neli ulrich, ms, phd huntsman cancer institute, salt lake...

February 3-5, 2016 | Lansdowne Resort, Leesburg, VA

Neli Ulrich, MS, PhD

Huntsman Cancer Institute,

Salt Lake City

“Using Genetics and Metabolomics

to Advance Precision Prevention”

Three Highlights

Aspirin precision prevention with genetics

Aspirin discovery with metabolomics

Colorectal cancer patients:

• Urinary biomarkers?

• Adipose tissue and metabolomics

Integration of –omics approaches

Flossman et al,. Lancet 2007

• Long-term follow up of British

Doctors Aspirin Trial and UK-TIA

Aspirin Trial

300, 500 or 1200 mg aspirin vs

control

• No risk reduction within first 5 years

• Risk reduction strongest:

10-14 years post randomization

• HR 0.51 (0.29-0.90)

Among compliant patients

Among long-term users (>5 years)

All of the above

• HR 0.26 (0.21-0.56)

Should we use aspirin for cancer

prevention?

• Aspirin is potent in cancer chemoprevention• However, GI symptoms, including in some cases serious

bleeding is a concern for the broad scale implementation

• Risk-benefit balance will differ by population Healthy, young individuals versus those with family

history, predisposing inflammatory or high-risk conditions, older, prior polyps, higher body mass index…

• Is pharmacogenetic tailoring possible?

• Personalized prevention:

Aspirin use

Benefits of pharmacogenetic tailoring

Being able to advice on aspirin, based on genetic predisposition

results in:

Risk minimization

Reduced concern among patients and treating clinicians

Reduced liability

Greater likelihood of response

Approaches to aspirin pharmacogenetics

Relevant targets:

Target pathways

• Efficacy and toxicity (prostaglandin synthesis,

leukotriene synthesis, thromboxane synthase)

• Metabolism and excretion (UGT, CYP2C9, MRP4-

for non aspirin NSAIDs)

Genome-wide association studies

Pharmacokinetic studies

All approaches are needed and complementary

>20 publications: Ulrich, Poole, Kulmacz et al

NSAID Pharmacogenetics

Meta-analysis & pooled analysis:colon & rectal cancer and adenoma,

NSAIDs and aspirin

• 4 individual study populations**

~10,000 individuals

• 2-stage approach based on

candidate pathways

Question: who benefits from NSAIDs

and who doesn’t?

** from Slattery, Potter, WHI, CCFR

Interaction between FLAP SNP and

NSAID use in risk of colorectal cancer

p for interaction

=0.038

Yes

No

0

0.2

0.4

0.6

0.8

1

1.2

GGCG/CC

Interaction between FLAP SNP and

NSAID use in risk of colorectal cancer

Ref

p for interaction

=0.038

Regular,

current use

of NSAIDs

FLAP genotype

0.97

1.02

0.68**Odds

Ratio

These signals are robust and replicate. When

can we take them to the clinic?

Discovery of new pathways: Metabolomics

in a randomized controlled trial of aspirin

• Randomized controlled trial

ABC Study (Johanna Lampe)

n=55

• Crossover study design

• 325mg ASS/d for 60d

• 3 month wash-out

• Reverse intervention

Each participant serves as their own control

Liesenfeld et al. Ca Epi Biom Prev 2016;25(1):180-7

Aspirin reduces

plasma 2-hydroxyglutarate

• ~30% reduction, p = 0.005 (robust)

• 2-HG is a known driver of carcinogenesis, particularly of brain

cancer (Losman et al. 2013)

Inte

ns

ity

Pla

ceb

o

Asp

irin

0

2 .01 04

4 .01 04

6 .01 04

8 .01 04

1 .01 05

U G T 1 A 6 g e n o ty p e

Inte

ns

ity

1*1

2*2

0

2 .01 04

4 .01 04

6 .01 04

8 .01 04

1 .01 05 P la c e b o

A s p ir in

2-hydroxyglutarate 2-hydroxyglutarate


Cell culture experiments suggest stereo-

specific reduction of 2-hydroxyglutarate with aspirin

• 500 µM aspirin (24 h):

reduction of (R)-2-HG in

6 of 8 colorectal cancer

cell lines;

Reduction: Ø = 20%,

p<0.01; range: 10 – 40%

• Effects achievable at

physiologic concentrations

• Salicylate inhibits the

HOT reaction

(R-2-HG formation)

Control ASS **: p < 0.01

All cell lines

S-2

-HG

R-2

-HG

0.0

0.5

1.0

1.5 **DLD1

S-2

-HG

R-2

-HG

0.0

0.5

1.0

1.5

s a lic y la te c o n c . [ lo g (n M ) ]

