Here's a Journal Club Presentation I made on the article 'Genetic Predisposition to Schizophrenia Associated with Increased Use of Cannabis' which appeared in the Nov 14 issue of Molecular Psychiatry. It explored the possibility of whether people with high genetic risk scores for Schizophrenia somehow had a propensity for cannabis, establishing a biological basis for 'reverse causality'.

1. Genetic Predisposition to Schizophrenia
Associated With Increased Use of Cannabis
RA Power
KJH Verweij
M Zuhair
GW Montgomery
AK Henders
AC Heath
PAF Madden
SE Medland
NR Wray
NG Martin
Molecular Psychiatry 19, 1201-1204 (November 2014) 1

2. Overview
• Introduction
• Cannabis and Schizophrenia
• Related studies
• Article
Aim
Material and Methods
Results
Discussion
Limitations
2

3. Molecular Psychiatry
• “Molecular Psychiatry publishes work aimed at elucidating biological
mechanisms underlying psychiatric disorders and their treatment. The
emphasis is on studies at the interface of pre-clinical and clinical
research, including studies at the cellular, molecular, integrative,
clinical, imaging and psychopharmacology levels”
• Impact Factor: 15.147*
• Rank:
1/135 Psychiatry
5/251 Neuroscience
4/291 Biochemistry & Molecular Biology
• *2013 Journal Citation Report (Thomson Reuters, 2014)
3

4. Introduction – Cannabis & Schizophrenia
• Case control studies have consistently demonstrated higher rates
of cannabis use among patients with schizophrenia as compared
with controls.
• A meta-analysis of prospective cohort studies found similarly high
rates of broadly defined psychosis outcome (psychotic disorder or
psychotic symptoms and assoc disability) in people who had at
some time used cannabis compared with those who had never
used
– Pooled Odds Ratio = 1.41
• Evidence to date suggests some specificity of effect for
schizophrenia and non-affective psychosis
Comprehensive Textbook of Psychiatry, 9th Ed Kaplan & Sadock
4

5. Introduction – Cannabis & Schizophrenia
• The association with prior cannabis use and subsequent psychosis
is a consistent finding across studies
• The effect is attenuated but remains statistically significant even
after adjusting for
– Other drug abuse
– Known risk factors for schizophrenia
– Socio-demographic variables
Comprehensive Textbook of Psychiatry, 9th Ed Kaplan & Sadock
5

6. • There is evidence that a specific
functional polymorphism in
catechol-O-methyl transferase
(COMT) gene moderates the
effects of adolescent cannabis use
– Indls homozygous for COMT
valine allele may be more likely
to exhibit psychotic symptoms /
develop schizophreniform
disorder in adult life after
cannabis use
– Indls homozygous for COMT
methionine allele do not show
this adverse effect
Comprehensive Textbook of Psychiatry, 9th Ed Kaplan
& Sadock

7. Cannabis and Schizophrenia
• Cannabis was
associated with an
increased risk of
developing
schizophrenia in a dose
dependent fashion
Zammit S, Allebeck P, Andreasson S, Lundberg I, Lewis G.
Self reported cannabis use as a risk factor for
schizophrenia in Swedish conscripts of 1969: historical
cohort study. BMJ 2002; 325: 1199
7

8. Cannabis and Schizophrenia
8
• “Despite a clear association between the two, the
possibility of reverse causation has not been
entirely excluded
– Is psychosis a risk factor for cannabis use?
• Those on a psychotic spectrum more likely to experiment with
drugs?
Spencer C, Castle D, Michie PT. Motivations that maintain substance use among individuals with psychotic disorders.
Schizophr Bull 2002; 28: 233–247
Power RA et al (2014) Genetic Predisposition to Schizophrenia Associated With Increased Use of
Cannabis Molecular Psychiatry

9. Related studies - 1
Cannabis use predicts future psychotic symptoms, and
vice versa
• AIM :
• To determine if cannabis use is a risk factor for future psychotic
symptoms, and vice versa, in adolescents and young adults from the
general population
• METHOD
• 14-year follow-up study of 1580 initially 4–16-year-olds who were
drawn randomly from the Dutch general population
• Life-time cannabis use and psychotic symptoms, assessed with the
Composite International Diagnostic Interview (CIDI)
• Ferdinand RF, et al. (2005) Cannabis use predicts future psychotic symptoms, and vice versa Addiction, 100, 612–618
9

