Road-map for translation of

Cancer Risk biomarkers into

clinical practice, individual

risk assessment and

prevention.

Stefano Bonassi, PhD, ERT

Department of Human Sciences and Quality of Life

Promotion San Raffaele University, and Unit of Clinical

and Molecular Epidemiology IRCCS San Raffaele Pisana;

Rome, Italy

Porto, Portugal 27 February 2020

2

A biomarker is any

substance, structure or

process that can be

measured in the body or

its products and

influence or predict the

incidence of outcome or

disease (AIRC, 1997)

3

A biomarker is any

substance, structure or

process that can be

measured in the body or

its products and

influence or predict the

incidence of outcome or

disease (AIRC, 1997)

Genetic background

+Exposure to Environmental Carcinogens

Cancer

BIOLOGICAL EVENT:e.g., symmetrical translocationin the target organ

BIOMARKER:Unstable aberrations in PBL

BIOMARKER:Micronucleated cells in PBL

Genetic background Environmental agents

Cancer

+

BMK Early detection, diagnosis

BMK Prevention

MN and Haematological Cancer

MN and Oral Cancer, Head and Neck Cancers

MN and Colorectal Cancer

MN and Cervical Cancer

MN and Bladder Cancer

MN and Lung Cancer

MN and Prostate Cancer

MN and Breast Cancer

MN and Skin Cancer

Bonassi & Fenech, Micronuclei and Their Association with Infertility, Pregnancy

Complications, Developmental Defects, Anaemias, Inflammation, Diabetes, Chronic Kidney

Disease, Obesity, Cardiovascular Disease, Neurodegenerative Diseases and Cancer

The Micronucleus Assay in Toxicology. Chapter 4. Royal Society of Chemistry, 2019

Meta-analysis of 22

epidemiological

studies of 32P-

postlabeling DNA

adducts and lung

cancer risk in

bronchial cells

according to

smoking habits.

(Ceppi et al., 2017)

Reverse causality occurs

when the probability of the

outcome is causally related

to the exposure being

studied.

Past Present

Cancer

cases

Biomarker

+

Biomarker

-

Hagmar et al., Cancer Res, 1998

European Study Group on Cytogenetic

Biomarkers and Health - ESCH

Chromosomal

aberration

frequency in

lymphocytes

predicts the

risk of cancer:

results

from a pooled

cohort study of

22 358

subjects in 11

countries

Bonassi et al.,

Carcinogenesis, 2008

Country Subject

s

Cancer

Cases

Person

Years

Period of

Test

CA frequency

mean (sd)

Croatia 1,320 24 11,148 1982-2000 3.5 (2.3)

Hungary 840 46 6,823 1978-2001 1.9 (2.1)

Lithuania 812 24 7,478 1981-2002 3.3 (2.0)

Slovakia 2,994 55 27,135 1985-2001 2.1 (1.7)

Poland 456 22 5,012 1981-2001 1.5 (1.8)

Cekia 11,991 363 114,712 1975-1999 2.4 (1.8)

Sweden 756 48 12,740 1970-1987 1.8 (1.8)

Norway 471 48 7,936 1970-1988 1.5 (1.3)

Denmark 197 6 2,456 1987 1.8 (1.5)

Finland 557 39 10,189 1974-1988 2.3 (2.1)

Total incidence 20,394 675 205,629 1970-2002 2.4 (1.9)

Italy 1,964 99 31,818 1965-1995 3.6 (4.2)

Total 22,358 774 237,447 1965-2002 2.5 (2.2)

Bonassi et al., Carcinogenesis, 2008

CA

frequency

Cases RR 95% CI p

LOW 152 1.0 -

MEDIUM 240 1.31 1.07-1.64 0.006

HIGH 283 1.41 1.16-1.78 <0.001

All cancers (140-208)

Bonassi et al., Carcinogenesis, 2008

WWW.HUMN.ORG

Link with the disease

Numerical and structural chromosomal instability is one of

the main hallmarks of cancer and MN assays are amongst

the best validated biomarkers of this disease.

Mechanism

The role of MN in cancer has been further supported by

recent studies showing that chromosomes entrapped in MN

are shattered and massively rearranged leading to the

localised hypermutation state often seen in cancer.

Link with the exposure

Furthermore, MN assays are internationally endorsed

methods for determining the mutagenic and carcinogenic

potential of chemicals in vivo and in vitro.

(Bonassi & Fenech, The Micronucleus Assay in Toxicology. Chapter 4.

2019)

Belgium a) 146 0 599 3.9 (2.3)Bulgaria 208 4 1,264 41.1 (28.6)Croatia 202 4 1,841 59.0 (41.5)Italy a, b) 2,654 56 25,47 64.6 (4.2)Japan 833 146 7,935 52.2 (14.6)Poland 205 11 1,839 14.3 (10.0)Slovakia 926 10 7,735 12.1 (6.1)Sweden 677 31 10,440 4.1 (2.7)Taiwan 203 2 1,443 13.0 (11.1)Yugoslavia 676 4 4,408 11.6 (9.3)

Total 6,730 268 62,980 15.5 (21.0)

a) Mortality Follow up;b) Incidence data were available for 253 subjects (1,460 person-years, 7 cancer cases):

Country Sbj. Cases PY MN ‰

Cases Subjects RR* 95% CI

MN frequency

Low 51 1,313 1 Ref.Medium 91 1,433 1.84 1.28-2.66High 77 1,435 1.53 1.04-2.25

* adjusted for age, gender, smoking habit, occupational exposure, and laboratory.

