The Pulmonary Absorption of Aerosolized and Intratracheally Instilled rhG-CSF and monoPEGylated rhG-CSF

Purpose. The objective of this study was to highlight differences in the pulmonary absorption of a monoPEGylated rhG-CSF and rhG-CSF after intratracheal instillation and aerosol delivery. Methods. Male Sprague Dawley rats (250 g) were anesthetized and intratracheally instilled (IT) with protein solution or were endotracheally intubated and administered aerosol for 20 min via a Harvard small animal ventilator. A DeVilbiss Aerosonic nebulizer containing 5 ml of protein solution at 3 mg/ml was used to generate aerosol. The volume of protein solution deposited in the lung lobes was estimated to be 13 µl after delivery of Tc-99m HSA solutions. The PEGylated proteins consisted of a 6 kDa (P6) or 12 kDa PEG (PI2) linked to the N-terminus of rhG-CSF. rhG-CSF also was administered IT in buffers at pH 4 and pH 7 and in dosing volumes ranging from 100 to 400 µl. Blood samples were removed at intervals after dosing and the total white blood cell counts (WBC) were determined. Plasma was assayed for proteins by an enzyme immuno assay. Results. The plasma protein concentration v. time profiles were strikingly different for aerosol v. IT delivery. The C _max values for rhG-CSF and P12 after aerosol delivery were greater than found after IT (Aerosol: 598 ± 135 (ng/ml) rhG-CSF; 182 ± 14 P12 v. IT: 105 ± 12 rhG-CSF; 65.9 ± 5 P12). Similarly, T_max was reached much earlier after aerosol administration (Aerosol: 21.7 ± 4.8 (min) rhG-CSF; 168 ± 31 P12 v. IT: 100 ± 17 rhG-CSF; 310 ± 121 P12). Estimated bioavailabilities (F_lung %) were significantly greater via aerosol delivery than those obtained after IT (Aerosol: 66 ± 14 rhG-CSF; 12.3 ± 1.9 P12 v. IT: 11.9 ± 1.5 rhG-CSF; 1.6 ± 0.1 P12). An increase in circulating WBC counts was induced by all proteins delivered to the lungs. The rate and extent of absorption of rhG-CSF was not influenced by the pH employed nor the instilled volume. Conclusions. Estimates of bioavailability are dependent upon the technique employed to administer drug to the lungs. Aerosol administration provides a better estimate of the systemic absorption of macromolecules.     

Improved survival in young women with breast cancer

Background: Young age has been hypothesized to be an adverse prognostic factor for women with breast cancer. This association, based on historical data, may not reflect recent advances in breast cancer management. Methods: A retrospective study was conducted of all women age 30 or younger who underwent definitive operation at our institution for primary operable breast carcinoma during one of two consecutive 20-year periods (1950–1969 or 1970–1989). All cancers were restaged according to current staging criteria. Actuarial survival and recurrence-free survival rates from the two patient eras were compared with each other and with published statistics for older breast cancer patients. Results: Eligibility criteria were met by 81 women from the 1950–1969 era and 146 women from the 1970–1989 era. Histologic diagnoses, tumor sizes, incidence of axillary nodal metastases, number of positive nodes, and American Joint Committee on Cancer stage at presentation were similarly distributed in the two eras. Despite these similarities, improved survival (p=0.009) was observed in the later era. Local recurrences were also more common (p<0.05) in the later era in association with less extensive resections. These local recurrences had an adverse impact on recurrence-free survival in the later era, but no concomitant decrease in overall survival was observed. Node-positive patients who received chemotherapy demonstrated a trend toward improved survival (p=0.06) compared with node-positive patients who did not. Survival for patients in the later era was similar to that for older women as reported in other published series. Conclusions: The stage of presentation of breast cancer in women 30 years or younger appears unchanged from prior decades, but survival has improved in association with the use of less extensive surgical resections and the introduction of cytotoxic chemotherapy. With current treatment, primary operable breast cancer in young women appears to have a similar prognosis to breast cancer in older women.Results of this study were presented at the 47th Annual Cancer Symposium of The Society of Surgical Oncology, Houston, Texas, March 17–20, 1994, and was judged Best Clinical Paper in the Resident/Fellow Essay Contest.     

Embryo development, pregnancy and twin delivery after microinjection of 'stump' spermatozoa

Intracytoplasmic sperm injection was performed with immotile spermatozoa affected by tail 'stump' defect, and resulted in normal fertilization, embryo transfer and pregnancy in a 35-year-old female. The husband had a consanguineous ancestry. Two healthy babies, a male and a female, were born and this confirms that male infertility due to certain genetic sperm defects can be overcome by the intracytoplasmic sperm injection-assisted reproduction technique. The likely genetic origin of this sperm defect and the probability of the male offspring inheriting this sperm defect should be considered. The fertilization ability of stump spermatozoa, microinjected into the oocyte, is explained on the basis of experience from our previous research.     

Interactive technology applications for behavioral counseling: issues and opportunities for health care settings

CONTENT: This article discusses the rationale for, and the potential benefits and limitations of, computer-based interactive health communication (IHC) programs for health behavior counseling. We describe common barriers to health behavior counseling in medical settings and show how IHCs can address these issues. Following an overview of current and likely near-future IHCs, the potential impact of IHCs on the patient-provider relationship is considered. Results from evaluations of IHCs are summarized and important and unique issues in evaluating IHCs are discussed. We conclude with recommendations for clinical applications, including recommendations for consumers considering purchase or adoption of IHCs and recommendations for future research.     

Interactions of Phosphodiester and Phosphorothioate Oligonucleotides with Intestinal Epithelial Caco-2 Cells

Purpose. Oral bioavailability for antisense oligonucleotides has recently been reported but the mechanistic details are not known. The proposed oral delivery of nucleic acids will, therefore, require an understanding of the membrane binding interactions, cell uptake and transport of oligonucleotides across the human gastro-intestinal epithelium. In this initial study, we report on the cell-surface interactions of oligonucleotides with human intestinal cells. Methods. We have used the Caco-2 cell line as an in vitro model of the human intestinal epithelium to investigate the membrane binding interactions of 20-mer phosphodiester (PO) and phosphorothioate (PS) oligonucleotides. Results. The cellular association of both an internally [³H]-labelled and a 5end [³²P]-labelled PS oligonucleotide (3.0% at 0.4 µM extracellular concentration) was similar and was an order of magnitude greater than that of the 5end [³²P]-labelled PO oligonucleotide (0.2%) after 15 minutes incubation in these intestinal cells. The cellular association of PS was highly saturable with association being reduced to 0.9% at 5 µM whereas that of PO was less susceptible to competition (0.2% at 5 µM, 0.1% at 200 µM). Differential temperature-dependence was demonstrated; PS interactions were temperature-independent whereas the cellular association of PO decreased by 75% from 37°C to 17°C. Cell association of oligonucleotides was length and pH-dependent. A decrease in pH from 7.2 to 5.0 resulted in a 2- to 3-fold increase in cell-association for both backbone types. This enhanced association was not due to changes in lipophilicity as the octanol:aqueous buffer distribution coefficients remained constant over this pH range. The ability of NaCl washes to remove surface-bound PS oligonucleotides in a concentration-dependent manner suggests their binding may involve ionic interactions at the cell surface. Cell-surface washing with the proteolytic enzyme, Pronase^®, removed approximately 50% of the cell-associated oligonucleotide for both backbone types. Conclusions. Binding to surface proteins seems a major pathway for binding and internalization for both oligonucleotide chemistries and appear consistent with receptor (binding protein)-mediated endocytosis. Whether this binding protein-mediated entry of oligonucleotides can result in efficient transepithelial transport, however, requires further study.     

The Effect of Conformation on Membrane Permeability of an Acyloxyalkoxy-linked Cyclic Prodrug of a Model Hexapeptide

Purpose. To determine the different conformations of the acyloxyalkoxy-linked cyclic prodrug 1 of the model hexapeptide 2 in solution and to investigate the relationship between these solution conformations and the cellular permeability characteristics of this prodrug. Methods. Two-dimensional Homonuclear Hartmann-Hahn spectroscopy, Rotating-Frame Overhouser effect spectroscopy, circular dichroism and molecular dynamics simulations were used to find the solution conformers of cyclic prodrug 1. Results. Our spectroscopic findings suggest that cyclic prodrug 1 exhibits a major and a minor conformer in solution. The major conformer appears to have a well-defined secondary structure, which involves a -turn and 4 1 intramolecular hydrogen bond, creating a compact structure with a reduced average hydrodynamic radius compared to the model hexapeptide 2. Conclusions. The increased ability of cyclic prodrug 1 to permeate membranes compared to the model hexapeptide 2 could be due to reduction in the average hydrodynamic radius of the molecule facilitating paracellular flux and/or the reduction in the hydrogen bonding potential facilitating transcellular flux.     

Current Status and Future Prospects of Transdermal Drug Delivery

Pharmaceutical Research -     

Solubilization and Stabilization of a Benzylpenicillin Chemical Delivery System by 2-Hydroxypropyl-β-cyclodextrin

A dihydropyridine pyridinium salt redox carrier-based chemical delivery system for benzylpenicillin (1) was complexed with 2-hydroxypropyl--cyclodextrin (HPCD). The solubility of the lipophilic 1, which is incompatible with aqueous formulations, was dramatically increased and showed a linear dependency on the HPCD concentration. The degree of incorporation was 20 mg of 1 per g of complex. The stability study of 1 in various pH buffers indicated the base-catalyzed hydrolysis of the acyloxyalkyl linkage and the hydration of the 5,6 double bond of the dihydropyridine as the main degradation processes. The overall loss of 1, which follows first-order kinetics, was not influenced by changes in ionic strength and elimination of oxygen from the reaction medium. The HPCD complex of 1, which has a stability constant of 720–940 M ^–1, stabilized the chemical delivery system. The influence of the temperature on the stability of 1 is also discussed.     

Hybrid Pharmacokinetic Models to Describe Anti-HIV Nucleoside Brain Disposition Following Parent and Prodrug Administration in Mice

Brain delivery of active anti-HIV compounds is important for successful treatment of the AIDS patient. As an initial step in predicting human brain drug concentrations, hybrid pharmacokinetic models were developed to characterize the disposition of anti-HIV nucleosides following parent and prodrug administrations in mice. Mouse data were obtained following intravenous administration of 3-azido-2,3-dideoxyuridine (AZddU or AZDU), 3-azido-3-deoxythymidine (AZT), and their dihydropyridine prodrugs (AZddU-DHP and AZT-DHP). Exponential equations were fitted to the serum concentration–time data for each species, including the pyridinium ion moieties, and subsequently used in differential mass balance equations describing the brain dynamics of each compound. Model parameters for the mass balance equations were estimated by various techniques, including the utilization of in vitro data. In general, model-predicted brain concentrations agreed with the observed data. Similar data in larger animals will permit scale-up of the current model to predict human brain drug concentrations.     

Neutrophil chemotaxis in infants delivered by caesarean section

We evaluated polymorphonuclear leucocyte (PMN) chemotaxis and cortisol levels in cord blood from 15 healthy term infants delivered by caesarean section and from 15 healthy vaginally delivered term infants. Mean neutrophil chemotaxis was significantly higher in infants delivered by caesarean section (78.3±23.4m) than in vaginally delivered infants (57.8±16.6 m;P=0.01). Mean blood cortisol level was significantly lower in infants delivered by caesarean section (9.14±2.76 g/dl) than in infants born by vaginal delivery (20.71±6.98 g/dl;P=0.0001). No relationship was found between PMN chemotaxis and blood cortisol level. The higher neutrophil chemotactic activity observed in infants delivered by caesarean section could be related to general maternal anaesthesia.