Fatty acid compounds and methods of use

Abstract

Disclosed are compounds, and pharmaceutically acceptable salts thereof, that can treat or prevent a skin cancer in a subject in need thereof.

Description

[0001] FATTY ACID COMPOUNDS AND METHODS OF USE

[0002] CROSS REFERENCE TO RELATED APPLICATIONS

[0003] This application claims the benefit of, and priority to, U.S. Provisional Patent Application No. 63 / 481,919 filed on January 27, 2023, the content of which is hereby incorporated by reference herein in its entirety.

[0004] BACKGROUND

[0005] Cutaneous squamous cell carcinoma (cSCC) is the most common keratinocyte-derived cancer with metastatic potential and it is associated with poor prognosis in the advanced stage (324 Nehal, K.S. 2018; 325 Venables, Z.C. 2019). Ultraviolet radiation (UVR) is the main causative agent involved in the development of skin cancer through multiple mechanisms, including induction of inflammation, oxidative stress, immunosuppression, DNA damage, and dysregulated signal transduction (327 Pihl, C. 2021; 326 Piipponen, M. 2021).

[0006] The progression of the skin carcinogenesis process is characterized by the occurrence of the epithelial-mesenchymal transition (EMT), a complex biological mechanism in which epithelial tumour cells tend to lose their differentiated properties (loss of E-cadherin, desmoplakin, and laminin- 1 expression) and redirect to a mesenchymal-like phenotype (increased expression of N-cadherin, vimentin, fibronectin, MMP-2), with the consequent increase of their migratory and invasive potential (279 Ciuciulete, A.R. 2021; 304 Hodorogea, A. 2019; 411 Nieto, M Angela 2017; 341 Saenz-Sarda, X. 2018; 280 Thiery, J.P. 2009; 305 Usman, S. 2021). Increasing evidence demonstrates that metabolic reprogramming is a hallmark of cancer and extensive metabolic dysregulation of cancer cells is related to the EMT program (355 Morandi, A. 2017; 356 Sciacovelli, M. 2017). The Warburg effect, facilitating the production of energy from glycolysis and less from oxidative phosphorylation, is the most thoroughly described metabolic change able to potentiate the aggressive proliferation of cancer cells (357 Liberti, M.V. 2016). Moreover, tumor cells show dysregulated lipid metabolism, including high lipogenic and lipolytic capacity, elevated membranous lipid synthesis, and upregulation of bioactive lipid production, which induces EMT processes (359 Kang, H. 2019; 358 Luo, X. 2017). Based on the studies on lipid metabolism in cancer, the most extensive changes in lipid metabolism pathways are fatty acids (FA) metabolism, cholesterol metabolism, and arachidonic acid metabolism (406 Hao, Y. 2019). Increased expression and activity of stearoyl CoA desaturase (SCD1), i.e., the enzyme converting saturated fatty acids to A9- monounsaturated fatty acids, is involved in increased cancer cells proliferation, growth, migration, EMT, metastasis, chemoresistance, and maintenance of cancer stem cells (360 Raeisi, M. 2022). An increase in cyclooxygenase-2 (COX-2) expression and a subsequent increase in intracellular prostaglandin E2 (PGE2) levels have been reported in malignant tumour cells, and many previous studies have indicated that these factors have roles in EMT induction (362 Che, D. 2017; 361 Li, Z.L. 2015).

[0007] Accordingly, there exists in the art a need for therapeutic agents such as fatty acids that can treat and / or prevent a skin cancer or pre-cancerous skin lesion such as from a keratinocyte- derived skin cancer.

[0008] BRIEF DESCRIPTION OF THE DRAWINGS

[0009] Fig. 1A shows quantitative real-time PCR analysis of CAT, HO-1, and NQO1 in NHKs treated with compound 1 (“Cmpd 1”) (10-50-90 pM) for 6 h. All mRNA values were normalized against the expression of GAPDH and were expressed relative to untreated control cells. The data in the graphs are the mean ± SD of three independent experiments (*p<0.05, **p<0.01 vs untreated control).

