The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

The Genetics of VHL

Xia Wang MD PhD

Oct, 2017

• How human cells and tissue grow and die?

• Normal tissue growth is regulated by many genetic

factors

• Safe traffic is maintained by traffic lights and all

responsible drivers.

Proper tissue growth - controlled traffic

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

Broken traffic light - predisposes to accident Additional traffic light malfunction -

Reckless driving – accident! When genetic factors are damaged

with time --

Tumor

CancerA

B

C

Mutated

A

B

C

Damaged

A

B

C

Defected

A

B

C

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

van Asselt et al. Cilia 2013, 2:3 http://www.ciliajournal.com/content/2/1/3

• VHL is a tumor

suppressor

• VHL mutations

predispose to

tumors

• VHL affects 1 in

36,000 births

To understand genes…

• Genes are DNA molecules – genomic blueprints

• DNA RNA

• RNA Protein

• Proteins are the building blocks of our body

Each cell has the same set of genomic blueprints of

~20,000 genes that we are born with

Socratic.org Greatbigcanvas.com

DNA RNA Protein

VHL protein

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

DNA genetic code, amino acid, and protein

www.bbc.co.uk Genome

Function of VHL protein:

control HIF1alpha

Normal oxygen levelLow oxygen level

Von Hippel–Lindau disease (VHL), modified from Wikipedia

VHL Tumors

Average age of onset Frequency in Patients, %

Hem

ang

iobla

sto

ma

Retina (Eye) 25 (0-68) 25-60

Cerebellum (Brain) 33 (9-78) 44-72

Brain stem 32 (12-46) 10-25

Spinal cord 33 (11-66) 13-50

Endolymphatic sac tumor 22 (12-50) 10-25

Renal cell carcinoma or cysts 39 (13-70) 25-75

Pheochromocytoma 27 (5-58) 10-25

Pancreatic Neuroendocrine Tumor (PNET)

36 (5-70) 11-17

Screening can find the tumor before it causes symptoms

Find the tumor early

Early intervention

Better outcomes

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

Scre

en

ing

Re

co

mm

en

da

tion

s

VH

L A

llian

ce

Age Screening and Frequency

From Birth

Inform pediatrician of family history of VHL.

Pediatrician to look for signs of neurological disturbance, nystagmus, strabismus,

white pupil, and other signs which might indicate a referral to a retinal specialist.

Routine newborn hearing screening.

0-4

Annually

Pediatrician to look for signs of neurological disturbance, nystagmus, strabismus,

white pupil, and abnormalities in blood pressure, vision, or hearing

Eye/retinal examination with indirect ophthalmoscope by an ophthalmologist skilled

in diagnosis and management of retinal disease, especially for children known to

carry the VHL mutation.

5-15

Annually

Physical examination and neurological assessment by pediatrician informed about

VHL, with particular attention to blood pressure taken (taken both while lying and

standing), hearing Issues, neurological disturbance, nystagmus, strabismus, white

pupil, and other signs which might indicate a referral to a retinal specialist.

Eye/retinal examination with indirect ophthalmoscope by ophthalmologist informed

about VHL, using a dilated exam.

Test for fractionated metanephrines, especially normetanephrine in a “plasma free

metanephrine” blood test or in a 24-hour urine test.

Abdominal ultrasonography annually from 8 years or earlier if indicated.

Abdominal MRI or MIBG scan only if biochemical abnormalities found.

Every 2-3 years

Complete audiology assessment by an audiologist. Annually if any hearing loss,

tinnitus, or vertigo is found

In the case of repeated ear infections, MRI with contrast of the internal auditory canal

using thin slices, to check for a possible ELST.

Scre

en

ing

Re

co

mm

en

da

tion

s

VH

L A

llian

ce

Age Screening and Frequency

From Birth

Inform pediatrician of family history of VHL.

Pediatrician to look for signs of neurological disturbance, nystagmus, strabismus,

white pupil, and other signs which might indicate a referral to a retinal specialist.

Routine newborn hearing screening.

0-4

Annually

Pediatrician to look for signs of neurological disturbance, nystagmus, strabismus,

white pupil, and abnormalities in blood pressure, vision, or hearing

Eye/retinal examination with indirect ophthalmoscope by an ophthalmologist skilled

in diagnosis and management of retinal disease, especially for children known to

carry the VHL mutation.

5-15

Annually

Physical examination and neurological assessment by pediatrician informed about

VHL, with particular attention to blood pressure taken (taken both while lying and

standing), hearing Issues, neurological disturbance, nystagmus, strabismus, white

pupil, and other signs which might indicate a referral to a retinal specialist.

Eye/retinal examination with indirect ophthalmoscope by ophthalmologist informed

about VHL, using a dilated exam.

Test for fractionated metanephrines, especially normetanephrine in a “plasma free

metanephrine” blood test or in a 24-hour urine test.

Abdominal ultrasonography annually from 8 years or earlier if indicated.

Abdominal MRI or MIBG scan only if biochemical abnormalities found.

Every 2-3 years

Complete audiology assessment by an audiologist. Annually if any hearing loss,

tinnitus, or vertigo is found

In the case of repeated ear infections, MRI with contrast of the internal auditory canal

using thin slices, to check for a possible ELST.

Physical examination with vigilance about VHL tumors

Abdominal ultrasound to be started at age 8

16+

Annually

Eye/retinal examination with indirect ophthalmoscope by

ophthalmologist informed about VHL, using a dilated exam.

