Dhaval R. Kalaria

Non-invasive iontophoretic delivery of peptides and proteins across the skin

Introduction: Peptides and proteins are playing an increasingly important role in modern therapy. Their potency and specificity make them excellent therapeutic agents; however, their physicochemical properties and stability requirements almost invariably necessitate their administration by subcutaneous, intramuscular or intravenous injection. Controlled non-invasive administration using more patient-friendly advanced delivery technologies may combine the precision afforded by parenteral administration with improved compliance and the potential for individualized therapy. Areas covered: Transdermal iontophoresis enables hydrophilic charged molecules to be administered through the skin in an effective, non-invasive, patient-friendly manner. This review presents the basic concepts and an analysis of the effect of iontophoretic parameters and molecular properties on electrotransport rates across the skin along with a summary of experimental studies with peptides and proteins. The last section covers other techniques used in conjunction with iontophoresis. Expert opinion: It has long been known that iontophoresis can administer therapeutic amounts of biologically active peptides into the body. More recent studies have shown that it is also capable of delivering structurally intact, functional proteins non-invasively into and across intact human skin. The next step is to develop cost-effective and easy-to-use iontophoretic patch systems that ensure biomolecule stability, optimize delivery efficiency and address unmet therapeutic needs.

Comparative in vitro and in vivo evaluation of lipid based nanocarriers of Huperzine A.

The purpose of the present investigation was to explore feasibility of nanocarrier based transdermal delivery of Huperzine A (HupA) for the treatment of Alzheimers disease. For this investigation, microemulsion (ME), solid lipid nanoparticles (SLNs), and nanostructured lipid carriers (NLCs) were formulated and characterized for physicochemical parameters. The pseudo-ternary phase diagrams for microemulsion region were developed using generally recognized as safe (GRAS) excipients. The SLNs and NLCs were prepared by microemulsion template technique. These nanodispersions were formulated into gels for transdermal application and evaluated for various physicochemical parameters. In vitro permeation profiles in rat skin exhibited zero-order kinetics. HupA loaded ME exhibited superior permeation than NLCs followed by SLNs and cumulative amount permeated after 24h was found to be 147.68±9.42 μg/cm(2), 129.11±32.76 μg/cm(2) and 10.74±0.68 μg/cm(2), respectively. Furthermore, optimized gels were subjected to primary skin irritation testing over a period of 48 h and were found to be safe for skin application. In vivo efficacy tested in scopolamine induced amnesia model indicated significant improvement in cognitive function in mice group treated with developed nanocarrier based formulations as compared to the control group.

Controlled iontophoretic delivery of pramipexole: Electrotransport kinetics in vitro and in vivo

The objective of the study was to investigate the anodal iontophoretic delivery of pramipexole (PRAM), a dopamine agonist used for the treatment of Parkinsons disease, in order to determine whether therapeutic amounts of the drug could be delivered across the skin. Preliminary iontophoretic experiments were performed in vitro using porcine ear and human abdominal skin. These were followed by a pharmacokinetic study in male Wistar rats to determine the drug input rate in vivo. Stability studies revealed that after current application (0.5 mA/cm(2) for 6h), the solution concentration of PRAM was only 60.2 ± 5.3% of its initial value. However, inclusion of sodium metabisulfite (0.5%), an antioxidant, increased this to 97.2 ± 3.1%. Iontophoretic transport of PRAM across porcine skin in vitro was studied as a function of current density (0.15, 0.3, 0.5 mA/cm(2)) and concentration (10, 20, 40 mM). Increasing the current density from 0.15 to 0.3 and 0.5 mA/cm(2), resulted in 2.5- and 4-fold increases in cumulative permeation, from 309.5 ± 80.2 to 748.8 ± 148.1 and 1229.1 ± 138.6 μg/cm(2), respectively. Increasing the PRAM concentration in solution from 10 to 20 and 40 mM resulted in a 2-fold increase in cumulative permeation (816.4 ± 123.3, 1229.1 ± 138.6 and 1643.6 ± 201.3 μg/cm(2), respectively). Good linearity was observed between PRAM flux and both the applied current density (r(2)=0.98) and drug concentration in the formulation (r(2)=0.99). Co-iontophoresis of acetaminophen showed that electromigration was the dominant electrotransport mechanism (accounting for >80% of delivery) and that there was no inhibition of electroosmotic flow at any current density. Cumulative iontophoretic permeation across human and porcine skin (after 6h at 0.5 mA/cm(2)) was also shown to be statistically equivalent (1229.1 ± 138.6 and 1184.8 ± 236.4 μg/cm(2), respectively). High transport and delivery efficiencies were achieved for PRAM (up to 7% and 58%, respectively). The plasma concentration profiles obtained in the iontophoretic studies in vivo (20 mM PRAM; 0.5 mA/cm(2) for 5h) were modelled using constant and time-variant input models; the latter gave a superior quality fit. The drug input rate in vivo suggested that PRAM electrotransport rates would be sufficient for therapeutic delivery and the management of Parkinsonism.