[0010] Fig. IB shows western blot analysis of catalase protein expression in NHKs treated with compound 1 (10-50-90 pM) for 24 h. Representative blots are shown.

[0011] Fig. 1C shows quantitative real-time PCR analysis of CAT, HO-1, and NQO1 in NHKs pre-incubated with compound 1 (10-50-90 pM) for 24 h and then irradiated with UVB 25 mJ / cm2. mRNA was extracted 6 h later the irradiation. All mRNA values were normalized against the expression of GAPDH and were expressed relative to untreated control cells. The data in the graphs are mean ± SD of three independent experiments (*p<0.05 vs untreated control;$p<0.05,$$p<0.01 vs UVB-treated cells).

[0012] Fig. ID shows western blot analysis of catalase, p53, and phospho-yH2AX protein expression in NHKs pre-incubated with compound 1 (10-50-90 pM) for 24 h and then irradiated with UVB 25 mJ / cm2. Proteins were extracted 24 h later the irradiation. Representative blots are shown. P-actin was used as an endogenous loading control for western blot analysis. Densitometric scanning of band intensities was performed to quantify the change in protein expression. Data represent the mean ± SD of three independent experiments and are expressed as fold change respect to untreated control cells (control value taken as 1-fold in each case).

[0013] Fig. 2A shows cell number analysis performed in A431 treated with 2,4,6-octatrienoic acid (“Octa”) or compound 1 (90 pM) for 72 h. The data are presented as the mean ± SD of three independent experiments and are expressed as the fold change respect to untreated control cells (*p<0.05 vs untreated control). Fig. 2B shows cell-cycle distribution evaluated by flow cytometric analysis on A431 treated with Octa or compound 1 (90 pM) for 48 h. The bar graph shows the distribution of cells among the different phases of the cell cycle. The data are expressed as the mean ± SD of three independent (*p<0.05 vs untreated control).

[0014] Fig. 2C shows western blot analysis of p21 and cyclin DI protein expression in A431 treated with Octa or compound 1 (90 pM) for 24 h and 48 h, respectively. Representative blots are shown.

[0015] Fig. 2D shows quantitative real-time PCR analysis of FLG, IVL, and LOR in A431 treated with Octa or compound 1 (90 pM) for 48 h. All mRNA values were normalized against the expression of GAPDH and were expressed relative to untreated control cells. The data in the graphs are mean ± SD of three independent experiments (*p<0.05, **p<0.01 vs untreated control).

[0016] Fig. 2E shows western blot analysis of filaggrin, involucrin, and loricrin protein expression in A431 treated with Octa or compound 1 (90 pM) for 72 h. P-actin and GAPDH were used as endogenous loading control for western blot analyses. Densitometric scanning of band intensities was performed to quantify the change in protein expression. Data represent the mean ± SD of three independent experiments and are expressed as fold change respect to untreated control cells (control value taken as 1-fold in each case).

[0017] Fig. 2F shows cell viability evaluated by MTT assay on NHKs treated with Octa or compound 1 (90 pM) for 72 h. The data in the graphs are mean ± SD of three independent experiments.

[0018] Fig. 3A shows quantitative real-time PCR analysis of FLG, IVL, and LOR in NHKs incubated with Octa or compound 1 (90 pM) for 48 h. All mRNA values were normalized against the expression of GAPDH and were expressed relative to untreated control cells. The data in the graphs are mean ± SD of three independent experiments (*p<0.05 vs untreated control).

[0019] Fig. 3B shows Western blot analysis of filaggrin, involucrin and loricrin protein expression in NHKs treated with compound 1 or Octa (90 pM) for 72 h. Representative blots are shown. GAPDH was used as an endogenous loading control for western blot analysis. Densitometric scanning of band intensities was performed to quantify the change in protein expression. Data represent the mean ± SD of three independent experiments and are expressed as fold change respect to untreated control cells (control value taken as 1-fold in each case).

