Hello! I need to calculate synergistic effect of 2 drugs, however, when I treat my cell culture with them, I can't get an IC50 value, as I can't use a dose higher than 10uM following manufacturer's recommendation. When I combine the 2 drugs, I get an IC50 lower than 10uM, but, as far as I know, I need the IC50 for individual drug treatment if I want to calculate synergism. Can I calculate synergism without using IC50 value?
Thank you very much!
Depending upon the model you use, you should be able to estimate IC50 without actually achieving 50% response. Would an estimated IC50 let you proceed the way you wanted to?
Dear Fernando, although it is conventional to estimate IC50 and evaluate synergy with these concentrations, you could eventually estimate IC75 or IC60 and evaluate synergy to get an idea of the performance of the combination. Why is it not possible to increase the concentration of one of the drugs above 10 uM? Is it possible that this imposition is for another cellular model?
Thank you for your answers! I cannot use more than 10uM because inhibitors (sorry I didn't say my drugs are inhibitors) are likely to target proteins non-specifically in higher concentrations. We've made a research and all authors use these drugs in concentrations below 10uM. I can extrapolate the IC50 but I don't know if the value is reliable enough to carry out a Chou-Talalay assay, as maybe, cell death is not due to the specific inhibition the drugs are supposed to do.
Dear Fernando, two responses back I had a mental lapse I meant that you could consider using IC less than IC50 (not higher) to preliminarily evaluate the presence of synergy. That is, if the maximum concentration you can use is something less than 10 uM and this affects, for example, 40% of cell viability, assess synergy in these circumstances.
The Chou-Talalay method considers the case in which there is interest in evaluating synergy when using an inhibitory concentration other than IC50. This is particularly useful when systems present hormesis. The equation of the combination index (CI) that applies in that case is the following:
CI = (D)1/(Dx)1 + (D)2/(Dx)2
This equation and the other details can be verified in the review article of Chou (2006) that I attach to this comment (see equation 19 within the document).
In any case, when calculating the Dx, it is necessary to know the IC50 (or Dm according to the Chou-Talalay nomenclature) of each drug even if you can not test one of them above 10 uM. The Dm value can be accessed with the Chou-Tatalay analysis of a single drug even if you have only inhibitions below 50%. The key is that you have a good fit of the data to the straight line
log (fu/fa) = m log (D) - m log (Dm)
That is, the value of R2 must be high, the closer to 1 the better it will be, it would mean that the estimate of Dm (IC50) is reliable and therefore the Dx of the problematic drug will be safe for the subsequent calculation of CI, the which would have to be less than 1 if the combination of drugs really involves synergy.
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