Nitric Oxide Synthase Inhibition Enhances the Antitumor Effect of Radiation in the Treatment of Squamous Carcinoma Xenografts

Abstract

This study tests whether the nitric oxide synthase (NOS) inhibitor, NG-nitro-L-arginine (L-NNA), combines favorably with ionizing radiation (IR) in controlling squamous carcinoma tumor growth. Animals bearing FaDu and A431 xenografts were treated with L-NNA in the drinking water. IR exposure was 10 Gy for tumor growth and survival studies and 4 Gy for ex vivo clonogenic assays. Cryosections were examined immunohistochemically for markers of apoptosis and hypoxia. Blood flow was assayed by fluorescent microscopy of tissue cryosections after i.v. injection of fluorospheres. Orally administered L-NNA for 24 hrs reduces tumor blood flow by 80% (p<0.01). Within 24 hrs L-NNA treatment stopped tumor growth for at least 10 days before tumor growth again ensued. The growth arrest was in part due to increased cell killing since a combination of L-NNA and a single 4 Gy IR caused 82% tumor cell killing measured by an ex vivo clonogenic assay compared to 49% by L-NNA or 29% by IR alone. A Kaplan-Meyer analysis of animal survival revealed a distinct survival advantage for the combined treatment. Combining L-NNA and IR was also found to be at least as effective as a single i.p. dose of cisplatin plus IR. In contrast to the in vivo studies, exposure of cells to L-NNA in vitro was without effect on clonogenicity with or without IR. Western and immunochemical analysis of expression of a number of proteins involved in NO signaling indicated that L-NNA treatment enhanced arginase-2 expression and that this may represent vasculature remodeling and escape from NOS inhibition. For tumors such as head and neck squamous carcinomas that show only modest responses to inhibitors of specific angiogenic pathways, targeting NO-dependent pro-survival and angiogenic mechanisms in both tumor and supporting stromal cells may present a potential new strategy for tumor control.

Pyranocoumarins from Root Extracts of Peucedanum praeruptorum Dunn with Multidrug Resistance Reversal and Anti-Inflammatory Activities

“…In particular, five compounds (1, 7, 8, 11, and 13) showed considerably significant activities compared to those of known MDR inhibitors (verapamil and cyclosporine A, Figure 5). Previous phytochemical investigations revealed that compounds 2 and 4 inhibited LPS-induced NO production in macrophages [12] and compounds 2-4 showed MDR reversal activities in cancer cells [42,46]. However, our study appears to be the first report of the anti-inflammatory potential of compound 16 and the potential MDR reversal activity of compounds 1, 7, 8, 11, and 13 in tumor cells.…”

“…NO is mainly produced from L-arginine by the inducible nitric oxide synthase (iNOS) and is known to play a role in the host defense system against bacterial or viral infections or both by inducing inflammatory condition [43]. However, prolonged or hyper-stimulated NO production not only has the propensity to damage host cells but also contributes to cancer development by regulating the expression of genes involved in tumorigenesis [44][45][46]. Based on these scientific observations, NOS inhibitors and compounds that From the spectral data obtained, compound 1 was found to be a new angular-type pyranocoumarin, 3 1 -angeloyl-4 1 -tigloylkhellactone, which was similar to (+)-praeruptorin B (3), (+)-cis-(3 1 S,4 1 S)-3 1 ,4 1 -diangeloylkhellactone, except for the tigloyl group.…”

The Role of Nitric Oxide Synthase Uncoupling in Tumor Progression

“…All cell lines were grown in 10%FCS in RPMI1640 plus penicillin-streptomycin. Subcutaneous tumor xenografts were created in the hind legs of athymicNCr-nu/nu mice (3). MCF-7 xenograft growth was supported by 17β-estradiol pellets (1.5 mg, 60 day release, Innovative Research of America) inserted subcutaneously on the backs of mice.…”

“…Clonogenic assays and analytical methods for biopterins and cGMP have been described (3, 19). OCT blocks of human colon tissue were dissolved in PBS and the tissue collected by centrifugation prior to extracting in 0.1 N HCl for HPLC analysis.…”

“…NO can have both pro- and anti-tumorigenic activities depending on concentration and levels of other ROS (2). For example, inhibition of endogenous NO synthases (NOS) blocks the growth of some but not all tumor cells by direct cytotoxic effects on the tumor cells or by inhibiting tumor endothelial cells (3, 4). In contrast, increased activity of inducible NOS (iNOS) in breast and colorectal cancer cells up-regulates tumor growth promoting Wnt/β-catenin signaling (5).…”