BASIC SCIENCE RESEARCH
Lecture 1: BSR Introduction & Scientific Fraud
I. TYPES OF MEDICAL RESEARCH
A. Basic Research
 Purpose: to advance scientific knowledge; immediate
practical application not being a direct objective
 Totally unpredictable
 Requires long term commitment
 Requires training in techniques in fields outside medicine
 No initial connection between the research and its medical
application
Reference parameters to ensure animal welfare:
 The Five (5) Basic Freedoms are the reference parameters
for animal welfare to be in place in an animal facility:
o Freedom from thirst, hunger and malnutrition
o Freedom from physical discomfort and pain – proper
anaesthesia and analgesia
o Freedom from injury and disease
o Freedom to conform to essential behaviour patterns
o Freedom from fear and distress 
Figure 1. Pain in Lab Mice: 0=Not present, 1=Moderate, 2=Severe
Using Animals in Research, Testing and Teaching
 The principles of:
o REDUCTION: Just enough sample size; reduce the
number to a number that is scientifically viable
o REFINEMENT : Refine the method to avoid harm
o REPLACEMENT: Make sure there is no in-vitro model
that can be used as replacement for an experiment
before using animals ; Once the animal is dead  there
is no need for (animal ethics) ethical clearance
B. Clinical Research
 Once you start to have human subjects  becomes clinical
research, regardless if healthy or with a disease
1. Epidemiological and behavioural studies determine:
o Prevalence ; Incidence; Distribution of Disease; Factors
that affect health;
o How people make health-related decisions
2. Outcomes and health services research: studies that
seek to identify the most effective and most efficient
interventions , treatments and services
3. Patient-oriented research: research that involves a
particular person or group of people (case series) or uses
materials from humans
o To understand the pathogenesis of human disease
o Studies on interventions for disease or therapeutics
o Develop biomedical devices
C. Clinical Trials
 Controlled studies on human subjects  evaluate the safety
and effectiveness of new drugs or devices or of behavioural
interventions
o "No approved therapeutic claims" - if not done  REMEMBER:
 Safety - adverse events
 Informed consent required
BASIC SCIENCE RESEARCH | Block #
BLOCK #
D. Translational Research
 Mantra is "bench to bedside and back"
 Combination of basic and clinical research
 Links the scientific advances to patient care
 Translates basic biomedical research into effective new
therapies - and to bring clinical observations back to the lab 
Types of Medical Important to remember
Research
Basic Research Unpredictable
Long term commitment
Does not require human subjects
Clinical Research Requires human subjects
regardless if healthy or with a
disease
Clinical Trials Requires human subjects to test
safety and effectiveness of new
drugs
Translational Research Combination of basic and clinical
research
II. RESEARCH AND DEVELOPMENT PROCESS
 Drug Discovery Phase – Before you reach Phase I, you
have to have enough pre-clinical evidence that warrants you
to go forward
 Clinical Trials
o Phase 1 : checks safety; tested on healthy individuals
with at least 6 months wash out period
o Phase 2 : checks efficacy
o Phase 3 : checks BOTH safety and efficacy
o Phase 4 : post marketing surveillance
 GAP: Standardization of the agricultural properties of the
plant; When and where the plant was harvested, what part of
the plant is used, what weather do you harvest the plant,
what level of processing
 GMP: Ensures anything given to humans is free from any
potential risk of causing harm
III. SCIENTIFIC FRAUD
Definition
 Intentional misrepresentation of the methods, procedures, or
results of scientific research 
 Includes: fabrication, falsification, or plagiarism in proposing,
performing, or reviewing scientific research, or in reporting
research results.
 Why is scientific fraud an issue? Because of research
integrity
A. Types of Fraud
1. Misdemeanors
o Improper credit to colleagues and collaborators
o Re-publishing content
o Plagiarism
o Unethical use of peer review 
2. Felonies
o Misrepresenting results
 Cooking: Retaining results that fit the data
 Trimming: Adjusting data to make it look extremely
accurate
 Forging: Fabricating data or entire experiments 
B. Causes of Scientific Fraud
 Stiff competition for research funds ; Fight for recognition
 Pressure to publish – to have a good position/ tenure
 Difficult experiments that cannot be repeated (funds)
 Conflicts of interest  (company-funded influence)
 Whenever you see a publication online, value where it is
indexed and not just rely on Google (not sure if
reliable/trustworthy)
 Make sure they are indexed somewhere that does the
proper screening
Page 1 of 8
 Lecture 2: Ethical Issues: The Pros and Cons will be protected and will