[0020] Fig. 4A shows quantitative real-time PCR analysis of NCAD, ECAD, SLUG, FIBRONECTIN, VIMENTIN, and MMP2 in A431 pre-incubated with Octa or compound 1 (90 pM) for 1 h and then treated with TGF-01 (15 ng / mL) for 48 h NHKs. All mRNA values were normalized against the expression of GAPDH and were expressed relative to untreated control cells. The data in the graphs are the mean ± SD of three independent experiments (*p<0.05, **p<0.01 vs untreated control;$p<0.05,$$p<0.01 vs TGF-01 -treated cells).

[0021] Fig. 4B shows western blot analysis of vimentin, E-cadherin, and fibronectin protein expression in A431 pre-incubated with Octa or compound 1 (90 pM) for 1 h and then treated with TGF-pi (15 ng / ml) for 72 h. GAPDH was used as an endogenous loading control for western blot analysis. Densitometric scanning of band intensities was performed to quantify the change in protein expression. Data represent the mean ± SD of three independent experiments and are expressed as fold change respect to untreated control cells (control value taken as 1-fold in each case).

[0022] Fig. 4C shows immunofluorescence and quantitative analysis of vimentin, fibronectin, and E-cadherin in A431 cells. Results are expressed as fold change of positive cells or mean fluorescence intensity respect to control. Nuclei were counterstained with DAPI. Bar: 20 pm. (*p<0.05, **p<0.01 vs control;$p<0.05 vs TGF-pi;$$p<0.01 vs TGF-pi).

[0023] Fig. 5 A shows quantification by GCMS of cell membrane MUFA / SFA ratio in A431 cells pre-treated with Octa and compound 1 (90 pM) for 1 h and then exposed to TGF-pi (15ng / ml) alone or in combination with Octa and compound 1 for 24 h and 72h.

[0024] Fig. 5B shows quantification by GCMS of extracellular C16: l / C16:0 ratio in A431 cells pre-treated with Octa and compound 1 (90 pM) for 1 h and then exposed to TGF-pi (15ng / ml) alone or in combination with Octa and compound 1 for 24 h and 72h.

[0025] Fig. 5C shows quantification by GCMS of extracellular C18: l / C18:0 ratio in A431 cells pre-treated with Octa and compound 1 (90 pM) for 1 h and then exposed to TGF-pi (15ng / ml) alone or in combination with Octa and compound 1 for 24 h and 72h.

[0026] Fig. 5D shows quantification by GCMS of cholesterol amount in A431 cells pre-treated with Octa and compound 1 (90 pM) for 1 h and then exposed to TGF-pi (15ng / ml) alone or in combination with Octa and compound 1 for 24 h and 72h.

[0027] Fig. 6A shows luciferase activity in MITO-Luc mouse-induced papillomas. BLI of MITO-Luc mice before treatment (pre-imaging), after treatment with DMBA-TPA treatments on the ventral abdomen (tO), after 8 treatments with placebo or compound 1 (tl), and after 15 treatments with placebo or compound 1 (t2). Images were collected on 5 animals for each treatment and showed a representative animal. Photon emission from the lesions was measured as photons per second per square centimeter per steradian (photons / s / cm2 / sr). Fig. 6B illustrates BLI signal intensities relative to DMBA-TPA drug treatment control (tO) after 15 treatments with placebo or compound 1 (tl). Each bar represents the mean value ± SEM of five animals.

[0028] Fig. 6C shows quantitative real-time PCR analysis of NCAD, ECAD, SLUG, FIBRONECTIN, VIMENTIN, a-SMA, xA MMB2 in samples of mouse skin collected in lesional and nonlesional areas at the end of the 15 treatments with placebo or compound 1 (3 different mice per treatment group). All mRNA values were normalized against the expression of GAPDH and were expressed relative to nonlesional skin (*p<0.05, **p<0.01 vs placebo).

