TRANSFERSOMES

RAJESH.T PhD SCHOLAR NIPER-HYD

INTRODUCTION
The concept of transfersome was introduced in 1991 by Cevc and coworkers. Transfersome is a term registered as a trademark by the German company IDEA AG, and used by it to refer to its proprietary drug delivery technology. The name means carrying body, and is derived from the Latin word transferre, meaning to carry across, and the Greek word soma, for a body.

Definition

A transfersome carrier is an articial vesicle designed to be like a cell vesicle or a cell engaged in exocytosis, and thus suitable for controlled and, potentially targeted, drug delivery. Transfersomes are highly efficient edge activator (EA)based ultraflexible vesicles capable of, noninvasively, trespassing skin by virtue of their high, self-optimizing deformability. Transfersomes are self-regulating, mixed lipid aggregates containing edge activators within a phospholipid matrix so as to drastically reduce the value of its elastic module. Transfersomes undergo a series of stress-dependant adjustments of the local carrier composition, to minimize the resistance to their motion through the otherwise

ADVANTAGES

Transfersomes are biocompatible, biodegradable and are capable of protecting the encapsulated drug from metabolic degradation. They are also able to transport therapeutic agents through very narrow pathways between most cells in the skin (5 to 10 times narrower than their own diameter) without signicant loss. The application of transfersomes does not involve any elaborate procedure and they can be applied by a non-occluded method, where by they pass through the multilayered lipid matrix of the stratum corneum as a result of the hydration or osmotic force within the skin. In addition to a non-invasive means of drug delivery, they also offer a novel approach for investigating skin histology. Delivery of proteins and nutraceuticals. Ease of scale up compared to other vesicular systems.

LIMITATIONS

Chemically unstable because of their predisposition to oxidative degradation. The lack of purity of natural phospholipids is another criterion that inuences acceptance of transfersomes as drug delivery systems.

COMPOSITION
CLASS % EXAMPLES USE PHOSPOLIPID 60-80 S Soya phosphatidyl choline, Vesicle forming Dipalmitoylphosphatidyl Components. choline, N-[1-(2,3-dioleyloxy)-propyl]N,N,N trimethylammonium chloride (DOTMA). Bile salts, Polysorbates, Glycolipids, Alkyl or acyl Polyethoxylenes and Dipotassium glycyrrhizinate. Saline pH(6.4). phosphate Responsible for the ultra-adaptive properties of transfersomes.

EDGE ACTIVATORS

10-25

SOLVENTS BUFFERING AGENTS DYES

3-10 Q.S Q.S

Ethanol, methanol, chloroform. As a solvent buffer As a hydrating medium . For CSLM study.

Rhodamine123, Rhodamine DHPE, Flouroesceins OHPE, Nile red.

Regulatory aspects of excipients used in transfersomes

Selection of an excipient during the preparation of a transfersome based formulation is restricted by the safety and toxicity concerns associated with these excipients. Regulatory agencies like USFDA, WHO, ICH, JMHW and IPEC have maintained a list of excipients classied as Generally Regarded as Safe (GRAS), which have been clinically categorized not to cause any toxicity. The FDA keeps a record entitled Inactive Ingredient Guide, which includes a catalogue of approved excipients. Soya phosphatidylcholine (SPC) is a GRAS listed phospholipid and also complies with specications of the Food Chemicals Codex. (http://www.NutriScienceUSA.com). Similarly, Sodium cholate (SC,) which is used as an EA, is reported to be nontoxic but has been kept in the hazardous category as it causes skin and eye irritations as well as respiratory sensitization. Surfactants can cause severe gastrointestinal discomfort when

Mechanism of permeation by transfersomes

Investigations indicate that the transport of transfersomes across the skin involves two methods. Depends on the affinity of the therapeutic agent towards water. Structure of transfersomes

The hydration or osmotic force has been reported to be involved in the transport of transfersomes across the stratum corneum. The ability of the skin to control water loss aids in establishing a difference in the water movement between the accessible part of the epidermis (75% water content) and the almost completely dry stratum corneum (15% water content), leading to formation of the water activity gradient.
Transfersomes are believed to be drawn into the body by hydrotaxis. Elastomechanics is considered to have a signicant role in the movement of transfersomes through the normally conning pores. It also offers a rationale for the development of ultradeformable vesicles as drug carriers. Drying and partial dehydration of vesicles are thought to be the initial events in skin permeation by the transfersome upon topical administration. As a consequence, the transfersome becomes compressed or curved. The phospholipid component of the transfersome has a tendency to evade a dry environment. Thus, to stay fully swollen, the transfersome follows the local hydration gradient and penetrates more strongly the hydrated layers of skin, reaching the epidermis and dermis.