receive recognition as
PROS CONS well as fair
Viral - use of informed - social stigma compensation and
Transmission consents - safety benefits.
Studies - Cohort studies - knowledge can be - secured and regulated
(observation of the misused research
course of the disease) - further mutations Prognostic - empowers the patient: - compromised
- used in vaccine testing Development provides more accurate reliability
- The need of viral details about their - not specific; too
transmission studies is condition and allows general
very great. them to decide on their - An ideal model is
- focus on the refinement treatment plan difficult to achieve.
of the methods used - time consuming
Embryonic - treat diseases - full moral status of and large data sets
stem cell - discover new the embryo are necessary
medicines and vaccines - use of terminology - expensive with no
without harming that is deliberately guarantee on return
thousands of people misused of investment
- does not necessarily - currently has
“kill life” major I. 3 MAJOR DOCUMENTS THAT REGULATE STUDIES THAT
disadvantages
INVOLVE HUMAN SUBJECTS
which outweigh its
promised benefits A. Nuremberg Code
Recombinant - fewer food shortages - with associated 1. The voluntary consent of the human subject is absolutely
DNA and safer foods health risks essential.
- used to breed plants of - trials have only 2. The experiment should be such as to yield fruitful results
better growth, survival released 3-month for the good society.
and availability long research 3. The e xperiment should be so designed and based on the
- increase diversity of period results of animal experimentation and knowledge of the
plant species - main ethical issue: natural history of the disease.
- reduced dependence of application of the 4. The experiment should be so conducted as to avoid all
plants on chemical technology itself unnecessary physical and mental suffering and injury.
products such as 5. No experiment should be conducted where there is a prior
pesticides reason to believe that death or disabling injury will occur.
- decrease or eliminate 6. The degree of risk to be taken should never exceed that
the allergenic proteins in determined by the humanitarian importance of the problem to
specific foods be solved by the experiment.
- helps in the prevention 7. Proper preparations should be made and adequate
of malnutrition and other facilities provided to protect the e xperimental subject against
associated diseases even remote possibilities of injury, disability or death.
- used in the production 8. The e xperiment should be conducted only by scientifically
of human insulin, qualified persons.
synthetic blood clotting 9. During the course of the e xperiment, the human subject
factor VIII and various should be at liberty to bring the experiment to an end.
antigens for diagnosis of
certain diseases B. Declaration of Helsinki
- used in Western blot to - Principles on:
screen for HIV  safeguarding research subjects
Reproductive - alternative way of - issues with value  informed consent
Cloning reproduction of life  minimizing risk
- capable of protecting - newborns  adhering to an approved research plan/protocol
the rights of clones developed genetic
defects, missing C. Belmont Report
body organs, - framework based on three discussions:
premature aging 1. Boundaries between practice and research
- emotional impact 2. Basic ethical principles
- possibility of a. respect for persons
abuse if successful b. beneficence
Synthetic - create enzymes for - unnatural c. justice
Cell bolstering biofuels - tends to mutate 3. Applications
- construction of more rapidly and
microorganisms deteriorate easily
- enzyme therapy - could disrupt the
- gene therapy local fauna through Lecture 3: Scientific Writing
- encapsulated cells competition or I. REASONS TO WRITE
- artificial oxygen carriers infection
- will not achieve its  What motivates you to write and publish?
potential until  What is the most common barrier/problem for writing?
scientists can  Aim of scientific communication: to put your messages
predict accurately across in a technical way.
how a new genetic  Other reasons to write (Martin Welch, 2004):
circuit will behave  To make a permanent, publicly accessible record of your
inside a living cell findings in a timely manner
Gene - protects therapeutic - may lead to  To avoid unwarranted repetition
Patenting proteins like insulin monopolization of  To convince funding agencies to provide more support for
- gives opportunities for genes your research
investment in generally - issue on patenting  Significant influence to your future career, job promotion. It
unprofitable research of genes adds to your credib ility if you give a lecture, you cite your
- Researchers are own findings or words.
assured that their work  Improve health outcomes

