Delivering advanced therapies: the big pharma approach.

After two decades of focused development and some recent clinical successes, cell and gene therapy (CGT) is emerging as a promising approach to personalized medicines. Genetically engineered cells as a medical modality are poised to stand alongside or in combination with small molecule and biopharmaceutical approaches to bring new therapies to patients globally. Big pharma can have a vital role in industrializing CGT by focusing on diseases with high unmet medical need and compelling genetic evidence. Pharma should invest in manufacturing and supply chain solutions that deliver reproducible, high-quality therapies at a commercially viable cost. Owing to the fast pace of innovation in this field proactive engagement with regulators is critical. It is also vital to understand the needs of patients all along the patient care pathway and to establish product pricing that is accepted by prescribers, payers and patients.

Viral hepatitis and liver cancer on the Island of Guam.

Patient records from the Guam Cancer Registry were compared with patients listed in a health department viral hepatitis case registry and the numbers of liver cancer and viral hepatitis cases were compared by ethnicity. Hepatitis C was the form of viral hepatitis most common among liver cancer cases on Guam (63.3% of viral hepatitis-associated liver cancer cases). Since viral hepatitis is an important cause of liver cancer, studies such as the present one may provide the information necessary to establish programs (screening of populations at risk and infant vaccination in the case of hepatitis B, for example) that may lessen the impact of liver cancer in the future.

Asymmetrical nostril axis angle in the post traumatic nose.

BACKGROUND: Nasal tip reconstruction is a challenging part of septorhinoplasty in post-traumatic patients. Aesthetic appearance of the nose is dependent on adequate tip definition, projection and rotation. Our aim is to establish if caudal septal dislocation (secondary to fracture or subluxation) is a significant factor in causing asymmetry in nostril axis angle (NAA) to effect surgical intervention. In addition, to assess if there is an association between the side of dislocation and the magnitude of NAA. METHODOLOGY: Photographic assessment of NAA was measured in a study group of 26 patients with caudal dislocation of the septum and a control group of 26 patients without caudal septal dislocation. All 52 patients were awaiting a septorhinoplasty procedure due to nasal trauma. RESULTS: Caudal septal dislocation causes a mean difference in NAA of 7.95o compared to 1.38o difference in the control group. We also found there is a greater degree of upward tip rotation on the side of the dislocation resulting in a larger NAA. CONCLUSION: Caudal dislocation of the septum causes a significant asymmetry in NAA, with a greater degree of rotation on the side of the dislocation. We should be aware of this fallacy prior to surgery.

Dynamics of a three-variable nonlinear model of vasomotion: comparison of theory and experiment.

The effects of pharmacological interventions that modulate Ca(2+) homeodynamics and membrane potential in rat isolated cerebral vessels during vasomotion (i.e., rhythmic fluctuations in arterial diameter) were simulated by a third-order system of nonlinear differential equations. Independent control variables employed in the model were [Ca(2+)] in the cytosol, [Ca(2+)] in intracellular stores, and smooth muscle membrane potential. Interactions between ryanodine- and inositol 1,4,5-trisphosphate-sensitive intracellular Ca(2+) stores and transmembrane ion fluxes via K(+) channels, Cl(-) channels, and voltage-operated Ca(2+) channels were studied by comparing simulations of oscillatory behavior with experimental measurements of membrane potential, intracellular free [Ca(2+)] and vessel diameter during a range of pharmacological interventions. The main conclusion of the study is that a general model of vasomotion that predicts experimental data can be constructed by a low-order system that incorporates nonlinear interactions between dynamical control variables.

Differential activation of ion channels by inositol 1,4,5-trisphosphate (IP3)- and ryanodine-sensitive calcium stores in rat basilar artery vasomotion.

