Highly potent visnagin derivatives inhibit Cyp1 and prevent doxorubicin cardiotoxicity.

Anthracyclines such as doxorubicin are highly effective chemotherapy agents used to treat many common malignancies. However, their use is limited by cardiotoxicity. We previously identified visnagin as protecting against doxorubicin toxicity in cardiac but not tumor cells. In this study, we sought to develop more potent visnagin analogs in order to use these analogs as tools to clarify the mechanisms of visnagin-mediated cardioprotection. Structure-activity relationship studies were performed in a zebrafish model of doxorubicin cardiomyopathy. Movement of the 5-carbonyl to the 7 position and addition of short ester side chains led to development of visnagin analogs with 1,000-fold increased potency in zebrafish and 250-fold increased potency in mice. Using proteomics, we discovered that doxorubicin caused robust induction of Cytochrome P450 family 1 (CYP1) that was mitigated by visnagin and its potent analog 23. Treatment with structurally divergent CYP1 inhibitors, as well as knockdown of CYP1A, prevented doxorubicin cardiomyopathy in zebrafish. The identification of potent cardioprotective agents may facilitate the development of new therapeutic strategies for patients receiving cardiotoxic chemotherapy. Moreover, these studies support the idea that CYP1 is an important contributor to doxorubicin cardiotoxicity and suggest that modulation of this pathway could be beneficial in the clinical setting.

Pharmacokinetic evaluation of visnagin and Ammi visnaga aqueous extract after oral administration in rats.

The furanochromone visnagin is one of the main compounds of Ammi visnaga L. (syn. Khella) with potential effects on kidney stone prevention. After determination of the pharmacokinetic properties of visnagin after intravenous bolus administration in rats, the aim of the present study was to evaluate the pharmacokinetic properties of visnagin and an aqueous Ammi visnaga extract after oral administration in rats. In two separate experiments, three doses of visnagin (2.5, 5, and 10 mg/kg) solubilized in 25 % Captisol® and three doses of Ammi visnaga extract (standardized on visnagin and containing equivalent amounts of visnagin) were administered by oral gavage to male Sprague-Dawley rats, respectively. Plasma samples were extracted and subsequently analyzed using a validated LC-MS/MS method. Plasma concentration-time profiles were explored by non-compartmental analysis. Visnagin plasma exposure (median AUClast and AUCinf) was significantly increased for all three doses (more than 10-fold for the low dose) when administered as an extract compared to the pure agent. For both the Ammi visnaga extract and the pure compound, AUClast and AUCinf increased disproportionately with an increase in dose. Visnagin resided significantly longer in the body when given in the form of AVE with up to a three times longer median MRTlast and MRTinf for the low dose. Cmax values after AVE administration were elevated and occurred at later time points in comparison to equivalent doses of pure visnagin. The terminal half-life increased with the dose for both AVE and pure visnagin, reaching a maximum value of 1.94 h for the 10 mg/kg pure compound group.In conclusion, the exposure of visnagin is enhanced after extract administration and could result in a superior efficacy of AVE compared to an equivalent dose of visnagin.

Nonlinear pharmacokinetics of visnagin in rats after intravenous bolus administration.

Ammi visnaga L. (syn. Khella, Apiaceae) preparations have traditionally been used in the Middle East for the treatment of kidney stone disease. Visnagin, a furanocoumarin derivative, is one of the main compounds of Ammi visnaga with potential effects on kidney stone prevention. To date, no information is available about the pharmacokinetic (PK) properties of visnagin. It was the aim of the study to characterize the PK properties of visnagin after intravenous (i.v.) bolus administration in rats and to develop an adequate model for the description of the observed data, including model parameter estimates. Therefore, three doses of visnagin (1.25, 2.5, and 5mg/kg) solubilized in 25% Captisol® were administered by i.v. bolus injection to male Sprague-Dawley rats. Plasma samples were extracted and subsequently analyzed using a validated LC-MS/MS method. Both non-compartmental and compartmental PK analyses were performed. A stepwise model building approach was applied including nonlinear mixed effect modeling for final model selection and to obtain final model estimates in NONMEM VI. The average areas under the curve (AUC(0-last)) after doses of 1.25, 2.5, and 5mg/kg were 1.03, 3.61, and 12.6 mg *h/l, respectively. The shape of the plasma concentration-time profiles and the observed disproportionate increase in AUC(0-last) with increasing dose suggested nonlinearity in the elimination of visnagin. A two-compartment Michaelis-Menten model provided the best fit with following typical values of the parameter estimates: 2.09 mg/(l*h) (V(max)), 0.08 mg/l (K(M)), 0.175 l (V(C)), 1.0 h⁻¹ (k12), and 1.22 h⁻¹ (k21). Associated inter-subject variability estimates (% CV) for V(max), K(M) and V(C) were 21.8, 70.9, and 9.2, respectively. Intra-subject variability (constant CV error model) was estimated to be 7.0%. The results suggest the involvement of a saturable process in the elimination of visnagin, possibly an enzyme or transporter system.

