Original Article Morphoproteomic Evidence of Constitutively Activated and Overexpressed mTOR Pathway in Cervical Squamous Carcinoma and High Grade Squamous Intraepithelial Lesions
Wei Feng, Xiuzhen Duan, Jinsong Liu, Jianguo Xiao and Robert E. Brown
Department of Pathology, The University of Texas Medical School at Houston, Houston, TX, USA and Department of Pathology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
Received 10 August 2008; Accepted 17 September 2008; Available online 2 October 2008
Abstract: Background: Human papillomavirus (HPV) infection of the uterine cervix is linked to the pathogenesis of cervical cancer. Preclinical in vitro and in vivo studies using HPV-containing human cervical carcinoma cell lines have shown that the mammalian target of rapamycin (mTOR) inhibitor, rapamycin, and epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor, erlotinib can induce growth delay of xenografts. Activation of Akt and mTOR are also observed in cervical squamous cell carcinoma and, the expression of phosphorylated mTOR was reported to serve as a marker to predict response to chemotherapy and survival of cervical cancer patients. Therefore, we investigated: a) the expression level of EGFR in cervical squamous cell carcinoma (SCC) and high-grade squamous intraepithelial lesions (HSIL) versus non-neoplastic cervical squamous epithelium; b) the state of activation of the mTOR pathway in these same tissues; and c) any impact of these signal transduction molecules on cell cycle. Design: Formalin-fixed paraffin-embedded tissue microarray blocks containing 20 samples each of normal cervix, HSIL and invasive SCC, derived from a total of 60 cases of cervical biopsies and cervical conizations were examined. Immunohistochemistry was utilized to detect the following antigens: EGFR; mTOR pathway markers, phosphorylated (p)-mTOR (Ser 2448) and p-p70S6K (Thr 389); and cell cycle associated proteins, Ki-67 and S phase kinase-associated protein (Skp)2. Protein compartmentalization and expression were quantified in regard to proportion (0-100%) and intensity (0-3+). Mitotic index (MI) was also assessed. An expression index (EI) for p-mTOR, p-p70S6K and EGFR, respectively was calculated by taking the product of intensity score and proportion of positively staining cells. Results: Plasmalemmal EGFR expression was limited to the basal/ parabasal cells (2-3+, EI = 67) in normal cervical epithelium (NL), but was diffusely positive in all HSIL (EI = 237) and SCC (EI 226). The pattern of cytoplasmic p-mTOR and nuclear p-p70S6K expression was similar to that of EGFR; all showed a significantly increased EI in HSIL/SCC versus NL (p<0.02). Nuclear translocation of p-mTOR was observed in all SCC lesions (EI = 202) and was significantly increased versus both HSIL (EI = 89) and NL (EI = 54) with p<0.015 and p<0.0001 respectively. Concomitant increases in MI and proportion of nuclear Ki-67 and Skp2 expression were noted in HSIL and SCC. Conclusion: Morphoproteomic analysis reveals constitutive activation and overexpression of the mTOR pathway in HSIL and SCC as evidenced by: increased nuclear translocation of p-mTOR and p-p70S6K, phosphorylated at putative sites of activation, Ser 2448 and Thr 389, respectively; correlative overexpression of the upstream signal transducer, EGFR, and increases in cell cycle correlates, Skp2 and mitotic indices. These results suggest that the mTOR pathway plays a key role in cervical carcinogenesis and targeted therapies may be developed for SCC as well as its precursor lesion, HSIL. (IJCEP808002 ).
Address all correspondence to: Robert E. Brown, M.D., Department of Pathology & Laboratory Medicine, University of Texas Health Science Center-Houston Medical School, 6431 Fannin Street, MSB 2.286, Houston, TX 77030, USA; Tel: 713-500-5332;Fax: 713-500-0695; E-mail: firstname.lastname@example.org