 Danish patients diagnosed with VHL before 18 years from a prospective cohort study from Denmark's National vHL Research Database (Binderup et al., 2013) were assessed. Parts of the Danish cohort have been previously assesed in the following papers PMID: 19863552, 25834951, 27966541, 27539272. Peripheral blood samples were isolated and sequenced coding exons and exon-intron boundaries using MLPA (Multiplex Ligation-dependent Probe Amplification). One family with one patient (Family ID-10) was found with a germline truncating mutation in the VHL gene (R113*; c.337C>T). Patient 1 was diagnosed at 15Y and had two pancreatic cysts at 15Y, one retinal hemangioblastoma at 17Y, and one epididymal cystadenoma at 17Y. The patient was 37Y by the end of the study.

This conference abstract presented a 9 year old, previously healthy, male with bilateral pheochromocytoma who was tested and revealed a p.V166F protein change. No specific nucleotide change was reported, however this could be deduced as p.V166F (c.496G>T). Testing of parents and siblings revealed that this was a de novo mutation, as they were esch negative for the mutation.

Of 65 VHL families from central Europe, 53 were identified with germline mutations using SSCP analysis of PCR products by peripheral blood. The Q164 (c.490C>T) mutation was detected in the patient, referred to in the paper by Q164 (c.703C>T) due to a difference in transcript sequence. This nonsense mutation was found in an Iranian, VHL type 1 patient with retinal angiomas, hemangioblastomas of the central nervous system, renal cell carcinoma, pancreatic cysts, and pancreatic islet cell tumors (family VHL 65). This study contains very strong evidence of pathogenicity in the form of a nonsense variant in a gene where loss-of-function is a known mechanism of disease (ACMG code: PVS1).

Of 65 VHL families from central Europe, 53 were identified with germline mutations. This missense mutation was found in 2 unrelated, German families. The mutation of V74G (c.221T>G) was referred to in the paper by c.434T>G due to a difference in transcript sequence. The VHL type 1 family of 4 affected individuals had 2 patients with retinal angiomas, 5 with hemangioblastomas of the central nervous system, and one with renal cyst and noted with a primitive neuroectodermal tumor (family VHL 53). The VHL type 2B family of 3 affected individuals had one patient with retinal angioma, 3 with hemangioblastomas of the central nervous system, one with pheochromocytoma, one with renal cell carcinoma and renal cysts, one with pancreatic cysts, and one with cystadenomas of the epididymis (family VHL 9). This study contains supporting evidence of pathogenicity due to cosegregation with disease in multiple affected family members in a gene definitively known to cause the disease (ACMG code: PP1).

Of 65 VHL families from central Europe, 53 were identified with germline mutations using SSCP analysis of PCR products by peripheral blood. This missense mutation was found in a German, VHL type 1 patient with retinal angiomas (family VHL 59). The H115Y (c.343C>T) mutation was detected in the patient, referred to in the paper by c.556C>T due to a difference in transcript sequence.

Of 65 VHL families from central Europe, 53 were identified with germline mutations using SSCP analysis of PCR products by peripheral blood. This mutation was found in 2 Croatian, VHL type 1 family members (family VHL 26). Both have hemangioblastomas of the central nervous system, one has renal cysts, and one with pancreatic cysts.

The authors report this as an AF010238.1:c.443insTCT mutation at codon 77 (insertion Ser). This does not match the associating c. and p. - NM_000551.3:c.230_231insTCT matches Cys77_Asn78insLeu., while c.231_232insTCT (p.Cys77_Asn78insSer) would give the ins Ser the authors write, however this doesn’t match the p.. Without confirmation from authors no assumption could be made.

Medical records of 31 Asian Indian genetically proven VHL patients with pheochromocytoma/paraganglioma were studied. A 55 year old female (Patient 18) presented with sympathetic paraganglioma (55Y). Genetic testing (PCR and Sanger sequencing) confirmed a missense mutation at c.584C>G of the VHL gene. She has a negative family history.

Medical records of 31 Asian Indian genetically proven VHL patients with pheochromocytoma/paraganglioma were studied. Two unrelated patients were genetically confirmed (PCR and Sanger sequencing) to have a germline frameshift mutation at c.224_226delTCT of the VHL gene. Patient 21 (M): presented with unilateral pheochromocytoma (29Y), pancreatic cysts (29Y), renal cell carcinoma (29Y) and a central nervous system hemangioblastoma (29Y). He has a negative family history. Patient 26 (F): presented with unilateral pheochromocytoma (45Y) and a negative family history.

A retrospective cohort study describes phenotypic variability within families with von Hippel-Lindau disease. Patients with genetically confirmed VHL were identified, although methods were not stated. 27 patients belonging to 5 families were discussed. The nonsense mutation S65* (c.189_192del) was found in both members belonging to family 4. Both had either a cerebral or spinal hemangioblastoma.

A retrospective study of metastatic (n = 10) or unresectable (n=5) PPGL patients who received Lu-DOTATATE therapy (registered at KEM Hospital, Mumbai, India) between January 2010 and December 2019 were examined. Two VHL mutations were found through genotyping based on the protocol from PMID: 27539324. One male aged 14Y when therapy started, had pheochromocytoma and pancreatic neuroendocrine tumor which were considered progressive and inoperable. He had a R167W (c.499C>T) VHL mutation. No family information was given.

Among patients prospectively followed by the members of the French VHL Study Group, 35 consecutive patients with well-documented endocrine pancreatic tumour (20 women and 15 men; median age,37 [21Y-57] years) were included. The data from 7 patients with EPT have been partially reported previously in Hammel et al., 1995;2000. All 35 patients had VHL germline mutations and were assessed for the presence of pheochromocytoma and renal carcinome in addition to pancreatic tumours. The presence of other phenotypes was not described. A germline splice mutation (c.340+1G>A) was found in patient no 23 from family 582. The patient presented with pancreatic endocrine tumour and renal carcinoma. The patient's phenotypes are highly specific to VHL disease (PP4) and a splice alternation may be supportive of PVS1.

Among patients prospectively followed by the members of the French VHL Study Group, 35 consecutive patients with well-documented endocrine pancreatic tumour (20 women and 15 men; median age,37 [21Y-57] years) were included. The data from 7 patients with EPT have been partially reported previously in Hammel et al., 1995;2000. All 35 patients had VHL germline mutations and were assessed for the presence of pheochromocytoma and renal carcinoma in addition to pancreatic tumours. The presence of other phenotypes was not described. A missense mutation (c.334T>C) was found in 2 members of the same family. Patient No. 10 and patient no. 17 from family 360 both presented with pancreatic endocrine tumour. Their familial relation and other family members were not described therefore there may not be sufficient evidence for PP1. However, patients' phenotypes are highly specific to VHL disease (PP4) and disease-causing missense variants are common in VHL disease (PP2).

Among patients prospectively followed by the members of the French VHL Study Group, 35 consecutive patients with well-documented endocrine pancreatic tumour (20 women and 15 men; median age,37 [21Y-57] years) were included. The data from 7 patients with EPT have been partially reported previously in Hammel et al., 1995;2000. All 35 patients had VHL germline mutations and were assessed for the presence of pheochromocytoma and renal carcinoma in addition to pancreatic tumours. The presence of other phenotypes was not described. Patient no. 20 from family 900 had a frameshift mutation in the VHL gene (c.284delC) and presented with pancreatic endocrine tumour. The patients' phenotypes are highly specific to VHL disease (PP4) and frameshift mutation supports PM4.

Among patients prospectively followed by the members of the French VHL Study Group, 35 consecutive patients with well-documented endocrine pancreatic tumour (20 women and 15 men; median age,37 [21Y-57] years) were included. The data from 7 patients with EPT have been partially reported previously in Hammel et al., 1995;2000. All 35 patients had VHL germline mutations and were assessed for the presence of pheochromocytoma and renal carcinoma in addition to pancreatic tumours. The presence of other phenotypes was not described. Patient no. 31 from family 463 had a missense mutation in the VHL gene (c.551T>A) and presented with pancreatic endocrine tumour and pheochromocytoma. The patient's phenotypes are highly specific to VHL disease (PP4) and disease-causing missense variants are common in VHL disease (PP2).