BASIC SCIENCE RESEARCH | Block # Page 2 of 8

  Contribute to evidence-based medicine o References – most relevant and most recent (20-35
 Where to publish references)
 Well respected and peer reviewed journal o Total pages: 12-20 pages
 Formal indicator of work performance  Writing style (Richard Smith, editor BMJ)
 Ultimate marker of research success o Keep it as simple as possible - No highfalutin words, no use
 Product, policy, translated knowledge of superlatives
 Greater chance to be published o Short words
 Study must have a rigorous design o Short paragraphs
o Stick to nouns and verbs
 Results must answer an important question
 What is written without effort is in general read without
 Your paper must be written well
pleasure (Samuel Johnson, 1709-1784)
 Give it a priority
o Scientific writing is a well-defined technique rather than a
 Good time management skills creative art.
 Ego. Your work is your epitaph
 Basic aspects:
o Thought – worthwhile results
II. TYPES OF PUBLICATION o Structure – right things in the right place
1. Original research articles o Style – fewest and most appropriate words, using the rule of
2. Review papers good grammar
3. Perspective commentaries and opinion pieces
• Planning stage
4. Rapid communications • Identify the questions to be answered, the analyses to be reported and the target
journals
Research Papers
 Original full-length research papers which have not been • Set framework for document
published previously, e xcept in preliminary form, and • (page size, beadings, etc.)
• put ideas on paper, plan topic: sentences, construct tables and figures
which should not exceed 7,500 words (including allowance
for no more than 6 tables and illustrations). Nowadays
3,000-3,500 words are set to b e followed but are not • Grotty first draft
enough for a thorough explanation of a paper. • use journal checklists and instructions to authors

Review Papers
 Synthesis of written papers; Areas of topical interest, • Presentable second draft
normally focus on literature published over the previous • circulate to coauthors

five years, and should not exceed 10,000 words (including

allowance for no more than 6 tables and illustrations).
• Good third draft
Perspective Commentaries and Opinion Pieces • circulate to papers and coauthors

 These should be concise, [focus] on hot topics and

describe cutting-edge developments and technologies.
• Excellent fourth draft
They should not exceed 2000 words. • polish up presentation and revisit checklists
Rapid Communications
 Short communications of up to 300 words, describing work
• Final document
that may be of a preliminary nature but which merits • submit to journal
immediate dissemination.

III. BARRIERS FROM A WRITING RETREAT

V. CHOOSING THE JOURNAL
 Making time to write  Who are your audience – general, clinical or specialty
 Distractions journal, basic science
 Getting started  New journals – more likely to be accepted but low impact
 Writing in chunks factor, limited circulation, not wide-reach audience.
 Perfectionism  High ranked – harder to get into, long wait times, read by
 Reworking a thesis experts in the field, maybe rejected, but received pertinent
 Permission to write
 Fear of rejection VI. AUTHORSHIP

IV. FORMAT OF MANUSCRIPT  Decide before the study will be implemented

 Vancouver guidelines on authorship:
1. Title
2. Abstract (structured) – required b y most journals nowadays; - Each author should have participated sufficiently in the
not in paragraph form anymore work to take full responsibility for the content
3. IMRAD:
a. Introduction Authorship Credit
b. Materials and Methods  Conditions a, b, and c must all be met. An y part of an article
c. Results critical to its main conclusions must be the responsibility of at
d. Discussion least one author. Editor may require authors to justify the
4. References assignment of authorship.
5. Acknowledgements a. Substantial contribution to concept and design, or analysis
and interpretation of data; and to
 You can turn a messy bunch of “academic stuff” into an article b. Drafting the article or revising it critically for important
quickly – but you must have: intellectual content
o Data, ideas or artefacts c. Final approval of the version to be published
o Preliminary analysis of thoughts
 Topics can be written in many ways depending how you First Author – most important
phrase the title  Takes primary for all aspects publishing the paper; main point-
 Basic Structure: person responsib le of the paper
o Introduction – why you did the study, the objectives  Conducts and supervises the data analyses and interprets the
o Methods – how you did it results
o Results – what you found  Writes the paper in consultation with co-authors
o Discussion – the implications of the findings  Maintains ownership of the master document
 Interpret in the context of the other literature  Submits the paper to a journal and deals with the
correspondence

BASIC SCIENCE RESEARCH | Block # Page 3 of 8

 Responsible for archiving and documenting all the data and o Following formatting rules
files
• Describe study sample
Co-authors Paragraph 1 • Who did you study
 Make early decisions about the aims of the paper
 Keep the paper on track in terms of the main messages
 Make intellectual contributions to the data analysis • Univariate analyses
• How many participants had w hat?
 Contribute to the interpretation of the results Paragraph 2

 Review each draft

 Take public responsibility for the content and results • Bivariate analysis
 JAMA and NEJM ask authors to testify that the y meet the Paragraph 3
• What is the relation betw een the outcome and the
to n-1 explanatory variables
Vancouver Criteria
 BMI and Lancet – requires a statement of each author‟s • Multivariate analyses
contribution • What is the result w hen the confounders and the effect
 Statement of no conflict of interest is also required
Final
paragraph/s modifiers have been taken into account

