Sustained Therapeutic Efficacy Research Articles

Adeno-Associated Viral Vectors at the Frontier between Tolerance and Immunity.

In recent years, the field of in vivo gene transfer with adeno-associated virus (AAV) vectors has seen an extraordinary expansion of applications and investments. Results emerging from clinical trials (1) and the recent market approval of a gene therapy drug for lipoprotein lipase deficiency (2) contributed to the hype around this vector system (3). Indeed, AAV vectors have several features that make them an ideal tool for gene transfer, for example, parental virions are replication deficient and non-pathogenic (4), and vectors can drive expression of a transgene for several years (5, 6) despite the fact that they do not integrate efficiently into the host genome. In recent years, a portfolio of natural AAV isolates (AAV serotypes) differing in tissue tropism has been developed as vectors. This toolbox has been further expanded with engineered AAV capsids developed to enhance efficiency and specificity of gene delivery, and to escape antibody neutralization (7). At the vector genome level, availability of potent promoter/enhancer sequences, codon-optimization of transgenes, and development of self-complementary AAV vectors (8) further enhanced efficacy of gene transfer. Finally, the availability of scalable processes to produce AAV vectors in GMP contributed significantly to the expansion of the field. As the AAV vector technology reached a more mature stage, it has become clear that a better understanding of the interactions of viral vectors with the host immune system is needed. In this Research Topic of Frontiers in Immunology, the editors present a collection of reviews and research articles discussing the two sides of immune responses triggered by in vivo gene transfer. These responses in fact can be desirable when they result in induction of tolerance to the therapeutic transgene (9), or when they are exploited for vaccine development, as discussed by Nieto and Salvetti in their review article (10). Conversely, immunogenicity of the viral capsid or the transgene product can be detrimental, as it may result in lack or loss of efficacy following vector-mediated gene transfer. Evidence for the critical role of tolerance induction in the achievement of sustained therapeutic efficacy following gene transfer comes from the work of Liao and colleagues, which provides evidence that glucocorticoid-induced TNF receptor (GITR) and its ligand GITR-L are of fundamental importance for the induction of immune regulatory responses in gene transfer and that lack of expression of GITR-L on antigen presenting cells results in impaired induction of regulatory T cells (Tregs) (11). Indeed, evidence of the key function of Tregs for successful in vivo gene therapy comes from several studies (12), and Liu and colleagues further demonstrate this concept in a model of plasmid gene transfer for hemophilia A, in which a combination of B cell depleting and Treg-enhancing drugs is used to successfully modulate transgene immunogenicity (13).

Preclinical evaluation of efficacy and safety of an improved lentiviral vector for the treatment of β-thalassemia and sickle cell disease.

A previously published clinical trial demonstrated the benefit of autologous CD34+ cells transduced with a self-inactivating lentiviral vector (HPV569) containing an engineered β-globin gene (βA-T87Q-globin) in a subject with β-thalassemia major. This vector has been modified to increase transduction efficacy without compromising safety. In vitro analyses indicated that the changes resulted in both increased vector titers (3 to 4 fold) and increased transduction efficacy (2 to 3 fold). An in vivo study in which 58 β-thalassemic mice were transplanted with vector- or mock-transduced syngenic bone marrow cells indicated sustained therapeutic efficacy. Secondary transplantations involving 108 recipients were performed to evaluate long-term safety. The six month study showed no hematological or biochemical toxicity. Integration site (IS) profile revealed an oligo/polyclonal hematopoietic reconstitution in the primary transplants and reduced clonality in secondary transplants. Tumor cells were detected in the secondary transplant mice in all treatment groups (including the control group), without statistical differences in the tumor incidence. Immunohistochemistry and quantitative PCR demonstrated that tumor cells were not derived from transduced donor cells. This comprehensive efficacy and safety data provided the basis for initiating two clinical trials with this second generation vector (BB305) in Europe and in the USA in patients with β-thalassemia major and sickle cell disease.

