Survey of X-ray induced Cherenkov excited fluorophores with potential for human use.

X-ray induced molecular luminescence (XML) is a phenomenon that can be utilized for clinical, deep-tissue functional imaging of tailored molecular probes. In this study, a survey of common or clinically approved fluorophores was carried out for their megavoltage X-ray induced excitation and emission characteristics. We find that direct scintillation effects and Cherenkov generation are two possible ways to cause these molecules' excitation. To distinguish the contributions of each excitation mechanism, we exploited the dependency of Cherenkov radiation yield on X-ray energy. The probes were irradiated by constant dose of 6 MV and 18 MV X-ray radiation, and their relative emission intensities and spectra were quantified for each X-ray energy pair. From the ratios of XML, yield for 6 MV and 18 MV irradiation we found that the Cherenkov radiation dominated as an excitation mechanism, except for aluminum phthalocyanine, which exhibited substantial scintillation. The highest emission yields were detected from fluorescein, proflavin and aluminum phthalocyanine, in that order. XML yield was found to be affected by the emission quantum yield, overlap of the fluorescence excitation and Cherenkov emission spectra, scintillation yield. Considering all these factors and XML emission spectrum respective to tissue optical window, aluminum phthalocyanine offers the best XML yield for deep tissue use, while fluorescein and proflavine are most useful for subcutaneous or superficial use.

Photo-inducible cytotoxic and clastogenic activities of 3,6-di-substituted acridines obtained by acylation of proflavine.

The cytotoxicity and photo-enhanced cytotoxicity of a series of 18 3,6-di-substituted acridines were evaluated on both tumour CHO cells and human normal keratinocytes, and compared to their corresponding clastogenicity as assessed by the micronucleus assay. Compounds 2f tert-butyl N-[(6-tert-butoxycarbonylamino)acridin-3-yl]carbamate and 2d N-[6-(pivalamino)acridin-3-yl]pivalamide displayed a specific cytotoxicity on CHO cells. These results suggested that the two derivatives could be considered as interesting candidates for anticancer chemotherapy and hypothesized that the presence of 1,1-dimethylethyl substituents was responsible for a strong nonclastogenic cytotoxicity. Compounds 2b and 2c, on the contrary, displayed a strong clastogenicity. They indicated that the presence of nonbranched aliphatic chains on positions 3 and 6 of the acridine rings tended to induce a significant clastogenic effect. Finally, they established that most of the acridine compounds could be photo-activated by UVA-visible rays and focussed on the significant role of light irradiation on their biological properties.

Quantifying RNA-peptide interaction by single-quantum dot-based nanosensor: an approach for drug screening.

The sequence-specific RRE RNA-Rev binding is essential for HIV-1 replication and provides a useful in vitro system for real-time evaluating the inhibitory effect of drugs on the RRE-Rev interaction. The rapid and sensitive detection of RRE-Rev interaction in complex biological systems represents a fundamental challenge. Here we report the development of a single-quantum-dot (QD)-based nanosensor for sensitively quantifying Rev peptide-RRE interaction and characterizing the potential inhibitors by virtue of single-molecule detection and QD-based fluorescence resonance energy transfer (FRET). We demonstrate that the stoichiometry of Rev peptide binding to RRE can be accurately determined by using this single-QD-based nanosensor. Importantly, this single-QD-based nanosensor can sensitively quantify the inhibitory efficacy of proflavin on the Rev peptide-RRE binding, even in the presence of substantial levels of interference fluorescence from high-concentration proflavin, which usually prevents the discrimination of FRET signals in ensemble measurements. The application of this nanosensor in the screening of libraries of small-molecule drugs will facilitate the development of new drugs against various diseases, cancers, and HIV.

Inhibition of promutagen activation by the antioxidants butylated hydroxyanisole and butylated hydroxytoluene.

Butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) exhibited antimutagenic activity in the Salmonella typhimurium reversion test. Both BHA and BHT reduced reversion induced by chemicals requiring metabolic activation for effectiveness. However, they did not affect reversion induced by direct-acting mutagens. These results suggested that BHA and BHT may inhibit the metabolic activation processes and demonstrated that the S. typhimurium reversion test may be used to identify inhibitors of the neoplastic process.

Ozonation of mutagenic and carcinogenic polyaromatic amines and polyaromatic hydrocarbons in water.