The decrease in ux observed upon wetting of the skin establishes the hydration theory. Hydrotaxis of these ultradeformable systems enables them to carry in excess of 50% of the administered therapeutic agent through the skin. Transfersomes can penetrate the intact stratum corneum spontaneously along two routes in the intracellular lipid so as to differ in their bilayers properties. When a transfersome reaches a pore, it is able to change its membrane composition reversibly as a result of its self-optimizing deformability. To pass through the pore, components of the transfersome responsible for its deformability start accumulating at the site of stress, whereas the less exible components undergo dilution, which signicantly reduces the active rate of membrane deformation and allows the highly exible particles to pass through the pores. The passage of transfersomes through the skin and the epithelial barrier is greatly inuenced by the exibility of their membrane, which can be achieved using an appropriate ratio of surfactants. The resulting exibility of the transfersomal membrane also reduces the possibility of its rupture in the skin.

Methods of preparation
ROTARY SHAKING

HAND SHAKING

PREPARATION

SONICATION

FREEZE THAW METHOD

Rotary evaporationsonication method

This method involves dissolution of phosphatidylcholine and EA in a blend of chloroform and methanol (2:1, v/v), followed by the removal of organic solvent using rotary evaporation under reduced pressure at 40 C. The lm deposited is hydrated with a solution of the therapeutic agent in a suitable aqueous phase while rotating the ask for one hour at room temperature. The vesicles produced are left to swell for 2 hours at room temperature, followed by 30 min of sonication in a bath sonicator so as to decrease their volume. Extrusion of vesicles then occurs through a sandwich of 220 and 450 nm polycarbonate membranes, with the resulting vesicles being stored at 40 C.

Vortexing-sonication method
In the vortexing-sonication method, mixed lipids (i.e. phosphatidylcholine, EA and the therapeutic agent) are blended in a phosphate buffer and vortexed to attain a milky suspension. The suspension is sonicated, followed by extrusion through polycarbonate membranes. Cationic transfersomes have also been prepared by this method, which involves mixing cationic lipids, such as DOTMA, with PBS to obtain a concentration of 10 mg/ml followed by the addition of sodium deoxycholate (SDC). The blend is vortexed and sonicated, followed by extrusion through a polycarbonate (100-nm)

Characterization of transfersomes
Vesicle size and distribution Charge on the vesicle Entrapment efficiency Membrane flexibility is generally observed by measuring the deformation of unilamellar vesicles near an adhesive lipid monolayer at the airwater interface using ellipsometry. Elasticity and/or deformability index estimated by extrusion measurement, which involves extrusion of vesicles through a 3050-nm polycarbonate filter at 2.5 bar for 10 min. The time involved in extrusion of 10 ml of the transfersomal formulation is taken as an index of its elasticity and is expressed in terms of a deformability index. Skin penetration study.

MOLECULES

ADVANTAGES

PROTIENS

The bioavaibility obtained from transfersomes is somewhat similar to that resulting from subcutaneous injection of the same protein suspension. Human serum albumin or gap junction protein was found to be effective in producing the immune response when delivered by transdermal route encapsulated in transfersomes. Heparin, Retinol, and Melatonin were also investigated using transfersomes and produced promising results.
Transfersomes containing proteins like integral membrane protein, human serum albumin, gap junction protein are used for this purpose. Advantages of this approach are injecting the protein can be avoided and higher IgA levels are attained. Transcutaneous hepatitis-B vaccine has given good results.

VACCINES

RETROVIRALS

12 times higher AUC was obtained for zidovudine as compared to normal control administration. Selectivity in deposition in RES (which is the usual site for residence of HIV) was also increased.
Studies have been carried out on Diclofenac and Ketotifen. Ketoprofen in a transfersome formulation gained

NSAIDS

MOLECULES STERIODS

ADVANTAGES Transfersomes improves the site specificity and overall drug safety of corticosteroid delivery into skin by optimizing the epicutaneously administered drug dose. Transfersomes based corticosteroids are biologically active at dose several times lower than the currently used formulation for the treatment of skin diseases flexible vesicles of ethinylestradiol showed significant anti-adulatory effects as compared to plain drug given orally and traditional liposomes given topically. Transfersome based formulations of local anesthetics lidocaine and tetracaine showed permeation equivalent to subcutaneous injections. Maximum resulting pain insensitivity is nearly as strong (80%) as that of a comparable subcutaneous bolus injection, but the effect of transferosomal anesthetics last longer. Anti cancer drugs like Methotrexate were tried for transdermal delivery using transfersomes technology. The results were favorable. Transfersomes of Capsaicin has been prepared by Xiao-Ying et al. have shown the better topical absorption in comparison to pure capsaicin.