P047 - Sustained benefit of natalizumab in pediatric multiple sclerosis with breakthrough activity: A case report

Objective Limited data are available on natalizumab in pediatric multiple sclerosis. This is to report the sustained therapeutic benefit of natalizumab over 32 months follow up in a 12-year old child with a breakthrough activity disease. Methods In February 2009, the diagnosis of childhood onset relapsing remitting multiple sclerosis was confirmed at age 9, based on the “International Pediatric Multiple Sclerosis Study Group”. A disease breakthrough activity with suboptimal response was considered after 26 months of a well tolerated treatment with beta-interferon-1b; 6 relapses, rapid neurological deterioration and an increase by 4 new plaques on average per year with at least one showing enhancement after gadolinium injection were observed on MRI follow up. Monthly natalizumab 300 mg intravenous infusion was initiated on February 2012, after written consent. Negative JCV antibodies from serum samples were confirmed using the “STRATIFT JCV” test before treatment initiation and at 6 months intervals; clinical evaluation and biological tests were performed on routine base every 3 months. Results The patient's neurological status improved dramatically with rapid recovery after the second natalizumab infusion. Presently, this 13-year-old patient has received a total of 32 infusions of natalizumab and is still asymptomatic with no occurrence of any clinical relapse or no new lesion. Treatment is still well tolerated. Conclusion This report highlights the tolerability and sustained therapeutic efficacy of natalizumab as second-line therapy in childhood multiple sclerosis with breakthrough disease activity when critical decision of shifting from first-line therapy has to be made in children with potential greater risk in reaching progressive neurological disability at younger age.

Novel mechanisms of anti-tumor action for therapeutic T-bet gene-modified dendritic cells (P4228)

Abstract Clinical implementation of Type-1 polarized dendritic cell (DC) in vaccine formulations holds significant promise in the cancer therapy setting. We have recently demonstrated that DC engineered to express the T cell Type-1 transactivator protein T-bet (TBX21) promote robust Type-1 T cell-mediated immunity via a unique mechanism that is largely interleukin (IL)-12p70-independent. In the current study, we investigated the anti-tumor mechanisms of action associated with intratumoral administration of DC.Tbet developed from syngenic IL-12p35 -/-, IL-12p40 -/-, IFNγ -/- or β2m -/- mice. We observed that Type-1 anti-tumor immunity associated with slowed tumor growth was effectively developed in animals receiving DC.Tbet deficient in their ability to produce IL-12 family cytokines (i.e. IL-12, IL-23, IL-35) or IFN-γ. Surprisingly, we observed that the ability of DC.Tbet to present MHC class I/peptide complexes (i.e. lost in DC developed from β2m -/- mice) was not important for early priming of anti-tumor CD8+ T cells, but was essential for the sustained therapeutic efficacy of DC.Tbet (and treatment-induced anti-tumor CD8+ T cells) within the tumor microenvironment (TME). In order to best translate this cancer treatment modality into the clinic, we are currently evaluating how DC.Tbet conditions the TME for improved function of Ag-primed Type-1 T effector cells and how these APC collaborate with host DC to diversify the therapeutic T cell repertoire.

Thermosensitive β-cyclodextrin modified poly(ε-caprolactone)-poly(ethylene glycol)-poly(ε-caprolactone) micelles prolong the anti-inflammatory effect of indomethacin following local injection

A novel biodegradable and injectable in situ gel-forming controlled drug delivery system based on thermosensitive β-cyclodextrin-modified poly(ε-caprolactone)–poly(ethylene glycol)–poly(ε-caprolactone) co-polymer (PCEC–β-CD) was studied in this work. The drug encapsulating capacity has been improved by introducing β-CD bound to the PCEC co-polymer. The prepared PCEC–β-CD co-polymers self-assembled in water to form micelles, and underwent a temperature-dependent gel–sol transition, which was in the form of a flowing injectable solution at low temperatures but became a non-flowing gel at around physiological body temperature. Furthermore, a small hydrophobic drug molecule indomethacin (IND) was successfully encapsulated in PCEC–β-CD micelles by dialysis at a high encapsulation efficiency and drug loading capacity. The IND-loaded micelles (IND-M) exhibited controlled release in vitro. Additionally, a pharmacodynamic study in vivo based on both the carrageenan-induced acute and complete Freund’s adjuvant-induced adjuvant arthritis models indicated that sustained therapeutic efficacy could be achieved through subcutaneous injection of IND-loaded micelles. A significant improvement in the anti-inflammatory effect of IND in rats occurred on encapsulation in PCEC–β-CD micelles.