All patients included in the study were VHL mutation carriers of 18 years or older examined between 1972 and 2012 in the VHL expertise centers University Medical Center Groningen (UMCG) and University Medical Center Utrecht (UMCU). Clinical data of 75 VHL mutation carriers were retrospectively retrieved from electronic and written patient record. Of these patients, one was found to harbour the c.208G>A germline mutation of the VHL gene. They presented with a retinal hemangioblastoma. Their age at the last follow-up was 20. Previously published in: PMID 24132471.

21 consenting patients treated for pheochromocytoma over a 10-yr period at the University of Freiburg Medical Center were affected by VHL and donated peripheral blood for germline DNA analyses. DNA was sequenced using single-strand conformation polymorphism (SSCP) analysis of all three exons of the VHL gene. Three of these 21 patients had a p.Leu188Val germline mutation, (c.775C>G legacy notation converted to c.562C>G). Patients 19 and 21 had a single benign pheochromocytoma while patient 20 had three benign pheochromocytomas. The three patients were related. No other patient data and demographics was available.

Direct sequencing and multiplex ligation dependent probe amplification were performed on thirteen unrelated subjects with VHL mutations. The clinical manifestations and family histories of the subjects were evaluated. A 38 year old Korean male patient presented with imbalance, gait disturbance and cerebellar hemangioblastoma, multiple pancreatic cysts and renal cysts. Genetic testing confirmed a splice mutation at c.464-1G > T of the VHL gene. Family history is unknown.

Through PCR purification and Sanger sequencing of peripheral blood (using an alcohol precipitation method), 121 consecutive, unrelated, index PCC/PGL patients underwent genetic testing for five PCC/PGL susceptibility genes (RET, VHL, SDHB, SDHD and SDHC) and were evaluated for clinical diagnosis of neurofibromatosis type1. A 20 year old female was genetically confirmed with a familial mutation at c.588_588dupA of the VHL gene. She presented with bilateral pheochromocytoma. Familial/syndromic (FS) association although further family history is unknown.

Through PCR purification and Sanger sequencing of peripheral blood (using an alcohol precipitation method), 121 consecutive, unrelated, index PCC/PGL patients underwent genetic testing for five PCC/PGL susceptibility genes (RET, VHL, SDHB, SDHD and SDHC) and were evaluated for clinical diagnosis of neurofibromatosis type1. A 15 year old male was genetically confirmed with a familial mutation at c.293A>C of the VHL gene. He presented with a sympathetic paraganglioma at age 12, and unilateral pheochromocytoma and retinal angioma (likely age 15). Further family history is unknown.

A retrospective study described polyglobulia in 19 patients precise germline VHL mutations and with hemangioblastomas. 10 other patients followed with VHL mutations without an exact location. 3 patients were reported with a p.Y98H (c.292T>C) mutation, described in the paper as c.505T>C due to a difference in transcript reference. Patient 8 was a 57Y female, patient 15 a 75Y female, and patient 16 a 56Y female. They each had a hemangioblastoma tumor 0.1, 13.6, and 0.4cm3 in size, respectively. Patients 8 and 15 also had an unspecified tumor cyst. No family information was provided; it is unclear if these three are related.

200 patients with intracranial aneurysms were molecularly analyzed for VHL to determine if it is a candidate gene for aneurysm formation using peripheral blood. One patient was analyzed in detail and determined to have the above mutation (341-6C>T). The patient was only recorded to have an intracranial aneurysm (or dilatation of the cerebral artery).

Tissue analysis from 61 VHL patients, 30 sporadic renal cell carcinoma (RCC) tumor samples, and 6 sporadic RCC cell lines revealed 22 variants within VHL patients, 10 variants in tumor samples and variants in all cell lines. All mutations cluster to the 3’ end of the VHL gene open reading frame, implicating this region as important for the tumor-suppressor function of the VHL protein. Northern blot, SSCP and RT-PCR analysis from normal tissue found a germline p.Q145Hfs*28 (c.435_436delGC) mutation, stated in the paper as c.648_649delGC due to an older transcript. It was seen in family [pedigree No] 9, with an unknown number of members described as ‘VHL-affected’ with RCC. Other manifestations are possible but not clear.

Tissue analysis from 61 VHL patients, 30 sporadic renal cell carcinoma (RCC) tumor samples, and 6 sporadic RCC cell lines revealed 22 variants within VHL patients, 10 variants in tumor samples and variants in all cell lines. All mutations cluster to the 3’ end of the VHL gene open reading frame, implicating this region as important for the tumor-suppressor function of the VHL protein. Northern blot, SSCP and RT-PCR analysis from normal tissue found a germline p.76delF (c.227_229delTTC) mutation, stated in the paper as c.437_439delF due to an older transcript. It was seen in two families: [pedigree No] 17 and 20, with an unknown number of members described as ‘VHL-affected’ with RCC. Other manifestations are possible but not clear.

Nine VHL patients (3 female, 6 male; mean age, 42 years; range, 29–71 years) with pancreatic cystic lesions were selected from the group of familial VHL patients followed on the Institutional Review Board-approved protocol at the National Cancer Institute of the National Institutes of Health. Patient No 7 was a 29Y male with pancreatic cyst; hemangioblastoma; retinal angioma; pheochromocytoma; renal cell carcinoma; pancreatic neuroendocrine tumour. This mutation of (c.486C>G; p.Cys162Trp) was referred to in the paper by c.699C>G due to a difference in transcript sequence. Family history was not described.

Thirteen patients from four families (A, B, C and D) with known VHL disease and known mutations in the VHL gene were examined. MLPA using venous blood showed 4 variants. Family D was found with a p.R64P mutation in the VHL gene, only confirmed in patient II-1 (female) and neither parent or sibling, suggested a de novo mutation. Patient II-1 presented with pheochromocytoma in the right adrenal gland at age 37, and by age 45, she showed no signs of metastases or other VHL-associated tumors.

Families with VHL disease type 2A were evaluated for mutations in the VHL gene. In four families, three different coding sequence substitutions causing a 15 amino-acid extension from the original stop codon of the VHL gene were described. Genomic DNA was extracted from peripheral blood lymphocytes using standard procedures. Sequence analysis consisted of polymerase chain reaction (PCR) amplification and automated fluorescence sequencing of all three VHL exons. Patient 1 (family 1), female, presented with a pheochromocytoma at age 11, a pheochromocytoma at age 20, bilateral retinal hemangioblastoma at age 20, and a nodal metastatic pancreatic neuroendocrine tumor at age 20. She had a family history of early onset pheochromocytoma. She was found to have a substitution mutation in the VHL gene causing a loss of the stop codon, and an extension of 14 amino acids on the C-terminal end of the protein c.641G>T (p.*214Lext*14). This mutation is noted as p.*214Lext*15 in the paper due to a difference in nomenclature.) Her mother was found to have the same mutation. Her mother had two pheochromocytomas at age 12, and was found to have bilateral retinal hemangioblastomas at age 40. Her brother was found to have a pheochromocytoma at age 13, but was not sequenced. Her maternal grandfather was noted to have had liver cancer at 72, her great grandfather had unspecified cancer, and her great aunt had Hodgkin's lymphoma at age 38. A second patient (patient 2) from an independent family was found to have the same mutation as patient 1. Patient 2 was also female and presented with bilateral retinal hemangioblastomas. Her last recorded age was 38 years old, and no age of diagnosis was provided.

A case study of a 14Y boy was found with a juxtapapillary retinal hemangioblastoma and secondary neovascularization. 16 months later, genetic analysis revealed an in-frame mutation in the VHL gene (p.Phe76del). The patient and his family was still being investigated for other manifestations during the time of the publication.

