Methods of Medical Research

Past and Present By M M Elshawwa MD

History • عالء الدين ابن النفيس

• The discoverer of greater circulation : Sir William Hatvey • Both use animal models and human dissections

Animal models in medicine:• Undergraduates: illustrating physiological

mechanism and effect of various known drugs on human body

• Postgraduate: elaborate the physiological mechanisms and actions

• Research to conduct screening for drugs, bioassay and preclinical testing of new drug

• Extensive toxicological studies in animals before the drug gets approval for clinical trials in humans

Animal Model

Medical Discovery

Dog Insulin

Monkey Polio vaccine

Mouse Rabies vaccine

Pig Skin grafts and R Insulin

Rabbit Corneal transplants

Rat Drug toxicity and carcinogen screening

BUT• There is no doubt that the best test species for humans

are humans. It is not scientific to generalize animal data directly to humans due to interspecies variation in anatomy, physiology and biochemistry.”

Continued but modified use of animals

• Russel and burch in 1959 proposed that “if animals were to be used in experiments, every effort should be made to have the 3R strategy which includes: • Refinement- refine experimental methods:

decrease pain and trauma to animals to neutralize the stress issue

• Reduction- reduce the number of animals used in these experiments

• Replacement- replace the animal experiments e.g. In-vitro methods and cell culture techniques.

Appropriate analgesics and anaesthetics for painful procedure

Proper handling technique for animals

Adequate training and adaptation prior to experiment

Ensure drug doses are correct and drugs are not expired

Aseptic surgery and procedures to prevent infection,

Methods of Refinement

Methods of Reduction • Use animals as their own controls e.g. Cross over

study. • Gather data for more than one experiment

concurrently. • Consult statistician and use minimum number of

animals. • Minimise variables such as disease, diet, stress

and genetics. • Use appropriate species of animals

Replacement

• Replace higher animals with lower animals.• Replace live animals with dummies for teaching and

dissection purposes.

•Alternatives

05/02/2023 10

ALTERNATIVE MODELS

• In vitro test methods models based on human stem cell and tissue cultures

• Micro dosing

• Volunteer studies

• Population research

• Organ on a chip• Computerized patient-drug databases and virtual drug trials• Computer assisted learning, models and simulations

• Non-invasive imaging techniques such as FMRIs and PET Scans

05/02/2023 11

Cell cultures

• Many types of human and animal cells can be grown in proper culture:• Cancers• Sepsis• Kidney disease and AIDS• Vaccine production and drug development.• Recently, printing of 3D human tumours and organs, providing more

realistic cell models on to test potential therapies.

05/02/2023 12

Stem Cells

 Embryonic Stem Cell Test (EST) 

• Technique used to find out if a drug harm the developing foetus.

• Replaces pregnant rats and rabbits animal test by mouse stem cell cultures.

• Involve use of microscope to classify toxic substance i.e. if they block the development of stem cells.

Statistical significance:

• 78% accurate, with close to 100% accuracy at detecting very toxic chemicals.

• Animal test detect 60% of toxicants.

Stem cell models • Disease genes are inserted into embryonic stem

cells, induced to differentiate into human disease tissue used for screening of drugs

e.g. 1-Genes from a Parkinsons patient were introduced in embryonic stem cells which grew into a model of Parkinsons disease and is used for screening potential drugs 2-Alzheimers and Diabetes stem cell models

05/02/2023 14

LIMITATIONS

• In vitro techniques focus on the cellular level and therefore cannot replace whole-body testing.

• It can’t completely ensure the drug safety and efficacy.

05/02/2023 15

TISSUES

Animal tissue:• Avian- chick embryos • Rodents- rats and mice: wild types and transgenic, embryonic and

adult. Human tissue:• Surgery e.g. biopsies, cosmetic surgery and transplants.• Neural progenitor cells from aborted foetuses and stem cell lines.*******Can be used in many forms, including isolated perfused organs, frozen tissue or in cell cultures.

05/02/2023 16

benefits• More reliable than animal tests.

• The use of human tissue in toxicity testing is more accurate than the animal models.

• More cost-effective, practical, and expedient.

• Less time is needed for completing the study compared to animal models and cost effective.

• But, they can not completely eliminate the need for animals in preclinical studies.

17

• Micro dosing and imaging studies allow human volunteers to share safely in these trials.

VOLUNTEER STUDIES

05/02/2023 18

• Includes studying and comparing illnesses in human populations

• Identify the risk factors and causes of human diseases

• Understanding the roles of genes, lifestyle, diet and occupation

E.G. These types of studies have led to the discovery of smoking related cancer and the association between high cholesterol and poor lifestyle choices with increase the risk of heart disease.

POPULATION RESEARCH

In silico models • Organ on chip • Microfluidic chips • DNA chips • Computer aided molecular drug design • Computer assisted learning • Computer or mathematical analysis

05/02/2023 20

ORGAN ON A CHIP

• Contain a series of tiny hollow chambers that are lined with real, living human cells from different parts of the body.

• Linked by channels through which a blood substitute flows, following same principle as in human body.

• Test drugs can be added to the blood substitute then circulates around the device while sensors within the chip feedback information for computer analysis.

05/02/2023 21

CONT..

05/02/2023 22

Cont..

• Use of human organ models for drug toxicity screening like heart-on-a-chip, and human disease models for development of novel drugs.

• Models to investigate routes for optimal drug uptake in the body.

• Capturing the genetic variation present in the normal e.g. pharmacogenomics and hypersensitivity

• To improve efficiency and reduce costs of drug development.

LIMITATIONS:

• Provides less information than whole-body testing

• For determining drug safety and efficacy requires testing in animals

05/02/2023 23

computer simulations

• Computer Models of disease and treatment.

• Translation of biological effect into a mathematical equation.• By collecting human research data points and carry out human

clinical trials.• Virtual human organs, virtual metabolism and Virtual

Physiology programmes can now predict effects in humans and interaction far more accurately with any given organ than animals can do.

• Computers design the molecular structure of drugs to target specific receptors.

Examples• Simulations of cancer are used to test drug

targets.• Mathematical models have helped to further

understanding of HCV dynamics and clinical trial in humans.

• Protease inhibitors were designed by computers and tested in tissue culture and computer models bypassing animal tests.

Computer assisted learning

• Deals with a range of software packages which simulate the animal experiments

Expharm• Software on: • Effect of drugs on the rabbit eye • Bio assay of histamine using guinea pig ileum • Effect of drugs on the frog heart • Effect of drugs on dog blood pressure and heart rate• Effect of drugs on the ciliary movement of

oesophagus• The user can conduct experiment and collect

data • Each program can be run in two modes-

a) tutorial mode , (b) examination mode

BIOPAC Systems for research

 Solutions for data acquisition and analysis : the student and /or researcher take data from animal and/or himself >>>>>• The hardware provides a flexible tool for research and teaching needs. • Amplifier modules, transducers, and electrodes to match research

design. • New modules are available for ECG, BP, noninvasive Electrical Bio-

impedance (Cardiac Output) and EEG with Brain Activity • Software included with each System is an interactive, intuitive program

that instantly view, measure, analyze, and transform data. • Perform complex data acquisition—no need to learn a programming

language.• Analysis Features: signal averaging, sophisticated integration, and

equation generation.

Conclusion• Could we stop animal use in medical research ?

PROBABLY NO,• Animal models just a step will be just a step in the future

research“WE cannot stop animal use in medical research.however, We can not move to the era of modern physiology with animal research alone……..Mohammad M Elshawwa