Diagnostic Odysseys

Dusica Babovic-Vuksanovic, MD
Professor of Genetics and Pediatrics
Chair, Department of Medical Genetics
Center for Individualized Medicine, Mayo Clinic

GENETICS…very
simple!
• Mendelian disorders
• Autosomal dominant
• Autosomal recessive
• X-linked

Autosomal dominant
inheritance
Neurofibromatosis
Tuberous sclerosis
I

II

III

Marfan syndrome
Ehlers-Danlos
syndrome
Familial cancer
syndromes
Achondoplasia

Autosomal recessive
inheritance

Cystic fibrosis
Metabolic
disorders
Syndromes…25%

Carrier

25%

25%

25%

Normal
Affected

X-linked
inheritance
Fragile X
Duchenne muscular dystrophy
Hemophilia A

Affected male
Normal male
Normal female
Carrier female

Risk for disease
CAD
70’s

Colon Ca
80’s

Lung Ca
75

55 y
Cleft
Palate
MR

45 y
BRCA

32 y

30 y

48 y
MI

20 y

Dementia

MVA
15 y

18 y

Dementia

Genetics of human disease
Is it so simple?
• Mitochondrial disorders
• Maternally inherited
mDNA

Epigenetics mechanisms
ANGELMAN SYNDROME PRADER WILLI SYNDROME
Paternal
Maternal
deletion/maternal UPD
deletion/paternal UPD
Hypotonia
Stiff ataxic gait
Developmental delay
Severe MR
Obesity
Absent speech
Hypogonadism
Almost never obese
OCD

Genetic
heterogeneity

Several genetic loci for
the
same phenotype
• Neuropsychiatric
disorders
• Autism
• Schizophrenia
• Hearing loss

Non-syndromic
>400 syndromes
Cell, Volume 141, Issue 2, 210-217, 16 April 2010

Allelic heterogeneity:
same gene-different
phenotypes
• FGFR3
Achondroplasia
• Crouzon syndrome/AN
• Thanatophoric
dysplasia

• LAMIN A
Progeria
• ED Muscular
dystrophy
• Cardiomyopathy
• Lipodystrophy

Variable
penetrance
• Incomplete penetrance
Deletion/duplication 22q
• Age dependent penetrance
Alzheimer disease
• Gender predisposition
X-linked disorders
Hip dysplasia

Anticipation
Trinucleotide repeat disorders
Huntington disease
Spinocerebellar ataxia
Myotonic Dystrophy
Fragile X syndrome

New Frontiers
Environment, including
microbiome

If the child looks like the father, it’s genetic
f the child looks like the neighbor, it’s environmenta

No rules? COMPLICATED
• Non-Mendelian (common) disorders
• Multifactorial disorders
• Genetics/environment complex
interactions
• Cleft lip/palate
• Cancers
• Diabetes
• Hypertension
• Aging…..

DNA
(A, G, T, or C)
is 6 billion bits in
every cell, with a
total length of
about 6 feet of
DNA in every cell

From Nature, Feb 2001

Information
content of human

Human genome







~ 25,000 genes
Over 3,000 monogenic disorders
Human genome variations (Craig Venter)
3,213,401SNPs
53,823 block substitutions (2–206 bp)
559,473 homozygous indels (1–82,711 bp)
62 CNVs
292,102 heterozygous insertion/deletion events
(indels)(1–571 bp)
• 90 inversions
Levy et al., PLoS Biol. 2007 October; 5(10): e254

Personalized medicine: Era of
genomic sequencing
• Risk for disease
• Predict response to
drugs
• Pre-symptomatic
diagnosis for
preventable and
treatable disease

• Translation to clinical
medicine
• Incidental findings
• Ethical concerns
• Emotional distress
• Cost of medical care

Gene screening may refine
prediction of heart attack risk,
researchers say
Dec. 3, 2010 | This Week at Mayo Clinic | Volume 2, Number 1

Testing for 11 specific genetic variations in hundreds of people
with no history of heart disease provided information that led to
revision of their estimated heart attack risk, say Mayo Clinic
researchers.
Early findings suggest benefit may be most valuable in people thought to
be at intermediate risk using the current model
Dr. Kullo adds, however, that these findings need to be replicated and validated
in prospective studies before they are used in patient care.