Eighteen VHL patients who underwent pancreatectomy for one or several endocrine tumors at Beaujon hospital from 1997 to 2008 were retrieved from the file of VHL patients. All patients had VHL germline mutations. Age is at the time of surgery. Patients #8 (male) was found with this germline VHL mutation (p.N131K (c.393C>A)) and pancreatic endocrine tumours. The last known age of the patient was 26Y. Other phenotypes and demographics were unknown. ACMG PP4

This study reports 1,548 germline and somatic mutations from 945 VHL families, including 30 previously unpublished kindreds from The Netherlands (six novel mutations). This mutation was detected by direct sequencing and MLPA. This missense mutation (c.407T>C) was found in a VHL patient with renal cell carcinoma (case no. 481). ACMG evidence codes: 'PP2' because they observe a missense variant in a gene where missense variants are a common mechanism of disease.

This study reports 1,548 germline and somatic mutations from 945 VHL families, including 30 previously unpublished kindreds from The Netherlands (six novel mutations). This mutation was detected by direct sequencing and MLPA. This frameshift mutation (c.351del) was detected as sporadic germline in case no. 198 with hemangioblastoma of the central nervous system. The case phenotypes are highly specific for a disease with a single genetic etiology (ACMG code: PP4).

A retrospective cohort study included all the VHL patients diagnosed at Peking University First Hospital (Beijing, China) prior to June 1 2016. A total of 291 patients from 115 different families were included for analysis. This familial mutation was confirmed in family #52 with 9 affected members who were all genetically confirmed with the mutation. A male proband was diagnosed with CNS hemangioblastoma at 28Y, RCC at 28Y, and multiple pancreatic lesions at 28Y. His son and daughter of unknown ages are asymptomatic. His mother was diagnosed with CNS hemangioblastoma at 66Y, RCC at 69Y, and multiple pancreatic lesions at 66Y. His first sister was diagnosed with CNS hemangioblastoma at 32Y, RCC at 36Y, and multiple pancreatic lesions at 36Y. His nephew and niece (son and daughter of first sister) of unknown ages are asymptomatic. His second sister was diagnosed with CNS hemangioblastoma at 39Y, RCC at 39Y, and multiple pancreatic lesions at 39Y. His nephew (son of second sister) of unknown age is asymptomatic. This cohort was also published in a paper by Wang et al., 2018 (PMID: 29330336). It is likely that both papers published the same family.

A retrospective cohort study included all the VHL patients diagnosed at Peking University First Hospital (Beijing, China) prior to June 1 2016. A total of 291 patients from 115 different families were included for analysis. This familial mutation was confirmed in family #50. Only the proband (patient 132) was genetically confirmed with the mutation. The proband (female) was diagnosed with a CNS hemangioblastoma at 26Y, renal cell carcinoma at 45Y, and multiple pancreatic cysts/tumors at 45Y. Her father (patient 135) was diagnosed with CNS hemangioblastoma at 25Y, but passed away before VHL disease diagnosis/ mutation confirmation. Her first brother (patient 133) was diagnosed with CNS hemangioblastoma at 29Y, RCC at 48Y, and multiple pancreatic cysts/tumors at 48Y, but no DNA sample was available for testing. Her second brother (patient 134) was diagnosed with CNS hemangioblastoma at 27Y, but passed away before VHL disease diagnosis/ mutation confirmation. This kindred was also published in a paper by Wang et al., 2018 (PMID: 29330336).

A retrospective cohort study included all the VHL patients diagnosed at Peking University First Hospital (Beijing, China) prior to June 1 2016. A total of 291 patients from 115 different families were included for analysis. This familial mutation was confirmed in family #43 with 2 affected members that were both genetically confirmed for the mutation. A female proband was diagnosed with CNS hemangioblastoma at 19Y and multiple pancreatic lesions at 26Y. Her father was diagnosed with CNS hemangioblastoma at 27Y, renal cell carcinoma at 48Y, and multiple pancreatic lesions at 60Y. This cohort was also published in a paper by Wang et al., 2018 (PMID: 29330336). It is likely that both papers published the same family.

A retrospective cohort study included all the VHL patients diagnosed at Peking University First Hospital (Beijing, China) prior to June 1 2016. A total of 291 patients from 115 different families were included for analysis. This familial mutation (p.P97Afs*35, c.287_288insA) was confirmed in family #33, with 2 affected members that were both genetically confirmed for the mutation. A male proband (patient 99) was diagnosed with CNS hemangioblastoma at 37Y, retinal angioma at 20Y, renal cell carcinoma at 20Y, and multiple pancreatic cysts or tumours at 29Y. His brother (patient 100) was diagnosed with retinal angioma at 32Y, renal cell carcinoma at 43Y, and multiple pancreatic cysts or tumours at 43Y. This cohort was also published in a paper by Wang et al., 2018 (PMID: 29330336).

A retrospective cohort study included all the VHL patients diagnosed at Peking University First Hospital (Beijing, China) prior to June 1 2016. A total of 291 patients from 115 different families were included for analysis. This familial mutation was confirmed in family #48. Only the male proband (patient 128) was genetically confirmed with the mutation. He was diagnosed with CNS hemangioblastoma at 32Y, RCC at 38Y, pheochromocytoma at 40Y and multiple pancreatic lesions at 38Y. His father (patient 129) was diagnosed with RCC at 58Y and his daughter (patient 130) was diagnosed with CNS hemangioblastoma at 13Y. This cohort was also published in a paper by Wang et al., 2018 (PMID: 29330336).

Molecular and clinical analysis of 126 French VHL families tested for renal involvement revealed 92 different mutations. 90 of these families had renal involvement. The p.S65P (c.193T>C) germline variant of the VHL gene was detected in a family (ID 185). Member 9359 has unknown renal involvement. Member 9357 presented with mild renal disease. Member 1052 had no renal involvement. Member 3979 has moderate renal disease (RCC). The paper states that this family is previously reported in PMID 12202531, where 2 patients with this mutation are reported but relation between patients is unknown.

Molecular and clinical analysis of 126 French VHL families tested for renal involvement revealed 92 different mutations. 90 of these families had renal involvement. The p.C162Y (c.485G>A) germline variant of the VHL gene was detected in a family (ID 17). Member 753 presented with moderate renal disease (RCC), member 11338 had unknown renal involvement, member 696 presented with severe renal disease (RCC) and members 2368 and 699 presented with mild renal disease.

Molecular and clinical analysis of 126 French VHL families tested for renal involvement revealed 92 different mutations. 90 of these families had renal involvement. The p.R69Afs*63 (c.204dup) germline variant of the VHL gene was detected in an individual (ID 6321, Family 343). This patient presented with renal cell carcinoma. No family information is provided.

Molecular and clinical analysis of 126 French VHL families tested for renal involvement revealed 92 different mutations. 90 of these families had renal involvement. The p.K159delinsI* (c.475_476insTTT) germline variant of the VHL gene was detected in a family (ID 43). Member 3207 had no renal involvement. Members 3206 and 3203 presented with mild renal disease. Member 3189 presented with severe renal disease (RCC). The paper states that this family is previously reported in PMID 12202531 and/or 9829912.

Molecular and clinical analysis of 126 French VHL families tested for renal involvement revealed 92 different mutations. 90 of these families had renal involvement. The p.N67_V74del (c.197_221del) germline variant of the VHL gene was detected in a family (ID 287). Member 6396 presented with mild renal disease. Member 6611 presented with severe renal disease (RCC). The paper states that this family is previously reported in PMID 9829912, where 1 patient with this mutation is reported but relation to this patient is unclear. The variant is reported as c.197_220del which was mapped to c.198_221del.

