Development of an immunohistochemical protein quantification system in conjunction with tissue microarray technology
Sho Isoyama1, Hisashi Yoshimi, Shingo Dan
1Division of Molecular Pharmacology, Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, Tokyo, Japan.
Abstract:
The phosphatidylinositol 3-kinase (PI3K) pathway is frequently activated in human cancers by gain-of-function mutations of phosphoinositide-3-kinase, catalytic, alpha polypeptide (PIK3CA) or dysfunction of phosphatase and tensin homolog deleted on chromosome 10 (PTEN). Therefore PI3K is thought to be a promising target for cancer therapy. Many agents targeting PI3K have been developed and some of them have been evaluated in clinical trials. In recent years, development of predictive biomarkers as companion diagnostics for molecular targeted drugs has become an important requirement for clinical development; however, no clinically established biomarkers that predict the efficacy of PI3K inhibitors have been found. We previously reported that expression of phosphorylated Akt determined by immunoblot analysis correlated with the antitumor efficacy of a PI3K inhibitor ZSTK474 in vitro and in vivo, suggesting that it might be used as a predictive biomarker. In this study, to evaluate biomarker candidates in in vivo tumor samples, we developed an immunohistochemical protein detection/quantification system in conjunction with the tissue microarray technology using a panel of 24 human tumor xenografts (JFCR24). We have clearly demonstrated that expression levels of phosphorylated v-akt murine thymoma viral oncogene homolog (Akt) and mitogen-activated protein kinase (MAPK) determined by this system significantly correlated with those determined by immunoblot analysis. As expected, PTEN status correlated with expression of phosphorylated Akt but not MAPK. Finally, we confirmed that phosphorylated Akt levels determined using this system correlated with the in vivo efficacy of ZSTK474. The present results indicate that the immunohistochemical protein detection/quantification system could be used to quantify expression of biomarker proteins in xenografted tumor tissues as well as in human tumor specimens to predict drug efficacy in future clinical trials.
Insights
Phosphatidylinositol 3-kinase (PI3K) pathway activation drives cancer. Researchers developed an immunohistochemical system to quantify phosphorylated Akt, a predictive biomarker for PI3K inhibitor efficacy in clinical trials.
Area of Science:
- Oncology
- Molecular Biology
- Biomarker Development
Background:
- The phosphatidylinositol 3-kinase (PI3K) pathway is frequently dysregulated in human cancers, making it a key therapeutic target.
- Development of predictive biomarkers is crucial for molecularly targeted cancer therapies, but none are clinically established for PI3K inhibitors.
- Previous work suggested phosphorylated Akt (p-Akt) expression correlates with PI3K inhibitor efficacy.
Purpose of the Study:
- To evaluate biomarker candidates for predicting PI3K inhibitor efficacy using in vivo tumor samples.
- To develop and validate an immunohistochemical (IHC) system for quantifying protein expression in xenografted tumors.
Main Methods:
- Developed an IHC protein detection/quantification system combined with tissue microarray technology.
- Utilized a panel of 24 human tumor xenografts (JFCR24) for validation.
- Correlated IHC results with immunoblot analysis for phosphorylated Akt (p-Akt) and mitogen-activated protein kinase (MAPK), and assessed PTEN status.
Main Results:
- IHC quantification of p-Akt and MAPK significantly correlated with immunoblot analysis.
- PTEN status correlated with p-Akt expression, as expected, but not MAPK.
- p-Akt levels quantified by the IHC system correlated with the in vivo efficacy of the PI3K inhibitor ZSTK474.
Conclusions:
- The developed IHC system can accurately quantify biomarker protein expression in xenografted and human tumor tissues.
- This IHC system holds promise for predicting PI3K inhibitor drug efficacy in future clinical trials.
- p-Akt is a validated predictive biomarker for PI3K inhibitor therapy.
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