APTAMERS FOR TOPICAL DELIVERY

BACKGROUND OF THE INVENTION

Aptamers are synthetically derived nucleic acid molecules having specific binding affinity to molecules through interactions other than classic Watson-Crick base pairing. Aptamers have a number of desirable characteristics for use as therapeutics and diagnostics including high specificity and affinity, biological efficacy, and excellent pharmacokinetic properties.

SUMMARY OF THE INVENTION

The skin is the major organ in the human body serving as the interface with the external environment, functioning as a semi permeable barrier to both protect the body from entrance of foreign substances while allowing for loss of excessive endogenous material like water. The skin is the most easily accessible organ in the human body, lending itself to topical and transdermal delivery of pharmaceutical agents. Some of the advantages of topical and drug delivery systems are the avoidance of first pass metabolism, convenience of application, ease of visual assessment, delivery of the pharmaceutical ingredients directly to the site of action, avoidance of the variations seen from systemic deliveries (e.g. gastric emptying, presence of enzymes, pH changes, etc.), ability to easily terminate the medications if safety signals arise, utilisation of highly potent compounds, and suitability for self-medication (Brown et al. Drug Deliv. (2006) 13(3):175-187). However, the skin presents a formidable barrier for topical delivery, especially for compounds that have a molecular weight above 500 daltons. Very few, if any, compounds with a molecular weight above 1,000 daltons have shown clinical efficacy from topical delivery without physical disruption of the barrier. Overcoming this size exclusion phenomenon could present a major breakthrough in dermatology and topical drug delivery as it could open up new areas of research and potentially targets that are not accessible using traditional small molecules.

In some aspects, the disclosure provides a method of treating a skin disease in a subject (e.g., a mammalian subject, e.g., a human), the method comprising topically administering (e.g., applying) a dose (e.g., effective dose, e.g., therapeutically effective dose) of an aptamer (e.g., a pharmaceutically acceptable composition comprising the aptamer) to skin of the subject that comprises the skin disease, (wherein the skin comprises epidermis and dermis), wherein at least 0.01% (e.g., at least 0.05%, at least 0.1%, at least 0.5%, at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, at least 30%, at least 40%, or at least 50%) of the applied dose enters (e.g., permeates) into the epidermis of the skin (e.g., within 15 minutes, 30 minutes, or within 1, 2, 3, 4, 6, 12, 15, 24, 48, or 72 hours of administration).

In some embodiments, the aptamer is present in a pharmaceutically acceptable composition (e.g., a vehicle) (e.g., a vehicle described herein, e.g., a vehicle presented in Table 4 or 9).

In some embodiments, the concentration of the aptamer in the pharmaceutically acceptable composition is between 0.001% and 20% (e.g., between 0.001% and 15%, between 0.001% and 10%, between 0.001% and 5%, between 0.001% and 2%, between 0.01% and 20%, between 0.01% and 10%, between 0.01% and 5%, between 0.01% and 2%, between 0.1% and 20%, between 0.1% and 15%, between 0.1% and 10% between, 0.1% and 5%, or between 0.1% and 2%).

In some embodiments, the pharmaceutically acceptable composition is administered as a spray (e.g., aerosol), a liquid, an ointment, a cream, a lotion, a solution (e.g., an aqueous solution), a suspension, an emulsion, a paste, a gel, a powder, a foam, a slow release nanoparticle, a slow release microparticle, a bioadhesive, a patch, a bandage, a semi-solid dosage form or a wound dressing.

In some embodiments, the pharmaceutically acceptable composition is administered as an aqueous solution.

In some embodiments, the pharmaceutically acceptable composition is administered as a semi-solid dosage form.

In some embodiments, the pharmaceutically acceptable composition is administered as a cream.

In some embodiments, the pharmaceutically acceptable composition does not contain a penetration enhancer (e.g., propylene glycol).

In some embodiments, the applied dose of the pharmaceutically acceptable composition to the skin is between 0.01 and 20 mg/cm²(e.g., between 0.01 and 15 mg/cm², between 0.01 and 10 mg/cm², between 0.01 and 5 mg/cm², between 0.01 and 2 mg/cm², between 0.05 and 20 mg/cm², between 0.05 and 15 mg/cm², between 0.05 and 10 mg/cm², or between 0.05 and 5 mg/cm², between 0.05 and 2 mg/cm², between 0.1 and 20 mg/cm², between 0.1 and 15 mg/cm², between 0.1 and 10 mg/cm², between 0.1 and 5 mg/cm², or between 0.1 and 2 mg/cm²).