VII. ABSTRACT
 100-200 words Reporting Numbers
 Condensed summary of the paper Rule Correct Expression
 Aims of the study: “The paper explores…” Numbers less than 10 are In the study group, eight
 Main argument: “In this paper we argue that…” words. paticipants underwent the
 Study design intervention.
 Methods (setting, participants, outcome measures) Numbers 10 or more are There were 120 participants in
 Main result – data and stat significance numbers the study.
- What‟s new?: “this paper contributes to the debates on…” Words not numbers begin a Twenty per cent of the
 Conclusion sentence participants had diabetes.
Be consistent in the lists of In the sample, 15 boys and 4
numbers girls had diabetes.
VIII. INTRODUCTION
Numbers less than 1 begin The P value was 0.013.
 One page with a zero
 Background – what we know Do not use a space between a In total, 35% of participants
number and its per cent sign had diabetes.
 Current knowledge of the research area
Use one space between a The mean height of the group
 What other people have done number and its unit was 170 cm.
- Limitations encountered: what we do not know / gaps in
Report percentages to only In our sample of 212 children,
the knowledge
one decimal place if the 10.4% had diabetes.
 Questions still unanswered sample size is larger than 100
 Last paragraph will answer what you did and why Do not use decimal places if In our sample of 44 children,
 Research problem / why we did the study the sample size is less than 10% had diabetes.
100
Paragraph Paragraph Paragraph
1 2 3 XII. RESULTS
• What we • what we • why we did
know don't know this study  Summary of results as per objective
 Limitations of the study
 How results agree/disagree with the body of knowledge
 Future directions
IX. REVIEW OF RELATED LITERATURE
 Impact on current knowledge
 Includes related lit which are the most relevant and most valid
studies
• What did this study show?
 Includes other related lit in the discussion Paragraph • Address the aims stated in the introduction
 Don‟t put textbook knowledge like definition of the disease 1

• Strengths and weaknesses of methods

Paragraph
X. METHODS 2

 2-3 pages
 Study design Paragraph
• Discuss how the results support the current literature or refute current knowledge

 Sampling design, selection of participants, sample size

3 to n-1

o Variables • Future directions

• "So what?" and "where next?"
o Data collection methods Final
paragraph • Impact on current thinking or practice
o Data plan of analysis
o Development of questionnaire
o Ethical considerations XIII. GRAMMAR AND SYNTAX
o Approval by an institutional ethics review board
o Informed consent How to Write Clear, Concise, and Direct sentences :

XI. RESULTS 1. UNLESS YOU HAVE A REASON NOT TO, USE THE
ACTIVE VOICE. Meaning use the 1 st person: “I”
 2-3 pages  At the heart of e very good sentence is a strong, precise verb;
 Sample coverage the converse is true as well--at the core of most confusing,
 Profile of study participants awkward, or wordy sentences lies a weak verb.
 Descriptive analysis and univariate statistics (means, SDs) There are sometimes good reasons to us e the passive voice:
 Bivariate analysis and multivariate analysis of associations
A. To emphasize the action rather than the actor.
 Tables and figures: 3-6 pages - E.g. After long debate, the proposal was endorsed by the
 Discussion: 2-3 pages
long-range planning committee.
 The whole manuscript is 2,000-2,500 words, 8-10 double- B. To keep the subject and focus consistent throughout a
spaced pages passage.
 Tables and Graphs
o Not more than 6 tables and/or graphs

BASIC SCIENCE RESEARCH | Block # Page 4 of 8

 - E.g. The data processing department recently presented Connecting Discrete Data Points
what proved to be a contro versial proposal to expand its staff.
After long debate, the proposal was endorsed by…
C. To be tactful by not naming the actor
- E.g. The procedures were somehow misinterpreted.
D. To describe a condition in which the actor is unknown or
unimportant.
- E.g. Every year, thousands of people are diagnosed as
having cancer.
E. To create an authoritative tone.
- E.g. Visitors are not allowed after 9:00 p.m.