Molecular and clinical analysis of 126 French VHL families tested for renal involvement revealed 92 different mutations. 90 of these families had renal involvement. The p.K159Efs*15 (c.473dup) germline variant of the VHL gene was detected in an individual (ID 5299, Family 235). They presented with mild renal disease (simple renal cyst, multicystic kidney[s], small lesion [less than 10 mm in size] of unknown significance, or small solid tumor without indication for surgical removal [less than 30 mm in size]). Individual RCC phenotype is noted as negative. The paper states that this individual is previously reported in PMID 12202531 and/or 9829912.

Molecular and clinical analysis of 126 French VHL families tested for renal involvement revealed 92 different mutations. 90 of these families had renal involvement. The p.Y156* (c.468T>G) germline variant of the VHL gene was detected in a family (ID 3). Member 37 had no renal involvement. Member 38 presented with moderate renal disease (RCC). Members 47 and 3 presented with severe renal disease (RCC). The paper states that this family is previously reported in PMID 12202531 and/or 9829912.

The aim of this study was to establish the characteristics of pheochromocytoma in a population of 58 patients between 4 and 20 years of age studied at Hospital de Niños Ricardo Gutiérrez. They represented a 23% of the total population of 255 pheochromocytoma patients studied. Two children belonging to the same family had VHL type 2A and presented with the missense mutation Arg167Trp. The children both had pheochromocytomas.

This paper shows the imaging result of an endolymphatic sac tumor found in the left temporal bone of an 8-year-old boy who presented with profound sensorineural hearing gloss, intermittent vertigo. VHL mutation analysis showed c.388G>C mutation (noted as G-to-C nucleotide 601 mutation in the paper).

216 patients with clinically expected VHL disease due to family history or presence of VHL typical tumours were routinely examined in an eye centre in Germany between January 2019 and January 2020, majority of which were of European descent (~95%). Since data on the genotype–phenotype correlation of the onset of Retinal Hemangioblastoma (RH) is scarce in relation to VHL disease, this study wanted to inversitage the relationship between VHL gene variants (truncating vs single amino acid substitutions) and the ocular phenotype of the disease. 42 different rare VHL gene variants in 148 patients were detected - some of whom had RH and some who did not. Although date of onset of RH, as well as data on central nervous system haemangioblastomas (CNSH), pheochromocytoma/paraganglioma (PPGL), clear cell renalcell carcinoma (ccRCC) and pancreatic neuroendocrine neoplasm (PNEN) were collected, it was not reported on a per patient basis and therefore could not be discussed per variant. There was 1 patient reported with the c.269A>T (p.N90I) mutation. This member presented with retinal hemangiomas. No family information was provided.

216 patients with clinically expected VHL disease due to family history or presence of VHL typical tumours were routinely examined in an eye centre in Germany between January 2019 and January 2020, majority of which were of European descent (~95%). Since data on the genotype–phenotype correlation of the onset of Retinal Hemangioblastoma (RH) is scarce in relation to VHL disease, this study wanted to inversitage the relationship between VHL gene variants (truncating vs single amino acid substitutions) and the ocular phenotype of the disease. 42 different rare VHL gene variants in 148 patients were detected - some of whom had RH and some who did not. Although date of onset of RH, as well as data on central nervous system haemangioblastomas (CNSH), pheochromocytoma/paraganglioma (PPGL), clear cell renal cell carcinoma (ccRCC) and pancreatic neuroendocrine neoplasm (PNEN) were collected, it was not reported on a per patient basis and therefore could not be discussed per variant. There were 2 patients reported with the p.V74Sfs*85 (c.220del) mutation. Both members presented with retinal hemangiomas, while no other clinical manifestations were discussed. Additionally, no family information was provided.

216 patients with clinically expected VHL disease due to family history or presence of VHL typical tumours were routinely examined in an eye centre in Germany between January 2019 and January 2020, majority of which were of European descent (~95%). Since data on the genotype–phenotype correlation of the onset of Retinal Hemangioblastoma (RH) is scarce in relation to VHL disease, this study wanted to inversitage the relationship between VHL gene variants (truncating vs single amino acid substitutions) and the ocular phenotype of the disease. 42 different rare VHL gene variants in 148 patients were detected - some of whom had RH and some who did not. Although date of onset of RH, as well as data on central nervous system haemangioblastomas (CNSH), pheochromocytoma/paraganglioma (PPGL), clear cell renal cell carcinoma (ccRCC) and pancreatic neuroendocrine neoplasm (PNEN) were collected, it was not reported on a per patient basis and therefore could not be discussed per variant. There were 3 patients reported with the p.F76del (c.227_229del) mutation. All 3 members presented with retinal hemangiomas, while no other clinical manifestations were discussed. Additionally, no family information was provided.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (S111R (c.333C>G)) was taken from the National Cancer Institute database, with an unknown number of members belonging to Family ID 4401NCI. It was reported in the paper as c.546C>G due to a difference in transcript reference. As reported in Table 3, this mutation is found in VHL families without pheochromocytoma, and with renal cell carcinoma.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (F136C (c.407T>G)) was taken from the Bristol Myers Squibb database, with an unknown number of members belonging to Family ID 10BMS. It was reported in the paper as c.620T>G due to a difference in transcript reference. As reported in Table 4, this mutation is found in VHL families with pheochromocytoma. Other phenotypes are likely, including renal cell carcinoma, however not detailed here.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (L135* (c.404T>A)) was taken from the National Cancer Institute database, and the Bristol Myers Squibb database. It had an unknown number of members belonging to Family ID 3710NCI, reported in Table 3, as being found in VHL families without pheochromocytoma, and with renal cell carcinoma. Additionally, it was reported in table 5, specified as families without detailed clinical information, under family ID 8BMS. It was reported in the paper as c.617T>A due to a difference in transcript reference.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (Y112* (c.336C>A)) was taken from the National Cancer Institute database, with an unknown number of members belonging to Family IDs 3759NCI and 3760NCI. It was reported in the paper as c.549C>A due to a difference in transcript reference. As reported in Table 3, this mutation is found in VHL families without pheochromocytoma, and with renal cell carcinoma. Later publications reference this cohort as being reported in PMID: 7728151.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (A56fs (c.165_166insA)) was taken from the Cambridge, Department of Pathology database, with an unknown number of members belonging to family ID 146CA. It was reported in the paper as c.378insA due to a difference in transcript reference. Table 5 lists all ID and mutations associated with individuals without detailed clinical information. Other phenotypes are possible, however not detailed here.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (c.468T>G) was taken from the Neuro Olcologie database in Paris, with an unknown number of members belonging to Family ID 41F. It was reported in the paper as c.681T>G due to a difference in transcript reference. As reported in Table 3, this mutation is found in VHL families without pheochromocytoma, and with renal cell carcinoma.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (L188fs (c.563delT)) was taken from the Cambridge, Department of Pathology database, with an unknown number of members belonging to Family ID 141CA. It was reported in the paper as c.776delT due to a difference in transcript reference. As reported in Table 3, this mutation is found in VHL families without pheochromocytoma, and with renal cell carcinoma.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (L153fs (c.457delC)) was taken from the Cambridge, Department of Pathology database. It had an unknown number of members belonging to Family ID 41CA, reported in Table 3, as being found in VHL families without pheochromocytoma, and with renal cell carcinoma. Additionally, it was reported in table 5, specified as families without detailed clinical information, under family ID 99CA. It was reported in the paper as c.670delC due to a difference in transcript reference.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (K159fs (c.473dupT)) was taken from the Bristol Myers Squibb database, with an unknown number of members belonging to Family ID 6BMS. It was reported in the paper as c.685insT due to a difference in transcript reference. Table 5 lists all ID and mutations associated with individuals without detailed clinical information. Other phenotypes are possible, however not detailed here.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (R82_V84del (c.243_251del)) was taken from the Cambridge, Department of Pathology database, with an unknown number of members belonging to Family ID 35CA. It was reported in the paper as c.456delRVV due to a difference in transcript reference. As reported in Table 3, this mutation is found in VHL families without pheochromocytoma, and with renal cell carcinoma.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (c.464-2A>C) was taken from the Bristol Myers Squibb database, with an unknown number of members belonging to Family ID 4BMS. It was reported in the paper as c.677-2A>C due to a difference in transcript reference. As reported in Table 3, this mutation is found in VHL families without pheochromocytoma, and with renal cell carcinoma.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (H115Y (c.343C>T)) was taken from the Munich, university of Freiburg and Institute of Pathology database, with an unknown number of members belonging to Family ID 59CE. It was reported in the paper as c.556C>T due to a difference in transcript reference. As reported in Table 3, this mutation is found in VHL families without pheochromocytoma, and with renal cell carcinoma.

