Inhibition of cyclic GMP-binding cyclic GMP-specific phosphodiesterase (Type 5) by sildenafil and related compounds.

Turko IV, Ballard SA, Francis SH, Corbin JD.

Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, Nashville, Tennessee, USA.

The cGMP-binding cGMP-specific phosphodiesterase (PDE5) degrades cGMP and regulates the intracellular level of cGMP in many tissues, including the smooth muscle of the corpus cavernosum of the penis. Sildenafil (Viagra), a specific PDE5 inhibitor, promotes penile erection by blocking the activity of PDE5, which causes cGMP to accumulate in the corpus cavernosum. In the present study, sildenafil, like other PDE5 inhibitors, stimulates cGMP binding to the allosteric sites of PDE5 by interacting at the catalytic site of this enzyme, but the drug does not compete with cGMP for binding at the allosteric sites. Both sildenafil and zaprinast are competitive inhibitors of PDE5, and double-inhibition analysis shows that these two inhibitors added together interact with the catalytic site of PDE5 in a mutually exclusive manner. After site-directed mutagenesis of each of 23 conserved amino acid residues in the catalytic domain of PDE5, the pattern of changes in the IC50 values for sildenafil or UK-122764 is similar to that found for zaprinast. However, among the three inhibitors, sildenafil exhibits the most similar pattern of changes in the IC50 to that found for the affinity of cGMP, implying similar interactions with the catalytic domain. This may explain in part the stronger inhibitory potency of sildenafil for wild-type PDE5 compared with the other inhibitors [sildenafil (Ki = 1 nM) > UK-122764 (Ki = 5 nM) > zaprinast (Ki = 130 nM)]. The affinity of each of these inhibitors for PDE5 is much higher than that of cGMP itself (Km = 2000 nM). It is concluded that residues such as Tyr602, His607, His643, and Asp754 may form important interactions for sildenafil in PDE5, but because these amino acids are conserved in all mammalian PDEs, the selectivity and potency of sildenafil is likely to be provided by a nonconserved residue or residues in the PDE5 catalytic domain.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10385692&dopt=Abstract sildenafil Viagra online



Interactions between cGMP- and cAMP-pathways are involved in the regulation of penile smooth muscle tone.

Uckert S, Hedlund P, Waldkirch E, Sohn M, Jonas U, Andersson KE, Stief CG.

Hanover Medical School, Department of Urology, 30625 Hannover, Germany. sue_de_99 yahoo.de

Nitric oxide (NO)/cyclic GMP (cGMP)-mediated mechanisms have a pivotal function in reducing the tone of the penile smooth musculature during normal erectile responses. The cyclic AMP (cAMP) signaling pathway is also involved in the adjustment of smooth muscle contractility, and suggestions for interactions between cGMP- and cAMP-mediated mechanisms have been presented. Using activators of the cGMP- or the cAMP-pathway, as well as inhibitors of protein kinase A (PKA; cAMP-dependent kinase) and protein kinase G (PKG; cGMP-dependent kinase), the present study was undertaken to further delineate the functional relation between these pathways in the penis. In addition, the distribution of PKA and some cAMP-binding phosphodiesterases (cAMP-PDEs) were investigated in human erectile tissue. Functional experiments were performed on isolated human corpus cavernosum (HCC). The effects of an inhibitor of the PKA, Rp-8CPT-cAMPS (10 microM), or the PKG, Rp-8-pCPT-cGMPS (10 microM), on relaxation induced by the cumulative administration of sodium nitroprusside (SNP), forskolin, sildenafil or tadalafil (IC351) were studied in preparations of HCC precontracted with 1 microM norepinephrine (NE). Using immunohistochemical procedures, the presence of immunoreactivity for cAMP-PDEs PDE3, PDE4, and PDE4A, as well as for PKA was investigated in specimens of HCC from which preparations were also used in the functional experiments. Forskolin, SNP, sildenafil, and IC 351 dose-dependently reversed NE-induced tension of isolated HCC preparations. The relaxing effects of SNP were significantly attenuated by Rp-8-pCPT-cGMPS, but not by Rp-8CPT-cAMPS. In contrast, relaxation induced by forskolin, sildenafil and tadalafil were significantly reversed by both Rp-8-pCPT-cGMPS and Rp-8CPT-cAMPS. Abundant immunoreactivity for PDE3 and PKA was observed in the corpus cavernosum smooth muscle cells. Immunoreactivity for PDE4 was also detected in the smooth musculature and in the cytoplasm of endothelial cells lining the cavernous sinusoids, as well as in nerve fibres interspersing the trabecular stroma. The present results support the hypothesis of interactions between cGMP- and cAMP-mediated signals in the HCC, and suggest that the effects of inhibitors of PDE5 on isolated erectile tissue may also partly or indirectly include actions of the cAMP second messenger system. The exact mechanism by which such an interaction occurs is not clear, but it may involve altered activity of the cGMP-inhibited PDE3 brought about by a change in the intracellular levels of cGMP by the inhibition of PDE5. This will in turn lead to increasing levels of cAMP, facilitating the interaction of cAMP with the PKA. The immunoreactivity specific for PDE3, PDE4, PDE4A and PKA registered in HCC section is also in support of an important role for the cAMP/PKA-system for penile smooth muscle function.

