Dr Trevor Cole's presentation from Osteoporosis 2016: From family history to epigenetics of osteoporosis. Find out more at: https://nos.org.uk/conference

1. From family history to theFrom family history to the
epigenetics of osteoporosisepigenetics of osteoporosis
National Osteoporosis Society ConferenceNational Osteoporosis Society Conference
Birmingham November 2016Birmingham November 2016
Dr Trevor ColeDr Trevor Cole
Consultant in Clinical and Cancer Genetics West Midlands RegionalConsultant in Clinical and Cancer Genetics West Midlands Regional
Genetics Service and Birmingham Health PartnersGenetics Service and Birmingham Health Partners
AndAnd
Honorary Reader in Medical Genetics University of BirminghamHonorary Reader in Medical Genetics University of Birmingham
Trevor.cole@bwnft.nhs.ukTrevor.cole@bwnft.nhs.uk

4. In Clinic ResourcesIn Clinic Resources
• Professor Neil Gittoes – ConsultantProfessor Neil Gittoes – Consultant
EndocrinologistEndocrinologist
• Sue Stewart – Clinical Nurse SpecialistSue Stewart – Clinical Nurse Specialist
• Dr Trevor Cole – Consultant Clinical GeneticistDr Trevor Cole – Consultant Clinical Geneticist
• DEXA scanning (not for all appointments)DEXA scanning (not for all appointments)

5. Types of OsteoporosisTypes of Osteoporosis
Primary Secondary
Type 1
Type 2
Type 3

6. Where do the genetic factorsWhere do the genetic factors
act on BMD and fragilityact on BMD and fragility
Bone Acquisition
Bone Loss

7. Genomic-Wide Association Studies (GWAS)Genomic-Wide Association Studies (GWAS)
and Heritability Studiesand Heritability Studies
Mitchell and Yerges-Armstrong Review 2011
Summarised 10 GWAS papers
29 Loci “robustly” associated with BMD
These single nucleotide polymorphism accounts for only about 2-6% of
variance in spine and femoral neck BMD
By definition each SNP associated with variance in one direction the
opposite SNPwill be associated with the opposite effect

8. What are Single nucleotide PolymorphismsWhat are Single nucleotide Polymorphisms
(SNPS)(SNPS)
A T C G T T T C G C T A ……………………….. GENE OF INTEREST
T A G C A A A G C G A T
A T C G T T G C G C T A ……………………….. GENE OF INTEREST
T A G C A A C G C G A T
“SLIGHTLY
GREATER
CHANCE”
“POPULATION
CHANCE OR
EVEN REDUCED
CHANCE
SNP’s may be away from the possible candidate gene or even if
within the gene may not have detectable effect on gene function

9. Will these SNP’s be clinically significant?Will these SNP’s be clinically significant?
Patient 1
Patient 2

10. Genomic-Wide Association Studies (GWAS)Genomic-Wide Association Studies (GWAS)
and Heritability Studiesand Heritability Studies
Slemenda et al 1992 Kelly et al 1993
111 Male Twin Pairs
Compared MZ and DZ twins
BMD modestly higher MZ twins
Most of heritability eliminated once
adjusted for non genetic factors
40 Twin pairs, predominantly female
Studied BMD changes femur and
lumber spine in MZ and DZ twins
Estimated 60-80% heritability
Wide confidence interval and only
lumber spine values significant

11. A lot of non genetic noiseA lot of non genetic noise
(how much is non genetic?)(how much is non genetic?)
• BMIBMI
• SmokingSmoking
• AlcoholAlcohol
• ActivityActivity
• Chronic DiseaseChronic Disease
• DrugsDrugs

12. Type 3 osteoporosisType 3 osteoporosis
• MalignancyMalignancy
• CushingsCushings
• Gastrointestinal disease, malabsorbtion and nutritionalGastrointestinal disease, malabsorbtion and nutritional
• Chronic renal failureChronic renal failure
• Inflammatory disease – especially RAInflammatory disease – especially RA
• Endocrinopathies – hypertyroidism, hypogonadismEndocrinopathies – hypertyroidism, hypogonadism
• Drugs Corticosteroids, chemotherapy, anti-Drugs Corticosteroids, chemotherapy, anti-
convulsants….convulsants….