Germline mutational data on VHL was gathered from various countries to assess variation in germline mutations that produce disease within different populations. Families were grouped according to three possible outcomes of disease: renal cell carcinoma without pheochromocytoma, renal cell carcinoma with pheochromocytoma, or pheochromocytoma alone. SSCP and denaturing gradient gel electrophoresis identified mutations in 300/469 (63%) of families affected with VHL. This germline familial VHL mutation (S72Pfs*87 (c.214delT)) was taken from the Cambridge, Department of Pathology database, with an unknown number of members belonging to Family ID 112CA. It was reported in the paper as c.427delT due to a difference in transcript reference. As reported in Table 3, this mutation is found in VHL family without pheochromocytoma, and with renal cell carcinoma.

A cohort of 64 pheochromocytoma and paragangliomas were screened for a BRAF mutation using direct Sanger sequencing and QRT-PCR. Genomic DNA was extracted from fresh or frozen peripheral blood. Germline mutations of VHL, SDHB, SDHD, and RET genes were also observed. The VHL mutation c.374A>C (p.His125Pro) was reported in patient #42, a 24Y male with bilateral pheochromocytoma. No family history was reported here.

4 paediatric patients presenting with pheochromocytomas and paragangliomas were reported. Upon genetic testing of peripheral blood leukocytes on the VHL, SDHB, SDHC, SDHC, and RET genes, 1 patient was found to have the c.374A>C germline mutation. At 13Y the patient had bilateral adrenal pheochromocytoma and retinal capillary hemangiomas. Now, 19Y, he remains in complete remission. Family screening showed the absence of the mutation in the asymptomatic mother; the father was not analyzed. Other family history was negative for VHL syndrome and PCCs.

One hundred and sixteen unrelated VHL disease patients have been investigated by molecular genetic analysis. Twenty-two of these were previously shown to have germline deletions on Southern analysis, and in the present study the remaining 94 patients were screened for intragenic mutations via SSCP and heteroduplex analysis of 855 bp of the VHL coding sequence and 800 bp of the 3' UTR. An intragenic mutation was identified in 55 (59%) out of 94 patients without germline deletions on Southern analysis. In all patients with multiple affected family members the missense mutation segregated with the disease. Large deletions or intragenic mutations predicted to cause a truncated protein were found in 36 of 53 families without pheochromocytoma (VHL type 1) and 2 of 12 families with pheochromocytoma (VHL type 2). This study also investigated the frequency of different mutation classes between kindreds with and without pheochromocytoma. The c.337C>T (p.R113) stop-gain mutation (noted as c.550C>T (p.R184) in the paper due to a difference in reference transcript used) was found in at least one patient in 2 kindreds (families): kindred no. 30 and 136. Pheochromocytoma was not observed in any members of kindred no. 136 (implying the affected patient(s) had VHL disease type 1). No phenotypic information was reported for members of kindred no. 30. ACMG evidence codes: none.

One hundred and sixteen unrelated VHL disease patients have been investigated by molecular genetic analysis. Twenty-two of these were previously shown to have germline deletions on Southern analysis, and in the present study the remaining 94 patients were screened for intragenic mutations via SSCP and heteroduplex analysis of 855 bp of the VHL coding sequence and 800 bp of the 3' UTR. An intragenic mutation was identified in 55 (59%) out of 94 patients without germline deletions on Southern analysis. In all patients with multiple affected family members the missense mutation segregated with the disease. Large deletions or intragenic mutations predicted to cause a truncated protein were found in 36 of 53 families without pheochromocytoma (VHL type 1) and 2 of 12 families with pheochromocytoma (VHL type 2). This study also investigated the frequency of different mutation classes between kindreds with and without pheochromocytoma. The p.Y98H (c.292T>C) variant (noted asp.Y169H (c.505T>C) in the paper due to a difference in reference transcript used) was found in at least one patient in kindred 22. No familial phenotypic information could be inferred from the paper.

One hundred and sixteen unrelated VHL disease patients have been investigated by molecular genetic analysis. Twenty-two of these were previously shown to have germline deletions on Southern analysis, and in the present study the remaining 94 patients were screened for intragenic mutations via SSCP and heteroduplex analysis of 855 bp of the VHL coding sequence and 800 bp of the 3' UTR. An intragenic mutation was identified in 55 (59%) out of 94 patients without germline deletions on Southern analysis. In all patients with multiple affected family members the missense mutation segregated with the disease. Large deletions or intragenic mutations predicted to cause a truncated protein were found in 36 of 53 families without pheochromocytoma (VHL type 1) and 2 of 12 families with pheochromocytoma (VHL type 2). The c.224_226del (p.F76del) in-frame deletion (noted as c.440delTCT (p.F147del) in the paper due to a difference in reference transcript used) was found in at least one patient in kindred no.48, and kindred no. 75. The affected patient(s) in both kindreds had VHL type 1 (no pheochromocytoma). Pheochromocytoma was not observed in any members of either kindred. ACMG evidence codes: none.

One hundred and sixteen unrelated VHL disease patients have been investigated by molecular genetic analysis. Twenty-two of these were previously shown to have germline deletions on Southern analysis, and in the present study the remaining 94 patients were screened for intragenic mutations via SSCP and heteroduplex analysis of 855 bp of the VHL coding sequence and 800 bp of the 3' UTR. An intragenic mutation was identified in 55 (59%) out of 94 patients without germline deletions on Southern analysis. In all patients with multiple affected family members the missense mutation segregated with the disease. Large deletions or intragenic mutations predicted to cause a truncated protein were found in 36 of 53 families without pheochromocytoma (VHL type 1) and 2 of 12 families with pheochromocytoma (VHL type 2). This study also investigated the frequency of different mutation classes between kindreds with and without pheochromocytoma. The p.R79P (c.236G>C) variant (noted as p.R150P (c.449G>C) in the paper due to a difference in reference transcript used) was found in at least one patient in kindred 150. No familial phenotypic information could be inferred from the paper.

One hundred and sixteen unrelated VHL disease patients have been investigated by molecular genetic analysis. Twenty-two of these were previously shown to have germline deletions on Southern analysis, and in the present study the remaining 94 patients were screened for intragenic mutations via SSCP and heteroduplex analysis of 855 bp of the VHL coding sequence and 800 bp of the 3' UTR. An intragenic mutation was identified in 55 (59%) out of 94 patients without germline deletions on Southern analysis. In all patients with multiple affected family members the missense mutation segregated with the disease. Large deletions or intragenic mutations predicted to cause a truncated protein were found in 36 of 53 families without pheochromocytoma (VHL type 1) and 2 of 12 families with pheochromocytoma (VHL type 2). This study also investigated the frequency of different mutation classes between kindreds with and without pheochromocytoma. The p.N131Pfs*3 (c.390_391delTA) deletion (noted as p.201FS (c.603delTA) in the paper due to a difference in reference transcript used) was found in at least one patient in kindred 64. Pheochromocytoma was not observed in any members of this kindred (implying the affected patient(s) had VHL disease type 1). No other familial phenotypic information could be inferred from the paper.