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A new era in the treatment of erectile dysfunction.

Padma-Nathan H.

Male Clinic, Urology, Beverly Hills, California 90212, USA.

The recent introduction of sildenafil has revolutionized the treatment of erectile dysfunction (ED). The availability of sildenafil, the first effective oral agent for ED, has also expanded the field of sexual healthcare to include general and primary care practitioners and other nonurology specialists. A new process-of-care algorithm has been developed to facilitate the evaluation and treatment of ED in these nonurology settings. Sildenafil has been demonstrated to be safe and effective in randomized, double-blind, placebo-controlled trials involving >3,000 men ages 19-87 years. Sildenafil is effective across a broad range of etiologies including diabetes. For patients in whom first-line treatment with sildenafil fails or ceases to be effective, second-line interventions with intracavernosal self-injection therapy or transurethral alprostadil may be indicated. In addition to sildenafil, other oral agents such as oral phentolamine and sublingual apomorphine are now in clinical trials. Drugs under development include second-generation phosphodiesterase type 5 (PDE5) inhibitors, endothelin antagonists, and agents that offer specific molecular targeting. Clinical studies are also being planned to examine the efficacy of combination oral drug regimens for ED.

Online source: www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=10503572&dopt=Abstract sildenafil Viagra online



Prehospital consideration of sildenafil-nitrate interactions.

Reed DB, Gough JE, Ho JD, Brown LH.

Department of Emergency Medicine, State University of New York Health Science Center at Syracuse, 13210, USA. reedd mailbox.hscsyr.edu

OBJECTIVE: To determine whether paramedics and on-line physicians screen patients for use of sildenafil citrate (Viagra) prior to prehospital administration of nitrates. METHODS: A prospective, observational study was performed over a one-month period in three EMS systems. Consecutive radio communications between on-line physicians and paramedics concerning male patients with cardiac complaints were monitored. Investigators observed the frequency with which on-line physicians screened for sildenafil use prior to ordering nitrates. After observation of the radio communications was completed, a written survey was distributed to all paramedics in the three EMS systems. RESULTS: Seventy-six physician-paramedic interactions were monitored. Nitrates were ordered by on-line physicians in 56 cases. No paramedic reported sildenafil use/nonuse, and no on-line physician inquired about the patient's potential use of the drug. Only half of the surveyed paramedics reported that they routinely screen for sildenafil use, and approximately a fourth reported that its use would not alter their management of chest pain patients. CONCLUSION: In this study, on-line physicians in three EMS settings did not screen for sildenafil use prior to ordering nitrates. While some paramedics do screen for sildenafil use, practice patterns among paramedics in these three systems were inconsistent.

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Development of human and rabbit vaginal smooth muscle cell cultures: effects of vasoactive agents on intracellular levels of cyclic nucleotides.

Traish A, Moreland RB, Huang YH, Kim NN, Berman J, Goldstein I.

Department of Urology, Center for Advanced Biomedical Research, Boston University School of Medicine, Massachusetts 02118, USA. atraish bu.edu

In this study, we subcultured and characterized human and rabbit vaginal smooth muscle cells and investigated the synthesis of second messenger cyclic nucleotides in response to vasodilators and determined the activity and kinetics of phosphodiesterase (PDE) type 5 (EC 3.1.4.35 3',5'-cyclic GMP phosphodiesterase). Cultured vaginal cells exhibited growth characteristics typical of smooth muscle cells and immunostained with antibodies against alpha smooth muscle actin. The cells retained functional prostaglandin E and beta-adrenergic receptors as demonstrated by increased intracellular cAMP synthesis in response to PGE1, or isoproterenol. The response to these vasoactive substances was augmented with forskolin, suggesting stabilization of G-protein-activated adenylyl cyclases. Treatment with the nitric oxide donor, sodium nitroprusside, in the presence of sildenafil, a PDE type 5 inhibitor, enhanced intracellular cGMP synthesis and accumulation. Incubation of rabbit vaginal tissue with sildenafil, sodium nitroprusside, and PGE1 or forskolin produced a marked increase in intracellular cGMP. These observations were similar to findings with cultured cells and suggest that subcultured cells retain functional characteristics exhibited in intact tissue. The cells retained phosphodiesterase type 5 expression as shown by specific cGMP hydrolytic activity. Sildenafil and zaprinast inhibited cGMP hydrolysis competitively and bound with high affinity (Ki = 7 and 250 nM, respectively). These observations suggest that cultured human and rabbit vaginal smooth muscle cells retained their metabolic functional integrity and this experimental system should prove useful in investigating the pathway of nitric oxide and PDE type 5 inhibitors in modulating vaginal smooth muscle tone.

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Sildenafil.

Rosenberg KP.

Sexual Disorders Clinic, Cornell University Medical Center, New York, New York, USA.

