Abstract 1508: Development of a strategy to knock down SDF-1 using shRNA as a means to prevent pulmonary metastasis

• Published in: Cancer research (Chicago, Ill.) Ill.), 2010-04, Vol.70 (8_Supplement), p.1508-1508
• Main Authors: Annan, Anand C., Preuss, Meredith A., Borovjagin, Anton V., Ugai, Hideyo, Curiel, David T.
• Format: Article
• Language: English
• ISSN: 0008-5472; 1538-7445
• DOI: 10.1158/1538-7445.AM10-1508

Metastasis, a poor prognostic indicator causing increased mortality, occurs in an organ specific pattern with lungs and lymph nodes most commonly involved. Cancer cells expressing ‘CXC'-chemokine family Receptor-4 (CXCR4) are chemo-attracted to the pulmonary tissue expressing its cognate ligand Stromal Derived Factor-1 (SDF-1), which aid in formation of metastasis. In this regard, we propose to modulate this axis through knock down of SDF-1 specifically in pulmonary vasculature to prevent lung metastasis. Our laboratory has previously shown that human adenovirus (Ad), when injected via the tail vein into human Coxsackie Adenovirus Receptor (hCAR) transgenic mice, mice that ubiquitously express the hCAR native Ad receptor, transduces pulmonary vasculature preferentially. To this end, we plan to knock- down SDF-1 using shRNA against SDF-1 employing Ad as the vector to deliver them in vivo to achieve pulmonary targeting. To establish a pulmonary metastatic model in the “hCAR” mouse, we injected B16-F10 melanoma cells stably expressing CXCR4 (to cause metastasis) and firefly luciferase (to track tumor cells) via tail vein. Presence of lung metastasis was confirmed by histopathological analysis, immunohistochemistry for CXCR4 and luciferase assay of the tissues. In order to identify a shRNA to deliver to the pulmonary vasculature to decrease SDF-1, we evaluated several plasmid shRNAs for their ability to suppress SDF-1 expression at the mRNA level using qRT-PCR in vitro. From this experiment, we identified one shRNA which gave significant decrease in mRNA of SDF-1 measured by qRT-PCR (>70%). We created an Ad vector expressing this particular shRNA and measured its efficiency of inhibiting SDF-1 expression of approximately 75% for mRNA by qRT-PCR and approximately 90% for protein by Western Blot. Future studies include demonstrating the functional knock-down of SDF-1 in vitro by transwell chamber assay and subsequently analyzing the biological effect of knocking down SDF-1 in vivo in the pulmonary metastatic model established in “hCAR” mouse. In conclusion, therapies available for prevention of metastasis are limited despite different treatment modalities available for cancer. Our approach of targeted knock down of SDF-1 would reduce the metastatic burden to the lungs with potential for clinical translation into a therapeutic approach to prevent lung metastasis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting