Anthem Biosciences, a Bangalore based Contract Research and Innovation Service Provider (CRISP) has created a group called “Platform Technologies” to perform innovative research that would result in proprietary assays/products. These technologies can either be offered as a service, or out-licensed to biotech, biopharmaceutical, pharmaceutical and industries in the diagnostics, nutraceutical, manufacturing, agricultural and animal healthcare sectors. The technologies are developed and validated at Anthem, and prior to release, these are also validated externally in other laboratories around the globe. Examples are two publications in peer-reviewed journals for the “ready to out-license” proprietary genotoxicity assay.
An alarming increase in the number of genotoxicity associated diseases worldwide warrants urgent measures to detect potential genotoxins. In addition, the genotoxicity profiling of potential drugs/cosmetics/ fertilisers /food additives will serve as an invaluable tool in early stages of drug discovery. Current technologies to detect Genotoxicity include AMES, Chromosomal aberration and Micro nucleus tests. However there are several disadvantages associated with these assays such as the following:
- Laborious
- Time consuming (2-3 weeks)
- Requires large amount of test compounds
- Higher percentage of false negatives
- Mostly involve bacteria or rodent cells
- AMES- Bacterial screen in relevance to mammalian genotoxicity is debatable
- Single sensor system will miss out on genotoxic signalling events that bypass the sensor
Anthem’s Genotox screen
The cellular response to DNA damage is complex, involves multiple pathways and activates DNA damage early-response promoters such as p21 and GADD153. These genotoxicity inducible genes participate in the cellular response to genotoxic stress by inhibiting proliferation, promoting repair, and/or apoptosis. Thus, by engineering these genes such that their activation results in luminescence provides for a novel human cell based high-throughput screen. The technology is a recipient of two highly coveted technology awards.
1. It is a human cell based screen that is high-throughput compatible.
2. The human cells are engineered such that they posses triple sensors of genotoxicity such that when the cell is exposed to a genotoxin, it luminescences. The triple sensors can be transferred to any cell line or cell type.
3. The screen is the first of its kind in that it provides invaluable insights into the cellular mechanisms underlying the genotype effect
4. The amount of compound required is minimal ~ 10 mg
5. The process time of the reaction is about four hours unlike days /weeks for any other assay
6. Extensive validations have been conducted in-house, at external sites and in collaboration with labs in Europe
7. The sensitivity and specificity values far exceed any of the standard assays currently in use.
Yet another screen that has been extensively used by clients is the Anthem Mitotox screen. A strategic combination of assays that can accurately identify a mitotoxin, this screen has been extensively used by clients. Mitochondria are responsible for two critical cellular functions, i.e. bioenergetics and programmed cell death (apoptosis).
Mitochondrial dysfunction has been recently been recognized as a critical event triggered by several drugs that have been issued black box warnings or been withdrawn from the marker. This warrants an urgent need to screen for mitotoxins and to integrate this screen into any drug toxicity screening program.
Current technologies include JC-1 dye staining method, cytochrome c release assay. The drawbacks associated with such assays include the following:
- Rely on one or two assays
- Do not reveal any mechanistic insights
- Functional assays rarely performed
- Isolated mitochondria studies not carried out to define mitotoxicity
Anthem’s Mitotox Assay
Anthem’s validated Mitotox screen incorporates multiple assays that reveal information about the following:
- mitochondrial function (ATP production)
- mitochondrial DNA damage using real time PCR cytochrome C release
- swelling assay,
- JC-1 staining and MTT assay
Anthem’s unique expertise in isolating functional mitochondria and incorporation of assays using this purified mitochondria sets standards that are unparalleled in this field. Anthem is in negotiations with universities and companies who are keen to use this platform.