Spontaneous, rhythmical contractions, or vasomotion, can be recorded from cerebral vessels under both normal physiological and pathophysiological conditions. Using electrophysiology to study changes in membrane potential, the ratiometric calcium indicator Fura-2 AM to study changes in [Ca(2+)](i) in both the arterial wall and in individual smooth muscle cells (SMCs), and video microscopy to study changes in vessel diameter, we have investigated the cellular mechanisms underlying vasomotion in the juvenile rat basilar artery. During vasomotion, rhythmical oscillations in both membrane potential and [Ca(2+)](i) were found to precede rhythmical contractions. Nifedipine depolarized SMCs and abolished rhythmical contractions and depolarizations. [Ca(2+)](i) oscillations in the arterial wall became reduced and irregular, while [Ca(2+)](i) oscillations in adjacent SMCs were no longer synchronized. BAPTA-AM, thapsigargin and U73122 hyperpolarized SMCs, relaxed the vessel, decreased basal calcium levels and abolished vasomotion. Chloride substitution abolished rhythmical activity, depolarized SMCs, increased basal calcium levels and constricted the vessel, while niflumic acid and DIDS abolished vasomotion. Ryanodine, charybdotoxin and TRAM-34, but not iberiotoxin, 4-aminopyridine or apamin, each depolarized SMCs and increased the frequency of rhythmical depolarizations and [Ca(2+)](i) oscillations. We conclude that vasomotion in the basilar artery depends on the release of intracellular calcium from IP(3) (inositol 1,4,5,-trisphosphate)-sensitive stores which activates calcium-dependent chloride channels to depolarize SMCs. Depolarization in turn activates voltage-dependent calcium channels, synchronizing contractions of adjacent cells through influx of extracellular calcium. Subsequent calcium-induced calcium release from ryanodine-sensitive stores activates an intermediate conductance potassium channel, hyperpolarizing the SMCs and providing a negative feedback pathway for regeneration of the contractile cycle.

Voltage independence of vasomotion in isolated irideal arterioles of the rat.

The cellular mechanisms underlying vasomotion of irideal arterioles from juvenile rats have been studied using electrophysiological methods, ratiometric calcium measurements and video microscopy. Vasomotion was not affected by removal of the endothelium. Spontaneous contractions were preceded by spontaneous depolarizations. Both were abolished by the intracellular calcium chelator, BAPTA AM (20 microM), but not by ryanodine (10 microM), suggesting a dependence on the cyclical release of calcium from intracellular stores, other than those operated by ryanodine receptors. Oscillations were little changed when the membrane potential of short segments of arteriole was either depolarized or hyperpolarized. When the segments were voltage clamped, oscillating inward currents were recorded, indicating that the changes in membrane potential were voltage independent. Vasomotion was preceded by intracellular calcium oscillations and both were abolished by inhibitors of phospholipase C (U73122, 10 microM), phospholipase A(2) (AACOCF(3), 30 microM) and protein kinase C (chelerythrine chloride, 5 microM, and myristoylated protein kinase C peptide, 10 microM). Inhibition of vasomotion by the dual lipoxygenase and cyclo-oxygenase inhibitor, NDGA (10 microM), the lipoxygenase inhibitor, ETI (1 microM) but not by the cyclo-oxygenase inhibitors, aspirin (10 microM) and indomethacin (10 microM), or the cytochrome P450 inhibitor 17-ODYA (10 microM), suggested an involvement of the lipoxygenase pathway. The observations suggest that vasomotion of iris arterioles is voltage independent and results from the cyclical release of calcium from IP(3)-sensitive stores which are activated by cross talk between the phospholipase C and phospholipase A(2) pathways in vascular smooth muscle.

Sibutramine pharmacokinetics in young and elderly healthy subjects.

OBJECTIVE: To investigate the pharmacokinetics of the pharmacologically active metabolites of sibutramine (metabolites 1 and 2) in healthy young and elderly volunteers following a single oral dose of sibutramine. METHODS: This was an open, parallel-group study completed by 12 young (six male, six female; mean age 24.0 years) and 12 elderly (six male, six female; mean age 70.3 years) healthy volunteers. Blood samples were taken at intervals up to 48 h post-dose. Plasma concentrations of metabolites were determined using HPLC-MS. Model-independent pharmacokinetic parameters of the two metabolites were compared for the two age groups. RESULTS: The similarity of the plasma profiles of the two desmethyl metabolites showed that despite the possibility of reduced hepatic function due to age, the rate and extent of formation of these was the same in both young and elderly, i.e. sibutramine metabolism was not impaired in elderly subjects. There were also no significant differences in elimination of metabolite 2 between groups, although the elderly group showed a slight trend for a reduction in k(el). CONCLUSIONS: The pharmacokinetics of the two pharmacologically active metabolites of sibutramine (metabolites 1 and 2) were not significantly different between the young and elderly groups in this study. Based on this information, a similar dosing regimen would be appropriate for both the young and elderly.

Measles outbreaks in Micronesia, 1991 to 1994.

BACKGROUND: Several islands in Micronesia experienced large measles outbreaks, during 1991 through 1994. Except for Guam, none of the islands had reported measles outbreaks during the previous 20 years. METHODS: To characterize the outbreaks, measles surveillance data, hospital records and death certificates were reviewed. Preoutbreak vaccination coverage rates were assessed by reviewing public health vaccination records. Viral isolates were genetically sequenced to determine the source of transmission. Linear regression analysis was performed to assess the effectiveness of outbreak control measures. RESULTS: Between 1991 and 1994 more than 1300 measles cases and 16 measles-related deaths were reported in Micronesia. Preoutbreak vaccination coverage rates among 2-year-old children were 55 to 94%. Genetic sequencing of the viral isolates and epidemiologic investigations suggested transmission between islands and new importations from outside of Micronesia. The highest attack rates were among children ages < 5 years (20/1000) and 10 to 19 years (38/1000). Compared with attack rates among children ages < 1 and 10 to 19 years, attack rates were lower among those ages 5 to 9 years, in whom 2-dose vaccination coverage rates were highest (P < 0.001). Early and rapid implementation of mass vaccination campaigns was significantly associated with shorter duration of outbreaks (P = 0.049). CONCLUSION: The measles outbreaks in Micronesia show that island populations may be highly susceptible to measles. High two-dose vaccination coverage levels must be maintained to prevent such outbreaks. Early and rapidly implemented mass measles vaccination campaigns were effective in control of island outbreaks. Strengthening public health infrastructure and surveillance is necessary for early identification of outbreaks and rapid implementation of mass campaigns.

Familial nature and continuing morbidity of the amyotrophic lateral sclerosis-parkinsonism dementia complex of Guam.

Chamorros suffer from two neurologic syndromes known as ALS and the parkinsonism-dementia complex (PDC) of Guam. We report mortality figures for these syndromes during 1991 to 1995 and compare them with those at 5-year intervals dating back to 1951. In contrast to predictions of disease disappearance, both syndromes remain prevalent. However, age of onset and age at death have increased for both syndromes, suggesting that shifting environmental factors are causing disease postponement. We also report the clinical, familial, neuropathologic, and immunohistochemical findings on a consecutive autopsy series of Guamanian Chamorro cases. Twelve cases were diagnosed as PDC, known locally as "bodig," and three as ALS, known locally as "lytico." All but three of these fifteen patients had a pronounced family history of similar illness. Eight of twelve boding patients had siblings who were also affected with bodig; two of three lytico cases had siblings afflicted with lytico. The family histories are compatible with genetic transmission of each syndrome. The neuropathology of bodig is characterized by severe and widespread neurofibrillary tangle (NFT) development. NFTs are surrounded by reactive microglia and reactive astrocytes, and complement proteins and other molecules connected with inflammation are associated with them. Similar inflammatory responses also occur in Alzheimer's disease (AD) but have been largely attributed to the presence of senile plaques. These data indicate that tangles, as well as plaques, generate inflammatory reactions that such reactions may exacerbate the fundamental pathology in bodig as well as in AD.

Histamine degradative uptake by cultured human pulmonary vascular endothelial cells utilizes an inflammatory cell diamine oxidase.

Neutrophil-endothelial interactions, altered clearance properties of the lung toward vasoactive mediators, and damaging effects of histamine that target the lung represent prominent elements of the inflammatory response at the systemic level. The pulmonary vasculature is unusual in that, unlike other tissues and vascular beds, it normally does not metabolize circulating histamine in vivo, although histamine-metabolizing enzyme activities have been detected in disrupted lung tissue. We have therefore explored the capability of human pulmonary artery endothelial cells in culture to express the receptor-mediated histamine degradative uptake system we previously defined in systemic endothelial cells (Haddock, R. C., Mack, P., Leal, S., and Baenziger, N. L. (1990) J. Biol. Chem. 265, 14395-14401). Pulmonary endothelial cells display all components of this system: histamine methyltransferase generating the proximal cell-associated metabolite tele-methylhistamine and receptors binding diamine oxidase which generates the distal product methylimidazoleacetic acid that is accumulated by the cells. A diamine oxidase released from human neutrophil granules by activation with Ca2+ ionophore binds pulmonary and systemic endothelial cell and fibroblast diamine oxidase receptors and, thereby, participates in histamine degradative uptake. This enzyme utilizes cell-associated tele-methylhistamine as a substrate, preferentially generating methylimidazoleacetic acid in addition to reactive oxygen species. Thus the enzymatic and interactive cellular machinery for histamine clearance is inherently present as a functional unit in two major human pulmonary cell types. It interacts with products of inflammatory host defense cells, and pulmonary endothelial-neutrophil interactions via this pathway may influence the progression of inflammation.

Metabolism and disposition of 14C-granisetron in rat, dog and man after intravenous and oral dosing.

1. The disposition and metabolic fate of 14C-granisetron, a novel 5-HT3 antagonist, was studied in rat, dog, and male human volunteers after intravenous and oral administration. 2. Complete absorption occurred from the gastrointestinal tract following oral dosing, but bioavailability was reduced by first-pass metabolism in all three species. 3. There were no sex-specific differences observed in radiometabolite patterns in rat or dog and there was no appreciable change in disposition with dose between 0.25 and 5 mg/kg in rat and 0.25 and 10 mg/kg in dog. Additionally, there were no large differences in disposition associated with route of administration in rat, dog and man. 4. In rat and dog, 35-41% of the dose was excreted in urine and 52-62% in faeces, via the bile. Metabolites were largely present as glucuronide and sulphate conjugates, together with numerous minor polar metabolites. In man, about 60% of dosed radioactivity was excreted in urine and 36% in faeces after both intravenous and oral dosing. Unchanged granisetron was only excreted in urine (5-25% of dose). 5. The major metabolites were isolated and identified by MS spectroscopy and nmr. In rat, the dominant routes of biotransformation after both intravenous and oral dosing were 5-hydroxylation and N1-demethylation, followed by the formation of conjugates which were the major metabolites in urine, bile and plasma. In dog and man the major metabolite was 7-hydroxy-granisetron, with lesser quantities of the 6,7-dihydrodiol and/or their conjugates.

An environmentally regulated receptor for diamine oxidase modulates human endothelial cell/fibroblast histamine degradative uptake.

Human vascular endothelial cells and fibroblasts express a cell-surface degradative pathway for the multifunctional mediator histamine, which employs a receptor for the metabolic enzyme diamine oxidase (DAO) and results in cellular accumulation of the final metabolite methylimidazoleacetic acid. We demonstrate recognition and regulatory properties of DAO receptors as a function of cellular environmental conditions. Fast and slow ligand binding receptor populations bind DAO at 4 degrees C maximally in 1 and 7 h, respectively; upon warming cells to 37 degrees C both populations participate in degradative uptake of histamine accumulated as methylimidazoleacetic acid. Bound DAO is displaced by heparin with 24-fold greater potency than dextran sulfate, implicating structural specificity of heparin-like glycosaminoglycan moieties as a critical factor in initial receptor/enzyme interaction at fast and slow sites. Receptor-bound DAO is retained under mildly acidic conditions characteristic of early to mid endocytic intracellular compartments and thus could recycle to the plasma membrane intact after internalization. DAO initially bound to receptors in whole cells is retained through cell disruption/membrane fractionation procedures, but DAO binds poorly to isolated membrane fractions or presolubilized receptors, suggesting that the geometry of DAO binding components is not readily maintained upon cell disruption unless DAO is already bound. Cells down-regulate their complement of DAO receptors upon prolonged exposure to DAO. In cells plated at high density, half of the bound DAO becomes nondisplaceable by heparin within 15 min at 37 degrees C, a time consistent with receptor internalization, whereas cells plated at low density retain all bound DAO in a heparin-sensitive state. The protein kinase C activator phorbol 12-myristate 13-acetate modulates DAO receptor number by 35% and total histamine degradative uptake by > 2-fold. Thus this pathway is subject to regulation at the levels of DAO receptor numbers, their state of cell-surface display, and additional cellular elements of the degradative pathway with which the DAO receptors interface.

Infant salmonellosis and vacuum cleaners.