One hundred and sixteen unrelated VHL disease patients have been investigated by molecular genetic analysis. Twenty-two of these were previously shown to have germline deletions on Southern analysis, and in the present study the remaining 94 patients were screened for intragenic mutations via SSCP and heteroduplex analysis of 855 bp of the VHL coding sequence and 800 bp of the 3' UTR. An intragenic mutation was identified in 55 (59%) out of 94 patients without germline deletions on Southern analysis. In all patients with multiple affected family members the missense mutation segregated with the disease. Large deletions or intragenic mutations predicted to cause a truncated protein were found in 36 of 53 families without pheochromocytoma (VHL type 1) and 2 of 12 families with pheochromocytoma (VHL type 2). This study also investigated the frequency of different mutation classes between kindreds with and without pheochromocytoma. The L153Cfs*6 (c.457del) deletion (noted as p.224fs (c.670delC) in the paper due to a difference in reference transcript used) was found in at least one patient in kindred 41, and at least one patient in kindred 99. Pheochromocytoma was not observed in any members of kindred 41 (implying the affected patient(s) had VHL disease type 1). No familial phenotypic information could be inferred from the paper for kindred 99.

Thirty-five unrelated patients suspected of having von Hippel-Lindau disease were analyzed and 32 VHL gene variants were found in 35 patients using PCR and SSCP by peripheral blood leukocytes. An association of clear cell renal carcinoma development with a relatively high loss of structural stability in VHL protein missense mutations was discovered. This missense variant (p.R167Q) was found in 4 patients (F8, F18, F56 and F71) all with retinal capillary hemangioma. Patient F8 was 35 years old at onset and noted of familial antecedents of phaeochromocytoma. Patient F18 was 33 years old at onset, had cerebellar hemangioblastoma, pancreatic cysts. Noted of familial antecedents of phaeochromocytoma and a total of 2 affected family members. Patient F56 was 45 years old at onset, had cerebellar hemangioblastoma and renal cyst. Also noted of familial antecedents of phaeochromocytoma and a total of 2 affected family members. Patient F71 was 25 years old at onset, had cerebellar hemangioblastoma, and unilateral pheochromocytoma. Noted a total of 2 affected family members, with the family member reported with clear cell renal cell carcinoma

Thirty-five unrelated patients suspected of having von Hippel-Lindau disease were analyzed and 32 VHL gene variants were found in 35 patients using PCR and SSCP by peripheral blood leukocytes. An association of clear cell renal carcinoma development with a relatively high loss of structural stability in VHL protein missense mutations was discovered. This missense mutation (p.C162Y) was found in a patient, whose age of onset was 15 years old, with retinal hemangioblastomas, unilateral pheochromocytoma, and renal cysts (patient no. F61). Patient has family history of clear cell renal carcinoma.

Thirty-five unrelated patients suspected of having von Hippel-Lindau disease were analyzed and 32 VHL gene variants were found in 35 patients using PCR and SSCP by peripheral blood leukocytes. An association of clear cell renal carcinoma development with a relatively high loss of structural stability in VHL protein missense mutations was discovered. This nonsense mutation was found in a patient with retinal hemangioblastoma, cerebellar hemangioblastoma, and pancreatic cysts (patient no. F3). Patient has family history of paragangliomas, epididymial cysts, and clear cell renal carcinoma. Paper gives variant by protein sequence: L135*. NM_000551.3:c.404T>A and NM_000551.3:c.404T>G both could result in this protein change. Unclear which is correct to the given patient.

Thirty-five unrelated patients suspected of having von Hippel-Lindau disease were analyzed and 32 VHL gene variants were found in 35 patients using PCR and SSCP by peripheral blood leukocytes. An association of clear cell renal carcinoma development with a relatively high loss of structural stability in VHL protein missense mutations was discovered. This missense variant (p.L128R) was found in a 22 year old patient with retinal and cerebellar hemangioblastomas, renal cysts, and pancreatic cysts (patient no. F41). Familial antecedents were found but unknown of any additional features than the index case.

53 patients with a molecularly confirmed VHL mutation (by PCR through EDTA-anticoagulated blood) and a pancreatic neuroendocrine tumor (ICT) were collected from the German NET-Registry and the German VHL-registry. An additional 296 patients were stated in this paper with a VHL associated germline mutation, but without pancreatic neuroendocrine tumors. 2 patients were found to have the above mutation (c.362A>G). 1 was a VHL patient with an ICT (pancreatic neuroendocrine tumor) taken from the VHL-registry. The other patient was stated to be in this study with the given mutation, however no ICT. Other clinical manifestations outside of a pancreatic neuroendocrine tumor are possible, but were not discussed. ACMG evidence codes: 'PP2' because they observe a missense variant in a gene where missense variants are a common mechanism of disease.

53 patients with a molecularly confirmed VHL mutation (by PCR through EDTA-anticoagulated blood) and a pancreatic neuroendocrine tumor (ICT) were collected from the German NET-Registry and the German VHL-registry. An additional 296 patients were stated in this paper with a VHL associated germline mutation, but without pancreatic neuroendocrine tumors. 3 patients were found to have the above mutation (c.240T>G). 1 was a VHL patient with an ICT (pancreatic neuroendocrine tumor) taken from the VHL-registry. The other 2 patients were stated to be in this study with the given mutation, however no ICT. Other clinical manifestations outside of a pancreatic neuroendocrine tumor are possible, but were not discussed. ACMG evidence codes: 'PP2' because they observe a missense variant in a gene where missense variants are a common mechanism of disease.

314 Chinese patients with Pheochromocytoma/paraganglioma (PPGL) from Peking Union Medical College Hospital, China, were included in this study for germline profiling. The majority of patients underwent surgery and were histopathologically diagnosed with PPGL. For patients who had an unresectable or metastatic tumor, PPGL diagnosis was determined based on clinical features (headache, palpitations, sweating, etc). Genomic DNA was extracted from peripheral blood of all 314 PPGL patients, and detect potential variants in 12 known genes associated with hereditary PPGL (SDHA, SDHB, SDHC, SDHD, SDHAF2, FH, VHL, RET, NF1, MAX, TMEM127, and KIF1B) using next-generation and Sanger sequencing. Germline variants were detected in 29% of the 314 PPGL patients. Germline genetic testing revealed a synonymous P138P(c.414A>G) variant in exon 2 of the VHL gene, in 2 patients, 2 years after their diagnosis. Patient #86 is a male PPGL patient who first presented with bilateral pheochromocytoma at age 15 and was also diagnosed at age 15. They also have a family history of PPGL, which was not detailed further. Patient #87 is a male PPGL patient who first presented with pheochromocytoma at age 36 and was also diagnosed at age 36. Patient 87 does not have family history of PPGL.

This study investigated the effectiveness of whole exome sequencing (WES) and an in-house hybridization based gene panel (ENDOGENE Panel v2.0) at detecting mutations in pheochromocytoma/paraganglioma (PPGL) related genes. The ENDOGENE panel included EGLN1, EPAS1, FH, KIF1B, MAX, MEN1, NF1, RET, SDHA, SDHB, SDHC, SDHD, SDHAF2, TMEM127, VHL, GOT2, MDH2 and SLC25A11. WES and the ENDOGENE panel were tested on 37 samples with known variants. After optimization of the bioinformatic pipeline, 61 additional samples were tested prospectively. VHL mutations were identified in 3 patients and all were confirmed by Sanger sequencing of blood. Patient 14 was found to have a F136C(c.407T>G) missense variant and diagnosed with VHL.

The proband (II/1), of Italian descent, was found to have the c.481C>T change (referred to in the paper as 8684 C->T due to a difference in transcript sequence) causing a p.R161* mutation in the VHL gene. This variant was detected using direct sequencing of PCR products obtained from genomic DNA extracted from the patients peripheral blood leukocytes using the DNA extractor BIO ROBOT EZ 1 (Qiagen). Genetic testing revealed that her parents did not have the VHL gene mutation. She presented with a cerebellar hemangioblastoma at age 33. ACMG evidence codes: 'PS2' since both parents are tested and do not present with the variant (de novo), 'PP4' since the patient presents phenotypes highly specific for a disease with a single genetic etiology.