[0029] SUMMARY

[0030] The disclosure is directed, at least in part, to novel fatty acid compounds that can be useful for treating or preventing a skin cancer in a subject in need thereof. Also disclosed herein are pharmaceutical compositions comprising at least one disclosed compound and a pharmaceutically acceptable excipient. The present disclosure should be understood to include compounds as described herein as well as methods of using the compounds for treatment or prevention of a skin cancer. Each of these different aspects is described more particularly by the various embodiments described herein, which embodiments can be equally applicable to the different aspects.

[0031] In one aspect, the present disclosure provides a compound of Formula (I): or a pharmaceutically acceptable salt thereof, wherein:

[0032] R1is selected from the group consisting of: CEE, CH, and NRa;

[0033] R2is selected from the group consisting of: H, Ci-Ce alkyl, and (Ci-Ce alkylene)phenyl, wherein the phenyl may be optionally substituted with one, two or three substituents selected from the group consisting of Ci-Ce alkyl, halogen, Ci-Ce haloalkyl, Ci-Ce alkoxy, and cyano;

[0034] (i) R3is selected from the group consisting of H, Ci-Ce alkyl, and oxo,

[0035] R4is selected from H and Ci-Ce alkoxy, and

[0036] R5is H; or (ii) R3and R4are taken together to form a 5-6 membered heteroaryl with the carbon atoms to which R3, R4, and R5are attached, and

[0037] R5is selected from H and Ci-Ce alkyl;

[0038] — is a double bond, or — is a single bond when (a) R1is NRaand R3is oxo or (b) R3and R4are taken together to form a 5-6 membered heteroaryl with the carbon atoms to which R3, R4, and R5are attached; and

[0039] Rais H or Ci-Ce alkyl; wherein at least one of R3, R4, and R5is not hydrogen.

[0040] In some embodiments, the present disclosure provides a pharmaceutical composition comprising a compound disclosed herein or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient.

[0041] Provided herein, in part, is a combination treatment comprising a compound described herein or a pharmaceutically acceptable salt thereof, and octatri enoic acid.

[0042] Also provided herein is a method of treating or preventing a skin cancer or a pre- cancerous skin lesion in a patient in need thereof comprising administering to the patient a therapeutically effective amount of a compound disclosed herein or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition disclosed herein.

[0043] Also provided herein is a compound disclosed herein or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition disclosed herein for use in treating or preventing a skin cancer or a pre-cancerous skin lesion in a patient.

[0044] In some embodiments, the skin cancer is keratinocyte-derived skin cancer.

[0045] DETAILED DESCRIPTION

[0046] The features and other details of the disclosure will now be more particularly described. Before further description of the present disclosure, certain terms employed in the specification, examples and appended claims are collected here. These definitions should be read in light of the remainder of the disclosure and as understood by a person of skill in the art. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by a person of ordinary skill in the art. Definitions

[0047] The term “alkyl,” as used herein, refers to a saturated straight-chain or branched hydrocarbon, such as a straight-chain or branched group of 1-6, 1-4, or 1-3 carbon atoms, referred to herein as Ci-Ce alkyl, C1-C4 alkyl, and C1-C3 alkyl, respectively. For example, “Ci- Ce alkyl” refers to a straight-chain or branched saturated hydrocarbon containing 1-6 carbon atoms. Examples of a Ci-Ce alkyl group include, but are not limited to, methyl, ethyl, propyl, butyl, pentyl, hexyl, isopropyl, isobutyl, ec-butyl, / e / 7-butyl, isopentyl, and neopentyl. In another example, “C1-C4 alkyl” refers to a straight-chain or branched saturated hydrocarbon containing 1-4 carbon atoms. Examples of a C1-C4 alkyl group include, but are not limited to, methyl, ethyl, propyl, butyl, isopropyl, isobutyl, sec-butyl and tert-butyl. Exemplary alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, 2 -methyl- 1 -propyl, 2- methyl-2-propyl, 2-methyl-l -butyl, 3 -methyl- 1 -butyl, 3-methyl-2-butyl, 2,2-dimethyl-l -propyl, 2-methyl-l -pentyl, 3 -methyl- 1 -pentyl, 4-methyl-l -pentyl, 2-methyl-2-pentyl, 3-methyl-2- pentyl, 4-methyl-2-pentyl, 2,2-dimethyl-l -butyl, 3, 3 -dimethyl- 1 -butyl, 2-ethyl-l -butyl, butyl, isobutyl, t-butyl, pentyl, isopentyl, neopentyl, and hexyl.

