Research

Percutaneous delivery provides advantages for skin diseases where the target site is present in the deep epidermis (e.g. fungal, bacterial, viral- infections, psoriasis, dermatitis and skin cancers like melanoma). In addition, percutaneous delivery offers several advantages over the conventional oral and intravenous dosage forms such as prevention of the first-pass metabolism, minimization of pain and possible controlled release of drugs. However, a foremost layer of the skin, stratum corneum (SC), acts as the main barrier between the body and the environment and limits the delivery of most drugs. To overcome SC barrier, different nanoparticles such as SLN, NLC, and polymeric nanoparticles were investigated for percutaneous delivery of various small molecules and peptides.

Bilayered Polymeric Nanoparticles for Combination Therapy

Bilayered polymeric nanoparticles (NPS) are prepared for combination therapy. These NPS are prepared using poly(lactic-co-glycolic acid (PLGA) and chitosan where one drug is encapsulated in PLGA core while another drug is encapsulated into chitosan coat. However, these nanoparticles were unable to reach to the deeper skin layers. Therefore the surface of NPS was modified with oleic acid (OA), FDA approved penetration enhancer. During our human skin permeation experiments, we have noted that the surface modification of the NPS with oleic acid (NPS-OA) is responsible for the significant improvement in the spantide (SP), and ketoprofen (KP) delivery to the deeper skin layers.

SP is an anti-inflammatory peptide (MW 1668.76) that antagonizes the neurokinin-1 receptors and thus inhibits inflammatory response associated with substance-P. KP is a potent non-steroidal anti-inflammatory drug (NSAID) which inhibits arachidonic acid metabolism by potent inhibitory action on cyclooxygenase.

Nanostructured Lipid Carriers for Percutaneous Drug Delivery

Various lipid carriers such as liposomes, solid lipid nanoparticles (SLN) have been well studied for percutaneous drug delivery. However, liposomes have limitations in their a) physical stability problems with liposomes and b) high costs for effective large-scale production. Similarly, SLNs have the limitation of drug expulsion from the carrier and limited drug loading capacity. To overcome this, the second generation of lipid nanoparticles, nanostructured lipid carriers (NLCs) have been developed using the blend of both solid lipids and liquid lipids (oils). NLCs has higher loading capacity and lower drug leaching in storage compared to SLN.

In our efforts to enhance the delivery of NLCs into the deep epidermis, we have already shown that a well-known Cell penetrating peptide (CPPs), transactivating transcriptional activator (TAT), when linked to nanoparticles, has the potential to carry the payloads across the skin layers. We are screening different CPPs for enhancing the drug delivery to the deep epidermis. Further, this approach was used to deliver a peptide, spantide II, and small molecule, ketoprofen together for the treatment of skin diseases like allergic contact dermatitis (ACD).

Allergic Contact Dermatitis (ACD)

Allergic contact dermatitis (ACD) is one of the most prevalent occupational skin diseases. ACD is may result from the interaction of xenobiotics with the immune system. ACD is regarded as an emergence of immunotoxicity in humans. Many chemicals have been shown to cause skin sensitization. In common with other forms of allergy, ACD develops in two phases which are defined operationally as induction and elicitation. Induction of skin sensitization is initiated by topical exposure of the chemical allergen to an individual sufficient to induce a cutaneous immune response of the necessary vigor, known as sensitization. If the sensitized individual is now exposed subsequently, at the same or a different skin site, to the inducing chemical allergen then a more vigorous secondary immune response will be provoked at the point of contact. This, in turn, initiates the cutaneous inflammatory reaction known as ACD. This model was sensitized and challenged using 2,4-dinitrofluorobenzene (DNFB).

Psoriasis-like Model

Psoriasis is a chronic inflammatory skin disease and is characterized by erythematous plaques, excessive growth and aberrant differentiation of keratinocytes along with increases in angiogenesis and inflammatory cell infiltrate. Most commonly used model for investigation of psoriasis is a xenograft model, in which immunodeficient mice are transplanted with human psoriasis-prone skin. However, these kinds of experiments are laborious, expensive and require considerable expertise and technical skills.Therefore a psoriasis plaque like the model was developed using topical application of imiquimod (IMQ) on mice back skin. This model showed the most features of human psoriasis where cytokines like IL-23 and IL-17 are involved.

This area involves the study of molecular mechanisms involved in anti-cancer therapy using mouse xenograft tumor models. Various lung xenograft models (orthotopic, i.v. administration of cells, s.c. model) are available in the laboratory. The study involves the use of novel PPAR-gamma agonists against non-small cell lung cancer and understanding their mechanism of action either alone or in combination with Taxotere. Novel approaches for the delivery of these anticancer drugs and are used with a brief outline:

Aerosols

Formulation of HFA based (134-a and 227) metered dose inhalers with anticancer drugs, proteins and Cox-2 inhibitors.

Nebulization of Cox-2 inhibitors to lungs using lung tumor models. A nose only chamber is available and has been extensively used to evaluate for deposition and efficacy of aerosols in mice.

Formulations of nasal formulations using various polymers and their assessment for droplet size, viscosity and surface tension using the statistical design of experiments.