Abstract 9178: Cell-Based Prostacyclin Gene Therapy Prevents Pulmonary Arterial Hypertension

Objectives: Intravenous prostacyclin is approved for the treatment of pulmonary arterial hypertension (PAH). However, it has limitations of a short half-life, the permanent placement of an intravenous catheter and substantial annual cost. We hypothesized that cell-based prostacyclin gene therapy may provide sustained therapeutic effects without the aforementioned limitations. Methods and Results: Since prostacyclin is produced from arachidonic acid by the enzymes cyclooxygenase (COX) and prostacyclin synthase (PGIS), we generated a vector expressing COX isoform 1 (COX1) and PGIS fusion protein that is capable of producing prostacyclin from arachidonic acid. Endothelium-like progenitor cells (ELPCs) from syngenetic Fisher 344 rats were transfected with COX1-PGIS expressing vector and lentivirus expressing nuclear localized red fluorescent protein (nuRFP). The engineered ELPCs expressing COX1-PGIS and nuRFP were tested in a monocrotaline (MCT) induced rat PAH model. Twenty-five days after cell transplantation, 48% of transplanted ELPCs resided in the lungs. MCT induced an increase in right ventricular systolic pressure (RVSP) compared to normal controls (51.9 ±2.2 mmHg vs. 26.3 ± 1.5 mmHg, p<0.01). The increase in RVSP was significantly attenuated by treatment with ELPCs alone (40.9 ±1.6 mmHg, p<0.01) or with ELPCs engineered to express the COX1-PGIS transgene (35.9 ±1.6 mmHg, p<0.01). Engineered ELPCs were significantly more effective than ELPCs alone in preventing the increase in RVSP (p<0.05). Right ventricle hypertrophy (right ventricle vs. left ventricle plus septum weight ratio) was reduced by treatment with ELPCs (28.8±0.8% vs. 36.5±3.1%, p <0.01) or engineered ELPCs (30.5±0.9% vs. 36.5±3.1%, p <0.01). Lung vessel thickness index increase (reflecting intima thickening and medial smooth muscle hypertrophy) was attenuated by treatment with engineered ELPCs (78.1±3.1% vs. 90.1±4.1%, p <0.05). Conclusions: COX1-PGIS expressing ELPCs prevented MCT-induced PAH and showed enhanced efficacy over ELPCs alone. The one time cell delivery offered sustained therapeutic efficacy for at least 25 days, which may provide a promising option for patients with PAH.

Lentiviral gene therapy of murine hematopoietic stem cells ameliorates the Pompe disease phenotype

AbstractPompe disease (acid α-glucosidase deficiency) is a lysosomal glycogen storage disorder characterized in its most severe early-onset form by rapidly progressive muscle weakness and mortality within the first year of life due to cardiac and respiratory failure. Enzyme replacement therapy prolongs the life of affected infants and supports the condition of older children and adults but entails lifelong treatment and can be counteracted by immune responses to the recombinant enzyme. We have explored the potential of lentiviral vector–mediated expression of human acid α-glucosidase in hematopoietic stem cells (HSCs) in a Pompe mouse model. After mild conditioning, transplantation of genetically engineered HSCs resulted in stable chimerism of approximately 35% hematopoietic cells that overexpress acid α-glucosidase and in major clearance of glycogen in heart, diaphragm, spleen, and liver. Cardiac remodeling was reversed, and respiratory function, skeletal muscle strength, and motor performance improved. Overexpression of acid α-glucosidase did not affect overall hematopoietic cell function and led to immune tolerance as shown by challenge with the human recombinant protein. On the basis of the prominent and sustained therapeutic efficacy without adverse events in mice we conclude that ex vivo HSC gene therapy is a treatment option worthwhile to pursue.

Development of a macromolecular prodrug for the treatment of inflammatory arthritis: mechanisms involved in arthrotropism and sustained therapeutic efficacy

