I am currently conducting experiments on checking combinatorial effect of two small molecule natural compound against cancer cell line. However, I am unable to determine how the combination of two drug should be done. How the ratio should be managed so that I can check the difference of combinational effect of drug and compare them with effect of individual drug through cytotoxicity assay, mRNA expression, etc.
Determining the ratio of two drugs for checking their combinatorial effect against cells can be a complex task. The optimal ratio may vary depending on the specific drugs, cell lines, and experimental conditions. Here are some general approaches and considerations that can help guide your experimental design:
Dose-response curves: Generate dose-response curves for each individual drug using a cytotoxicity assay, such as the MTT or CellTiter-Glo assay. This will help you determine the effective concentration range for each drug and identify the concentrations that result in 50% cell viability (IC50)
Fixed ratio method: Start with a fixed ratio of the two drugs, such as 1:1 or 1:10, and perform a dose-response experiment using various concentrations of the drug combination. Evaluate the cytotoxicity of the drug combination and compare it to the cytotoxicity of the individual drugs at their respective IC50 concentrations. If the fixed ratio does not yield the desired effect, try different ratios to optimize the combination
Isobologram analysis: Isobologram analysis is a graphical method used to determine whether a drug combination is additive, synergistic, or antagonistic. This analysis involves plotting the concentrations of the two drugs on the x and y axes, respectively, and connecting the points that correspond to the IC50 values for each drug. If the line connecting the points is below the line of additivity, the combination is considered synergistic; if it is above the line of additivity, the combination is considered antagonistic; and if it is on the line of additivity, the combination is considered additive
Combinatorial index (CI) The CI is a quantitative measure of the interaction between two drugs in a combination. It is calculated using the formula CI = (D)1/(Dx)1 + (D)2/(Dx)2, where (D)1 and (D)2 are the doses of drugs 1 and 2 in the combination that produce a certain effect (e.g., 50% growth inhibition), and (Dx)1 and (Dx)2 are the doses of drugs 1 and 2 alone that produce the same effect. A CI value less than 1 indicates synergy, a CI value equal to 1 indicates additivity, and a CI value greater than 1 indicates antagonism
Time-dependent effects: Consider the time-dependent effects of the drugs and their combination. Some drug combinations may have a delayed or prolonged effect on cell viability, which may require longer incubation times or multiple time points for analysis
You could take the easy route and use CalcuSyn software.
https://calcusyn.software.informer.com/2.0/
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