What is a Diagnostic
Odyssey?
• A protracted and arduous journey for
patients and families to find answers about
a rare medical condition they suffer from
• A long sequence of medical evaluations
and referrals until patients have an
eventual diagnosis
• Rare diseases affect about 25 million
people in the US
• Cumulatively, rare diseases are not rare

Diagnostic Odyssey in the
Era of Precision Medicine
• 25% of patients on a diagnostic odyssey
identified a gap of 5 to 30 years between
getting their first symptoms and receiving
diagnosis
• With advancements in genome sequencing
more patients are finding answers to their
diagnostic dilemmas
• Some
are
treatments

even

finding

lifesaving

Genetic testing
Cytogenetic methods (karyotype, FISH etc)
Chromosome analysis
Chromosome microarray
Targeted microdeletion/duplication testing
Molecular methods (sequencing, UPD, methylation
assay etc)
Single gene analysis
Gene panels
Exome analysis
Genome analysis

21

From Gregory, Nature Reviews Genetics 6, 699-70

Genome sequencing

23

6 billion
bases!
lgvrdlmnqvtthequickababcmfxlqbrownfoxjulrcsmped
overthelazyyyzpolfdogjjirttiythedoglayhhbeldquetly
dreaminghwwiqldnsofdinnerplwosiucnd

25

Whole Exome Sequencing
(WES)
Analysis of all exons
180,000 exons arranged in ~22,000 genes
Portion of the human genome that contains proteincoding
sequences
2-3% of human genome

26

Whole Exome Sequencing
(WES)
• Genes related to clinical phenotype
• Medically actionable diagnosis (ACMGG guidelines)
• Carrier status for AR conditions (CF, Tay-Sachs, Sickle
Cell, Familial Dysautonomia, Canavan disease, G6PD
deficiency)
• Pharmacogenetic information (CYP2C9 and
CYP2C19), warfarin and Plavix metabolism
• Not included adult onset neurodegenerative
disorders
Optional expended report includes deleterious
mutations and unclassified variants in genes
unrelated to disease phenotype and deleterious
mutations in genes with no currently known function
27

Whole exome sequencing
• Need to have
parental samples
• Complex test and
difficult interpretation
• Variants of
unknown
significance (VUS)
• Incidental findings

We’re not all the same
28

Individualized Medicine (IM)
Clinic
• IM Clinic Workgroup established in January
2012
• Multidisciplinary, collaborative effort
• Designed/implemented two IM Clinic service
lines

Sept 30th, 2012:
• Advanced Cancer
• Diagnostic Odyssey

IM Clinic: Diagnostic Odyssey
• Undiagnosed patients with suspected
genetic component
• Whole Exome Sequencing of patient
genomic DNA and affected or unaffected relatives - performed to find
genes contributing to disease
• Dx provides:
• Comfort, resolve, closure
• Information for family planning
• Possible therapy

Diagnostic Odyssey
IM Clinic
eConsult

Referral into
IM Clinic

Referral to
Mayo Clinic

••

Whole
Whole Exome
Exome Sequencing
Sequencing
of
of germ-line
germ-line

•• Genomic
Genomic counseling
counseling
•• IM
IM Clinic
Clinic Consultant
Consultant meets
meets patient
patient

•• Bioinformatics
Bioinformatics
analyzes,
analyzes, generates
generates
report
report
•• Results
Results presented
presented to
to
Genomic
Genomic Board
Board
•• Results
Results and
and possible
possible
treatment
treatment options
options
returned
returned to
to patient
patient

What is Standard Lab
Order?

IM Clinic
Genomic Odyssey Board (GOB)
Review new cases and test results.
Members include:

Diagnostic Odyssey
Case 1
Holoprosencephaly
Central incisor
Facial clefting
Diabetes insipidus
Ectrodactyly
Developmental delay

ECC (TP63 gene)
negative
Normal chromosome
array
Odyssey case!

Hartsfield syndrome

35

Hartsfield syndrome
Autosomal disorder
Only sporadic cases known
Recurrence risk low?