Comparison of the cutaneous iontophoretic delivery of rasagiline and selegiline across porcine and human skin in vitro

The objective was to investigate the anodal iontophoresis of the MAO-B inhibitors rasagiline (RAS) and selegiline (SEL) across porcine and human skin in vitro. Passive delivery of RAS and SEL from aqueous solution was minimal; however, increasing current density from 0.1 to 0.3 and 0.5 mA/cm(2) produced a linear increase in steady-state iontophoretic flux (J(ss,RAS)=49.1i(d)+27.9 (r(2)=0.96) and J(ss,SEL)=27.8i(d)+25.8 (r(2)=0.98)). In the absence of background electrolyte, a four-fold change in donor concentration (10, 20 and 40 mM) did not produce a statistically significant increase in cumulative permeation of either drug after iontophoresis at 0.5mA/cm(2) for 7h. Co-iontophoresis of acetaminophen confirmed that electromigration was the dominant transport mechanism for both drugs (∼90%). Total iontophoretic delivery of RAS and SEL across porcine and human skin in vitro was statistically equivalent (RAS: 1512.7 ± 163.7 and 1523.6 ± 195.9 μg/cm(2), respectively, and SEL: 1268.7 ± 231.2 and 1298.3 ± 253.3 μg/cm(2), respectively). Transport efficiencies for RAS and SEL were good (ranged from 6.81 to 8.50 and 2.86 to 3.61%, respectively). Furthermore, the delivery efficiency, i.e., the fraction of the drug in the formulation that was delivered was very high (>56% at 0.5 mA/cm(2)). Cumulative permeation of RAS and SEL from carbopol gels, potential drug reservoirs for iontophoretic systems, was 891.5 ± 148.3 and 626.6 ± 162.4 μg/cm(2), respectively; this was less than from solution and was tentatively attributed to either different partitioning or slower drug diffusion in the gel matrix. The results demonstrated that therapeutic amounts of rasagiline and selegiline could be easily delivered by transdermal iontophoresis with simple gel patches of modest surface area.

Controlled iontophoretic transport of huperzine A across skin in vitro and in vivo: effect of delivery conditions and comparison of pharmacokinetic models.

The aim of this study was to investigate constant current anodal iontophoresis of Huperzine A (HupA) in vitro and in vivo and hence to evaluate the feasibility of using electrically assisted delivery to administer therapeutic amounts of the drug across the skin for the treatment of Alzheimers disease. Preliminary experiments were performed using porcine and human skin in vitro. Stability studies demonstrated that HupA was not degraded upon exposure to epidermis or dermis for 12 h and that it was also stable in the presence of an electric current (0.5 mA · cm(-2)). Passive permeation of HupA (2 mM) was minimal (1.1 ± 0.1 μg · cm(-2)); iontophoresis at 0.15, 0.3, and 0.5 mA · cm(-2) produced 106-, 134-, and 184-fold increases in its transport across the skin. Surprisingly, despite the use of a salt bridge to isolate the formulation compartment from the anodal chamber, which contained 133 mM NaCl, iontophoresis of HupA was shown to increase linearly with its concentration (1, 2, and 4 mM in 25 mM MES, pH 5.0) (r(2) = 0.99). This was attributed to the low ratio of drug to Cl¯ (in the skin and in the receiver compartment) which competed strongly to carry current, its depletion, and to possible competition from the zwitterionic MES. Co-iontophoresis of acetaminophen confirmed that electromigration was the dominant electrotransport mechanism. Total delivery across human and porcine skin was found to be statistically equivalent (243.2 ± 33.1 and 235.6 ± 13.7 μg · cm(-2), respectively). Although the transport efficiency was ∼ 1%, the iontophoretic delivery efficiency (i.e., the fraction of the drug load delivered) was extremely high, in the range of 46-81% depending on the current density. Cumulative permeation of HupA from a Carbopol gel formulation after iontophoresis for 6 h at 0.5 mA · cm(-2) was less than that from solution (135.3 ± 25.2 and 202.9 ± 5.2 μg · cm(-2), respectively) but sufficient for therapeutic delivery. Pharmacokinetic parameters were determined in male Wistar rats in vivo (4 mM HupA; 0.5 mA · cm(-2) for 5 h with Ag/AgCl electrodes) using two-compartment models with either constant or time-variant input rates. A superior fit was obtained using the time-variant model, and the input rate in vivo was significantly greater than that in vitro. Based on these results and the known pharmacokinetics, it was estimated that therapeutic amounts of HupA could be delivered for the treatment of Alzheimers disease using a reasonably sized patch.

Cutaneous iontophoretic delivery of CGP69669A, a sialyl Lewis(x) mimetic, in vitro.