IntroductionThe purpose of the present manuscript is to test the hypothesis that arthrotropic localization and synovial cell internalization account for the unique capacity of N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer-dexamethasone conjugate (P-Dex, a macromolecular prodrug of dexamethasone) to induce sustained amelioration of joint inflammation and inhibition of tissue damage in an animal model of inflammatory arthritis.MethodsRats with adjuvant-induced arthritis (AA) were treated with P-Dex, free dexamethasone, saline or HPMA homopolymer. To define the biodistribution of P-Dex, conjugates with different imaging labels were given to AA rats and analyzed. Isolated joint tissues were evaluated by fluorescence-activated cell sorting (FACS) and immunohistochemical staining. Cellular uptake of P-Dex and its effects on apoptosis and production of proinflammatory cytokines were examined using human monocyte-macrophages and fibroblasts.ResultsA single systemic administration of P-Dex completely suppressed AA for >20 days. Magnetic resonance imaging demonstrated higher HPMA copolymer influx into the inflamed joints than the normal joints. Immunohistochemistry and FACS analyses of arthritic joints revealed extensive uptake of the polymer conjugate by synovial fibroblasts and myeloid lineage cells. The capacity of P-Dex to suppress inflammation was confirmed in monocyte-macrophage cultures in which P-Dex treatment resulted in suppression of lipopolysaccharide-induced IL-6 and TNFα release. Similarly, TNFα-induced expression of matrix metalloproteinases (MMP1 and MMP3) in synovial fibroblasts from a rheumatoid arthritis patient was suppressed by P-Dex. P-Dex showed no detectable effect on monocyte apoptosis.ConclusionsP-Dex provides superior and sustained amelioration of AA compared with an equivalent dose of free dexamethasone. The arthrotropism and local retention of P-Dex is attributed to the enhanced vascular permeability in arthritic joints and the internalization of P-Dex by synovial cells. The uptake and processing of P-Dex by macrophages and fibroblasts, and downregulation of proinflammatory mediators, provides an explanation for the sustained anti-inflammatory efficacy of P-Dex in this model of inflammatory arthritis.

Long-Term Durability of Percutaneous Tibial Nerve Stimulation for the Treatment of Overactive Bladder

The Overactive Bladder Innovative Therapy Trial during phase 1 was a randomized trial demonstrating comparable effectiveness of percutaneous tibial nerve stimulation and extended-release tolterodine during 12 weeks of therapy for frequency, nocturia, urgency, voided volume and urge incontinence episodes. In this second phase of the Overactive Bladder Innovative Therapy Trial we assessed the sustained therapeutic efficacy of percutaneous tibial nerve stimulation in subjects with overactive bladder during 1 year. After 12 weeks subjects randomized to weekly percutaneous tibial nerve stimulation with Urgent((R)) PC were offered an additional 9 months of treatment with assessments at 6 and 12 months from baseline. Outcome measures included voiding diary data, overactive bladder questionnaires, global response assessments and safety assessments. A total of 33 percutaneous tibial nerve stimulation responders continued therapy with 32 and 25 subjects completing 6 and 12 months of therapy, respectively. Subjects received a mean of 12.1 treatments during an average of 263 days, with a mean of 21 days (median 17) between treatments. Subject global response assessments showed sustained improvement from 12 weeks at 6 and 12 months, with 94% and 96% of responders, respectively. At 12 months mean improvements from baseline included a frequency of 2.8 voids daily (p <0.001), urge incontinence of 1.6 episodes daily (p <0.001), nocturia with 0.8 voids (p <0.05) and a voided volume of 39 cc (p <0.05). Overactive bladder questionnaire symptom severity was significantly improved from 12 weeks to 12 months (p <0.01) as well as from 6 to 12 months (p <0.01). No serious adverse events occurred. Statistically significant overactive bladder symptom improvement achieved with 12 weekly percutaneous tibial nerve stimulation treatments demonstrates excellent durability through 12 months. The durability of response demonstrates the effectiveness of percutaneous tibial nerve stimulation as a viable, long-term therapy for overactive bladder.

Active immunization to tumor necrosis factor-α is effective in treating chronic established inflammatory disease: a long-term study in a transgenic model of arthritis