10. Related studies - 1 (contd…)
• Results
– Cannabis use, in individuals who did not have psychotic symptoms before they
began using cannabis, predicted future psychotic symptoms (hazard ratio =
2.81; 95% confidence interval = 1.79–4.43)
– However, psychotic symptoms in those who had never
used cannabis before the onset of psychotic symptoms
also predicted future cannabis use (hazard ratio = 1.70;
95% confidence interval = 1.13–2.57)
• Conclusion
• The results imply either
– a common vulnerability with varying order of onset
– a bi-directional causal relationship between cannabis use and
psychosis
Ferdinand RF, et al. (2005) Cannabis use predicts future psychotic symptoms, and
vice versa Addiction, 100, 612–618
10

11. Related studies - 2
Tests of causal linkages between cannabis use
and psychotic symptoms
• AIM : To examine possible causal linkages between cannabis use and
psychosis using data gathered over the course of a 25-year
longitudinal study
• METHOD
• A 25-year longitudinal study of the health, development and
adjustment of a birth cohort of 1265 New Zealand children
– Data on cannabis use and psychotic symptoms were available on at least one
occasion from 18, 21 and 25 years
– Structural equation models permitted evaluation of reciprocal relationships
between cannabis use and psychosis and using these models to provide a guide
to probable patterns of causation
• Fergusson DM et al (2005), Tests of causal linkages between cannabis use and psychotic symptoms Addiction,100,
354–366
11

13. Related studies - 2 (contd…)
• Results
– For both models, the effect of psychotic symptoms on cannabis use was
negative and, for Model 2, statistically non-significant.
– it was unlikely that the development of psychotic symptoms led to
increased use of cannabis and that, if anything, the development of these
symptoms may have inhibited rather than encouraged cannabis use.
– Daily users of cannabis had rates of psychotic symptoms that were
between 1.6 and 1.8 times higher (P<0.001) than non-users of cannabis
• Conclusion
– the association between cannabis use and psychotic symptoms is unlikely
to be due to confounding factors
– the direction of causality is from cannabis use to psychotic symptoms
13

14. • Schizophrenia is a highly heritable condition, with upto 80%
of the variance explained by additive genetic effects
• Sullivan PF, Kendler KS, Neale MC. Schizophrenia as a complex trait—Evidence from a meta-analysis of
twin studies. Arch Gen Psychiatry 2003; 60: 1187–1192.
• Cannabis use itself has been reported to be heritable
(although no genetic risk variants have been identified)
Agrawal A, Lynskey MT. The genetic epidemiology of cannabis use, abuse and dependence. Addiction
2006; 101: 801–812.
Genetic Risk for Schizophrenia → Association with cannabis use
• Such an association would suggest that those genetically
predisposed to schizophrenia use cannabis more frequently

15. • Cannabis use itself is not associated with the COMT valine
or methionine allele, suggesting an underlying
gene–environmental interaction (rather than a gene–
environment correlation) is operating to increase risk
• Comprehensive Textbook of Psychiatry, 9th Ed Kaplan & Sadock
• “This would mean that the association between
schizophrenia and cannabis use is not simply one of an
environmental risk factor, but rather involves gene–
environment correlation, as individuals choose and shape
their own environment based on their own innate
preferences”
• Power RA et al (2014) Genetic Predisposition to Schizophrenia Associated With Increased Use of
Cannabis Molecular Psychiatry

16. Related studies - 3
Genome-wide Association Analysis Identifies
13 New Risk Loci for Schizophrenia
• AIM : To perform a Genome Wide Association Study (GWAS) to
identify associations between SNPs/Loci and Schizophrenia
• METHOD
– GWAS performed on 13833 cases and 18310 controls
• Ripke S, O'Dushlaine C, Chambert K, Moran JL, Kahler AK, Akterin S et al. Genomewide association analysis identifies 13
new risk loci for schizophrenia. Nat Genet 2013; 45: 1150–1159.
16

17. 17

18. Related studies - 3
• First unit for reporting effect is Odds Ratio
– Odds of Disease in indls having the allele
– Odds of Disease in indls not having the allele
↓
• Chi-squared Test gives p-value of a certain significance
18

19. 19

20. • Pa = Freq of Allele a occurring at a locus
• Pb = Freq of Allele b occurring at a locus
• Pab = Freq of a and b occurring together in the same gamete
• If Pa x Pb ≠ Pab there is a non-random association between a & b
and there is Linkage Disequilibrium in between these alleles
• Coeff of LD = Dab = Pab – (Pa x Pb)
• Many GWAS findings implicate an extended region containing
multiple SNPs with significant association
– These are not independent associations but result because of high LD between
associated SNPs
– These associations are summarized in terms of the index SNP with the
strongest association and other SNPs in high LD with the index SNP 20