Quality ultrasound, and at least every other year MRI scan of

abdomen with and without contrast to assess kidneys, pancreas,

and adrenals, but not during pregnancy.

Physical examination by physician informed about VHL.

Test for fractionated metanephrines, especially normetanephrine

in “plasma free metanephrines” blood test or 24-hour urine test. Abdominal MRI or MIBG scan if biochemical abnormalities found.

Every 2-3 years

MRI scans should be ordered as no less than a 1.5T MRI with and

without contrast of brain, cervical, thoracic, and lumbar spine, with

thin cuts through the posterior fossa, and attention to inner

ear/petrous temporal bone to rule out both ELST and

hemangioblastomas of the neuraxis.

Audiology assessment by an audiologist.

Preg-nancy

Regular retinal checkup to anticipate potentially more rapid

progression of lesions.

Test for pheo early, mid, and again late pregnancy to ensure no

active pheo during pregnancy or especially labor and delivery.

During the 4th month of pregnancy, MRI—without contrast—to

check on any known lesions of the brain and spine. If known retinal,

brain, or spinal lesions, consider C-section.

Scre

en

ing

an

d e

arly

de

tectio

n

of tu

mors

Who to be screened?

• If a parent has VHL, how do we know which child will

be affected with VHL tumors?

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

• 20% of the individuals affected with VHL did not inherit

it from the parents.

• The first case of VHL in the family is due to a new, i.e.

de novo mutation.

Who to be screened? VHL Inheritance Pattern – Autosomal Dominant

Von Hippel–Lindau disease (VHL), From Wikipedia

VHL Clinical Diagnostic Criteria

• Simplex case with two or more of the following:

–Two or more hemangioblastomas

–A single hemangioblastoma with multiple kidney or

pancreatic cysts

–Renal cell carcinoma

–Pheochromocytomas

–Endolymphatic sac tumors, papillary cystadenomas

of the epididymis or broad ligament, or

neuroendocrine tumors of the pancreas

VHL Clinical Diagnostic Criteria

• Simplex case with two or more of the following:

–Two or more hemangioblastomas

–A single hemangioblastoma with multiple kidney or

pancreatic cysts

–Renal cell carcinoma

–Pheochromocytomas

–Endolymphatic sac tumors, papillary cystadenomas

of the epididymis or broad ligament, or

neuroendocrine tumors of the pancreas

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

VHL diagnosis based on VHL gene testing

VHL gene structure

Human genome -- 3,000,000,000 base pairs (bps)

VHL gene – 12,633 bps, 639 bps in exons, 213 amino

acids

VHL gene on the short arm of chromosome 3 Types of VHL gene changes

• Deletion of a segment of the chromosome

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

• Base pair (DNA code) changes

• Deletion of a segment of the gene

Types of VHL gene changes

• The technology for genetic testing has changed

dramatically in the past 10 years.

Sanger sequencing reading the raw sequences of

VHL gene takes 3-4 days

Large deletions detection by Southern Blot takes

4-5 days

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

Next-generation sequencing can detect single code

change, small/large deletions or duplications. It

takes 3 days to get the raw sequences of the entire

human genome!

Types of VHL variants

Classifications and Terms Meaning

Mutation

Pathological variant

Disease causing variant

Dysfunction

Loss of function

Benign variant

PolymorphismNormal function

Variant of uncertain significance

(VUS)Function unknown

VHL gene Variants on VHL gene

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

Variants on VHL gene Interpretation of a variant is not always

straight forward J Genet Counsel (2015) 24:882–889, PMID 26323595

c.463+4C>T

VHL GRCh38

Chr3:10,146,640

Next gen

sequencing

???

significance

Affect

intron

splicing

In silico

prediction

model

RNA analysis

In test tubes

Confirmed

exon 2

skipping in

some RNA

products

One patient with

Hemangioblastoma

A 2nd degree

relative with Pheo

Current variant classification: c.463+4C>T

VUS (variant of uncertain significance)

• Blood or Skin cells To find the germline mutation

(mutation present at the beginning of life – a fertilized

egg)

• Tumor tissue May reveal the germline and/or the

somatic mutation (mutation occurred in later stage of

life)

Samples for Genetic Testing

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

Genetic mutations lead to kidney cancer

(Renal cell carcinoma)

By Dr. Wong Ming Ho, Edmond, Urology, HK

An ethnic group in the mid-Volga region of Russia

Chuvash population

Chuvash population,

• A unique condition is known in Chuvash population

• It is caused by VHL gene mutations

• But it does not cause VHL tumors

Familial Erythrocytosis Type 2 (ECYT2) Familial Erythrocytosis Type 2 (ECYT2)

• Increased red blood cell production

• Increased levels of erythropoietin

• Normal oxygen affinity

• Thrombosis and/or hemorrhage

• Two copies of mutations on VHL, inherited in an

autosomal recessive fashion.

• No individuals with one copy of the mutation have

developed VHL-related tumors [Gordeuk et al 2004].

The Genetics of VHL Xia Wang, MD, PhD

VHL Alliance, 2017 Tampa, FL

• R200W (Chuvash)

• H191D

• Russell et al. (2011)

• R200W -- Not associated with tumors seen in VHL.

Familial Erythrocytosis Type 2 (ECYT2)