[0048] The term “alkylene” refers to the diradical of an alkyl group.

[0049] The term “alkoxy,” as used herein, refers to an alkyl group attached to an oxygen atom (alkyl-O-). Alkoxy groups can have 1-6 or 2-6 carbon atoms and are referred to herein as Ci- Ce alkoxy and C2-C6 alkoxy, respectively. Exemplary alkoxy groups include, but are not limited to, methoxy, ethoxy, propyloxy, isopropoxy, and tert-butoxy.

[0050] The term “cyano,” as used herein, refers to the radical -CN.

[0051] The terms “halo” and “halogen,” as used herein, refer to fluoro (F), chloro (Cl), bromo (Br), and / or iodo (I).

[0052] The term “haloalkyl” as used herein refers to an alkyl group substituted with one or more halogen atoms, for example, -CH2F, -CHF2, -CF3, CH2CF3, and -CF2CF3.

[0053] The term “heteroatom,” as used herein, refers to an atom of any element other than carbon or hydrogen and includes, for example, nitrogen (N), oxygen (O), silicon (Si), sulfur (S), phosphorus (P), and selenium (Se).

[0054] The terms “heteroaryl” as used herein refers to a monocyclic aromatic 4-6 membered (e.g., 5-6 membered) ring system containing one or more heteroatoms, for example one to three heteroatoms, such as nitrogen, oxygen, and sulfur. Where possible, said heteroaryl ring may be linked to the adjacent radical though carbon or nitrogen. Examples of heteroaryl rings include but are not limited to furan, thiophene, pyrrole, thiazole, oxazole, isothiazole, isoxazole, imidazole, pyrazole, triazole, pyridine, and pyrimidine.

[0055] The terms “hydroxy” and “hydroxyl,” as used herein, refer to the radical -OH.

[0056] The term “oxo,” as used herein, refers to the radical =0 (double bonded oxygen).

[0057] The term “compound,” as used herein, refers to the compound itself and its pharmaceutically acceptable salts, hydrates, esters and N-oxides and its isotopically-labelled forms, unless otherwise understood from the context of the description or expressly limited to one particular form of the compound, i.e., the compound itself, a specific stereoisomer and / or isotopically-labelled compound, or a pharmaceutically acceptable salt, a hydrate, an ester, or an N-oxide thereof. It should be understood that a compound can refer to a pharmaceutically acceptable salt, a hydrate, an ester, or an N-oxide of the compound and / or an isotopically- labelled compound.

[0058] The disclosure also embraces isotopically-labeled compounds which are identical to those compounds recited herein, except that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of isotopes that can be incorporated into compounds described herein include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, fluorine and chlorine, such as2H (“D”),3H,13C,14C,15N,180,170,31P,32P,35S,18F, and36C1, respectively. For example, a compound described herein can have one or more H atoms replaced with deuterium.

[0059] Certain isotopically-labeled compounds (e.g., those labeled with3H and14C) can be useful in compound and / or substrate tissue distribution assays. Tritiated (i.e.,3H) and carbon- 14 (i.e.,14C) isotopes can be particularly preferred for their ease of preparation and detectability. Further, substitution with heavier isotopes such as deuterium (i.e.,2H) can afford certain therapeutic advantages resulting from greater metabolic stability (e.g., increased in vivo half-life or reduced dosage requirements) and hence can be preferred in some circumstances. Isotopically-labeled compounds can generally be prepared by following procedures analogous to those disclosed herein, for example, in the Examples section, by substituting an isotopically- labeled reagent for a non-isotopically-labeled reagent.