21. • Correlation between a pair of Loci
• Value = r Freq = r2
r2 = D2 / (p1p2 x q1q2)
• If r2 = 0 there is complete Linkage Equilibrium
• If r2 = 1 there is complete Linkage Disequilibrium
• R2=0.25, high degree of LE
• Polygenic Schizophrenia Risk Scores
– Number of Schizophrenia risk alleles weighted by Logistic Regression
– PLINK program
– (P = 0.0001, 0.001, 0.01. 0.05, 0.1, 0.2, 0.3, 0.4, 0.5 and 1.0)
21

22. Related studies - 3 (contd…)
• Results
– Identified 22 loci associated at genome-wide significance; 13 of
these are new, and 1 was previously implicated in bipolar disorder.
– Examination of candidate genes at these loci suggests the
involvement of neuronal calcium signaling.
– Estimate that 8,300 independent, mostly common SNPs (95%
credible interval of 6,300–10,200 SNPs) contribute to risk for
schizophrenia and that these collectively account for at least 32%
of the variance in liability.
• Conclusion
– Common genetic variation has an important role in the etiology of
schizophrenia, and larger studies will allow more detailed
understanding of this disorder
22

23. Genetic Predisposition to Schizophrenia
Associated With Increased Use of Cannabis
RA Power
KJH Verweij
M Zuhair
GW Montgomery
AK Henders
AC Heath
PAF Madden
SE Medland
NR Wray
NG Martin
Molecular Psychiatry 19, 1201-1204 (November 2014) 23

24. Introduction
AIM
To discern the direction of causation between
cannabis use and schizophrenia
by
Studying the association between cannabis use and
Polygenic Risk for Schizophrenia
24

25. Materials & methods – Subjects & Clinical Assessment
• Both groups participated in telephonic interviews
• Based on Semi-Structured Assessment of the Genetics of
Alcoholism
25
Sample 1 Sample 2
Number 6265 9688
Age 23 to 39 yrs (mean = 29.9) 18 to 91 yrs (mean 46.3)
Years 1996 to 2000 2001 to 2005
Born betn 1964 to 1971 1895 to 1964
1964 to 1971

26. • Collaborative Study on the Genetics of Alcoholism (COGA) funded by
the NIAAA
• Objectives
– Characterize the familial transmission of alcoholism and related phenotypes
– Localize susceptibility genes for alcoholism using genetic linkage.
• 600 nuclear families were recruited
– All available biologically related individuals were personally interviewed.
– A subset of families were selected for more intensive study using standard
biological markers, and neurophysiological and neuropsychological protocols
Polydiagnosis Comorbidity Phenotype
ADS based on numerous
diagnostic systems
Depression
ASPD
Age of onset, Comorbidity
Drinking patterns
Severity of disorder

27. Materials & methods – Subjects & Clinical Assessment
• 14 087 individuals, of whom 7172 were genotyped
– (1) did you ever use marijuana?
– (2) how old were you the very first time you tried marijuana (not counting the
times you took it as prescribed)?
– (3) how many times in your life have you used marijuana (do not count times
when you used a drug prescribed for you and took the prescribed dose)
• Ever use was measured on a dichotomous scale (ever versus never)
• Age at initiation and quantity of use were open questions
27
Checking For Pruning for relatedness
Ancestry outliers
Hardy–Weinberg equilibrium
Mendelian errors
Call rate
Genotypic missingness (>5%)
Individual missingness (>5%)
Minor allele frequency
Removing one individual from each
pair with relatedness >0.05, as
determined from genetic data

28. 28

29. 29

30. Materials & methods – Subjects & Clinical Assessment
• Final sample comprised 2082 ‘unrelated’ indls
30

31. Results
• Participant Characteristics
– Males showed a higher rate of use than females
• 53.5% vs 43.9% (P<0.001)
• Polygenic Risk Scores for Schizophrenia
– Positive associations for
• ever versus never use of cannabis across all P-value thresholds
– Strongest association for those SNPs with P-values of 0.01 or below in
the original schizophrenia GWAS
– Positive association for
• analysis of quantity of cannabis use for 9 of the 10 SNP cutoffs
– top association seen for those SNPs with P ⩽0.05 for schizophrenia (R2
= 0.85%, P = 0.003)
31