IntroductionPassive blockade of tumor necrosis factor-alpha (TNF-α) has demonstrated high therapeutic efficiency in chronic inflammatory diseases, such as rheumatoid arthritis, although some concerns remain such as occurrence of resistance and high cost. These limitations prompted investigations of an alternative strategy to target TNF-α. This study sought to demonstrate a long-lasting therapeutic effect on established arthritis of an active immunotherapy to human (h) TNF-α and to evaluate the long-term consequences of an endogenous anti-TNF-α response.MethodshTNF-α transgenic mice, which spontaneously develop arthritides from 8 weeks of age, were immunized with a heterocomplex (TNF kinoid, or TNF-K) composed of hTNF-α and keyhole limpet hemocyanin after disease onset. We evaluated arthritides by clinical and histological assessment, and titers of neutralizing anti-hTNF-α antibody by enzyme-linked immunosorbent assay and L929 assay.ResultsArthritides were dramatically improved compared to control mice at week 27. TNF-K-treated mice exhibited high levels of neutralizing anti-hTNF-α antibodies. Between weeks 27 and 45, all immunized mice exhibited symptoms of clinical deterioration and a parallel decrease in anti-hTNF-α neutralizing antibodies. A maintenance dose of TNF-K reversed the clinical deterioration and increased the anti-hTNF-α antibody titer. At 45 weeks, TNF-K long-term efficacy was confirmed by low clinical and mild histological scores for the TNF-K-treated mice. Injections of unmodified hTNF-α did not induce a recall response to hTNF-α in TNF-K immunized mice.ConclusionsAnti-TNF-α immunotherapy with TNF-K has a sustained but reversible therapeutic efficacy in an established disease model, supporting the potential suitability of this approach in treating human disease.

Nonvirally modified autologous primary hepatocytes correct diabetes and prevent target organ injury in a large preclinical model.

BackgroundCurrent gene- and cell-based therapies have significant limitations which impede widespread clinical application. Taking diabetes mellitus as a paradigm, we have sought to overcome these limitations by ex vivo electrotransfer of a nonviral insulin expression vector into primary hepatocytes followed by immediate autologous reimplantation in a preclinical model of diabetes.Methods and ResultsIn a single 3-hour procedure, hepatocytes were isolated from a surgically resected liver wedge, electroporated with an insulin expression plasmid ex vivo and reimplanted intraparenchymally under ultrasonic guidance into the liver in each of 10 streptozotocin-induced diabetic Yorkshire pigs. The vector was comprised of a bifunctional, glucose-responsive promoter linked to human insulin cDNA. Ambient glucose concentrations appropriately altered human insulin mRNA expression and C-peptide secretion within minutes in vitro and in vivo. Treated swine showed correction of hyperglycemia, glucose intolerance, dyslipidemia and other metabolic abnormalities for ≥47 weeks. Metabolic correction correlated significantly with the number of hepatocytes implanted. Importantly, we observed no hypoglycemia even under fasting conditions. Direct intrahepatic implantation of hepatocytes did not alter biochemical indices of liver function or induce abnormal hepatic lobular architecture. About 70% of implanted hepatocytes functionally engrafted, appeared histologically normal, retained vector DNA and expressed human insulin for ≥47 weeks. Based on structural tissue analyses and transcriptome data, we showed that early correction of diabetes attenuated and even prevented pathological changes in the eye, kidney, liver and aorta.ConclusionsWe demonstrate that autologous hepatocytes can be efficiently, simply and safely modified by electroporation of a nonviral vector to express, process and secrete insulin durably. This strategy, which achieved significant and sustained therapeutic efficacy in a large preclinical model without adverse effects, warrants consideration for clinical development especially as it could have broader future applications for the treatment of other acquired and inherited diseases for which systemic reconstitution of a specific protein deficiency is critical.

Physicochemical characterization, in vitro, and in vivo evaluation of indomethacin‐loaded nanocarriers self‐assembled by amphiphilic polyphosphazene

Copolymeric nanocarriers assembled by amphiphilic polyphosphazene bearing poly(N-isopropylacrylamide) (PNIPAAm) and ethyl glycinate (EtGly) as substitutes, were investigated as drug vehicles for indomethacin (IND). The physicochemical characteristics of the novel nanocontainers were studied, including lower critical solution temperature (LCST), critical micelle concentration (CMC) and drug loading capacity. LCST measurements revealed that copolymer is more sensitive to the introduction of salts into aqueous solution compared with homopolymer. A significant decrease in CMC was observed when the temperature increased above LCST. As evidenced by transmission electron microscopy (TEM) measurement, morphological transformation from multicompartment into spherical nanoparticles was observed when nanocarriers with higher IND content were concerned. In vitro release tests suggested that IND-loaded nanocontainers exhibited pH dependent release profiles. In vivo pharmacokinetic study after subcutaneous administration provided a relatively sustained release behavior. Additionally, compared with free drug solution at the same dose, IND concentration in rat plasma showed a prolonged retention in experimental group treated with IND-loaded micelles. In vivo pharmacodynamic study based on both carrageenan-induced acute and complete Freund's adjuvant (CFA) induced adjuvant-arthritis models indicated that sustained therapeutic efficacy could be achieved through intraarticular injection of IND-loaded micelles. Most importantly, local delivery of IND can avoid the severe gastrointestinal stimulation, which is frequently associated with oral administration.