17 patients from 10 families affected with VHL syndrome, particularly with retinal capillary hemangioblastomas (RCHs) were examined. In 2019, a 14-year-old boy (patient 8) from family 4 was diagnosed with visual impairment, due to visual acuity of 20/20 in the right eye and 20/640 in the left eye. Four peripheral RCHs were detected in the right eye. The visual impairment of the left eye was due to the severe exudative process involving the macula. Moreover, numerous microhemangioblastomas were revealed by fluorescein angiography. Laser therapy was performed in both eyes, but vitreous hemorrhage occurred in the left eye. Bevacizumab was given three times, as well simultaneous cryotherapies were performed under general anesthesia. Many laser sessions were performed for regression of the RCHs to occur. Further testing revealed a cerebellar hemangioblastoma. Genomic DNA was extracted from whole blood for sanger sequencing of the VHL gene and multiplex ligation-dependent probe amplification was performed to identify genomic rearrangements. A nonsense R113(c.337C>T) mutation was identified in exon1 of the VHL gene. The parents of patient 8 were molecularly tested and neither possessed the mutation; thus, the mutation was identified as de novo. Neither parent had VHL syndrome characterized by RCHs. A 32-year-old woman (patient 9) from family 5 had been diagnosed in 2000 with multiple cerebellar and spinal hemangioblastomas. In 2005, she was examined at the eye clinic, and were visual acuity was 20/20 and 20/25 in the right and left eye, respectively. No abnormalities were found in the right eye, but a peripheral RCH that was already partially treated with laser was observed in the left eye. Many tiny RCHs that surrounded the major lesion were also detected in this eye. Further laser treatments were scheduled, but the patient did not follow up and instead came to the ophthalmic emergency service three years later. Visual acuity of the left eye worsened to 20/640, and she presented with retinal detachment. The partially treated RCH of the left eye worsened and more tiny RCHs were detected. Cryotherapy was performed in the left eye but retinal detachment was still present at follow-up. On follow-up, no RCHs were detected in the right eye, and vision loss developed in both eyes. The same nonsense R113(c.337C>T) mutation was identified in exon1 of the VHL gene in this patient. Neither parent of patient 9 had VHL syndrome characterized by RCHs.

This paper analyzed 8 patients with clinical or imaging features consistent with VHL, but who tested negative for an abnormality by Sanger sequencing and deletion analysis. All eight patients were subsequently analyzed by next generation sequencing using peripheral blood lymphocytes. The missense mutation, c.500G>A, was found in Patient 1 in mosaic with a frequency of the mutated allele of 5.7%. This patient presented with CNS hemangioblastoma at the age of 16 and developed pheochromocytoma, pancreatic neuroendocrine tumour, and adrenal pheochromocytoma at the age of 17. The patient had no family history of VHL. This patient is believed to be the same as a patient published in 30731206. That paper offers more updated information on this patient.

Genomic DNA was extracted from whole blood samples for genetic testing of SDHB, SDHC, SDHD, and VHL. 27 patients with urinary bladder paragangliomas, revealed 17 germline mutations, three of which were in the VHL gene. PCR–based bidirectional sequencing was performed along with MLPA on large deletions. This nonsense mutation (p.R113*) was found in a 37-year-old female (patient no. 21). Family history of pheo or paragangliomas was said to be positive.

At age 14 the patient presented with a pancreatic neuroendocrine tumor that was resected. At age 19 she had hemangiomas of the fourth ventricle and cervical spinal cord which were operated. At age 27 it was noted that multiple solid tumors in the pancreas had gradually increased in size (the tumor in pancreatic head had a max size of 4.1cm) and she underwent a total pancreatectomy where pathological investigation of these tumors supported a diagnosis of well-differentiated neuroendocrine carcinoma. At age 30 she was found to have renal cell carcinoma and she underwent a nephrectomy. DNA from peripheral blood cells found a G to A point mutation at neucleotide 500 in exon 1 (Arg167Glu). Her father and elder brother were also diagnosed with VHLD. Her father had a history of two surgical resections for pancreatic neuroendocrine tumors and associated metastatic recurrence in the liver.

This study shows the clinical characteristics of three large Chinese families with VHL disease and evaluates the consequence of the genetic test for the diagnosis of VHL disease and clinical screening of the family members. DNA is extracted from peripheral blood in 43 members from three large families with VHL disease and amplified by PCR to three exons of the VHL gene. The PCR products were directly sequenced and the mutations compared with the Human Gene Mutation Database. The ages of the patients who are given the initial diagnosis ranged from 16 to 47 years (mean: 31 years), and the mean time was 17.3 months (2-30 months) from the emergence of the symptom to patients' first visit. Furthermore, the gender distribution was 20% female (4) and 80% male (16). VHL phenotypes of the disease is seen in 8 members of a Chinese family. Molecular diagnosis is confirmed in only 3 family members, as 5 died prior to genetic testing. These 5 could be assumed due to pedigree showing future linear relations of this mutation. The mutation p.L116V (c.346C>G) was referred to in the paper by c.559C>G due to a difference in transcript sequence. Manifestations in this family include CNS hemangioblastoma, and renal cysts. The proband (female) had renal cysts and hemangioblastoma, and 2 of her brothers and their 3 children (2 male; 1 female) had hemangioblastomas. The clinical manifestations were characterized by early morbidity, rapid progress, high relapse rate, and high mortality rate. There are two members who are asymptomatic, but they are young (likely before the age of onset). ACMG evidence codes: 'PP1' based on segregation results, 'PP2' because they observe a missense variant in gene where missense variants are common mechanism of disease, 'PP4' because the patients phenotypes and family history are highly specific for a disease with single genetic etiology.

This study shows the clinical characteristics of three large Chinese families with VHL disease and evaluates the consequence of the genetic test for the diagnosis of VHL disease and clinical screening of the family members. DNA is extracted from peripheral blood in 43 members from three large families with VHL disease and amplified by PCR to three exons of the VHL gene. The PCR products were directly sequenced and the mutations compared with the Human Gene Mutation Database. The ages of the patients who are given the initial diagnosis ranged from 16 to 47 years (mean: 31 years), and the mean time was 17.3 months (2-30 months) from the emergence of the symptom to patients' first visit. Furthermore, the gender distribution was 20% female (4) and 80% male (16). Von Hippel-Lindau disease is seen in 12 members of a Chinese family. Molecular diagnosis is confirmed in only 7 family members. The proband (male; deceased) has retinal capillary hemangioma; hemangioblastoma, 2 of his brothers (deceased) have pancreatic cancer, a third brother has renal cancer and hemangioblastoma, and 5 nephews have hemangioblastoma. There are three members who are asymptomatic carriers, but are young and may be before the age of onset. The mutation p.S168T (c.503G>C) was referred to in the paper by c.716G>C due to a difference in transcript sequence. At least 5 additional members had VHL phenotypes, but had all passed away before genetic testing. It could be assumed their genetic mutation due to pedigree showing future linear relations of this mutation all in generation III. ACMG evidence codes: 'PP1' based on segregation results, 'PP2' because they observe a missense variant in gene where missense variants are common mechanism of disease, 'PP4' because the patients phenotypes and family history are highly specific for a disease with single genetic etiology.

A study of 34 Polish families revealed germline mutations in 30 families using DNA isolated from peripheral blood luykocytes to sequence through PCR and Quantitative Southern analysis. Mutations were not detected in 4 probands fulfilling clinical criteria for von Hippel-Lindau disease (VHLD). This mutation was found in a VHL patient and 2 additional families. The patient had hemangioblastomas of the central nervous system and retina (family no. 6). In a family with two affected family members, 2 had hemangioblastomas of the central nervous system and 1 had retinal hemangioblastomas (family no. 7). A VHL family of 3 had 2 with hemangioblastomas of the central nervous system and 3 with retinal hemangioblastomas (family no. 5). Two members were published in a future publication: PMID: 32179488.