2. PUT THE ACTION OF THE SENTENCE IN THE VERB.

 Don't bury the action in a noun or blur it across the entire
sentence.
 Watch out especially for nominalizations (verbs that have been
made into nouns by the addition of -tion)
3. REDUCE WORDY VERBS.
4. USE EXPLETIVE CONSTRUCTIONS SPARINGLY.
("It is”, "There is”, "There are")
5. TRY TO AVOID USING VAGUE, ALL-PURPOSE NOUNS,
WHICH OFTEN LEAD TO WORDINESS.
(“factor”, “aspect”, “area”, “situation”, “consideration”, “degree”,
“case”)
6. UNLESS YOUR READERS ARE FAMILIAR WITH YOUR
TERMINOLOGY, AVOID WRITING STRINGS OF NOUNS (OR
NOUN STRINGS).
7. ELIMINATE UNNECESSARY PREPOSITIONAL PHRASES.
8. AVOID UNNECESSARILY INFLATED WORD.
9. PUT WORDY PHRASES ON A DIET.
 Usual CONTENT errors:
1. Nonessential data are included
2. Redundancy (text, table, graph)
3. Too many significant figures in tables (excessive precision)
4. Inadequate definition of abbreviations and symbols
5. Not self-explanatory (when a graph cannot be fully interpreted
when isolated from the main text)
Regression Line Beyond the Range of Data
 Extrapolation of regression line beyond the set of measured
data should not be done. Never extend the line. It is not a b ell
curve.
 Regression line has a starting and an end point.

XIV. TABLES, GRAPHS, FIGURES

 Well-structured and organized tables, graphs, & figures:
o to display data and trends, and
o to summarize information

Advantages of Graphs over Texts

1. Facilitates the understanding and interpreting of complex data
& relationships
2. Decreases the reading time by highlighting and summarizing
major trends or findings
Typeface
3. Reduces the overall word count of the article

A. What NOT to do
 Usual DESIGN errors:
1. Too simple tables
o information could be included in the text
2. Too large tables
o readers would have a hard time to follow
3. Failure to use bordering and shading in tables
o these techniques could improve readability of the tables
4. Wrong choice of graphical scale and/or format to depict data
5. Use of 3D graphs instead of 2D
o even though the use of 2D graphs would be s ufficient
o 3D graphs are hard to interpret.
6. Design elements obstruct with the clearness of a figure or
graph.

3D vs 2D graph B. What to do
 Authors should ask themselves
o Is it worth it to put any data into a figure, graph, or table?
o Will the graph, table or figure contribute something to the
article?
 All the illustrations should add to or supplement the text and
should deliver the necessary information.
 Avoid repetition
o Creating a graph/table that just duplicate the text or contain
extraneous data will only mess up the manuscript and burden
the readers/reviewers
 Make a decision
o What data are essential
o Best way to depict the data
 GRAPHS: better used at showing/illustrating complex
relationships
 TABLES: better used when exact values are important
 “The distribution of the results should be the author‟s guides
when choosing the graphic format that displays the optimal