(R)-

2-H

G f

orm

ed

[nm

ol

* m

g-1 *

h-1]

0 2 4 6

0 .0

0 .5

1 .0

1 .5

2 .0


Aspirin pharmacometabolomics

• Resulted in the discovery of a potential new biologic

mechanism of aspirin in cancer chemoprevention

2-hydroxyglutarate is a known oncometabolite

• Confirmation in vitro and mechanism (HOT

inhibition)

ColoCare Patient Cohort

International Consortium Cohort Study

• Fred Hutchinson Cancer Research Center

• Moffitt Cancer Center

• Huntsman Cancer Institute

• NCT Heidelberg

Inclusion of patients

• With colorectal carcinoma stages I-IV

• n = 5000 in 5 years

Inclusion criteria:

• Women and men from the age of 18

• Newly diagnosed CRC of all stages

ColoCare as a resource

for novel scientific projects on cancer prognosisExamples:

Genomics GAME-ON, Oncoarray, gene expression, mutations

Metagenomics Next Gen Sequencing of the gut microbiome

Metabolomics Metabolic profiling of blood, adipose tissue, urine

Epigenomics Tumor methylation and miRNA

Proteomics Protein Array

Other biomarkers Prostaglandins, 25(OH)-vitamin D, 8-oxo-dG, …

Quality of Life Physical and emotional functioning and other QoL

Nutrition Dietary patterns and supplements

Physical activity State of the art physical activity (accelerometry)

Lifestyle factors NSAIDs, smoking, etc.

The ColoCare Study

Results to date –

Urine metabolomics

Metabolite map for the therapeutic effects on

colorectal cancer patients’ metabolomes.

Metabolites were mapped using the NIH WCMC

MetaMapR (https://github.com/dgrapov/MetaMapR/)

Liesenfeld et al. 2015. Metabolomics 11(4). 998-1012

Aromatic

compounds

gut microbiota

associated

• Pre-surgery vs. post-surgery:

metabolites presumably from

microbial origin were lower

• Several amino acids and

downstream products of

tryptophan metabolism were

altered

• Dopamine levels lower in

patients pre-surgery

PLS-DA model enabled good separation of groups (pre-surgery

vs. post-surgery)

The ColoCare Study

Results to date – Urine metabolomics

Receiver operator characteristic (ROC) curve and score plot from a

partial least squares-discriminant analysis (PLS-DA) model

containing 20 metabolites for all patients (red line) and a subset of

fasted patients (blue line)

Liesenfeld et al. 2015. Metabolomics 11(4). 998-1012

More than a storage organ

Composed of:

• Adipocytes

• Macrophages

• Fibroblasts

• Endothelial cells

Fully functioning endocrine organ that produces and secretes

adipokines, cytokines, hormones

Biology of adipose tissue

Ouchi, Nature Reviews, 2011

The ColoCare Study

Adipose tissue metabolomics in colorectal cancer patients

SAT

VAT

n=59 CRC patients

(stage I to IV)

LC-qTOF lipidome

GC-TOF primary metabolites

Microarray transcriptome

Pilot Study with NIH West Coast Metabolomics Center (O. Fiehn)

Goal – to understand metabolomics and transcriptomic differences

between the visceral and subcutaneous adipose tissue

Gene Expression Analysis: VAT / SAT

VAT

SAT

(n = 83, paired)

Principal Component Analysis

VAT

inflammatory

profile:

• Arachidonic acid

higher compared to

SAT

• Increased liberation

of arachidonic acid

through PLA2

• Based on gene

expression:

metabolic flux into

prostaglandins

anticipated

Liesenfeld, Grapov, Fahrmann et al. Am J Clin Nutr. 2015;102:433-43.

The ColoCare Study

Adipose tissue metabolomics & transcriptomics

illustrates inflammatory character of VAT

Adipose tissue metabolomics

– ratio of triglycerides to membrane lipids:

a marker for adipocyte size


• 9 unknown metabolites showed a linear trend of

increasing/decreasing concentration with increasing tumor stage

(raw p < 0.05, Kendall’s τ > 0.25 or < -0.25) when comparing

stages I to IV

• None of which remained significant after FDR correction…

ColoCare adipose tissue metabolomics suggests

biomarkers to be investigated further


Summary and outlook into the future

Precision prevention is important to translate chemoprevention

to the clinic

Metabolomics advances our understanding of prevention

• Discovery of new pathways

• Biomarkers for exposures

Power of integrative analysis

neli ulrich, ms, phd huntsman cancer institute, salt lake...

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