Human α-defensin-1 inhibits growth of human lung adenocarcinoma xenograft in nude mice

Abstract

Human A-defensin-1 (HNP1), a small antimicrobial peptide, shows cytotoxicity to tumor cells in vitro and inhibitory activity for pathologic neovascularization in vivo. Here, we did a gene therapy with a plasmid that expresses a secretable form of HNP1 for assaying its antitumor activity. The expression and secretion of HNP1 were determined by reverse transcription-PCR and ELISA in vitro. We found that expression of HNP1 in A549 tumor cells caused significant growth inhibition. This effect is most likely cell autonomous, as a significant amount of recombinant HNP1 protein was found to be accumulated in the cytoplasm by immunohistochemical staining using an anti-HNP1 antibody and the supernatant containing secreted HNP1 failed to produce any noticeable antitumor activity. Flow cytometry and Hoechst 33258 staining showed that the number of apoptotic cells among the A549 cells expressing recombinant HNP1 proteins was significantly greater than that of the nontransfected control cultures, suggesting that this growth-inhibitory activity was due to an apoptotic mechanism triggered by the intracellular HNP1. The antitumor activity of intracellularly expressed HNP1 was also shown in vivo. Decreased microvessel density and increased lymphocyte infiltration were observed in tumor tissue from HNP1-treated mice through histologic analysis. These results indicate that intracellularly expressed HNP1 induces tumor cell apoptosis, which inhibits tumor growth. The antiangiogenesis effect of HNP1 may contribute to its inhibitory activity in vivo, and HNP1 might involve the host immune response to tumor. These findings provide a rationale for developing HNP1-based gene therapy for cancer. [Mol Cancer Ther 2008;7(6):1588 -97]

Intratumoral expression of mature human neutrophil peptide-1 potentiates the therapeutic effect of doxorubicin in a mouse 4T1 breast cancer model

“…ABC transporters confer a drug-resistant phenotype by decreasing intracellular accumulation of chemotherapeutic drugs via ATP-dependent efflux mechanisms (40). Previously, we found that HNP1 increased plasma membrane permeability in A549 cells (17). In the present research, the intracellular expression of HNP1 caused enhancement of Dox accumulation in 4T1 cells and exhibited a synergistic apoptosis-inducing effect in combination therapy.…”

“…Previously, we observed that intracellularly expressed HNP1 increased plasma membrane permeability in human A549 lung carcinoma cells (17). We also noted that the intratumoral expression of HNP1 mediated specific angiogenesis inhibition in vivo (17,18).…”

“…Previously, we observed that intracellularly expressed HNP1 increased plasma membrane permeability in human A549 lung carcinoma cells (17). We also noted that the intratumoral expression of HNP1 mediated specific angiogenesis inhibition in vivo (17,18). In addition, HNP1 can cause VEGF-dependent endothelial cell proliferation inhibition by forming a ternary complex with fibronectin and α 5 β 1 integrin (19).…”

“…Doxorubicin hydrochloride (Meiji Co.) was freshly prepared before each experiment. A recombinant plasmid pSecTag-HNP1 (pHNP1) that expresses a secretable form of mature HNP1 was previously constructed (17) and used in the present study. The concentration of MTT (Sigma) solution was 5 mg/ml dissolved in phosphate-buffered saline (PBS).…”

Intratumoral Expression of Mature Human Neutrophil Peptide-1 Mediates Antitumor Immunity in Mice

“…Furthermore, upregulation of HNP1-3 in tumor tissue and plasma has been thought to be a potential marker for prognostic assessment (14,15). We recently also reported that intratumoral expression of HNP1 within human A549 lung carcinoma xenografts resulted in increased cell apoptosis and inhibition of angiogenesis in nude mice (16).…”