17 patients with genetic variants in SDHB, VHL, EPAS1, or RET with pheochromocytoma were identified retroactively for this study. 5 pathogenic germline variants were confirmed in patients with clinical criteria of von Hippel Lindau syndrome. Next generation sequencing (NGS) re-confirmed these germline mutations. One of which was a p.*214G (c.640T>G) mutation in patient 6, a 46Y male with pheochromocytoma. No further manifestation details were provided, nor family information.

Germline mutations in 45 unrelated Japanese VHL patients were determined from blood samples using PCR-SSCP or Southern blot analyses. The authors reported a nucleotide deletion (i.e. 571insAC) which coincides with legacy variant nomenclature (AF010238.1) and was mapped to c.358_359insAC (NM_000551.3). PCR-SSCP or Southern blot analyses of the DNA from family members showed that mutations were segregated with disease phenotype in VHL families, although this data was not shown. Data on the presence of retinal angioma, CNS hemangioblastoma (Hb) renal cell carcinoma (RCC), and pheochromocytoma was available. Case 14 had p.Arg120fs ( c.358insAC) mutation. 2 members were affected with renal cell carcinoma. Other demographics of the members were not described.

Germline mutations in 45 unrelated Japanese VHL patients were determined from blood samples using PCR-SSCP or Southern blot analyses. The authors reported a nucleotide deletion (i.e. 697T>C) which coincides with legacy variant nomenclature (AF010238.1) and was mapped to c.484T>C (NM_000551.3). PCR-SSCP or Southern blot analyses of the DNA from family members showed that mutations were segregated with disease phenotype in VHL families, although this data was not shown. Data on the presence of retinal angioma, CNS hemangioblastoma (Hb) renal cell carcinoma (RCC), and pheochromocytoma was available. Case 4 had a C162R (c.484T>C) mutation. 2 members were affected: both had RCC, 1 had retinal angioma, and 1 had CNS Hb. We were not able to determine which member had more than one phenotype. Other demographics of the members were not described.

The 10 year old female patient presented with sustained hypertension, headaches, and vomiting for several weeks. She had no signs of hypertensive retinopathy or papilledema. However, in the right eye, a small, white-yellowish nodular lesion in the retinal periphery was found. Luoresceinangiography showed early and late leakage and marked hyperfluorescence compatible with retinal hemangioblastoma. A heterozygous mutation 488T>A (Leu163His) in exon 3 of the VHL gene was identified (methods unspecified). Subsequently, bilateral pheochromocytomas were found. After surgical resection of the pheochromocytomas the patient was asymptomatic. No family history was mentioned.

Of 16 patients presenting with clinical VHL features that underwent genetic testing, patient 1 presented with the c.203C>G mutation of the VHL gene (stated in the paper as c.416C>G which used the Latif sequence as reference). Peripheral blood lymphocytes were collected and products of the polymerase chain reaction performed on this sample were analyzed with single-stranded conformational polymorphism analysis (SSCP), PCR sequencing and Southern analysis. This missense mutation was confirmed using restriction fragment length polymorphism or allele-specific oligonucleotide analysis. This patient has a family history of multiple tumor types indicative of VHL disease but the family did not undergo genetic testing. This patient presented with a pheochromocytoma.

Germline mutations were found in 20 Brazilian, VHL probands and their families. Peripheral blood samples were taken and genomic DNA was sequenced using PCR and Southern Blotting. The three exons of the VHL gene were amplified by PCR, sequenced in both strands with Big Dye terminators (Applied Biosystems), and collected and analysed on an ABI Prism 377 (Applied Biosystems). Germline rearrangements involving part or the entire VHL gene were detected by quantitative Southern blotting using g7 cDNA11 and a β globin probe. This mutation, H115Pfs*44 (c.344delA) , was found in a VHL type 1 family of 3 affected individuals (VHL family 9). This mutation caused a frameshift to occur leading to the protein terminating at residue 157. One had hemangioblastomas of the central nervous system, one with retinal angiomas, and one with renal carcinoma. Within this family a second mutation was also discovered in the VHL gene, P61= (183C>G), which was assumed to not be deleterious. ACMG codes as follows: A frameshift mutation in a gene where loss-of-function is a known mechanism of disease (PVS1). Cosegregation with disease in multiple affected family members in a gene definitively known to cause the disease (PP1).

42 CNS hemangioblastoma samples were obtained from 31 patients, who were consecutively treated between 1993 and 1997 at the Department of Neurosurgery of the Freiburg University Medical Centre were analyzed for VHL mutations. One patient of 17 presenting with VHL germline mutations presented with this mutation. It was state din the paper (which used the Latif sequence as reference) as p.106insR (c.529insGCC). This which automaps to the updated NM_000551.3:c.322_324dupCGC. Single strand conformation polymorphism and southern blotting for variant identification in the DNA found in blood samples. The patient was established as having VHL disease by standard of Neumann at al., 1987. The patient presented with one cerebellum tumor.

Recently, Lenglet et al., 2018 identified a new VHL cryptic exon termed E1' deep in intron 1 that is naturally expressed in many tissues, and found E1' mutations in eight families with either erythrocytosis or VHL disease. The goal of this study was to determine the prevalence of E1' germline mutations from two cohorts, 221 patients with a single or multiple tumours, suggestive of VHL disease, and 946 patients with a Pheochromocytoma/paraganglioma (PPGL) but without an identified mutation in the three VHL exons or in the main PPGL susceptibility genes. They identified a rare germline genetic variant (maf <1%) in the E1’ VHL cryptic exon in 1% of patients (12/1167). Patient 12 was diagnosed at the age of 34 due to Pheochromocytoma, and was found to have a rare germline c.340+866C>A E1' variant.

Recently, Lenglet et al., 2018 identified a new VHL cryptic exon termed E1' deep in intron 1 that is naturally expressed in many tissues, and found E1' mutations in eight families with either erythrocytosis or VHL disease. The goal of this study was to determine the prevalence of E1' germline mutations from two cohorts, 221 patients with a single or multiple tumours, suggestive of VHL disease, and 946 patients with a Pheochromocytoma/paraganglioma (PPGL) but without an identified mutation in the three VHL exons or in the main PPGL susceptibility genes. They identified a rare germline genetic variant (maf <1%) in the E1’ VHL cryptic exon in 1% of patients (12/1167). Patient 1 was diagnosed at the age of 47 with follow up of a 5 year duration due to a Carotid body paraganglioma (PGL) and was found to have a germline c.340+563C>T E1' variant. Tumor analysis was performed for patient 1 since there was a paraffin embedded sample. Patient 1 did not have loss of heterozygosity, but the mutated allele was lost in the tumor. Immunostaining was negative for detection of CA9 expression. CA9 is a protein reported to influence regulation cell proliferation and tumor progression and is known to be expressed at the membrane of tumour cells in the case of VHL inactivation.

Recently, Lenglet et al., 2018 identified a new VHL cryptic exon termed E1' deep in intron 1 that is naturally expressed in many tissues, and found E1' mutations in eight families with either erythrocytosis or VHL disease. The goal of this study was to determine the prevalence of E1' germline mutations from two cohorts, 221 patients with a single or multiple tumours, suggestive of VHL disease, and 946 patients with a Pheochromocytoma/paraganglioma (PPGL) but without an identified mutation in the three VHL exons or in the main PPGL susceptibility genes. They identified a rare germline genetic variant (maf <1%) in the E1’ VHL cryptic exon in 1% of patients (12/1167). Patient 11 was diagnosed at the age of 12 due to multiple head and neck paragangliomas, and was found to have a rare germline c.340+725A>T E1' variant.

Through blood sample testing, ten affected individuals from 7 families in Singapore were identified with VHL disease. One patient of Chinese ethnicity was found with the c.393C>G (p.Asn131Lys) mutation in the VHL gene and presented with CNS hemangioblastoma, retinal angioma, renal cell carcinoma, renal cysts, and pancreatic cysts. ACMG: PP2, PP4