INTRODUCTION In drug development, preclinical development, also named preclinical studies and nonclinical studies, is a stage of research that begins before clinical trials (testing in humans) can begin, and during which important feasibility, iterative testing and drug safety data are collected, typically in laboratory animals. Preclinical pharmacology and toxicology are essential elements of the drug discovery and development process and are critical in enabling the translation of findings from the laboratory and the clinic. The drug discovery process is complex and involves numerous iterative steps designed to optimize the pharmacological and drug-like properties of a candidate molecule, a New Chemical Entity (NCE), and minimize the potential for side effects and toxicities. Key concepts addressed in this record include: compound identification; lead optimization; pharmaceutical profiling; the use of animal models to predict efficacy and safety and toxicological assessment as these relate to the regulatory requirements for Phase I trial initiation. Commentary is also provided on the current challenges associated with translational medicine as it applies to the effective evolution of candidate NCEs into viable clinical candidates. The main goals of preclinical studies are to determine a starting, safe dose for first-in-human study and assess potential toxicity of the product, which typically include new medical devices, prescription drugs, and diagnostics. On average, only one in every 5,000 compounds that enters drug discovery to the stage of preclinical development becomes an approved drug.

1. WELCOME
Priclinical Pharmacological
Experiments
Mr. Akash Anil kale
(B. Pharm, VII Sem)
Department of pharmaceutical sciences
Rashtrasant Tukadoji Maharaj Nagpur University,
Nagpur 440033
2020 – 2021
Presented By
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2. In drug development preclinical Experimental studies , is a
stage of research that begins before clinical trials (testing in
humans) can begin, and during which important feasibility,
iterative testing and drug safety data are collected, typically in
laboratory animal.
The main goals of preclinical studies are to determine a
starting, safe dose for first-in-human study and assess
potential toxicity of the product, which typically include
new medical devices, prescription drugs, and diagnostics.
2
Introduction

3. Objective of the Preclinical studies
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• Preclinical studies aim at providing information about safety and
efficacy of a drug candidate before testing it in humans.
• The purpose of pre-clinical study is to develop adequate data to
decide that it is reasonably safe to proceed with human trials of the
drug
• Means, a laboratory test of a new drug or a new medical device,
usually done on animal subjects, to see if the treatment really works
and if it is safe to test on humans.
• However, the main objective is to collect the data to submit to the
FDA for IND filing.
• Screening of new dosage form and formulation.

4. 4
Basic Terminologies
• IN VIVO :- Studies that are in vivo are those in which the effects of
various biological entities are tested on whole, living organisms or cells,
usually animals, including humans, and plants, as opposed to a tissue
extract or dead organism.
• IN VITRO :- means studies or experiments conducted on
microorganisms and cells outside of their normal biological
environment.
• Ex vivo:- In this type of experiment the living tissue are not created
artificially but directly taken from living organism.
• In situ:-. In this the experiment is carried out in their natural habit or
environment.
• In silico:- In silico technique the experiment is carried out bythe use of
computational methods.

5. Animals used
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NAME OF Animals USE OF Animals
1. RAT AND
MICES.
• Used For toxic study , bioassay of
drug , testing of analgesics etc.
• Tissue Used for the study are colon ,
uterus , ceacum , vas deference ,
abdominal stomach muscle etc.
2. Guine Pig
(Cavia
procellus)
• They are very sensitive to
Tuberculosis and anaphylaxis.
• The commonly used tissue is ileum
for study.
3. Rabbits
(Oryctolagus
Cuniculus)
• Used for pyrogen testing, Bioassay of
anti-diabetics, curare derivatives
and sex hormones.
• Used to study of meiotic and
mydriatics.

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Name of Animals Use OF Animals
4. Frog
(RanaTigrina)
• Isolated Heart, rectus abdominal
muscles used to study of drugs.
• Used for evolution of local anaesthetic.
• Use as research model form many
years.
5.Transgenic Animals
A transgenic animal is one
whose genome has been
altered by the transfer of a
gene or genes from another
species or breed.
• Transgenic rodents play a
number of critical roles in drug
discovery and development.
• Importantly, they enable scientists
to study the function of specific
genes at the level of the whole
organism which has enhanced the
study of physiology and disease
biology and facilitated the
identification of new drug targets.

7. 1. Behavioural Models of Animals
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• Animal model’s:- The animal model is living organism in
which normative biology or behaviour can be studied , or in
which a spontaneous or induced pathological processes can
be investigated.
• Types of behavioural animal models:-
A. Elevated plus maze
B. Eddy’s hot plate
C. Rota rod
D. Elecroconvulsiometer
A. Elevated plus maze. :-
• The elevated plus maze task is a simple
method to assess anxiety-like behaviors in
rodents.

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B. Eddy’s hot plate
• The hot plate test is used in basic pain
research and in testing the effectiveness of
analgesics by observing the reaction to pain
caused by heat.
C. Rota rod :-
• Rotarod test is widely used to evaluate
drug effects on motor coordination,
balance and motor learning in rodents.
D. Elecroconvulsiometer :-
• This instrument is used to study the anti-
convulsant activity of phenytoin against
maximal electro-shock induced convulsions
in rat or mice.