In some embodiments, the percentage of the applied dose of topically administered aptamer that enters the epidermis is determined in an in vitro assay using ex vivo human skin (e.g., in a Franz cell set up or flow through diffusion cell) (e.g., at 32° C.) (e.g., as described herein).

In some embodiments, the aptamer enters (e.g., permeates) into the dermis of the skin; and wherein at least 0.01% (e.g., at least 0.05%, at least 0.1%, at least 0.5%, at least 1%, at least 5%, or at least 10%) of the applied dose enters (e.g., permeates) into the dermis (e.g., within 15 minutes, 30 minutes, or within 1, 2, 3, 4, 6, 12, 15, 24, 48, or 72 hours of administration).

In some embodiments, the percentage of the applied dose of topically administered aptamer that enters the dermis is determined in an in vitro assay using ex vivo human skin (e.g., in a Franz cell set up or flow through diffusion cell) (e.g., at 32° C.) (e.g., as described herein).

In some embodiments, less than 5% (e.g., less than 1%, less than 0.5%, less than 0.1%, less than 0.05%, less than 0.01%, less than 0.001%, or below the limit of detection), of the topically applied the aptamer reaches systemic circulation (e.g., after 2, 3, 4, 6, 12, 15, 24, 48 or 72 hours of administration).

In some embodiments, the length of the aptamer is less than 100 (e.g., less than 95, 90, 85, 80, 75, or 70) nucleotides.

In some embodiments, the length of the aptamer is greater than 30 (e.g., greater than 35, 40, 45, 50, 55, 60, or 65) nucleotides.

In some embodiments, the aptamer is between 30 and 90 nucleotides (e.g., between 35 and 85 nucleotides) in length.

In some embodiments, at least one nucleotide of the aptamer comprises a chemical modification.

In some embodiments, all nucleotides of the aptamer comprise a chemical modification.

In some embodiments, the chemical modification comprises a modification on the 2′ position of the sugar.

In some embodiments, the chemical modification comprises a 2′-O-methoxyethyl addition.

In some embodiments, the chemical modification comprises a 2′fluoro addition.

In some embodiments, the aptamer comprises a single inverted deoxythymidine residue (idT) on its 3′ terminus.

In some embodiments, the pharmaceutically acceptable composition is administered once daily.

In some embodiments, the pharmaceutically acceptable composition is administered twice daily.

In some embodiments, the pharmaceutically acceptable composition is administered once weekly.

In some embodiments, the skin disease comprises an exacerbation of asthma, an exacerbation of rheumatoid arthritis, an exacerbation of psoriatic arthritis, an exacerbation of Sjogren's Syndrome, an exacerbation of uveitis, an exacerbation of Graft vs. Host disease (GVHD), an exacerbation of chronic obstructive pulmonary disease (COPD), an exacerbation of arthralgia, or an exacerbation of Islet Cell Transplant inflammation.

In some embodiments, the skin disease comprises psoriasis, dermatitis (e.g., atopic dermatitis, contact dermatitis, eczematous dermatitis, or seborrhoic dermatitis), hidradentis supporativa, Oral Lichen Planus, a chronic blistering (bullous) disorder, acne, seborrhoeic cutaneous manifestations of immunologically-mediated disorders, alopecia, alopecia greata, scleredoma, scar formation (e.g., a keloid scar or a hypertrophic scar), urticaria, rosacea, melanoma, or inflammation from kidney transplant.