[0060] “2,4,6-octatrienoic acid,” “(2E,4E,6£)-octa-2,4,6-trienoate,” or “Octa” has the chemical formula:

[0061] The phrases “pharmaceutically acceptable” and “pharmacologically acceptable,” as used herein, refer to compounds, molecular entities, compositions, materials, and / or dosage forms that do not produce an adverse, allergic or other untoward reaction when administered to an animal, or a human, as appropriate. For human administration, preparations should meet sterility, pyrogenicity, general safety and purity standards as required by FDA Office of Biologies standards.

[0062] As used herein, “pharmaceutically acceptable excipient” refers to a substance that aids the administration of an active agent to a subject and can be included in the compositions of the present invention without causing a significant adverse toxicological effect on the subject. Non-limiting examples of pharmaceutically acceptable excipients include water, NaCl, normal saline solutions, a phosphate buffered saline solution, emulsions (e.g., such as an oil / water or water / oil emulsions), lactated Ringer’s, normal sucrose, normal glucose, binders, fillers, disintegrants, lubricants, coatings, sweeteners, flavors, salt solutions (such as Ringer’s solution), alcohols, oils, gelatins, carbohydrates such as lactose, amylose or starch, fatty acid esters, hydroxypropylmethylcellulose, polyvinyl pyrrolidine, and colors, and the like. Such preparations can be sterilized and, if desired, mixed with auxiliary agents such as lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, coloring, and / or aromatic substances and the like that do not deleteriously react with the compounds of the disclosure.

[0063] As used herein, the term “pharmaceutically acceptable carrier” refers to any of the standard pharmaceutical carriers, such as a phosphate buffered saline solution, water, emulsions (e.g., such as an oil / water or water / oil emulsions), and various types of wetting agents. The compositions also can include stabilizers and preservatives. For examples of excipients, carriers, stabilizers and adjuvants, see Martin, Remington's Pharmaceutical Sciences, 15th Ed., Mack Publ. Co., Easton, PA (1975).

[0064] The phrase “pharmaceutical composition,” as used herein, refers to a composition comprising at least one compound as disclosed herein formulated together with one or more pharmaceutically acceptable excipients, for example, pharmaceutically acceptable carriers. The pharmaceutical compositions can also contain other active compounds providing supplemental, additional, or enhanced therapeutic functions.

[0065] The terms “individual,” “patient,” and “subject,” as used herein, are used interchangeably and include any animal, including mammals, preferably mice, rats, other rodents, rabbits, dogs, cats, swine, cattle, sheep, horses, or primates, and more preferably, humans. The compounds described in the disclosure can be administered to a mammal, such as a human, but can also be administered to other mammals such as an animal in need of veterinary treatment, for example, domestic animals (e.g., dogs, cats, and the like), farm animals (e.g., cows, sheep, pigs, horses, and the like) and laboratory animals (e.g., rats, mice, guinea pigs, and the like). The mammal treated in the methods described in the disclosure is preferably a mammal in which treatment, for example, of a skin cancer, is desired.

[0066] The term “treating,” as used herein, includes any effect, for example, lessening, reducing, modulating, ameliorating, or eliminating, that results in the improvement of the condition, disease, disorder, and the like, including one or more symptoms thereof. Treating can be curing, improving, or at least partially ameliorating the disorder.

[0067] The term “disorder” refers to and is used interchangeably with, the terms “disease,” “condition,” or “illness,” unless otherwise indicated.

[0068] The phrase “therapeutically effective amount,” as used herein, refers to the amount of a compound (e.g., a disclosed compound) that will elicit the biological or medical response of a tissue, system, animal or human that is being sought by the researcher, veterinarian, medical doctor or other clinician. The compounds described in the disclosure can be administered in therapeutically effective amounts to treat a disease. A therapeutically effective amount of a compound can be the quantity required to achieve a desired therapeutic and / or prophylactic effect, such as an amount which results in lessening of a symptom of a disease such as a skin cancer.