BASIC SCIENCE RESEARCH | Block # Page 5 of 8

 amount of detail necessary to accurately tell the story of the
experiment.” - Cooper RJ et al. 2002
C. Some important Reminders
1. Authors utmost concern should be reader understanding.
2. Authors must consider the publication requirements, limits, and
color-printing costs for illustrations set by journal.
3. Authors should consider that GTF should be fully
understandable even outside of the context of the paper or
abstract.
4. Authors should do a thorough, precise titling and labeling of all
components and units (graphs).
5. Align the entries, use minimum significant figures, and include
statistical values when applicable (tables).
6. Figures, which include flowcharts, photographs, diagrams, or
line drawings, should only include simple text.
7. Aim for individual figures, tables, and graphs that fit onto 1
page, with readable scales.
8. Read the requirements of the journal.
Lecture 4: Biomarker Technology
I. HISTORY OF BIOMARKERS
 Term introduced by Karpetsky, Hymphrey and Levy in April
1977 edition of J National Cancer Institute
 Early Biomarkers include uroscopy, blood pressure, ECG,
hematology, blood and urine chemistry
 Genomics, Proteomics, Metabolomics – foundations for the
molecular basis of disease
a) 1980s – 1990s: Molecular Biomarkers were used as viral
and immune indicators of disease. They were used to
distinguish responders to targeted therapies.
b) S&T Advances in Biomarker Research include completion
of the human genome project
c) Biomarkers Consortium in 2006 was a public-private
initiative with industry and government to spur biomarkers
development and validation projects in cancer, CNS and
metabolic disorders
II. DEFINITION OF TERMS
 Biomarker - A biologic indicator of health or disease; A
characteristic that is measured and evaluated objectively as an
indicator of normal biologic processes, pathogenic processes,
or pharmacologic response to therapeutic intervention; Any
parameter of a patient that can be measured: mutations,
mRNA expression profiles, lipids, proteins,imaging methods,
electrical signals
 Surrogate endpoint marker- A biomarker that can substitute
for a clinical endpoint and is expected to predict clinical benefit
or harm, or lack of benefit or harm, based on epidemiologic,
therapeutic, pathophysiologic, or scientific evidence.
 Uses of Biomarkers: Diagnosing, classifying or grading the
severity of disease in both clinical and laboratory settings ;
Provide efficacy, toxicity, and mechanistic info for the
preclinical and clinical phases of drug discovery; May be
applied to produce commercial tests that aid patient selection
or drug dosing (personalized medicine)
 The best biomarkers are accurate, relatively non-invasive,
easy-to-perform tests done at bedside or outpatient setting and
can be measured serially
 Genetic Biomarkers-Genetic variation gives rise to unique
DNA profiles of each individual
 Protein Biomarkers- Protein alterations in disease may occur
in many different ways that are not predictable from genomic
analysis
 Proteome – the entire protein complement in a given cell,
tissue or organism
 Proteomics- A global approach to the molecular biology study
of the overall distribution of proteins in cells, identification and
characterization of individual proteins of interest, and the
elucidation of their relationships and functional roles
 Functional Proteomics – studies function of diff proteins in
vivo and in vitro
 Differential Proteomics – uses sub tractive comparison
 Bioinformatics can be used for profiling of disease tissues,
serum biomarkers, differential in-gel electrophoresis
BASIC SCIENCE RESEARCH | Block #
 Requirements for Biomarker Research: Innovations in
technology applications, New statistical methods and clinical
research designs, Data management and informatics, Clinical
registries, Repositories of biological specimens, Imaging files,
Common reagents
 Requirements for Successful Biomarker Development:
Access to sufficient, well-characterized sample→ biobanks,
Understanding of the entire complex biomarker development
process, Use of multi-disciplinary team approach, validation of
every step of both assay performance and diagnostic utility
III. BIOMARKER DEVELOPMENT PATHWAY
Step 1. Understand and Define the Disease (requires detailed
knowledge of the disease and a reliable case definition)
Step 2. Frame the question: What critical information will the
biomarker provide?
Step 3. Desired site of clinical measurement (Choice is driven
by a balance between clinical relevance, ease of collection and
stability versus disease specificity and analytical simplicity of the
discovery step)
Step 4. Devise a strategy for the discovery process
Step 5. Which samples should be used for the discovery
phase? (Samples should match the clinical question as closely