Case reports have documented the utility of sildenafil for sexual dysfunction caused by selective serotonin-reuptake inhibitors (SSRIs) and have suggested its potential utility for women and men with various iatrogenic sexual dysfunctions. This brief review summarizes the psychopharmacology of sildenafil, discusses possible interactions with SSRIs, reviews side effects and risks, highlights the need for concomitant psychological counseling, reviews sildenafil's possible mechanisms of action for iatrogenic sexual dysfunctions, and suggests areas for future research.

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Interaction of sildenafil and indinavir when co-administered to HIV-positive patients.

Merry C, Barry MG, Ryan M, Tjia JF, Hennessy M, Eagling VA, Mulcahy F, Back DJ.

Department of Pharmacology and Therapeutics, Trinity Centre of Health Sciences, St James's Hospital, Dublin, Ireland.

OBJECTIVES: The prevalence of erectile dysfunction in HIV-infected men is estimated to be 33%. Sildenafil citrate (Viagra; Pfizer Ltd, Sandwich, Kent, UK) is the first oral drug for this condition. Since sildenafil and the protease inhibitors are both metabolized by, and act as inhibitors of cytochrome P450 3A4, we evaluated the pharmacokinetics of the combination sildenafil plus indinavir in HIV-infected patients. DESIGN AND METHODS: Six patients at steady state in treatment with indinavir participated in the study. On the first day blood samples for indinavir assay were drawn at times 0, 1, 2, 3, 4, 6 and 8 h after dosing. On the second study day patients received a single dose of 25 mg of sildenafil in addition to their routine morning medication. Blood samples were taken as described. Separated plasma was stored at -80 degrees C until analysis by high performance liquid chromatography. In a parallel study, the effect of indinavir, ritonavir, saquinavir and nelfinavir on the in vitro hepatic metabolism of sildenafil was assessed. RESULTS: The geometric mean area under the concentration curve for 0-8 h (AUC0-8h) and maximum plasma concentration (Cmax) for indinavir were 19.69 microg/ml h (range, 9.19-31.99 microg/ml h) and 7.02 microg/ml (range, 2.33-16.17 microg/ml), respectively, on the first study day. In the presence of sildenafil, the mean AUC0-8h and Cmax of indinavir were 22.37 microg/ml h [range, 10.08-37.25 microg/ml h; 95% confidence interval (CI) for difference between means, -15 to 13.25) and 9.11 microg/ml (range, 3.41-22.78 microg/ml; 95% CI, -13 to 6.37), respectively. The geometric mean AUC0-8h and Cmax for sildenafil were 1631 ng/ml h (range, 643-2970 ng/ml h) and 384 ng/ml (range, 209-766 ng/ml) respectively. The AUC for sildenafil was 4.4 times higher than data from historical controls given either 50 mg or 100 mg of sildenafil and dose normalized to 25 mg. Indinavir was a potent inhibitor of sildenafil hepatic metabolism in vitro [concentration producing 50% inhibition of control enzyme activity (IC50) = 0.39 +/- 0.17 microM, mean +/- SD]. CONCLUSIONS: Co-administration of sildenafil 25 mg did not significantly alter the plasma indinavir levels. However, plasma sildenafil AUC was markedly increased in the presence of indinavir compared with historical controls. From the in vitro data, the mechanism of increase is indinavir inhibition of the hepatic metabolism of sildenafil. The magnitude of this interaction suggests a lower starting dose of sildenafil may be more appropriate in this clinical setting.

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Evaluation of the safety of sildenafil for male erectile dysfunction: experience gained in general practice use in England in 1999.

Boshier A, Wilton LV, Shakir SA.

Drug Safety Research Unit, Bursledon Hall, Blundell Lane, Southampton SO31 1AA, UK. andrew.boshier dsru.org

OBJECTIVE: To examine the safety of sildenafil, the first of the phosphodiesterase type 5 inhibitors licensed for the treatment of male erectile dysfunction (ED), as used in general medical practice in England, quantifying the incidence of a range of events in patients treated with sildenafil, and identifying any previously unrecognized adverse drug reactions. METHODS: In a postmarketing observational cohort study using prescription-event monitoring (PEM), exposure data were derived from dispensed prescription details for patients who started treatment between April and August 1999. Outcome data were derived from "green form" questionnaires (GFs) returned by general practitioners (GPs). RESULTS In all, 24 835 (54.7%) of GFs posted to GPs were returned, of which 22 473 contained useful data for 22 471 male and two female patients. The major primary indications/clinical context of prescribing were impotence (16 583, 73.8%) and diabetes mellitus (183, 0.8%); 145 events were reported as adverse drug reactions to sildenafil. GPs recorded 3951 reasons for stopping sildenafil, and ischaemic heart disease (IHD) in 135 patients was the commonest clinical reason reported. The clinical condition reported most frequently in the first month of observation was diabetes mellitus and/or hyperglycaemia (in 99 events). A standardized mortality ratio (SMR) for deaths caused by IHD in the first 8893 of 22 473 patients was 31.41 (95% confidence interval 18.29-50.29), using the comparator population of males in England in 1998. CONCLUSION: This study identified the safety profile of sildenafil as used in the community, showing no unexpected events. The SMR analysis of deaths from IHD provided no evidence to suggest a higher incidence of deaths in the study cohort than in the male population in England.

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