Under the microscope: an introduction to biomarkers

1
Under the microscope an introduction to
biomarkers

• Marc van de Vijver
• Netherlands Cancer Institute, AmsterdamThe
  Netherlands

2
What is a biomarker?

• Biomarkers
• anatomical, physiological, biochemical or
  molecular parameters associated with the presence
  and severity of specific disease states
• detectable and measurable by a variety of methods
  including physical examination, laboratory assays
  and medical imaging

3
What is a useful biomarker?

• A useful marker should meet two criteria
• can be measured reproducibly by means of a
  reliable and widely available assay
• conveys information about the disease that is
  meaningful to the physician and the patient

Source www.cancerdiagnosis.nci.nih.gov
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Potential uses for biomarkers in oncology
Screen for disease
Distinguish between benign versus malignant
processes
Assess risk of developing disease
Biomarker
including staging
Monitor disease status before and after therapy
Predict response to therapy
Determine prognosis independent of therapy
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Prognostic versus predictive an important
distinction

• Prognostic markers
• indicate the likelihood of outcome (tumour
  recurrence or patient survival) regardless of the
  specific treatment the patient receives
• Predictive markers
• indicate the likelihood of response to a specific
  therapy

Source www.cancerdiagnosis.nci.nih.gov
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Molecular biomarkers

• Gene expression
• microarray technology
• quantitative reverse transcription-polymerase
  chain reaction (RT-PCR)
• Gene amplification
• fluorescent/chromogenic in-situ hybridisation
  (FISH/CISH)
• Gene sequence
• DNA sequencing (other methods possible for known
  changes)
• Protein expression
• immunohistochemistry (IHC)
• enzyme-linked immunosorbent assay (ELISA)
• Only IHC, FISH and ELISA are widely established
  in clinical pathology laboratories
• other methods are limited to clinical research
  laboratories

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Molecular diagnostics in cancer current and
future technologies
21st century (?)
mRNA expression arrays Multiplex PCR
19th century
1980s
2000
Proteomics
Tumour type Histology Patient characteristics
Single-gene/protein predictors
Multi-gene/protein predictors
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Example of IHC EGFR
Coloured product
Substrate
Secondary antibody specific for primary antibody
type and conjugated to enzyme
Formalin-fixed, paraffin-embedded tumour tissue
Primary antibody specific for EGFR
EGFR
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Example of FISH testing HER2
Formalin-fixed, paraffin-embedded tumour tissue
Hybridisation with fluorescent HER2 gene
probe (CEP17 probe)
Microscopicevaluation offluorescent signals
(counting gene copies)
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Challenges in developing clinical biomarkers

• Tissue availability
• Assay methodology
• Clinical validation

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Practical example HER2 testing in breast cancer

• HER2-positive status has both prognostic and
  predictive value in breast cancer
• shorter overall survival time
• earlier relapse after adjuvant chemotherapy
• required for response to Herceptin1

1Slamon DJ, et al. N Engl J Med 200134478392
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The benefit of using diagnostic testsin clinical
trials Herceptin in MBC
1.0 0.8 0.6 0.4 0.2 0
1.0 0.8 0.6 0.4 0.2 0
Herceptin paclitaxel Paclitaxel
Herceptin paclitaxel Paclitaxel
Probability of survival
Probability of survival
40
17.9
24.8
0 12 24 36 48
0 5 10 15 20 25 30 35 40
Months
Months
Herceptin combination pivotal trial overall
survival (IHC 3)
Simulation of effect of Herceptin in unscreened
population
MBC metastatic breast cancer
13
HER2 testing algorithm
Adapted from Bilous M, et al. Mod Pathol
20031617382
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General issues IHC

• IHC is widespread in pathology laboratories
• Important to ensure that laboratory is using
  correct test and that this has undergone adequate
  quality control

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General issues FISH

• More widespread in pathology laboratories since
  HER2 testing required
• not as much experience as with IHC, but is
  increasing
• FISH is generally only performed in IHC 2 cases
  (approximately 10)
• in a centre dealing with 200 breast cancers per
  year, this equates to approximately two FISH
  tests per month

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Interlaboratory concordance of HER2 testing

• Local IHC versus central IHC
• 77.5 (95 CI 0.460.55, n1,536)
• Local FISH versus central FISH
• 92 (95 CI 0.730.93, n131)

Standardisation, validation and quality control
required
Press MF, et al. Clin Cancer Res 2005116598607
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Summary

• To date, relatively few markers have been
  accepted for routine clinical use by regulatory
  authorities or major oncology groups
• reflects the lengthy process of assay development
  and validation
• Despite 5 years of extensive study, HER2 testing
  in breast cancer has still not been fully
  optimised or standardised
• False results have major implications
• may deny patient access to a life-extending
  treatment
• patients receive a therapy that they are unlikely
  to benefit from
• When well defined, biomarkers can play a
  significant role in the management of cancer

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Conclusion the benefits of biomarkers

• Despite the challenges involved, biomarkers have
  the potential to deliver great benefits to our
  patients

Screening
Who has cancer?
Prognostic markers
Helps guide treatment
Predictive markers
Which treatment is best?
- rapid access to most appropriate therapy -
improved cost-effectiveness