Abstract:  The aim was to investigate the feasibility of using iontophoresis for the cutaneous delivery of the E‐selectin antagonist CGP69669A, a sialyl Lewisx‐glycomimetic with potential activity against inflammatory skin diseases. The effects of current density and formulation on iontophoretic transport were evaluated in porcine and human skin in vitro. Cumulative permeation of CGP69669A increased with current density (69.73 ± 9.51, 113.97 ± 26.80 and 160.44 ± 13.79 μg/cm2 at 0.1, 0.3 and 0.5 mA/cm2, respectively) and drug concentration (37.42 ± 13.13, 78.96 ± 23.13 and 160.44 ± 13.79 μg/cm2, at 1, 3 and 5 mg/ml, respectively). In contrast, passive delivery was negligible. Although permeation from a 2% hydroxyethyl cellulose gel was lower than that from aqueous solution, skin deposition – more relevant for the local treatment of dermatological conditions – was 3‐fold higher. The results demonstrated that although CGP69669A cannot be delivered passively into the skin it is an excellent candidate for transdermal iontophoresis, a technique that is ideally suited to the delivery of glycomimetics.

Development and validation of a HPAE-PAD method for the quantification of CGP69669A, a sialyl Lewisx mimetic, in skin permeation studies

A simple, rapid, precise and specific isocratic HPAE-PAD method for quantification of CGP69669A was developed and validated. CGP69669A is a glycomimetic of sialyl Lewis(x) and an antagonist of E-selectin with potential application in the treatment of inflammatory skin disease. Quantification was performed using a Dionex CarboPac(TM) PA-200 anion-exchange column (3 × 250 mm) with 100 mm NaOH solution as mobile phase, a flow rate of 0.50 mL/min and an injection volume of 10 μL. A quadruple potential waveform was used to detect the carbohydrate (+0.1 V from 0.00 to 0.40 s, -2.0 V from 0.41 to 0.42 s, +0.6 V at 0.43 s and -0.1 V from 0.44 to 0.50 s with current integrated between 0.20 and 0.40 s for detection) and rafinose was employed as an internal standard. The optimized conditions enabled rapid elution of CGP69669A (at 3.0 min) without interference from solvent peaks or substances present in the skin. The method showed good intra- and inter-day precision and accuracy and the response was linear from 1.0 to 25 µg/mL. This is the first validated direct method for the quantification of CGP69669A. It will now be employed in studies investigating the topical and transdermal delivery of CGP69669A in vitro and in vivo and it should also be of use for other applications of this molecule.

Simultaneous controlled iontophoretic delivery of pramipexole and rasagiline in vitro and in vivo: Transdermal polypharmacy to treat Parkinson’s disease

Graphical abstract Figure. No caption available. &NA; Effective treatment of Parkinsons disease (PD) involves administration of therapeutic agents with complementary mechanisms of action in order to replenish, sustain or substitute endogenous dopamine. The objective of this study was to investigate anodal co‐iontophoresis of pramipexole (PRAM; dopamine agonist) and rasagiline (RAS; MAO‐B inhibitor) in vitro and in vivo. Passive permeation of PRAM and RAS (20 mM each) across porcine skin after 6 h was 15.7 ± 1.9 and 16.0 ± 2.9 &mgr;g/cm2, respectively. Co‐iontophoresis at 0.15, 0.3 and 0.5 mA/cm2 resulted in statistically significant increases in delivery of PRAM and RAS; at 0.5 mA/cm2, cumulative permeation of PRAM and RAS was 613.5 ± 114.6 and 441.1 ± 169.2 &mgr;g/cm2, respectively – corresponding to 38‐ and 27‐fold increases over passive diffusion. Electromigration was the dominant mechanism for both molecules (>80%) and there was no effect on convective solvent flow. Statistically equivalent delivery was observed with human skin. The co‐iontophoretic system showed high delivery efficiency with 29% and 35% of the applied amounts of PRAM and RAS being delivered. Preliminary pharmacokinetics studies in rats confirmed that the input rate in vivo was such that therapeutic amounts of the two drugs could be co‐administered to humans by transdermal iontophoresis using reasonably sized patches and moderate current densities.

Iontophoretic Delivery of Glycomimetics-A New Approach for the Treatment of Inflammatory Skin Diseases

A key event in the inflammatory response is the recruitment of leukocytes from the microcirculation to the inflamed tissue through endothelial-dependent mechanisms that include leukocyte tethering and rolling, activation, firm adhesion and diapedesis to the interstitium. Leukocyte tethering and rolling along the endothelial surface is mediated by a family of carbohydrate-binding proteins (E-, P- and L-selectins). While L-selectin is constitutively expressed on the surface of leukocytes, expression of E- and P-selectin is upregulated on the endothelial surface during inflammation.