Local Delivery of Indomethacin to Arthritis-Bearing Rats through Polymeric Micelles Based on Amphiphilic Polyphosphazenes

Preparation, in vitro and in vivo evaluation of indomethacin-loaded polymeric micelles based on amphiphilic polyphosphazene. Amphiphilic polyphosphazenes (PNIPAAm/EAB-PPPs) with poly (N-isopropylacrylamide) (PNIPAAm) and ethyl 4-aminobenzoate (EAB) as side groups were synthesized through thermal ring-opening polymerization and subsequent substitution reactions. Indomethacin (IND) loaded polymeric micelles based on PNIPAAm/EAB-PPPs were prepared by dialysis procedure. In vitro IND release kinetics was investigated in 0.1 M PBS (pH 7.4), while in vivo pharmacokinetics was performed in Sprague-Dawley rats. In vivo pharmacodynamic study was carried out based on two animal models, i.e. carrageenan-induced acute paw edema and complete Freund's adjuvant (CFA) induced ankle arthritis model. Drug loading capacity of micelles based on this type of amphiphilic copolymers was mainly determined by copolymer composition and the chemical structure of drug. In addition to the compatibility between drug and micellar core, hydrogen bonding interaction between drug and hydrophilic corona may significantly influence drug loading as well. In vitro drug release in PBS suggested that there was no significant difference in release rate between micelles based on copolymers with various EAB content. Compared with the rats administered with free IND aqueous solution, IND concentration in rats' plasma showed a prolonged maintenance in experimental group treated with IND-loaded polymeric micelles. In vivo pharmacodynamic study indicated that sustained therapeutic efficacy could be achieved through topical injection of the aqueous solution of IND-loaded micelles. Local delivery of IND can avoid the severe gastrointestinal stimulation, which was frequently associated with oral administration as evidenced by ulceration evaluation. The promising results of current preliminary study suggest that this type of amphiphilic copolymers could be used as injectable drug carriers for hydrophobic drugs.

Indomethacin-loaded polymeric nanocarriers based on amphiphilic polyphosphazenes with poly ( N-isopropylacrylamide) and ethyl tryptophan as side groups: Preparation, in vitro and in vivo evaluation

The effects of copolymer composition, drug structure and initial drug feed on drug loading of polymeric micelles based on amphiphilic polyphosphazenes were investigated. It was found that the drug loading capacity of micelles based on this type of amphiphilic copolymers was mainly determined by copolymer composition and the chemical structure of drug. In addition to the compatibility between drug and micellar core, hydrogen bonding interaction between drug and hydrophilic corona may significantly influence drug loading as well. In vitro drug release in 0.1 M PBS (pH 7.4) suggested that indomethacin (IND) in the micelles was released through Fickian diffusion, and no significant difference in release rate was observed for micelles based on copolymers with various EtTrp content. Compared with in vitro IND release profile, in vivo pharmacokinetic study after subcutaneous administration provides a more sustained release behavior. Additionally, in comparison with free drug solution at the same dose, IND concentration in rat plasma showed a prolonged retention when the drug was delivered through polymeric micelles. In vivo pharmacodynamic study based on both carrageenan-induced acute and complete Freund's adjuvant-induced adjuvant arthritis model indicated that sustained therapeutic efficacy could be achieved through intraarticular injection of IND-loaded micelles. Most importantly, local delivery of IND can avoid the severe gastrointestinal stimulation, which was frequently associated with oral administration.