In some embodiments, the skin disease comprises a skin disorder of persistent inflammation, cell kinetics, and differentiation (e.g., psoriasis, psoriatic arthritis, exfoliative dermatitis, pityriasis rosea, lichen planus, lichen nitidus, or porokeratosis); a skin disorder of epidermal cohesion, vesicular and bullous disorders (e.g., pemphigus, bulluous pemphigoi, epidermamolysis bullosa acquisita, or pustular eruptions of the palms or soles); a skin disorder of epidermal appendages and related disorders (e.g., hair disorders, nails, rosacea, perioral dermatitis, or follicular syndromes); a skin disorder such as an epidermal and appendageal tumors (e.g., squamous cell carcinoma, basal cell carcinoma, keratoacanthoma, benign epithelial tumors, or merkel cell carcinoma); a disorder of melanocytes (e.g., pigmentary disorders, albinism, hypomelanoses and hypermelanoses, melanocytic nevi, or melanoma); a skin disorder of inflammatory and neoplastic disorders of the dermis (e.g., erythema elavatum diutinum, eosinophils, granuloma facilae, pyoderma gangrenosum, malignant atrophic papulosis, fibrous lesions of dermis and soft tissue, or Kaposi sarcoma); a disorder of the subcutaneous tissue (e.g., panninculitis or lipodystrophy); a skin disorder involving cutaneous changes of altered reactivity (e.g., urticaria, angiodererma, graft-vs-host, allergic contact dermatitis, autosensitization dermatitis, atopic dermatitis, or seborrheic dermatitis); a skin change due to mechanical and physical factors (e.g., thermal injury, radiation dermatitis, corns, or calluses); photodamage (e.g., acute and chronic UV radiation, or photosensitization); or a skin disorder due to microbial agents (e.g., leprosy, lyme borreliosis, onychomycosis, tinea pedra, rubella, measles, herpes simplex, EBV (Epstein-Barr virus), HPV (Human papillomavirus) (e.g., HPV6 &7), warts, or prions).

In some embodiments, the skin disease comprises psoriasis.

In some embodiments, the psoriasis is mild psoriasis.

In some embodiments, the psoriasis is moderate psoriasis.

In some embodiments, the psoriasis is severe psoriasis.

In some embodiments, the skin disease comprises dermatitis (e.g., atopic dermatitis, contact dermatitis, eczematous dermatitis, or seborrhoic dermatitis).

In some embodiments, the pharmaceutically acceptable composition is administered in combination with second agent that comprises a treatment for the skin disease.

In some embodiments, the aptamer binds to a first target and/or a target of the first target, which can include interactions with other biologic or chemical entities that can be subject to modulation (including, e.g., agonism, antagonism, stabilization, destabilization, enhanced clearing from cellular, organ, or whole body systems by aptamers), including soluble ligands and/or their receptors, extracellular, transmembrane, and intracellular enzymes, scaffolding/structural proteins, RNA, DNA, lipids, fatty acids, co-factors and the respective specific and/or non-specific interactions.

In some embodiments, the aptamer binds to a target (e.g., protein) in the skin, e.g., IL2, IL12, IL17, IL22, IL23, IFNγ, TNFα, TSLP, CCR1, CXCR4, CRTH2, aTLR, p53, RAS, MEK, PATCH1, a mediator of sonic hedgehog signaling, miRNA (e.g., mir21), PI3K/AKT, PTEN, FAS, DGAT, SCD1, EGFR, EGF, HB-EGF, TGF-α, EPR, BTC, NRG1-4, a β-integrin, CD44, E-cadherin, CLA, HPV1, HPV5, HPV8, HPV14, HPV16, HPV17, HPV20, HPV31, HPV47, VEGFC, VEGFD, VEGFEGF, FGF, GMCSF, IL-1, PDGF, or TGF-β1.

In some embodiments, the aptamer binds to a target in the epidermis. In some embodiments, the aptamer binds to a target in the dermis.

In some embodiments, the skin is normal skin.

In some embodiments, the skin is compromised skin.

In some embodiments, the compromised skin is diseased skin.

In some embodiments, the compromised skin comprises an altered barrier.

In some embodiments the aptamer comprises cargo (e.g., attached (e.g., conjugated) thereto), e.g., a small molecule, an imaging dye, a fluorophore, or a radiolabel.

In some aspects, the disclosure provides a method for topically administering an aptamer to a subject (e.g., a mammalian subject, e.g., a human), the method comprising:

administering a dose (e.g., effective dose, e.g., therapeutically effective dose) of aptamer (e.g., a pharmaceutically acceptable composition comprising the aptamer) to skin of the subject (wherein the skin comprises epidermis and dermis);

wherein the aptamer enters (e.g., permeates) into the epidermis of the skin; and

wherein at least 0.01% (e.g., at least 0.05%, at least 0.1%, at least 0.5%, at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, at least 30%, at least 40%, or at least 50%) of the applied dose enters (e.g., permeates) into the epidermis (e.g., 15 minutes, 30 minutes, or within 1, 2, 3, 4, 6, 12, 15, 24, 48, or 72 hours of administration);

thereby topically administering the aptamer to the subject.