Case Study: Measuring ADP/ATP ratio and testing a custom compound.

This case study details a recent project that Cellomatics worked on for an academic and biotech collaboration client.

The Client

The client was an academic and biotech collaboration who had recently published a paper which illustrated that one of the compounds that specific colleagues had designed protected mitochondria in the presence of GOX (glucose oxidase) treatment where GOX induces mitochondrial dysfunction.

 

The Objective

The main objective was to test if the client’s compounds reduce the ADP/ATP ratio in the presence of GOX.

 

The Project

Cellomatics was required to optimise an assay that measures ADP/ATP ratio and test the client’s compounds with this assay.

The basis of the project was to test compounds that were similar in structure to the one published by the client, the compounds are designed to improve mitochondrial function and cell viability. The increase in mitochondrial function will be hypothetically achieved through the increase of ATP production and through the protection of mitochondrial integrity by compound treatment.

The increased rate of ATP production by mitochondria impacts the ratio of ADP molecules to ATP molecules (ADP/ATP ratio). The alteration of this ratio to favour ATP production increases the cell viability and thus can polarise the cells exhibiting mitochondrial dysfunction away from apoptosis.

 

Did this project relate to a specific therapeutic area, if so, what was it?

Mitochondrial related diseases.

 

What were the outcomes of this project?

The project is almost completed. Cellomatics has successfully tested 7/7 compounds in the C2C12 cell line and 3/7 compounds in the primary endothelial brain cells.

The outcomes so far seem to suggest that the designed compounds all reduced the ADP/ATP ratio, which indicates a higher ATP production and therefore a greater cell viability.

Graphs A and B represent the ADP/ATP ratio of primary brain endothelial cells post treatment with both GOX and the client’s compound. The data suggests that compounds X and Y are reducing the ADP/ATP ratio which is an indicator of an increased viability of the cells under stress.
Statistical significance was determined via an ANOVA (Sidak multiple comparisons test), *** Represents comparisons between the glucose oxidase concentrations and the vehicle control (* P < 0.0332/ ** P<0.0021 /*** P < 0.001)
The two Vehicle controls are 0.1% sodium acetate and 0.1% DMSO. The concentrations refer to the concentrations of the client’s compound used.

Why did the client come to Cellomatics?

The client approached Cellomatics for access to their high-quality plate reader and for their high throughput approaches.

 

Where there any challenges to overcome during the project?

There were a few challenges to the assay, mainly the optimisation of the assay. To achieve this, Cellomatics recommended a multi-stage plan to the client, whereby they performed pilot assays to determine the optimal cell density and the optimal GOX concentration. The stages were as follows:

Stage 1 – determine optimum cell density

Stage 2a – determine optimum GOX concentration

Stage 2b – determine optimum drug concentration range

Stage 3 – combine the results of the above and test the clients’ compounds

 

How long did Cellomatics work on this project?

The initial quote, which will finish in the coming weeks, has taken approximately 3 months. However, the client has requested more quotes and a variety of future collaborations with Cellomatics.

 

A comment from Cellomatics

‘The work that the client had performed had yet to be proved and tested outside of an academic environment until now, which made the project exciting and very rewarding. The client’s compounds have a lot of potential and we look forward to working with them in the future.’

Request a consultation with Cellomatics Biosciences today

Our experienced team of in vitro laboratory scientists will work with you to understand your project and provide a bespoke project plan with a professional, flexible service and a fast turnaround time.

To request a consultation where we can discuss your exact requirements, please contact Cellomatics Biosciences.