ANAESTHETIC S

ANTI CANCER

HERBAL DRUGS

DRUG Terbinafine

EXCIPIENTS SPC SODIUM CHOLATE

INVESTIGATION/CONCLUSION Transfersome tested against dermatophytes with respect to plain terbinafine and commercial terbinafine spray. Test showed potent activity against dermatophyte strains with a MIC range of 0.000030.015 mg/ml. values were found to be 8 and 60 times lower than those of naked terbinafine and terbinafine spray, respectively. Transfersomes prepared and characterized for in vitro drug permeation, drug deposition and antitumour activity in albino mice. Permeation of cisplatin loaded transfersome gel across mice skin was found to be significantly higher than in controls (P < 0.001).

Cisplatin

SOYA LECITHIN, SDC

Amp B

SPC, SDC.

Efficacy of AmB transfersomes was studied against resistant clinical isolates of L. donovani in comparison to conventional liposomal formulation and free AmB. AmB transfersomes showed 1.5-fold higher flux compared with conventional liposomes and free AmB because of better partitioning in skin layers. Transfersomes were found to be more effective against insensitive and resistant clinical isolates of Leishmania donovani.
In vitro permeation studies were performed using rat skin and in vivo pharmacokinetic studies for optimized TF formulation was performed on male white rabbits. Transfersome formulation containing indinavir sulphate showed a transdermal flux which was 812 times higher than conventional formulations. The transfersome gel, liposomal gel, and plain drug gel exhibited

Indinavir

SPC, SDC, SPAN 80, TWEEN 80, TRITON X-100.

DRUG Meloxicam

EXCIPIENTS PC, CHOLESTEROL, SC, SODIUM OLEATE DIACETYLPHOSPHATE.

INVESTIGATION/CONCLUSION The in vitro permeation was compared to the MX-loaded liposomes by varying the MX concentration from 2.5 to 70 w/w% of PC. The presence of sodium oleate and sodium cholate in the transfersome resulted in higher entrapment efficiency. Liposomal formulation containing 2.5 w/w% of PC had the highest entrapment efficiency but the lowest loading efficiency, whereas the MX-loaded liposomes containing 70 w/w% of PC showed the opposite results. Transfersomes showed significantly higher skin permeation of MX than did liposomes because of greater disruption of stratum corneum lipids by the transfersomes. Skin permeation studies of transfersome formulation showed an enhanced transdermal flux and a decrease in lag time. The flux through the transfersome was 7.512.04 times higher than by conventional liposome and other formulations bearing stavudine. Single application of 25 mg, as well as multiple doses up to 100 mg ketoprofen were compared with oral ketoprofen and placebo for the treatment of pain induced by eccentric muscle contractions. A single dose of Diractin was found to be superior to placebo and oral ketoprofen. Curcumin-loaded transfersomes prepared using varying concentration of surfactants, such as Tween 80 and span 80, were evaluated. Entrapment efficiency and permeation of transfersomes were dependent on the lecithin:EA ratio (Tween 80 and span 80). Low permeability of curcumin through the skin by conventional formulation was circumvented by transfersome use.

Stavudine

SDC, SPC, SC, SPAN 80 , TWEEN 80, TRITON X-100. SPC, POLYSORBATE.

Ketoprofen

Curcumin

LECITHIN, TWEEN 80, SPAN 80 , TRITON X-100.

DRUG Docetaxel (DTX)

