Oncolytic viruses, including vaccinia virus (VACV), are a promising alternative to classical mono-cancer treatment methods such as surgery, chemo- or radiotherapy. However, combined therapeutic modalities may be more effective than mono-therapies. In this study, researchers enhanced the effectiveness of oncolytic virotherapy by matrix metalloproteinase (MMP-9)-mediated degradation of proteins of the tumoral extracellular matrix (ECM), leading to increased viral distribution within the tumors.
Protein expression levels were determined using Neuromics' Goat Polyclonal MMP-9 antibody: Simon Schäfer, Stephanie Weibel, Ulrike Donat, Qian Zhang, Richard J Aguilar, Nanhai G Chen and Aladar A Szalay. Vaccinia virus-mediated intra-tumoral expression of matrix metalloproteinase 9 enhances oncolysis of PC-3 xenograft tumors. BMC Cancer 2012, 12:366 doi:10.1186/1471-2407-12-366.
Images and Figures: MMP-9 expression in PC-3 tumor sections and collagen IV quantification. Intratumoral expression of MMP-9 (red) was visualized using MMP-9 labeled PC-3 tumor sections. Nuclei (white) were stained with Hoechst 33258 dye. GFP (green) is a VACV reporter gene. Tumors were obtained at 7 days p.i. from PC-3 tumor-bearing mice injected with PBS, GLV-1h68 or GLV-1h255. All images are representative examples. (B) Quantification of MMP-9 expression was done by microscopic analysis. Mean fluorescence intensities were measured with ImageJ. (C) Quantification of collagen IV 7 days p.i. in GLV-1h68 or GLV-1h255 infected areas of PC-3 tumor sections. Images were taken at a 100× magnification (Leica MZ16 FA) and converted from RGB to grayscale using Photoshop. For image analysis ImageJ was used, the threshold value was 8/255.
Figure and Images: Expression of functional MMP-9 by GLV-1h255-infected tumor cells. (A) Expression cassettes of GLV-1h68 and GLV-1h255. In GLV-1h255 the insert in the Tk locus was replaced by the human mmp-9 gene under control of the PSE promoter. PSEL, synthetic early/late promoter; PSE, synthetic early promoter; P7.5, VACV p7.5 K early/late promoter; P11, VACV p11 late promoter; Tk, thymidine kinase locus, Ha, hemagglutinin locus. (B) Expression of virus-encoded MMP-9 (92 kDa) in GLV-1h255 infected PC-3 cells and supernatants in vitro, β-actin (42 kDa) was used as a loading control. (C) Activity of the MMP-9 protein was tested by gelatin zymography. Lysates and supernatants of infected A549 cells were isolated and separated by non-reducing SDS-PAGE. In zymography, cleavage of the substrate by MMP-9 resulted in a clear band.
This study revealed that the degradation of collagen IV (ECM) by VACV-encoded MMP-9 may represent a new option to significantly enhance the oncolytic effect of rVACV in PC-3 xenografts. We confirmed that the degradation of collagen IV facilitated viral infection of the tumor tissue, represented by significantly higher viral tumor titers and an accelerated tumor regression. Furthermore, both oncolytic viruses, parental GLV-1h68 and mmp-9-encoding GLV-1h255, significantly reduced the size of lumbar and renal lymph node metastases, indicating that MMP-9 enhances both virotherapy of the primary tumor and sustains the rVACV-metastasis reducing effect.
I will keep you posted on further studies.
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