Treatment of vitiligo with khellin liposomes, ultraviolet light and blister roof transplantation.

BACKGROUND: Various surgical and non-surgical methods are available to treat vitiligo. Surgical techniques such as epidermal blister graft transplantation may be effective for the re-pigmentation of stable, but refractory vitiligo areas. Khellin has phototherapeutic properties that are similar to those of the psoralens, but with substantially lower phototoxic effects and DNA mutation effects. Its penetration into the hair follicles is enhanced by encapsulating it into liposomes. Subsequent activation of the khellin with UV light stimulates the melanocytes in the hair follicles. OBJECTIVE: The first objective was to evaluate the additional value of combining blister roof transplantation (BRT) with khellin in liposomes and ultraviolet light (KLUV) in the treatment of recalcitrant vitiligo patches. The second objective was to assess patients' satisfaction. MATERIALS AND METHODS: Nineteen patients with vitiligo lesions which did not respond to KLUV treatment for at least a year were treated with BRT followed by KLUV. The transplantation was performed by creating blisters with a suction device, preparing the target site with Erbium laser ablation and the actual transplantation. Locations where randomly assigned. A blinded observer established the results. RESULTS: Seventy-five percent of the patients were satisfied with the cosmetic result. All of the patients would recommend the treatment to other vitiligo patients. More than 75% re-pigmentation of the vitiligo areas was noted in 47% of the patients according to the blinded evaluation of photographs taken before and after the treatment.

Prevention of renal crystal deposition by an extract of Ammi visnaga L. and its constituents khellin and visnagin in hyperoxaluric rats.

In Egypt, teas prepared from the fruits of Ammi visnaga L. (syn. "Khella") are traditionally used by patients with urolithiasis. The aim of this study was to evaluate whether oral administration of an aqueous extract prepared from the fruits of A. visnaga as well as two major constituents khellin and visnagin could prevent crystal deposition in stone-forming rats. Hyperoxaluria was induced in male Sprague-Dawley rats by giving 0.75% ethylene glycol and 1% ammonium chloride via the drinking water. The Khella extract (KE; 125, 250 or 500 mg/kg) was orally administered for 14 days. The histopathological examination of the kidneys revealed that KE significantly reduced the incidence of calcium oxalate (CaOx) crystal deposition. In addition, KE significantly increased urinary excretion of citrate along with a decrease of oxalate excretion. Comparable to the extract, khellin and visnagin significantly reduced the incidence of CaOx deposition in the kidneys. However, both compounds did not affect urinary citrate or oxalate excretion indicating a mechanism of action that differs from that of the extract. For KE, a reasonably good correlation was observed between the incidence of crystal deposition, the increase in citrate excretion and urine pH suggesting a mechanisms that may interfere with citrate reabsorption. In conclusion, our data suggest that KE and its compounds, khellin and visnagin, may be beneficial in the management of kidney stone disease caused by hyperoxaluria but that it is likely that different mechanism of action are involved in mediating these effects.

Monochromatic excimer light 308 nm in monotherapy and combined with topical khellin 4% in the treatment of vitiligo: a controlled study.

Vitiligo is an acquired depigmentation disorder affecting 1-4% of the world's population. Conventional therapies include steroids, photosensitive topical agents, surgical treatments, and phototherapy. The aim of the study was to evaluate the efficacy of monochromatic excimer light 308 nm (MEL), both as a monotherapy and in combination with khellin 4% ointment in vitiligo. Forty-height patients (36 male and 12 female) affected with vitiligo were enrolled in this open prospective study. Patients were selected and divided into three groups: group I included 16 patients treated with MEL 308 nm once-weekly and oral vitamin E; group II included 16 patients treated with MEL 308 nm once-weekly combined with khellin 4% ointment (MEL-K) and oral vitamin E; group III (control group) included 16 patients treated only with oral vitamin E. Efficacy was assessed at the end of 12 weeks based on the percentage of repigmentation. Group I (MEL-group) showed a moderate repigmentation in 2/16 (12.5%) patients, good repigmentation in 10/16 (62.5%), and excellent repigmentation in 4/16 (25%) patients. Group II (MEL-K group) presented moderate repigmentation in 2/16 (12.5%) patients, good repigmentation in 5/16 (31.25%), and excellent repigmentation in 9/16 (56.25%). Group III (control group) showed a moderate repigmentation in 3/16 patients (18.75%), a good repigmentation in 1/16 (6.25%) patient, while 10/16 (62.5%) patients did not show signs of repigmentation. The clinical response achieved in group I and II was higher compared with group III (control group) without showing significant differences. MEL 308 nm, alone and/or combined with khellin 4% offered encouraging results and it may be considered a valid therapeutic option worthy of consideration in the treatment of vitiligo.

A case study to evaluate the treatment of vitiligo with khellin encapsulated in L-phenylalanin stabilized phosphatidylcholine liposomes in combination with ultraviolet light therapy.

