Cancer is characterized by uncontrolled cell proliferation that has transformed from normal cells. Several screening methods are used to test potential cancer treatments in vitro and in vivo. In vitro methods include tetrazolium salt assays, sulphorhodamine B assays, and thymidine uptake assays to test cell viability. In vivo methods include inducing tumors in mice and rats through chemicals like DMBA and testing whether treatments reduce tumor incidence and size. DMBA is used to induce skin papillomas in mice and mammary gland carcinomas in rats, with the drug's efficacy measured by decreased tumor rates compared to controls. Various assays then measure cell viability and proliferation after treatment to screen for potential anti-cancer effects.

1. SCREENING METHODS OF
CANCER
By
Arijit Chakraborty
M.Pharm
(Pharmacology)

2. What is Cancer?
Cancer is a disease which characterized by
uncontrolled proliferation of cell that have
transformed from the normal cells of the body.
CAUSE:
• External Factors – chemicals, radiation, viruses, and
lifestyle
• Internal Factors – hormones, immune
conditions, and inherited mutations
• Theories
– Cellular change/mutation theories
– Carcinogens
– Oncogenes/ protooncogenes

3. Factors Believed to Contribute to
Global Causes of Cancer

4. Methods
In Vitro:
1. Tetrazolium salt assay.
2. Sulphorhodamine B assay.
3. 3H-Thymidine uptake.
4. Dry exclusion test.
5. Clonogenic test.
6. Cell counting assay.
In Vivo:
1. DMBA-induced Mouse Skin Papillomas.
2. DMBA-induced Rat Mammary Gland
Carcinogenic.
3. MNU-induced Tracheal Squamous cell
Carcinoma in Hamster.

5. Tetrazolium salt assay (IN VITRO)
It is performed to determine the Enzymatic properties.
The cell from a particular cell live when in log phase a
trysinized.
It is counted in a homocytometer and adjusted multiwell
plates (96 well plates)
The cells are treated with a various concentration of
drug for specified duration

6. CONT….
After MIT dye is added in each well and plates are
incubated at 37 C for 4 hrs in a CO2 incubator.
The plates are taken out the incubator and dark-blue
colored formazan crystal are thoroughly dissolved in
DMSO in room temperature.
The plates are then read on a ELISA reader at 570
nm.
To calculate the percent cell viability with respect to
control is calculated using,
% cell viability =(OD of treated cells/ OD of control

7. Sulphorhodamine B Assay
The Sulphorhodamine B assay measures whole-
culture protein content, which should be proportional
to the cell number.
Cell culture are stained with a protein staining dye,
Sulphorhodamine B.
SRB is a bright pink anionic dye that binds to basic
amino acid of cell.
Unbound dye is then removed by washing with acetic
acid.

8. DMBA-induced Mouse Skin
Papillomas
This is a classical to stage experimental carcinogenesis
model.
Mouse skin is generally most sensitive to epidermal
carcinogenesis.
SENCAR mice are highly sensitive to DMBA induced
skin tumor.
DMBA acts as a inhibitor and 12- O- tetradecanoyl-
pharbol- 13 acetate is used as a promoter to induce
skin papillomas and Squamous cell carcinomas.

9. CONT…
Mice are topically applied a single dose of 2.5 µg
DMBA in acetone on the shared back, followed by 5-
10 µg of TPA in 0.2 ml acetone twice weekly on the
same site starting one week after DMBA application.
Percent tumor incidence and multiplicity of treatment
groups is compared with DMBA control group.
Drug under test can be administered either topically or
oral route.
The tumor incidence in this model is usually about
100% DMBA controls.

10. DMBA-induced Rat Mammary
Gland Carcinogenic
Female Sprague-Dawley are use for this method.
Rats are given single intragastric injection of 12 mg/kg
DMBA at 50 days of age.
This dose results in 80-100% incidence of total mammary
tumors within 120 days post carcinogen.

11. This model can detect the agents/drugs inhibiting
carcinogen activation.
DNBA produced capsulated tumors with high
incidence.
Drug efficacy is measured as percent reduction in
adenoma incidence, percent increase in
adenocarcinoma latency compared with that of
carcinogen control.