[0069] As used herein, the term “pharmaceutically acceptable salt” refers to any salt of an acidic or a basic group that may be present in a compound of the present disclosure, which salt is compatible with pharmaceutical administration. As is known to those of skill in the art, “salts” of the compounds of the present disclosure may be derived from inorganic or organic acids and bases.

[0070] Examples of salts include, but are not limited to: acetate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphorsulfonate, cyclopentanepropionate, digluconate, dodecyl sulfate, ethanesulfonate, fumarate, flucoheptanoate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate, 2- naphthalenesulfonate, nicotinate, oxalate, palmoate, pectinate, persulfate, phenylpropionate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, tosylate, undecanoate, and the like. Other examples of salts include anions of the compounds of the present disclosure compounded with a suitable cation such as Na+, NHZ, and NWZ (where W can be a C1-4 alkyl group), and the like. For therapeutic use, salts of the compounds of the present disclosure can be pharmaceutically acceptable. However, salts of acids and bases that are non- pharmaceutically acceptable may also find use, for example, in the preparation or purification of a pharmaceutically acceptable compound.

[0071] Compounds included in the present compositions that are basic in nature are capable of forming a wide variety of salts with various inorganic and organic acids. The acids that can be used to prepare pharmaceutically acceptable acid addition salts of such basic compounds are those that form non-toxic acid addition salts, i.e., salts containing pharmacologically acceptable anions, including but not limited to, malate, oxalate, chloride, bromide, iodide, nitrate, sulfate, bisulfate, phosphate, acid phosphate, isonicotinate, acetate, lactate, salicylate, citrate, tartrate, oleate, tannate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucaronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, -toluenesulfonate and pamoate (i.e., 1, l'-methylene-bis-(2-hydroxy-3-naphthoate)) salts.

[0072] Compounds included in the present compositions that are acidic in nature are capable of forming base salts with various pharmacologically acceptable cations. Examples of such salts include alkali metal or alkaline earth metal salts and, particularly, calcium, magnesium, sodium, lithium, zinc, potassium, and iron salts.

[0073] Compounds included in the present compositions that include a basic or acidic moiety can also form pharmaceutically acceptable salts with various amino acids. The compounds of the disclosure can contain both acidic and basic groups; for example, one amino and one carboxylic acid group. In such a case, the compound can exist as an acid addition salt, a zwitterion, or a base salt.

[0074] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains. Throughout the description, where compositions and kits are described as having, including, or comprising specific components, or where processes and methods are described as having, including, or comprising specific steps, it is contemplated that, additionally, there are compositions and kits of the present disclosure that consist essentially of, or consist of, the recited components, and that there are processes and methods according to the present disclosure that consist essentially of, or consist of, the recited processing steps.

[0075] In the application, where an element or component is said to be included in and / or selected from a list of recited elements or components, it should be understood that the element or component can be any one of the recited elements or components, or the element or component can be selected from a group consisting of two or more of the recited elements or components.

[0076] Further, it should be understood that elements and / or features of a composition or a method described herein can be combined in a variety of ways without departing from the spirit and scope of the present disclosure, whether explicit or implicit herein. For example, where reference is made to a particular compound, that compound can be used in various embodiments of compositions of the present disclosure and / or in methods of the present disclosure, unless otherwise understood from the context. In other words, within this application, embodiments have been described and depicted in a way that enables a clear and concise application to be written and drawn, but it is intended and will be appreciated that embodiments can be variously combined or separated without parting from the present teachings and disclosure(s). For example, it will be appreciated that all features described and depicted herein can be applicable to all aspects of the disclosure(s) described and depicted herein.

[0077] The articles “a” and “an” are used in this disclosure to refer to one or more than one (i.e., to at least one) of the grammatical object of the article, unless the context is inappropriate. By way of example, “an element” means one element or more than one element.

[0078] The term “and / or” is used in this disclosure to mean either “and” or “or” unless indicated otherwise.

