Dear Thomanai!

Different IC50 values ​​in cancer cell tests depend on the sensitivity of the methods used. The IC50 values ​​in the MTT test turned out to be higher, since the sensitivity of this test was lower for the TKI, and the sensitivity of tests based on enzymes was higher. The reason for this is the mechanism or principle used that underlies these methods. In the MTT test, cytotoxicity is based on the metabolism of MTT to pharmacan by mitochondrial dehydrogenases. Your tyrosine kinase inhibitors, having cytotoxicity on cancer cells, do not affect the activity of these dehydrogenases, but they inhibit the activity of other enzymes more strongly (therefore, the IC50 values ​​are lower).

Hello

Cell-based assay involves many variables. For instance, cell is a biological entity and therefore its response to the drug molecule is bound to vary each time. Moreover, there are additional steps involved in cell-based assay. For instance in MTT, steps such as addition of MTT followed by solubilizing solution could also introduce more variations. More number of steps will introduce human error. This could lead to higher IC50 value for cell-based assays.

On the other hand, enzyme inhibition assay is a straight forward biochemical reaction in vitro without the involvement of cells. So the drug molecule like TKIs can directly act on the enzyme in the tube or a plate format without the need of any additional step except for the addition of the substrate followed by incubation and then recording the absorbance.

So naturally, since enzyme inhibition assay is just a simple straight forward assay without the involvement of cells which could make a lot of difference. IC50 value could therefore be lower than what is obtained from cell-based assays.

I hope this is helpful.

Best Wishes.

It is very​ helpful, thank you.