The Salmonella-microsome assay for mutagenesis was used to determine the effect of ozone on the mutagenesis of selected carcinogens and mutagens in water. Short periods of ozonation were shown to completely inactivate the mutagenicity of several polyaromatic amine mutagens including acriflavine, proflavine, and beta-naphthylamine. Selected polyaromatic hydrocarbons were also sensitive to ozonation. Kinetic studies revealed that the mutagenicity of benzo(a)pyrene, 3-methylcholanthrene, and 7,12-dimethylbenz(a)anthracene was destroyed after short periods of ozonation. To correlate loss of mutagenicity with loss of carcinogenicity, two polyaromatic hydrocarbons were treated with ozone, extracted from water with hexane, and tested for carcinogenicity in mice. When 7,12-dimethyl-benz(a)anthracene and 3-methyl-cholanthrene were treated with ozone, there was a substantial reduction in carcinogenicity compared to control groups treated with oxygen alone. However, a small number of tumors developed in the group of animals receiving a hexane extract of ozonated 7,12-dimethylbenz(a)anthracene. This activity may be due to breakdown products of 7,12-dimethylbenz(a)anthracene that are not mutagenic.

Treatment of genital herpes simplex virus infection with photodynamic inactivation.

A double-blind randomized study to evaluate the effect of proflavine in the treatment of genital herpesvirus infection was conducted. One hundred fifty-seven women were studied, of whom 75 were treated with proflavine (treated women) and 82 were treated with placebo (control group). There were 62 women with primary disease and 95 with recurrent infection. Under the conditions by which this study was conducted, there was no apparent difference in the time of healing of lesions, development of recurrences, or virus isolation following treatment in the proflavine-treated and control groups.

Fractionation of of proteolytic enzymes by affinity chromatography on sepharose aminocaproyl proflavin.

The serine proteinases trypsin, chymotrypsin, elastase, and acrosin bind to the proflavin resin, the sulfhydryl proteinases ficin, bromelain, and papain are retarded by the resin, whereas most proteins and enzymes tested are not bound. Elution of the bound activities is accomplished NaCl or by variation from the pH optimum of the enzyme. Commercially available enzymes that are bound or retarded are easily further purified by the column. The acrosin activity of sperm acrosomal extracts is separated into bound and unbound activities. Acrosin is purified 120-fold from sperm acrosomal extracts in a single step, yielding a specific activity of 96.

Proflavine photodynamic viral inactivation in herpes simplex keratitis.

We used the photodynamic inactivation technique with proflavine as the photoactive dye to treat herpetic epithelial keratitis in a preliminary study of patients who had idoxuridine toxicity or resistance. A comparative study with idoxuridine in treating dendritic ulcerations of the cornea showed a good therapeutic effect. But the investigation was suspended when adverse reactions, consisting of a generalized epithelial keratitis and an anterior uveitis, possibly of phototoxic origin, developed in a few patients receiving treatment. The ulcers treated by photodynamic inactivation apparently healed by a process of "debridement" followed by subsequent re-epithelialization.

Comparison of the treatment of herpes genitalis in men with proflavine photoinactivation, idoxuridine ointment, and normal saline.

36 male patients with genital infection by HSV confirmed by culture were each allocated to one of three treatment groups: (1) Proflavine photoinactivation, (2) 0.5 per cent. idoxuridine ointment (IDU), (3) Normal saline. They were assessed objectively at each attendance by measurement of the lesions with an operating microscope fitted with a measuring grid in one eyepiece. Material for culture for HSV was taken at each visit; the presence of symptoms (pain, discomfort, and irritation) was noted. The areas of lesions in the proflavine photoinactivation group remained larger significantly longer than in the other groups, the healing time was slower, and HSV could be isolated for longer. It is concluded that proflavine photoinactivation is of no greater value than 0.5 per cent. IDU or normal saline in the treatment of genital infection by HSV in the male.

Anti-viral treatment of warts.

Several anti-DNA agents have been tested for their value in the treatment of warts in humans. A pilot study was carried out on patients with warts resistant to other forms of treatment. The agents used were hydroxyurea, idoxuridine and photodynamic inactivation. Idoxuridine 20% in a cream base appears to offer some possibilities as a treatment when other measures have failed.

Genital herpes: Treatment with methylene blue and light exposure.

Fifty-six patients with genital herpes were treated by photodynamic inactivation of herpesvirus by methylene blue (36 patients), proflavine (101 or neutral red (10). The first 35 patients were randomly selected for treatment with one of the three dyes. All the others were initially treated with methylene blue. The disease was temporarily eradicated in about 70% of instances by exposure of the lesions to light plus topically applied methylene blue, neutral red or proflavine. Relapses, however, were not prevented or appreciably reduced.