as possible and should mirror the disease process being
investigated)
Step 6. Determine which discovery method to use
Step 7. Review the significance and feasibility
Step 8. Perform the experiments and prioritize the hit list
Step 9. Develop a robust clinical assay and initial clinical
evaluation to detect existing disease
Step 10. Evaluate the clinical utility
Step 11. Combine biomarker with clinical data and other
biomarkers.
 Advantages of Biomarker Development: opportunity to have
an impact on patient health in a more economical manner,
provide an opportunity to speed up the drug development
process, earlier disease detection and prediction of which
patients will respond to therapies
 Biomarker Identification: Comparative analysis of protein
expression, analysis of secreted proteins in cell lines and
primary cultures, direct serum protein profiling by mass
spectrometry and identification of tumor antigens
 Biomarker Validation: Prospective, well controlled clinical
studies of diverse patients across multiple institutions, with
well-established standards for all steps in the process,
Reproducibility within and among laboratories)
 Chronology of Development: A biomarker is first identified,
then evaluated for a particular clinical indication → analytical
and clinical validations performed → submitted to US FDA for
approval (bypassed if for research purposes only)→ Center for
Medicaid and Medicare Services (CMS) determines if it is
reasonable and necessary therefore, reimbursable
 Regulatory Bodies: FDA for „safety and effectiveness‟ of an
IVD (refers to the consequences expected from reliance on it
to make „clinically significant‟ diagnostic or treatment
decisions); Center for Medicaid and Medicare Services for
Analyte-specific reagents (biomarkers used in in-house clinical
laboratory for restricted „for research only‟ purposes)
 Challenges: Biological variability among patient samples,
huge dynamic range of biomarker concentrations,
reproducibility and validation of tumor biomarkers,
standardization of techniques, integration of data, cost and the
need to develop multiplex assay systems
 Biomarkers in the future will be used for understanding the
molecular basis of tumor characteristics for development of
personalized cancer therapy. Discovery of new highly sensitive
and specific biomarkers for early disease detection and risk
stratification along with personalized therapy will be the first
line treatment for cancer.
Page 6 of 8
 Lecture 5: Cardiac Biomarkers
I. CORONARY HEART DISEASE
 Major cause of death worldwide
 Atherosclerosis, the underlying cause, starts early in life and
progresses slowly and silently for decades
o Aside from lipid deposition, systemic inflammation also plays
a role in athero-thrombotic inception and progression
o Mononuclear cells, macrophages, and T-lymphoc ytes are
prominent in arterial wall plaques.
o Monocytes  Macrophages  Foam cells
o The shoulder region where it tapers off to the sides is the
most vulnerable site for rupture, and is heavily infiltrated with
inflammatory cells
o Cytokines ha ve been shown to increase in acute coronary
syndromes, even in the absence of myocardial necrosis 
de-novo hepatic production of acute phase reactants (C-
reactive protein). Cytokines also inhibit collagen prod‟n.
o Pro-inflammatory cytokines provide a chemotactic stimulus
for leukocytes to migrate into the intima
o Possible biomarkers: T-l ymphocytes, cytokines, and C-
reactive proteins
 Cholesterol screening used to identify individuals at risk of
developing future coronary events.
o Fails to identify 50% of MI patients who have either normal
or moderately increased serum cholesterol levels
Trichrome stain: luminal thrombus and intraplaque
hemorrhage are red, collagen is blue.
IV. LIPOPROTEINS
 Primary target of therapy: LDL-C
 HDL-C: childhood levels perform as well as Apolipoprotein B in
predicting subclinical atherosclerosis in adulthood
 Apolipoprotein B
o Superior to LDL-C in predicting atherosclerotic risk
o Present on chylomicrons, VLDL, IDL, LP(a), which are
atherogenic particles
o Measure of atherogenicity
o <90 mg/dL – for diabetics or with 2 CVD risk factors
o <80 mg/dL – with known CVD or DM with additional CVD
risk factor
o Independent predictor of: endothelial vasodilatory function,
increased carotid IMT, arterial stiffness in healthy patients
and those with familial combined hyperlipidemia
 Apo B/Apo A-1 Ratio: associated with increased CAD
V. INFLAMMATORY MARKERS
A. Myeloperoxidase (MPO)
 May have a role in CVD protection
 Stored in WBC granules and released w/ neutrophil activation
B. Lipoprotein-associated Phospholipase A2 (Lp-LPA2)
 Elevated = CVD and increased ischemic stroke risk
 Pro-inflammatory substance produced by monocytes,
lymphocytes, mast cells
 80% bound to LDL-C, 20% to HDL-C