CytotoxicActivity of PBL

Cases RR 95% CI p

HIGH 45 1.0 -

MEDIUM 41 1.70 1.15-2.50 <0.01

LOW 68 1.59 1.09-1.32 <0.01

All cancers (140-208)

Imai et al., Lancet 2000

Natural Cytotoxic activity of PBL and cancer

incidence (11 years FU study of a general

population)

The hCOMET cohort

Overall Mortality

LogRank test p<0.02

Total Cancer Incidence

Covariate Hazard

Ratio

95% CI P

value

Tail Intensity

2 Tertile (Medium)

3 Tertile (High)

1.11

1.06

0.78-1.57

0.76-1.48

0.55

0.75

Sex

Males 1.16 0.83-1.60 0.38

Age

5 years increase 1.10 1.08-1.11 0.001

Smoking habit

Current smoker 1.41 1.02-1.94 0.039

Genotoxic exposure

Exposed 3.73 2.08-6.69 0.001

hCOMET cohort: Cox Regression analysis

What now besides mechanistic implications ?

Surrogate endpoint of cancer at individual level, i.e., Clinical trials, Chemoprevention, Individual Risk Assessment, etc.

Cancer Risk prediction in groups exposed to genotoxins, mutagens, or bearing a susceptible genotype or phenotype.

1 2

To establish a biomarker as a Cancer Risk

Biomarker useful for individual risk assessment

some basic requirements have to be addressed:

Mechanisms known/

Association with the

disease established

Measurement of the

biomarker valid,

reproducible, repeatable,

standardized

The predictive link with

the disease is well

established and

quantitatively strong

enough to be effective at

individual level

An early identification of

individual at risk because

of altered levels of the

biomarker may contribute

to modify the prognosis

of the disease

Chromosomal

Instability/DNA

damage

Total

Cholesterol

Cancer CHD

Slide Source:

Lipids Online Slide Librarywww.lipidsonline.org

Relative Risks of CHD by Quintiles of LDL-C and HDL-C in ARIC Men

*Adjusted for age and race.

4.50

2.85

1.80

1.15

0.75

LDL-C (mg/dL)

Rela

tive R

isk o

f CH

D* 1.00

0.60

0.35

0.20

0.15

Adapted from Sharrett AR et al. Circulation 2001;104:1108-1113.©2001 Lippincott Williams & Wilkins. www.lww.com

Rela

tive R

isk o

f CH

D*

HDL-C (mg/dL)

90.9

110.2

130.0 168.2

149.3 187.5

28.6

36.3

44.1 59.6

51.8

Framingham Heart Study TG Levels and Relative Risk of CHD

Castelli WP. Ann Epidemiol 1992;2:23-28.

0.0

0.5

1.0

1.5

2.0

2.5

3.0

50 100 150 200 250 300 350 400Triglycerides (mg/dL)

Rela

tive R

isk

Men

Women

Total Cholesterol Distribution: CHD vs Non-CHD Population

35% of CHD

Occurs in People

with TC<200 mg/dL

150 200

Total Cholesterol (mg/dL)

250 300

No CHD

CHD

Framingham Heart Study—26-Year Follow-up

Road-map for translation of

Risk biomarkers into clinical

practice, individual risk

assessment and prevention.

Lesson learned

from the

Cholesterol story !

32

1° LEVEL

To discriminate between groups of subjects affected /healthy

• Provide useful hints and mechanistic insight and may indicate

potential therapeutic target.

Possible use of Biomarkers of DNA damage in individual

risk assessment/clinics

33

1° LEVEL

To discriminate between groups of subjects affected /healthy

• Provide useful hints and mechanistic insight and may indicate

potential therapeutic target.

2° LEVEL

From population to individuals

• geno/fenotipic susceptibility to cancer and other NCD’s

• Subject with risk of adverse events (safety)

• Enrichment designs (es HerB2)

• Early identification of responders

Possible use of Biomarkers of DNA damage in individual

risk assessment/clinics

Our systematic review indicates a potential

usefulness of the buccal micronucleus assay in the

prescreening and follow up of patients at risk for oral

and head and neck cancer.

Clinical application of micronucleus test in exfoliated

buccal cells: A systematic review and metanalysis,

Bolognesi et al., Mutat Res Review, 2015

Our systematic review indicates a potential

usefulness of the buccal micronucleus assay in the

prescreening and follow up of patients at risk for oral

and head and neck cancer.

A main limitation of the assay, which needs to be

addressed in the perspective of a practical

application, is the large variability in the MN

frequency observed across the laboratories in

patients as well as in control groups. The main

source for this variability is related to the different

experimental protocols and scoring criteria applied

Clinical application of micronucleus test in exfoliated

buccal cells: A systematic review and metanalysis,

Bolognesi et al., Mutat Res Review, 2015

Clinical and Genomic safety of treatment with Gingko

Biloba: A Randomized Clincal Trial. Bonassi et al.,

BMC Complementary and Alternative Medicine, 2018

37

1° LEVEL

To discriminate between groups of subjects affected /healthy

• Provide useful hints and mechanistic insight and may indicate

potential therapeutic target.

2° LEVEL

From population to individuals

• geno/fenotipic susceptibility to cancer and other NCD’s

• Subject with risk of adverse events (safety)

• Enrichment designs (es HerB2)

• Early identification of responders

3° LEVEL

Validate the use of Biomarkers in real life

Cohort studies, Intervention studies

Randomized Clnical Trials

Possible use of Biomarkers of DNA damage in individual

risk assessment/clinics

Gould AL et al. Circulation. 1998;97:946-952.

Clinical Benefits of Cholesterol Reduction• A meta-analysis of 38 trials demonstrated

that for every 10% reduction in Total

Cholesterol

–CHD mortality decreased by 15%

(P<0.001)

– total mortality decreased by 11%

(P<0.001)

• Decreases were similar for all treatment

modalities

• Cholesterol reduction did not increase non-

CHD mortality

Thank you