Vitiligo destroys the melanocytes in the epidermis; the inactive melanocytes in the outer root sheaths are not affected. Phosphatidylcholine liposomes are able to target molecules contained in them into the hair follicles. Khellin is activated by UVA and previous studies have shown that a combination of khellin and UVA (KUVA) can be effective in the treatment of vitiligo. The aim of this study was to determine in an open trial the efficacy and safety of treatment with khellin encapsulated in L-phenylalanine stabilized phosphatidylcholine liposomes in combination with UVA/UVB light therapy(KPLUV) in 74 subjects with vitiligo. After a mean treatment period of 12 months (range 10-14 months) 72% of the treated locations had a repigmentation response of 50% to 100%. Repigmentation of 75-100% was achieved on the face in 63%, the back in 59%, the arms in 58%, the trunk in 57%, the legs in 56% and on the hands in 4% of the patients. Side effects were not seen with KPLUV. The patients in the control group, only treated with UVA/B-light, hardly showed any repigmentation. This indicates that the exposure of the skin to UV light alone is not responsible for the results of KPLUV.

In-vitro release characteristics of tetracycline HCl, khellin and nicotinamide adenine dineculeotide from halloysite; a cylindrical mineral.

The use of halloysite clay as a low cost alternative to more traditional microencapsulation systems is reported. Halloysite is an alumino-silicate clay which demonstrates a predominately cylindrical geometry, uniquely characterized by a hollow core or series of voids with diameters ranging from 16-50 nm. These nanoscale-to-mesoscale microcylinders are capable of entrapping active agents within the core lumen as well as within any void spaces contained in the multilayered walls of the cylinder. Some of the active agents associated with the clay are also bound to the external surfaces of the clay. Delivery of the active agent is first by desorption of the active agent from the exterior surfaces and exposed ends of the microcylinders, and is followed by a second more prolonged phase dominated by pore diffusion from the ends of the cylinders. Halloysite is capable of retaining and releasing a range of active ingredients. Both hydrophilic and hydrophobic agents may be entrapped following appropriate pre-treatment of the clay to render it lipophilic. Here, a unique low cost alternative microcylindrical delivery system: the clay mineral halloysite, is investigated.

Treatment with topical khellin in combination with ultraviolet A or solar-simulated radiation is carcinogenic to lightly pigmented hairless mice.

Khellin is used together with either UVA irradiation or sun exposure in the treatment of vitiligo. The purpose of this study was to investigate the carcinogenic effect of topically applied khellin together with UVA or solar simulated radiation (SSR) in lightly pigmented C3H/Tif mice. For comparison purposes a 0.1% 8-methoxypsoralen (8-MOP) cream was also tested in combination with SSR. Fifty microliters of a 5% khellin cream, a 0.1% 8-MOP cream, or a cream without active substances were spread uniformly on the back of the mice 30 minutes before UVA or SSR irradiation. All mice were irradiated 3 times a week until age or skin tumor development necessitated killing. Treatment with topical khellin and UVA irradiation was carcinogenic to lightly pigmented hairless mice, time to 50% of the mice had developed one tumor (t50) was 507 days. This indicates that the combination of topical khellin and UVA radiation, formerly expected to be rather innocuous, is carcinogenic to mice. Also the combination of khellin and SSR (t50 = 268 days) enhanced skin tumor development significantly compared with control cream and SSR (t50 = 330 days), P < 0.05. In addition, the combination of khellin and SSR was found to have the same carcinogenic effect as treatment with 0.1% 8-MOP and SSR (t50 = 262 days). This study shows that topically applied khellin increases the carcinogenic effect of both UVA and sunlight.

Photochemotherapy with topical khellin and sunlight in vitiligo.

In order to evaluate the efficacy of topical khellin the vitiligo macules of one side only were painted in 41 patients with a 2% solution of khellin in acetone and propylene glycol (90 and 10%, respectively) and exposed to sunlight for a period of 4 months with 3 weekly applications and with exposure times up to 90 min. The macules of the other side were treated in 36 of the 41 patients with acetone and propylene glycol only and sun-exposed with the same schedule, while in the remaining 5 patients they were neither treated with khellin or placebo nor sun-exposed. No significant difference was evidenced between the khellin and placebo-treated sides: no excellent result (repigmentation more than 75% of the affected area) was found, and good results (repigmentation more than 50%) were found in 24.9% of khellin- plus sunlight-treated macules and in 22.3% of placebo- plus sunlight-treated macules.

Preliminary report on the therapeutic effect of khellin in psoriasis.

A double-blind study has been performed to search for the possible effect of khellin in psoriasis. 10 patients were treated orally with khellin and subsequently exposed to sunlight for 4 months; a total of 8 cases responded positively with variable degrees of clearance. None of 10 controls had any response. A relapse-free duration of 3 years was recorded for 6 out of the 8 responders. This study would introduce the nonpsoralen compound khellin as a possible agent for the therapy of psoriasis.