32. Results
• Secondary analysis
– Mean polygenic risk score for within 990 twin pairs (608 dizygotic
and 382 monozygotic) where data on cannabis use of both twins
was available; ordinal regression to predict whether cannabis was
being used by
• neither (n = 272)
• one (n = 273)
• or both twins (n = 445)
• Twin pairs where
– both reported using cannabis had the greatest burden of
schizophrenia risk alleles
– pairs with only one user were found to have an intermediate level
– lowest burden was found in pairs where neither twin reported use
32

33. 33

34. Results
– Positive association of polygenic risk score with ever
versus never use of cannabis across all P-value
thresholds
– Positive association for analysis of quantity of cannabis
use for 9 of the 10 SNP cutoffs
– Twin pairs where
• both reported using cannabis had the greatest burden of
schizophrenia risk alleles
• pairs with only one user were found to have an intermediate
level
• lowest burden was found in pairs where neither twin reported
use
34

35. Discussion
• Results suggest that indls with an increased genetic
predisposition to schizophrenia are both
– More likely to use cannabis
– More likely to use greater quantities of cannabis
• This does NOT refute a causal relationship between use of
cannabis and risk of Schizophrenia
• Establishes that part of the association betn cannabis and
schizophrenia may be due to a causal relationship in the
opposite direction
35

36. Discussion
• Variance in cannabis use explained by Schizophrenia
Polygenic Risk Score is small
– Consistent with other cross-phenotype analyses
• Autism, MDD, BPAD
• Further research is reqd to see whether the
illustrated genetic overlap is specific to cannabis or
present across illicit drug use / addiction
phenotypes
• The possibility that the association may be
bidirectional in causation has been supported
– Risks of cannabis are being overestimated??
36

37. Limitations
• Schizophrenia GWAS sample is likely to include
more cannabis users among cases than controls
– ?? An excess of SNPs associated with cannabis use may
mistakenly have been identified as Schizophrenia risk
alleles
37

38. Limitations
High Polygenic Risk Score for Schizophrenia
↓↓
??????????
↓↓
Cannabis use
38

39. Checklist for assessing the quality of
quantitative studies
Criteria Yes(2) Partial(1) No(0) N/A
1 Question/ objective
sufficiently described? √
2 Study design evident &
appropriate? √
3 Method of
subject/comparision
group selection or
source of information/
input variable describe
and appropriate?
√
4 Subject ( and
comparison group, if
applicable)
characteristics
sufficiently described?
√
39

40. Yes (2) Partial (1) No (0) NA
5 If interventional and random allocation
was possible, was it described? √
6 If interventional and blinding of
investigators was possible, was it
reported?
√
7 If interventional and blinding of subjects
was possible, was it reported? √
8 Outcome and (if applicable) exposure
measures well-defined and robust to
measurement/misclassification bias?
Means of assessment reported
√
40

41. Yes (2) Partial (1) No (0) NA
9 Sample size appropriate?
√
10 Analytic methods
described/justified and
appropriate?
√
11 Controlled for confounding?
√
12 Results reported in sufficient
detail? √
13 Conclusions supported by the
results? √
41

42. Take Home Message
• A specific set of SNPs confer a significantly
heightened risk for Schizophrenia
• There is no refuting the forward association
between cannabis use and Schizophrenia
• This study has provided evidence that a heightened
genetic risk for Schizophrenia is associated with
higher incidence of cannabis use
• This suggests a reciprocal relationship between
Schizophrenia risk and cannabis
• We might have overrated the risk of psychosis
assigned to cannabis
42

43. References
• Comprehensive Textbook of Psychiatry, 9th Ed Kaplan & Sadock
• Zammit S, Allebeck P, Andreasson S, Lundberg I, Lewis G. Self
reported cannabis use as a risk factor for schizophrenia in Swedish
conscripts of 1969: historical cohort study. BMJ 2002; 325: 1199
• Fergusson DM et al (2005), Tests of causal linkages between cannabis
use and psychotic symptoms Addiction,100, 354–366
• Ripke S, O'Dushlaine C, Chambert K, Moran JL, Kahler AK, Akterin S et
al. Genomewide association analysis identifies 13 new risk loci for
schizophrenia. Nat Genet 2013; 45: 1150–1159.
Agrawal A, Lynskey MT. The genetic epidemiology of cannabis use,
abuse and dependence. Addiction 2006; 101: 801–812. 43

44. Thank you
44