Susceptibility of nanoparticle-encapsulated paclitaxel to P-glycoprotein-mediated drug efflux

Overexpression of P-glycoprotein (P-gp) is a key factor contributing to the development of multidrug resistance (MDR) in cancer cells. The objective of the study is to investigate whether a P-gp substrate, paclitaxel, delivered to MDR tumor cells in poly( d, l-lactide- co-glycolide) (PLGA) nanoparticles is susceptible to P-gp – mediated drug efflux. Paclitaxel-loaded nanoparticles were formulated by emulsion-solvent evaporation technique. Nanoparticles had a mean hydrodynamic diameter of about 195 nm, and demonstrated sustained release of paclitaxel. In vitro cell culture studies indicated that paclitaxel nanoparticles result in sustained, dose-dependent and significant cytotoxicity in drug-sensitive MCF-7 tumor cells but not in drug-resistant NCI-ADR/RES cells. Resistance to nanoparticle-encapsulated paclitaxel was reversed by verapamil, a P-gp inhibitor. Further, sustained inhibition of P-gp was necessary for sustaining the cytotoxicity of nanoparticle-encapsulated paclitaxel in drug-resistant cells. Inhibition of P-gp by verapamil did not significantly affect the uptake or retention of nanoparticles in drug-resistant cells. In conclusion, our studies suggest that P-gp substrates, such as paclitaxel, delivered to MDR cells by PLGA nanoparticles, are susceptible to efflux by P-gp. Inhibition of P-gp restores sensitivity to paclitaxel; however, sustained inhibition of P-gp is required for sustained therapeutic efficacy of nanoparticle-encapsulated drug.

Long-term drug treatment of benign prostatic hyperplasia — results of a prospective 3-year multicenter study using Sabal extract IDS 89

A 3-year prospective multicenter study on Sabal extract IDS 89 was carried out to evaluate its potential and limitations as phytotherapy in the treatment of benign prostatic hyperplasia (BPH). The results obtained from 435 patients confirm its sustained therapeutic efficacy. A marked symptomatic improvement, that included a 50% reduction in residual urine and a 6.1 ml/sec increase in peak urinary flow rate due to therapy was observed. Both physicians and patients voted its efficacy as good or very good in over 80% of the cases, and the drug was well tolerated by 98% of the patients. The clinical status of BPH and thus the quality of life of 4 out of 5 patients was markedly improved by long-term IDS 89 therapy. Furthermore, the deterioration rate at the end of the 3-year treatment period was significantly lower than in untreated BPH subjects. It is therefore concluded that long-term IDS 89 therapy can also reduce the incidence of surgery.

Sustained therapeutic efficacy and safety of oral propafenone for treatment of chronic ventricular arrhythmias: A 2-year experience

Thirty-two men with chronic ventricular arrhythmias responded to propafenone, a new potent antiarrhythmic agent, in short-term trials with 85% or greater reduction of total ventricular premature complexes (VPCs) per hour, 95% or greater reduction of ventricular couplets (VCs) per hour, and 100% abolition of ventricular tachycardia (VT) beats per 24 hours. These patients were continued on long-term propafenone therapy to assess sustained therapeutic efficacy and safety. Thirty patients completed 1 year and 26 patients completed 2 years of testing with this agent; one patient died of sudden death and another died of a noncardiac cause. Although there were significantly fewer patient responders at 1 and 2 years, the majority of patients (>79%) continued to respond optimally to propafenone. Side effects were minor and included bitter taste, dizziness, congestive heart failure, fatigue, and significant prolongation of the PR and QRS intervals. Propafenone has sustained antiarrhythmic efficacy after 2 years without serious toxicity.

Pyrimethamine sustained release systems based on bioresorbable polyesters for chemoprophylaxis of rodent malaria

Pyrimethamine-containing microspheres and implants were prepared, from various bioresorbable polymers and copolymers derived from lactic acid enantiomers and from glycolic acid. Release patterns of the temporarily entrapped drug were examined in vitro for the various systems. The efficacy of these polymer-drug systems against Plasmodium berghei was also investigated using mouse as the model animal. Both in vitro sustained release and therapeutic efficacy depend on the drug content, the respective amounts of l- and d-repeating units in lactic acid stereocopolymers and the composition in lactic and glycolic repeating units in lactic/glycolic copolymers. For some of the pyrimethamine-containing implants, protection over a period as long as 6 months has been observed which is consistent with long-term chemoprophylaxis of rodent malaria.

H-1 Blockers—Classical Antihistamines

The H1-blocking antihistamines pharmacology, pharmacokinetics, uses, toxicity and special precautions are presented. Newer agents may offer advantages of longer half-life and lower levels of sedation and motor impairment. Therapeutic considerations are listed with attention to sustained therapeutic efficacy.