Microbiological examination of the contents of vacuum cleaner bags collected from case and control homes demonstrated a statistically significant association (OR = 3.13, CL = 1.32-7.50) between infant salmonellosis cases and Salmonella contamination of the vacuums used in their homes. This suggests that some cases of infant salmonellosis may result from contact with contamination in the home environment and that steps taken to protect infants from potentially contaminated dust or dust aerosols may reduce the risk of contracting this infection.

The origin of non-outbreak Vibrio parahaemolyticus infections on Guam.

A study of Vibrio parahaemolyticus infected patients not associated with known outbreaks and controls matched for sex, ethnicity and age (+/- 5 years) was conducted on Guam. Cases were asked if they had eaten seafood within the 24-hour period preceding onset of illness and controls were asked if they had eaten seafood within the 24-hour period preceding their interview. Cases were more likely than controls to have eaten seafood (OR = 37.59, CL [8.30-220.24]).

The effect of selective serotonin re-uptake inhibitors on cytochrome P4502D6 (CYP2D6) activity in human liver microsomes.

Inhibition of human cytochrome P4502D6 (CYP2D6)-catalysed metabolism can lead to clinically significant alterations in pharmacokinetics. Since there is evidence that the selective serotonin reuptake inhibitor (SSRI) class of antidepressant drugs might inhibit CYP2D6, the effects of five SSRIs on human liver microsomal CYP2D6 activity were compared with each other and with three tricyclic antidepressant drugs. On a molar basis, paroxetine was the most potent of the SSRIs at inhibiting the CYP2D6-catalysed oxidation of sparteine (Ki = 0.15 microM), although fluoxetine (0.60 microM) and sertaline (0.70 microM) had Ki values in the same range. Fluvoxamine (8.2 microM) and citalopram (5.1 microM) also inhibited CYP2D6 activity. The major circulating metabolites of paroxetine in man produced negligible inhibition. In contrast, norfluoxetine the active metabolite of fluoxetine, was a potent CYP2D6 inhibitor (0.43 microM). CYP2D6 activity was also diminished by the tricyclic antidepressant drugs clomipramine (2.2 microM), desipramine (2.3 microM) and amitriptyline (4.0 microM). These findings suggest that compounds with SSRI activity are likely to interact with human CYP2D6 in vivo with the potential of causing drug interactions.

Are the endemic motor neuron diseases of Guam really disappearing?

Death certificates were reviewed to observe trends in the number of persons on Guam reported to have chronic degenerative motor neuron diseases (amyotrophic lateral sclerosis, parkinsonism-dementia or Parkinson's disease) at the time of death. Additional data, including age and race of the deceased and the name of the certifying physician were also collected. The number of persons having been diagnosed with amyotrophic lateral sclerosis is apparently decreasing while their mean age is increasing suggesting that the etiologic agents or factors causing this disease are less prevalent on Guam today than they have been in the past. Trends with regard to parkinsonism-dementia and Parkinson's disease are less clear.

The role of cytochrome P4502D6 in the metabolism of paroxetine by human liver microsomes.

Paroxetine is a selective serotonin reuptake inhibitor possessing anti-depressant activity. Demethylenation of the methylenedioxy phenyl group is the initial step in its metabolism, the liberated carbon appearing in vitro as formate. A radioassay involving [14C-methylenedioxy] paroxetine was developed and used to examine the role of cytochrome P4502D6 in paroxetine metabolism by human liver microsomes. The rate of formate production was much higher in microsomes from an extensive metaboliser of debrisoquine than from a poor metaboliser. Also, demethylenation of paroxetine was inhibited by the quinidine and quinine isomer pair in microsomes from the extensive metaboliser only. These observations strongly suggested that the process was catalysed by the enzyme cytochrome P4502D6. Metabolism could not be completely inhibited by quinidine, the residual activity representing the contribution of at least one other enzyme. The ability of microsomes from a poor metaboliser of debrisoquine to demethylenate paroxetine provided further evidence for the involvement of an enzyme distinct from P4502D6. This was confirmed by kinetic analysis of the process in microsomes from both poor and extensive metabolisers. It is concluded that, in man, the initial step of paroxetine metabolism is performed by at least two enzymes, one of which is cytochrome P4502D6.