Liposomal/Nanoparticle Drug Delivery

Development of various anticancer drugs-antibody conjugates, a liposome containing drugs, liposome-antibody conjugates and nanoparticle formulations for cancer drug delivery in combination with immunotoxins.

Another component of research in Dr. Sachdeva’s laboratory is the use of various 3D in vitro cultures as an alternative to animals. In this area, two projects are being pursued. The role of 3D cultures as models to study skin irritation has already been demonstrated and various publications are already available in this area.

In Vitro 3D Culture Model for Wound Healing:

Presently there is no wound healing model available in vitro and most studies are being conducted in vivo. In our laboratory, we are developing a 3D human wound healing model using the EFT-300 cultures from Mattek Corp. Currently, the model is being developed using either various corrosive chemicals (e.g. Sulphuric acid, acrylic acid, potassium hydroxide) to induce a wound or heat to induce a burn wound. Various histological changes along with the role of various growth factors and cytokines are also being monitored to validate such a model. EFT-300 model consists of epidermis and dermis which are very important to understand the natural mechanism of wound healing to establish normal equilibrium of the skin.

In Vitro Tumor 3D Culture Model

This project involves the development of in-vitro tumor model which is more predictive of disease states and drug responses. Algimatrix obtained from Invitrogen is currently being used as the matrix to grow tumor cells as spheroids which will then be treated with various drugs/formulations and to perform various mechanistic studies. The objective is to show a correlation between in vitro and in vivo studies and currently studies are in progress with lung tumor cells. Algimatrix is pure, non-toxic, has a better nutrient delivery without damage to cells.

Dry eye Disease (DED) is an ocular disease caused by hyperosmolarity of tears which is manifested by discomfort and visual disturbances consequently damaging the ocular surface. Topical administration is the most favored route for the treatment of DED, however, frequent drainage from the ocular surface upon administration and poor corneal permeability requires frequent administration of formulations. So there is a need for innovative formulation strategies that can overcome these shortcomings. We are designing of cholecalciferol PEG conjugates which can function as an ester prodrug for cholecalciferol as well as self-assemble into nanomicelles due to hydrophilic PEG2000 for the ocular delivery of various drugs such as tacrolimus to treat DED. A mouse model of DED was developed using benzalkonium chloride treatment and therapeutic efficacy of cholecalciferol conjugates was evaluated. Progression of DED was monitored by assessing tear pH and corneal staining scores. We observed significant improvement in DED in terms of corneal grading score (1 vs 4) and reduction of inflammatory cells.

The formulation of dosage forms for the therapeutic delivery of drugs to the systemic circulation is currently seeing advancements in the pharmaceutical industry today. Variations of fabrication techniques are actively being researched with one of the major players being the additive manufacturing of drugs using 3D printing techniques. In our lab, we are currently working on optimal designs and formulations for both the novel microneedle arrays (MNs) and the traditional tablet drug dosage forms. We have been successful in achieving well defined 3D printed MNs, however we are now working on optimizing the composition of the MNs to demonstrate and advance therapeutic drug dissolution rates in addition to favorable skin permeation rates. Our goal of 3D printing a suitable tablet with favorable dissolution rates to bypass the harsh conditions of the GIT and the liver's tendency to metabolize drugs before reaching the systemic circulation is also coming along with favorable results. Our main driving force in developing this novel form of tablet manufacturing stems from a desire to enable ease of manufacturing as well as multi-drug tablets. We are using DLP, and SLA printing technologies for this purpose and have formulated MNs and tablets by using several model drugs.

"(a): “Partially Polymerized Membrane” effect on the base on the iμ NA due to background printing from over exposure which peels off as the polymer matrix is not completely polymerized in the portions of the structure between the μN bases. (b) “Pancaking” effect on the edges of the iμ NA base due to overexposure. (c) Optimally printed iμ NA. (d-l) Optimization of the aspect ratio (AR) of the iμ NA to achieve a desired R_OC to successfully penetrate human skin. (m) iμ NA dyed in methylene blue showing that the polymer matrix retains its hydrogel properties post printing which would allow for “intelligent” release. (n) SEM image of an optimal aspect ratio iμ NA having a R_OC of ∼20 μm (o). (p) Dyeing of the tips with Gentian Violet to study the effect of penetration on an artificial skin model. (q) Photomicrographs showing successful penetration of an entire 10×10 array with a close-up image in (r)." Kundu et al (2020)

"(a): Schematic of PEGDA (blue) being mixed with 2.5 wt% Diphenyl (2, 4, 6-trimethylbenzoyl) phosphine oxide (TPO) (green) and 0.25wt% therapeutic cargo of diclofenac sodium (pink). (b) DLP 3D printing of the prepared polymer matrix with a UV light source of 385 nm onto the build platform. The iμNA printed consist of a sacrificial base plate and a raft to improve adhesion of the 3D printed device onto the build platform. (c) Removal of the iμNA from the build platform to have the final iμNA which may be used for applications including ocular, acute and chronic drug delivery via transdermal route and allergen testing on human torso among other applications. (d) Close-up of a singular API loaded μNA in the intelligent polymer matrix showing stimulus-based release of the drugs upon sensing its external environment change while retaining the non-drug PEGDA matrix." Kundu et al (2020)