Dear Zhen, 

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Article Intracellular Drug Concentrations and Transporters: Measurem...

Article Rapid Measurement of Intracellular Unbound Drug Concentrations

Hello,

I suppose that you refer to a Cmax measured in plasma ? If I am right, you have to check plasma protein binding of the drug, in order to translate the in vivo plasma concentration, which is a total (bound +unbound) concentration into an in vitro concenration, which is a free concentration (generally the medium contains much less plasma proteins than plasma itself).

Then you could first test in your cellular experiments if you observe an effect when the medium contains a concentration corresponding to the Cmax (unbound).

If no effect, you can anticipate that nothing will happen in the clinical setting at the dose corresponding to the Cmax. (I assume that you cannot increase the dose in vivo ; if yes, then you could test higher concentrations).

If there is an effect, you could go foward, by exploring a concentration-response curve, for example.

Regards

Alain

1. Is the Cmax in vivo roughly at a level where it has activity in vitro? If it is out by orders of magnitude you are likely to be wasting your time

2. Whilst the remarks of Alain are correct, do check the method of measuring Cmax: sometimes they measure free drug not just total. The degree of adjustment may not be very much depending also on the protein concentration of your culture system: human serum is *roughly* 6-8% protein, plasma a bit more because of presence of clotting factors. 

3. You should also not just use a single dose in your experiments but a range centred around the Cmax to take into account the biological variation in Cmax that will exist in a population, and in an individual over time as drug is excreted/metabolized.

Hi Zhen,

1)     Clinical Cmax for one disease cannot be transposed to other indication. Is the clinical Cmax (2.45uM) you cited related to efficacy? There should be a range for “therapeutic window” for efficacious exposure (Cmax or AUC or both can drive the efficacy). So once you know the efficacy range for plasma Cmax (e.g., 1.5 to 3.5 uM, a guess for your case), you need to find value of unbound drug in plasma (and preferably in target tissue).

2)     Once you know target unbound Cmax range in plasma or target tissue, you can pick-up few concentrations higher and lower to cover that range in your in vitro experiments you are planning. Say for your purpose, you would try 0.5, 1.0, 2.0, 4.0, 6.0 and 8.0 uM to cover a wide range to maximize your chances to see measurable effect in your new cellular disease model.

3)     Remember, cellular (in vitro) data cannot be directly translated to in vivo situation, because in your culture plate you will have longer and continuous exposure of your drug to cells vs in target tissue (in vivo) the drug concentration will be dynamically changing as per the PK profile! But this doesn’t invalidate usefulness of in vitro testing to find effect of a test compound in relatively simple system.

Best luck!