Cadherin 13 overexpression as an important factor related to the absence of tumor fluorescence in 5-aminolevulinic acid–guided resection of glioma

Abstract

Object Gliomas contain aggressive malignant cancer, and resection rate remains an important factor in treatment. Currently, fluorescence-guided resection using orally administered 5-aminolevulinic acid (5-ALA) has proved to be beneficial in improving the prognosis of patients with gliomas. 5-ALA is metabolized to protoporphyrin IX (PpIX) that accumulates selectively in the tumor and exhibits strong fluorescence upon excitation, but glioma cells do not always respond to 5-ALA, which can result in incomplete or excessive resection. Several possible mechanisms for this phenomenon have been suggested, but they remain poorly understood. To clarify the probable mechanisms underlying the variable induction of fluorescence and to improve fluorescence-guided surgery, the authors searched for key negative regulators of fluorescent signal induced by 5-ALA. Methods A comprehensive gene expression analysis was performed using microarrays in 11 pairs of tumor specimens, fluorescence-positive and fluorescence-negative tumors, and screened genes overexpressed specifically in fluorescence-negative tumors as the possible candidates for key negative regulators of 5-ALA–induced fluorescence. The most possible candidate was selected through annotation analysis in combination with a comparison of expression levels, and the relevance of expression of the selected gene to 5-ALA–induced fluorescence in tumor tissues was confirmed in the quantified expression levels. The biological significance of an identified gene in PpIX accumulation and 5-ALA–induced fluorescence was evaluated by in vitro PpIX fluorescence intensity analysis and in vitro PpIX fluorescence molecular imaging in 4 human glioblastoma cell lines (A1207, NMCG1, U251, and U373). Knockdown analyses using a specific small interfering RNA in U251 cells was also performed to determine the mechanisms of action and genes working as partners in the 5-ALA metabolic pathway. Results The authors chose 251 probes that showed remarkably high expression only in fluorescent-negative tumors (median intensity of expression signal > 1.0), and eventually the cadherin 13 gene (CDH13) was selected as the most possible determinant of 5-ALA–induced fluorescent signal in gliomas. The mean expression level of CDH13 in the fluorescence-negative gliomas was statistically higher than that in positive ones (p = 0.027), and knockdown of CDH13 expression enhanced the fluorescence image and increased the amount of PpIX 13-fold over controls (p < 0.001) in U251 glioma cells treated with 5-ALA. Comprehensive gene expression analysis of the CDH13-knockdown U251 cells demonstrated another two genes possibly involved in the PpIX biosynthesis: ATP-binding cassette transporter (ABCG2) significantly decreased in the CDH13 knockdown, while oligopeptide transporter 1 (PEPT1) increased. Conclusions The cadherin 13 gene might play a role in the PpIX accumulation pathway and act as a negative regulator of 5-ALA–induced fluorescence in glioma cells. Although further studies to clarify the mechanisms of action in the 5-ALA metabolic pathway would be indispensable, the results of this study might lead to a novel fluorescent marker able to overcome the obstacles of existing fluorescence-guided resection and improve the limited resection rate.

Tumor detection with 5-aminolevulinic acid fluorescence and Gd-DTPA–enhanced intraoperative MRI at the border of contrast-enhancing lesions: a prospective study based on histopathological assessment

“…An important step could be the identification of overexpression of cadherin 13 as a predictor of absent fluorescence in gliomas. 27 Yet, so far no histological or clinical predictors for a positive protoporphyrin IX accumulation have been identified in METs. However, from the clinical perspective a lesion that is suspected of being a MET might turn out to be a GBM intraoperatively.…”

In vivo visualization of GL261-luc2 mouse glioma cells by use of Alexa Fluor–labeled TRP-2 antibodies

“…Although the extent to which this may or may not occur is somewhat controversial, there is a need for the development of instruments with increased sensitivity for the detection of intraoperative fluorescence and a need for methods to increase specificity. 21,28 Additional studies are also needed to identify therapeutic agents and intrinsic molecular characteristics that affect the metabolism of 5-ALA, such as has been seen with phenytoin, 16 cadherin 13 expression, 34 aquaporin, 33 and ferrochelatase expression. 36 Other dyes, such as methylene blue, are being studied for their ability to enhance the morphologic image; 46 however, the utility of this approach relies on the surgeon's ability to recognize sometimes subtle morphological characteristics.…”

Optical technologies for intraoperative neurosurgical guidance

“…151 Multiple mechanisms and actuators have been posited as contributors to the selective accumulation of PpIX in tumors, including enzymatic modifications (e.g., ferrochelatase, porphobilinogen deaminase), transporter regulation, varying cellular states like hypoxia and acidic environments, increased cellular growth, glucose levels, metabolic and structural changes. 6,20,39,50,94,101,103,138,151,153,159,160 …”

“…132 In a subsequent reanalysis of the data to look at morbidity, no statistically significant difference in long-term morbidity was found between the fluorescence and white light groups. 102,133,136,138 …”