13. Sounds a mess? Don’t DespairSounds a mess? Don’t Despair

14. A way forwardA way forward
For Primary Osteoporosis – a simple solution
For Secondary Osteoporosis – a few illustrative exemplars

15. London Morphology DatabaseLondon Morphology Database
>250 conditions associated with osteoporosis>250 conditions associated with osteoporosis
• Complex syndromes – eg Barakat 1996Complex syndromes – eg Barakat 1996
• Metabolic - aromatase deficiencyMetabolic - aromatase deficiency
• Mechanical – arthrogryphosis syndromesMechanical – arthrogryphosis syndromes

16. Array-CGH methodArray-CGH method
Test
Genomic
DNA
Reference
Genomic
DNA
Ratio
Position on Sequence
Cot-1
DNA
Genomic
clones or
oligos
spotted on
glass slide
DNA gain
DNA loss
“Dye-flip” experiments commonly employed

20. Hadju-Cheney Syndrome and response toHadju-Cheney Syndrome and response to
BisphosphonatesBisphosphonates

24. Examples of Disorders in 2016Examples of Disorders in 2016
• Stuve-WiedemannStuve-Wiedemann
• Schimke Immuno-osseous dystrophySchimke Immuno-osseous dystrophy
• Osteoporosis – pseudogliomaOsteoporosis – pseudoglioma
• Kenny-CaffeyKenny-Caffey
• TMEM38B OITMEM38B OI
• OI/Bone fragilityOI/Bone fragility
• Skeletal DysplasiasSkeletal Dysplasias
• Hyperparathyroid disordersHyperparathyroid disorders

25. The effect of next generation sequencingThe effect of next generation sequencing
Why has this happenedWhy has this happened The effect of NGSThe effect of NGS
• MEN1MEN1
• CDC73CDC73
• CASRCASR
• CDKN1ACDKN1A
• CDKN1BCDKN1B
• CDKN2BCDKN2B
• RET (exons 8,10-11,13-RET (exons 8,10-11,13-
16)16)
• AP2S1 (pArg15)AP2S1 (pArg15)

27. Large MEN1 pedigreeLarge MEN1 pedigree

28. Menin and CASR mutation studies normalMenin and CASR mutation studies normal
2015 Oxford hyper-
parathyroid panel
Confirmed CDC73
mutation

29. Why is it important to recognise CDC73Why is it important to recognise CDC73
hyperparathyroidism (HPRT2)?hyperparathyroidism (HPRT2)?
• Non PenetranceNon Penetrance
• Different spectrum of tumoursDifferent spectrum of tumours
• Therefore not just managing HPTTherefore not just managing HPT
• Parathyroid carcinomaParathyroid carcinoma
• Jaw tumoursJaw tumours
• Uterine tumoursUterine tumours
• Renal tumoursRenal tumours

32. CDC73 can be variable and the geneticsCDC73 can be variable and the genetics
doesn’t always give the answerdoesn’t always give the answer
Heterozygous c26delG(pArg9fs)

33. Biochemically and Genetically both haveBiochemically and Genetically both have
hypophosphatasiahypophosphatasia
Very low AlkPhosVery low AlkPhos
Compound heterozygous mutations TNALPCompound heterozygous mutations TNALP
Long term Paediatric ITU and Asfotase alphaLong term Paediatric ITU and Asfotase alpha
Very low AlkPhosVery low AlkPhos
Homozygous mutations TNALPHomozygous mutations TNALP
AsymptomaticAsymptomatic

34. Patient seen in Joint EndocrinePatient seen in Joint Endocrine
Genetic ClinicGenetic Clinic
Osteoporosis
On Bisphosponate

35. History Obtained in the EndocrineHistory Obtained in the Endocrine
Genetic ClinicGenetic Clinic
OI and Osteoporosis
On Bisphosponate
a few # as child
Age 45
a few # as a child
Age 41
History of #
Childrens Hospital
History of #
Childrens Hospital

37. Family Members at risk of OIFamily Members at risk of OI
271
140
127
44
2
0
50
100
150
200
250
300
Definitely
Affected
Status
Uncertain
at 50% risk Possibly
Affected
Obligatory
carriers

39. OI phenotypes and genotypesOI phenotypes and genotypes
OI Type Inheritance Phenotype Gene Defect
Classical –Silence
Type
I
II
III
IV
AD
AD
AD
AD
Mild
Lethal
Progressive
Moderate
Nul Col1A1
Col7A1/Col1A2
Col1A1/Col 1A2
Col1A1/Col1A2
Other V
VI
AD
?AR
Distinctive
histology/callus
Mineralisation
defect
IFITM5
SERPINF1
3 hydroxylation
defects
VII
VIII
IX
AR
AR
AR
Hypomorphic (S)
null (L)
Severe to lethal
Moderate –
severe
CRTAP
LEPRE1
PP1B
Chaperone
Defects
X
XI
AR
AR
Severe to lethal
BRUCK
progressive
SERPINH1
FKBP10
Unclassified types
Bruck Type II AR Contractures PLOD2
Caffey Disease AD Cortical
hyperostosis
Col1A1
Osteoblast
maturation defect
AD Moderate SP7