36

Second child
Ectrodactyly
Holoprosencephaly
Cleft lip and palate
Heart defect

Hartsfield syndrome
No other reported familial cases
WES: FGFR1 gene novel variant c.1880G>C
(p.R627T)
2 affected siblings with same mutation, parents
negative
Recurrence risk?

Third child
Ectrodactyly
Holoprosencephaly
Cleft palate
Encephalocele

3 affected siblings with same mutation, parents
negative, Odyssey continues!
Targeted testing of sperm: 23% of cells with the
mutation

Diagnostic Odyssey Case 2



Age : 26
Sex : Male
Progressive muscle weakness onset age 6 or 7
Difficulty sitting and standing up from sitting
position and difficulty climbing stairs
• Difficulty swallowing solid foods
• Mild form of ophthalmoplegia
• Elevated CK (643)

Previous Genetic Testing
Prior muscle biopsies not specific
No genetic testing (insurance problems)
Referred for WES

IM evaluation
Personal history
Pedigree analysis
Physical examination
Review of previous evaluations
Differential diagnosis
Review of literature

44

Results

• Progressive myopathy, suspicious family history,
Hutterite ancestry
• Limb-girdle muscular dystrophy type 2H Sarcotubular myopathy
• Autosomal recessive disorder previously described
in Hutterite population
• Targeted gene testing

IM Clinic – Diagnostic Odyssey
Consecutive Cases of 18 Months in
Operation

MedGen
Non
MedGen
Total

All Referrals
Declined
Proceeded with
WES
WES* testing
testing
42
23
8
50

Total
65

9

17

32

82

IM Clinic – Diagnostic Odyssey
Consecutive Cases of 18 Months in Operation
Reason for Referral
Age
Neuro

5<5
18

>18

Total

All referred 17 26

7

50

All tested with WES 10 15

6

31

All positive 1

5

3

9

All referred 2

6

24

32

All tested with WES 1

4

14

19

All positive 0

1

3

4

Non-neuro

IM Clinic – Diagnostic Odyssey
Consecutive Cases of 18 Months in
Operation
Cases
Total Complete

50

Positive

15

Negative

35

Success Rate

30%

IM Clinic – Diagnostic Odyssey
Consecutive Cases of 18 Months in Operation
Results – from Clinical Focused report
Tota
l

Range per
patient

22

0-3

Medically actionable*

2

0-1

Carrier status

3

0-1

76

0-4

2

223

0-16

5.6

Pathogenic/likely pathogenic
mutations

**PGX
***VUS possible related to
phenotype

* Medically actionable -- MUTYH (het), DSC2 Carrier - CFTR, G6PD, FANCC
** PGX: Pharmacogenomics
***Variants of unknown clinical significance

Averag
e

Cost for Evaluating a Patient on
a
Diagnostic Odyssey
Success
Rate

Cost

Conventional Genetic
Testing*

15%

~$25,0
00

Whole Exome Sequencing

30%

$8,000

Tests

* Shashi V et al., Genet Med 16;176-2;2014

IM Clinic – Diagnostic Odyssey
Party Responsible for Payment
%

Category
Patients received complete or
31 partial commercial insurance
coverage
27

Medicaid

18 CIM
6

International patients

18 Other

WES and Still No Diagnosis:
What
is
Next
Associated
Variant Type
Phenotype(s)

Repetitive DNA, including
trinucleotide repeats

Fragile X syndrome
Huntington’s disease

Copy-number variants

DiGeorge syndrome
Charcot-Marie-Tooth disease type 1A

Long insertion-deletion variants*

Resistance to HIV infection

Structural variants

Chromosomal translocations
associated with spontaneous
abortions

Aneuploidy

Down’s syndrome
Turner’s syndrome

Epigenetic alterations

Prader-Willi syndrome, BeckwithWiedemann syndrome

Acknowledgeme
nts
Center for Individualized Medicine
Leadership
Konstantinos N. Lazaridis, M.D.

Department of Medical Genetics
David R. Deyle, M.D.
Pavel Pichurin, M.D.
Ralitza Gavrilova, M.D.
Geoffrey Beek, CGC
Jennifer Kemppainen, CGC
Marine Murphree, CGC

Questions

We’re not in the “post-genome” era yet!

From http://www.infosci.coh.org/corelab/genomics.htm

Sign up to vote on this title
UsefulNot useful