II. RISK PREDICTION FOR CVD C. High-Sensitivity C-reactive Protein (hsCRP)

 Predictive of CVD events (MI, ischemic stroke, CV death, DM)
 Based on demographic and clinical variables
o Good performance on population basis  Prognostic for peripheral arterial disease (PAD)
o May misclassify some individuals  Acute-phase reactant; reflects low-grade inflammation
o May underestimate long CV risk  Produced primarily by liver in response to IL-1, IL-6, TNF-α
 Traditional risk factors  Might reflect vulnerability of atheromatous lesions and
o Demographic – age, gender likelihood of plaque rupture
o Clinical – cholesterol, HTN, smoking, blood pressure,  >15ug/mL, CRP significantly reduced endothelial progenitor
diabetes (EPC) cell number (↓ vascular wall viability), inhibited
o Less prognostic value in secondary prevention expression of EC-specific markers (Tie-2, EC-lectin, VE-
 Multi-marker risk prediction models cadherin), increased EPC apoptosis, impaired EPC-induced
o Several biomarkers independently predict CV events when angiogenesis
added individually to models containing traditional  CRP  decreased eNOS mRNA expression  direct inhibition
demographic and clinical variables of EPC differentiation, survival, and function
 Blood-based biomarkers  Rosiglitazone (PPAR agonist) inhibits CRP negative effects
o Lipoproteins (Apolipoproteins, triglycerides, LDL, HDL)
o Inflammatory markers (CRP, IL-6) D. Interleukin-6 (IL-6)
o Coagulation markers (fibrinogen)  Inflammation, bone metabolism, immunity, reproduction, neural
o Cardiac troponin-I development, hematopoiesis
o Brain Natriuretic Peptide  Major regulator of acute phase reactant synthesis in the liver
o Cystatin C  Source maybe from activated macrophages, with contributions
from fibroblasts and endothelial cells, and adipose tissue
 Has more central role than CRP or fibrinogen in CHD
III. BIOMARKER ASSAYS  Two polymorphisms in promoter region: -174 G>C, -572 G>C
 Genotype effect on CHD risk is largely unexplained by blood
 Specimen collection: pressure
o Blood samples are drawn in the fasting state o Smokers are more predisposed for all genotypes for
o Serum and plasma – frozen; either tested immediately or development of myocardial events
aliquoted and stored at -70 to -800C in biofreezers o Significant differences in survival; poorest for heterozygous
 Clinical manifestation: MI, stroke, angina, or sudden death at GC genotype
ages 50-60 y/o (men) and 60-70 y/o (women).
VI. OTHER MARKERS
Analyte Specim en Method Target A.Plasma Fibrinogen – increased CVD risk in men > women
Process B.Cardiac Troponins – MI diagnosis, prognosis of acute
NT-proBNP Plasma, Serum ECL Cardiac coronary syndrome patients, and selection of those who would
Immunoassay, Hormone benefit most from early invasive management
point of care - Levels below the threshold for MI may signal presence of
(POC) tests CAD and increased future CVD risk
Cystatin C Serum Nephelometry Renal C.Brain Natriuretic Peptide
dysfunction
- Major source is ventricular myocardium
and CV risk
Albumin 24-H urine Nephelometry - Stimulus for release: myocardial wall stress
hsCRP Serum Integra and Inflammatory - BNP and NT-proBNP: predictors for CVD risk or death
extended marker - Higher NT-proBNP levels = increased long and short-term
range assays mortality in stable CAD cases
Apolipo-protein B Atherogenecity - Strength: rule out heart failure in patients with shortness of
IL-6 Serum Quantikine Inflammatory breath in the emergency department
immunoassay marker - Provide prognostic info in patients with heart failure, CAD,
Fibrinogen Plasma Clauss assay Coagulation and valvular heart disease
marker

BASIC SCIENCE RESEARCH | Block # Page 7 of 8

D.Cystatin C
- More sensitive than serum creatinine in diagnosing mild
reductions in glomerular filtration rates, and in assessing
renal function.
o Patients with chronic kidney diseases are at high risk
for developing CVD and CV events
- Plasma Cystatin C concentration is influenced by factors
such as age, BMI, sex, smoking, and high CRP conc.
- Associated with increased CV risk (all-cause mortality, CV
events, and incident heart failure)
- Better predictor of coronary artery calcium progression
than serum creatinine or GFR
- Shown to be directly involved in atherosclerotic process
- (Shi et al., 2014) Cystatin C severely reduced in
atherosclerotic and aneurysmal aortic lesions.
- Cystatin C deficiency = vascular disease
- High conc associated with hypermetabolic status

VII. TRANSLATION FROM LAB TO CLINIC

 Biomarkers must satisfy the following issues:
o Availability of population-based cut off points for
interpretation and risk assessment
o Existence of potential therapeutic and risk assessment
o Reliability of the analytical systems used for measurement
 Biomarkers must change management
 Assessment of CV risk vs. extending this to modifying
treatment as a result of biomarker data
o Rosiglitazone
o Individuals with an increased CV risk due to elevated
hsCRP level may benefit from Rosuvastatin treatment
 Multi-Biomarker Approach
o Whether use of multiple biomarkers improves CV risk
stratification in the outpatient setting of CAD cases remains
unknown (Shlipak et al., 2009)
o Ideal scheme might combine traditional risk factors known
to promote atherosclerosis (hyperglycemia, dyslipidemia,
etc.) with:
 Measures of inflammation (CRP, IL-6)
 Myocyte necrosis (troponins)
 Hemodynamic stress (BNP or NT-proBNP)
 Renal dysfunction/vascular damage (creatinine, cystatin
C, microalbuminuria)
o Prognostic Significance - useful in:
 Evaluating the risk/benefit tradeoff of possible
intervention strategies
 For counselling patients about their prognosis
 For making decisions about non-CV prevention
strategies, such as cancer screening
 Presence of elevated NT-proBNP → earlier initiation of
ACE inhibitors and beta blockers
 Kidney damage by either increased albuminuria or
cystatin C → trigger use of RAAS inhibitors or more
aggressive systolic blood pressure control
 Inflammatory biomarkers like CRP and IL-6 have been
consistently predictive of CV outcomes, but may not be
modifiable in either primary or secondary prevention
settings

BASIC SCIENCE RESEARCH | Block # Page 8 of 8