40. The Reality – the answer to theseThe Reality – the answer to these
questions was mostly NO!questions was mostly NO!
• Did we know where were they now?Did we know where were they now?
• Did we know how many more children had beenDid we know how many more children had been
born?born?
• Did we know the type / severity of their OI?Did we know the type / severity of their OI?
• Did we need expensive genetic testing?Did we need expensive genetic testing?
• Did we know if they were on any treatment?Did we know if they were on any treatment?
• Was anybody monitoring adults?Was anybody monitoring adults?
• Did we need a very complex / expensive overhaulDid we need a very complex / expensive overhaul

41. Types of OsteoporosisTypes of Osteoporosis
A very simple suggestionA very simple suggestion
Primary Secondary
Type 1
Type 2
Type 3

42. 3 Simple actions3 Simple actions
• Take a family historyTake a family history
• Assess if medical history in patient or relative isAssess if medical history in patient or relative is
indicative of an underlying disorderindicative of an underlying disorder
• See if a joint clinic with a genetics colleague isSee if a joint clinic with a genetics colleague is
realisticrealistic
• But has genetics anything to really add?But has genetics anything to really add?

43. Audit of referrals to clinical genetics 2006Audit of referrals to clinical genetics 2006
Figure 1. Referral source – OI
Patients Referred to Genetics
Figure 2. Referral source – CAH
Patients Referred to Genetics
Figure 1. Referral source – OI
Patients Referred to Genetics
Figure 2. Referral source – CAH
Patients Referred to Genetics

44. Important Gene PathwaysImportant Gene Pathways
• Wnt / Beta CateninWnt / Beta Catenin
• RANKL/RANK/OPGRANKL/RANK/OPG
• Oestrogen and TestosteroneOestrogen and Testosterone
• PTH Calcium homeostasisPTH Calcium homeostasis

46. Fascinating science but just do notFascinating science but just do not
know how to apply clinicallyknow how to apply clinically but :-but :-
• Wnt / Beta CateninWnt / Beta Catenin
• RANKL/RANK/OPGRANKL/RANK/OPG
• Oestrogen and TestosteroneOestrogen and Testosterone
• PTH Calcium homeostasisPTH Calcium homeostasis

48. Epigenetics and Future TherapyEpigenetics and Future Therapy
• Change of gene expression without changing the codingChange of gene expression without changing the coding
sequencesequence
• Three main mechanismsThree main mechanisms
– Post transcriptional histone modificationPost transcriptional histone modification
– MicroRNA’s (miRNA’s) in regulation of gene expressionMicroRNA’s (miRNA’s) in regulation of gene expression
– DNA methylation in regulaion of gene expressionDNA methylation in regulaion of gene expression

50. Vrtanik, Marc and Ostanek 2014. Epigeneticc mechanisms in bone; Clin Chem Lab Medicine 52; 589-608

53. From Family History to EpigeneticsFrom Family History to Epigenetics
oror
A Holistic Approach made SimpleA Holistic Approach made Simple
• Different clinical structures to match clinical serviceDifferent clinical structures to match clinical service
• A route to consider the unusual cases or rare disordersA route to consider the unusual cases or rare disorders
with secondary osteoporosiswith secondary osteoporosis
• A mechanism to identify and follow up extended familyA mechanism to identify and follow up extended family
• Identifying cohorts for future researchIdentifying cohorts for future research

54. My Thanks to :-My Thanks to :-
• Professor Neil GittoesProfessor Neil Gittoes
• Sue Stewart – Clinical nurse specialistSue Stewart – Clinical nurse specialist
• Lisa Cooper-Charles –W Mids Regional GeneticsLisa Cooper-Charles –W Mids Regional Genetics
LaboratoryLaboratory
• Authors of 2 excellent reviews –Authors of 2 excellent reviews –
• Vrtacnik et al (2014) Epigenetics mechanisms in bone. ClinVrtacnik et al (2014) Epigenetics mechanisms in bone. Clin
ChemLab Med 52:589-608ChemLab Med 52:589-608
• Mitchell and Streeten (2013). Clinical impact of recent geneticMitchell and Streeten (2013). Clinical impact of recent genetic