EXCIPIENTS PC, SC, TRYPAN BLUE AND TRITON X-100

INVESTIGATION/CONCLUSION The effect of the combination of microneedle pretreatment and transfersomes on the permeability of DTX was evaluated using porcine skin. Transfersomes loaded with DTX were found to enhance transdermal delivery of DTX without microneedle pretreatment. Lag time following the application of transfersomes through microneedle pretreated skin was decreased by approximately 70% compared with that observed with conventional liposomes. VCR-loaded transfersomes were evaluated for tissue distribution and PK in Swiss albino rats. In vitro lymph targeting of the formulations was evaluated in rats to treat lymphocytic leukaemia./ Average particle size of VCR-loaded transfersomes was found to be 63 2 nm with an entrapment efficiency of 59%. Compared with the intravenous injection of VCR solution, the retention time of VCR in blood was extended 12-fold using VCR-loaded transfersomes, and the targeting index in rat lymph was found to increase by 2.75-fold. In vivo permeation studies were carried out. Transfersome formulation for transdermal delivery of propranolol hydrochloride provided a higher transdermal flux of 16.19 mg/cm2/h, a higher entrapment efficiency of 60.2%, capability as a self penetration enhancer and increased effectiveness for transdermal delivery, compared with liposomes and other conventional preparations. Capsaicin transfersomes were prepared and investigated for in vitro skin penetration and in vivo tissue distribution. In vitro capsaicin cumulative penetration was found to be higher in transfersomes than cream and suspension using rat abdominal skin. Tissue distribution as observed by multiple topical applications of capsaicin transfersomes on to thighs of rats was found to be in the order: bone > skin > plasma > muscle.

Vincristine (VCR)

LECITHIN

Propranolol HCl

SPC, CHOLESTEROL, SEPHADEX G-50, TRITON X-100

Capsaicin

LECITHIN

Case study 1

Role of edge activators and surface charge in developing ultradeformable vesicles with enhanced skin delivery, International Journal of Pharmaceutics 397 (2010) 164172.
Vortexing Sonication, Rotary evaporatio n Sonication.

DS , Egg PC stearylamine, sephadex-G-100, SDC, SC, sodium azide, Tween 80,span 85, chloroform, methanol and isopropyl alcohol, Cellophane membrane.

Measurement of elasticity Entrapment efficiency DSC, In vitro drug release, Vesicle size analysis, TEM, Rotary evaporation Sonication EE% 62%. Physical stability studies. Effect of drug concentration 0.25 to 1 g %= 25 to 61.92 %. Effect of total lipid concentration fraction of lipid taking part in encapsulation was reduced upon increasing the total lipid concentration. Effect of vesicle composition on EE% and relative deformability of Transfersomes optimum deformability PC:EA ratio was 85:15%. Effect of edge activator type TW80-3 showed the highest deformability (4 s).

Case study 2

This case study report the effects of edge activators on the formation of ultradeformable liposomes (UL) for gene delivery. Sodium cholate, sodium deoxycholate, and Tween 80 were tested as edge activators. Of the edge activators, sodium cholate and sodium deoxycholate resulted in the smaller sizes of UL and more positive zeta potentials than did Tween 80. Moreover, sodium deoxycholate-based UL showed the highest positive zeta potentials, which might lead to the firmer binding with negatively charged DNA. Following topical application onto mice, DNA complexed with UL containing either sodium cholate or sodium deoxycholate showed substantial transdermal absorption. In contrast, DNA complexed with Tween 80-based UL did not show in vivo transdermal absorption. These data suggest that UL might be of use as a transdermal delivery system of plasmid DNA, and that the choice of edge activators may play an important role in the transdermal delivery of plasmid DNA via UL.

CASE STUDY 3: New, highly efficient formulation of diclofenac for the topical, transdermal administration in ultradeformable drug carriers, Transfersomes. Biochimica et Biophysica Acta 1514 (2001) 191205. The relative advantage of diclofenac delivery by means of ultradeformable carriers increases with the treated muscle thickness and with decreasing drug dose, as seen in mice, rats and pigs; this can be explained by assuming that the drug associated with carriers is cleared less efficiently by the dermal capillary plexus. In pigs it suffices to use 0.3 mg of diclofenac in highly deformable vesicles per kg body weight, spread over an area of 25 cm2, to ensure therapeutic drug concentration in a 5-cm thick muscle specimen, collected under the agent application site. When the drug is used in a

Conclusion

The transdermal route of drug delivery does not allow transport of high-molecular-weight therapeutic agents and drugs because of the barrier properties of the stratum corneum layer of the skin. Transfersomes are specially designed vesicles capable of responding to external stress by squeezing themselves through skin pores that are many times narrower than they are, leading to increased transdermal flux of the therapeutic agents. Given the interest of researchers in these ultradeformable vesicles, it is evident that these systems hold great potential for circumventing the barrier properties of the stratum corneum for the delivery of both macromolecular therapeutic agents and drugs. However, there are only two transfersome based formulations currently on the market and the reported clinical studies mainly involve ketoprofen and insulin. It is essential to explore new pharmaceutical excipients that can minimise the oxidative degradation of these vesicular systems and that are more consistent with regards to their purity, to realize completely the potential of these versatile systems in transdermal drug delivery.