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In situ drug-receptor binding kinetics in single cells: a quantitative label-free study of anti-tumor drug resistance.

Wang W, Yin L, Gonzalez-Malerva L, Wang S, Yu X, Eaton S, Zhang S, Chen HY, LaBaer J, Tao N - Sci Rep (2014)

Bottom Line: Many drugs are effective in the early stage of treatment, but patients develop drug resistance after a certain period of treatment, causing failure of the therapy.We further showed that the steric hindrance of Mucin-4, a membrane protein, was responsible for the altered drug-receptor binding.This effect of a third molecule on drug-receptor interactions cannot be studied using traditional purified protein methods, demonstrating the importance of the present intact cell-based binding kinetics analysis.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, China.

ABSTRACT
Many drugs are effective in the early stage of treatment, but patients develop drug resistance after a certain period of treatment, causing failure of the therapy. An important example is Herceptin, a popular monoclonal antibody drug for breast cancer by specifically targeting human epidermal growth factor receptor 2 (Her2). Here we demonstrate a quantitative binding kinetics analysis of drug-target interactions to investigate the molecular scale origin of drug resistance. Using a surface plasmon resonance imaging, we measured the in situ Herceptin-Her2 binding kinetics in single intact cancer cells for the first time, and observed significantly weakened Herceptin-Her2 interactions in Herceptin-resistant cells, compared to those in Herceptin-sensitive cells. We further showed that the steric hindrance of Mucin-4, a membrane protein, was responsible for the altered drug-receptor binding. This effect of a third molecule on drug-receptor interactions cannot be studied using traditional purified protein methods, demonstrating the importance of the present intact cell-based binding kinetics analysis.

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Top: Under “bypass hypothesis”, in situ binding kinetics of Herceptin with resistant cell was not affected. Bottom: “Alternation hypothesis” suggests an altered binding kinetics due to the influences of relevant proteins surrounding Her2.
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f1: Top: Under “bypass hypothesis”, in situ binding kinetics of Herceptin with resistant cell was not affected. Bottom: “Alternation hypothesis” suggests an altered binding kinetics due to the influences of relevant proteins surrounding Her2.

Mentions: We hypothesized that in situ binding kinetics analysis in intact cells would provide a direct answer to this question, because the “alternation hypothesis” would mean a large change in the binding behaviors of drugs with the receptors in drug-resistant cells while the “bypass hypothesis” would imply no change in the binding kinetics (Fig. 1). Such information cannot be obtained by traditional binding kinetics studies that measure the interactions of drugs with purified membrane receptors immobilized on a solid surface. This is because the purification process isolates the receptors from the native cellular environment where binding modulators are located or bypass pathway developed, and the immobilization may affect the drug-receptor interactions.


In situ drug-receptor binding kinetics in single cells: a quantitative label-free study of anti-tumor drug resistance.

Wang W, Yin L, Gonzalez-Malerva L, Wang S, Yu X, Eaton S, Zhang S, Chen HY, LaBaer J, Tao N - Sci Rep (2014)

Top: Under “bypass hypothesis”, in situ binding kinetics of Herceptin with resistant cell was not affected. Bottom: “Alternation hypothesis” suggests an altered binding kinetics due to the influences of relevant proteins surrounding Her2.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4196117&req=5

f1: Top: Under “bypass hypothesis”, in situ binding kinetics of Herceptin with resistant cell was not affected. Bottom: “Alternation hypothesis” suggests an altered binding kinetics due to the influences of relevant proteins surrounding Her2.
Mentions: We hypothesized that in situ binding kinetics analysis in intact cells would provide a direct answer to this question, because the “alternation hypothesis” would mean a large change in the binding behaviors of drugs with the receptors in drug-resistant cells while the “bypass hypothesis” would imply no change in the binding kinetics (Fig. 1). Such information cannot be obtained by traditional binding kinetics studies that measure the interactions of drugs with purified membrane receptors immobilized on a solid surface. This is because the purification process isolates the receptors from the native cellular environment where binding modulators are located or bypass pathway developed, and the immobilization may affect the drug-receptor interactions.

Bottom Line: Many drugs are effective in the early stage of treatment, but patients develop drug resistance after a certain period of treatment, causing failure of the therapy.We further showed that the steric hindrance of Mucin-4, a membrane protein, was responsible for the altered drug-receptor binding.This effect of a third molecule on drug-receptor interactions cannot be studied using traditional purified protein methods, demonstrating the importance of the present intact cell-based binding kinetics analysis.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, China.

ABSTRACT
Many drugs are effective in the early stage of treatment, but patients develop drug resistance after a certain period of treatment, causing failure of the therapy. An important example is Herceptin, a popular monoclonal antibody drug for breast cancer by specifically targeting human epidermal growth factor receptor 2 (Her2). Here we demonstrate a quantitative binding kinetics analysis of drug-target interactions to investigate the molecular scale origin of drug resistance. Using a surface plasmon resonance imaging, we measured the in situ Herceptin-Her2 binding kinetics in single intact cancer cells for the first time, and observed significantly weakened Herceptin-Her2 interactions in Herceptin-resistant cells, compared to those in Herceptin-sensitive cells. We further showed that the steric hindrance of Mucin-4, a membrane protein, was responsible for the altered drug-receptor binding. This effect of a third molecule on drug-receptor interactions cannot be studied using traditional purified protein methods, demonstrating the importance of the present intact cell-based binding kinetics analysis.

Show MeSH
Related in: MedlinePlus