9. 2. Irwin test (Battery Test)
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• The Irwin test is an observational screening that is comprised of a
battery of tests used to assess a mouse or rat’s neurobiological and
physiological state.
• Parameters that are evaluated include autonomic and sensorimotor
functions, convulsive behaviour, and other activities produced a drug after
administration.
• The Irwin Test is use to evaluate the qualitative effects of the New
chemical entity on behaviour and physiological function, from the first dose
that has observable effects up to doses that induce clear behavioural
toxicity or even death.

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3. Biochemical assays
• Biochemical assays are analytical methods used to measure and quantify metabolic
reactions and cellular processes such as cell signalling and apoptosis. Biochemical
assays helps to analyse the biomolecules and their functions during drug discovery and
development.
• There are so many techniques like ELISA and Western blotting techniques which are
used to quantify metabolic activities and measuring functional behaviour of biomolecules
such as proteins, enzyme, small molecules, cofactors.
A. WESTERN Blotting technique.
B. Enzyme-linked immunosorbent assay (ELISA)
C. ENZYMATIC ASSAY
A. WESTERN Blotting techniques :-
In this method labelled antibody against particular protein is used identify the desired protein, so
it is a specific test. Western blotting is also known as immunoblotting because it uses antibodies
to detect the protein.
B. ELISA :- In ELISA, various antigen-antibody combinations are used, always
including an enzyme-labeled antigen or antibody, and enzyme activity
is measured colorimetrically.

11. I. Continuous assays :-
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C. ENZYMATIC ASSAY
The enzymes or proteins present in the human body exhibit catalytic activity, the
ability to perform tasks resulting in reactions. Hence, it is necessary for enzymes
synthesized industrially to exhibit the correct functions.
There are two main types of enzymatic assays based on sampling
methods. I. Continuous assays.
II. Discontinuous assays.
This is the type of enzymatic assay which give the continuous readings of
activity. Methods Used :-
1. Spectrophotometric.
2. Fluorimetric.
II. Discontinuous assays:-
In this type of enzymatic assay the samples is taken after the specific time interval
from the enzyme reaction at intervals and the product production and substrate
consumption is measured in the sample by different chemicals methods.
• Methods used :-
1. Chromatographic
2. Radiometric

12. 4.Protein Estimation :-
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Proteins estimations or protein assays are done to estimate the
concentration or amount of protein present in product.
A. Biuret method:-
Principle :-
Protein reacts with this alkaline copper complex and colour changes to violet.
The protein can then be estimated by reading the absorption at 540nm. This
method takes 20-30 minutes to complete.
Principle :-
The principle behind the Lowry method of determining protein Concentrations lies in
the reactivity of the peptide nitrogen[s] with the copper[II] ions under alkaline
conditions.
B. Lowry Method :-

13. D. Bradford assay(Dye Binding method):-
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C. BCA assay(Bicinchonic acid
Method):-
• This method is highly sensitive and detects proteins at a low concentration of 1 µg. In
this method, Copper ions bind to Nitrogens in protein and the complex is then bound to
bicinchoninic acid resulting in the change of colour to purple depending on protein
concentration.
• This is a very sensitive method and simple dye binding assay. This method uses
Coomassie brilliant blue-250 dye that binds with negatively charged protein molecules.
• The dye colour changes based on protein concentrations and the absorption is measured
at 595nm
E. UV absorption:-
In this method, no reagents are required, the liquid protein sample is monitored
under UV absorption at OD 280nm.

14. 5.Nucleic acid estimation
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• In nucleic acid estimation blotting techniques are very widely used as analytical
tools for the specific identification of desire DNA and RNA fragments from
thousands of molecule.
• Blotting refers to process of immobilization of sample nucleic acid on solid support.
• The blotted nucleic acid then used as target in the hybridization experiments for
their specific detection.
• Types of blotting:-
A) Northern blotting ( FOR RNA)
B)Dot Blotting ( for RNA)
C) Southern blotting ( for DNA)

15. • Toxicology and safety studies identify potential target organs for
adverse effects and define the therapeutic Index to set the initial
starting doses in clinical trials.
• Preclinical studies can be designed to identify a lead candidate from
several hits; develop the best procedure for new drug scale-up, select
the best formulation, determine the route, frequency, and duration of
exposure, and ultimately support the intended clinical trial design.
• It plays an important role in development of new drug or chemical
entity.
IMPORTANCE OF PRECLINICAL EXPERIMENTS
IN DRUG DISCOVERY PROCESS

16. References :-
• https://en.wikipedia.org/wiki/Wikipedia
• Granchelli, J.A., Pollina, C. and Hudecki, M.S., 2000. Pre-clinical screening of
drugs using the mdx mouse. Neuromuscular Disorders, 10(4-5), pp.235-239
• Badyal, Dinesh K., and Chetna Desai. "Animal use in pharmacology education
and research:The changing scenario." Indian journal of pharmacology 46,
no. 3 (2014): 257.
• https://en.m.wikipedia.org
• http://www.researchgate.net
• http://www.slideshare.net
• http://www.exploabiolabs.com

17. THANK YOU
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