[0079] It should be understood that the expression “at least one of’ includes individually each of the recited objects after the expression and the various combinations of two or more of the recited objects unless otherwise understood from the context and use. The expression “and / or” in connection with three or more recited objects should be understood to have the same meaning unless otherwise understood from the context.

[0080] The use of the term “include,” “includes,” “including,” “have,” “has,” “having,” “contain,” “contains,” or “containing,” including grammatical equivalents thereof, should be understood generally as open-ended and non-limiting, for example, not excluding additional unrecited elements or steps, unless otherwise specifically stated or understood from the context.

[0081] It should be understood that the order of steps or order for performing certain actions is immaterial so long as the present disclosure remain operable. Moreover, two or more steps or actions can be conducted simultaneously.

[0082] At various places in the present specification, substituents are disclosed in groups or in ranges. It is specifically intended that the description include each and every individual subcombination of the members of such groups and ranges. For example, the term “Ci-6 alkyl” is specifically intended to individually disclose Ci, C2, C3, C4, C5, Ce, Ci-Ce, C1-C5, C1-C4, Ci- C3, C1-C2, C2-C6, C2-C5, C2-C4, C2-C3, C3-C6, C3-C5, C3-C4, C4-C6, C4-C5, and C5-C6alkyl. By way of other examples, an integer in the range of 0 to 40 is specifically intended to individually disclose 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, and 40, and an integer in the range of 1 to 20 is specifically intended to individually disclose 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, and 20. Additional examples include that the phrase “optionally substituted with 1-5 substituents” is specifically intended to individually disclose a chemical group that can include 0, 1, 2, 3, 4, 5, 0-5, 0-4, 0-3, 0-2, 0-1, 1-5, 1-4, 1-3, 1-2, 2-5, 2-4, 2-3, 3-5, 3-4, and 4-5 substituents.

[0083] The use of any and all examples, or exemplary language herein, for example, “such as” or “including,” is intended merely to illustrate better the present disclosure and does not pose a limitation on the scope of the disclosure unless claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the present disclosure.

[0084] Further, if a variable is not accompanied by a definition, then the variable is defined as found elsewhere in the disclosure unless understood to be different from the context. In addition, the definition of each variable and / or substituent, for example, Ci-Ce alkyl, R2, Rb, w and the like, when it occurs more than once in any structure or compound, can be independent of its definition elsewhere in the same structure or compound. Definitions of the variables and / or substituents in formulae and / or compounds herein encompass multiple chemical groups. The present disclosure includes embodiments where, for example, i) the definition of a variable and / or substituent is a single chemical group selected from those chemical groups set forth herein, ii) the definition is a collection of two or more of the chemical groups selected from those set forth herein, and iii) the compound is defined by a combination of variables and / or substituents in which the variables and / or substituents are defined by (i) or (ii).

[0085] Various aspects of the disclosure are set forth herein under headings and / or in sections for clarity; however, it is understood that all aspects, embodiments, or features of the disclosure described in one particular section are not to be limited to that particular section but rather can apply to any aspect, embodiment, or feature of the present disclosure.

[0086] Compounds

[0087] Disclosed herein, for example, are novel fatty acids and related compounds that can be useful for treating or preventing a disease or disorder, e.g., a skin cancer, in a subject in need thereof.

[0088] In one aspect, the present disclosure provides a compound of Formula (I): or a pharmaceutically acceptable salt thereof, wherein:

[0089] R1is selected from the group consisting of CH2, CH, and NRa;

[0090] R2is selected from the group consisting of H, Ci-Ce alkyl, and (Ci-Ce alkylene)phenyl, wherein the phenyl may be optionally substituted with one, two or three substituents selected from the group consisting of Ci-Ce alkyl, halogen, Ci-Ce haloalkyl, Ci-Ce alkoxy, and cyano;

[0091] (i) R3is selected from the group consisting of H, Ci-Ce alkyl, and oxo,

[0092] R4is selected from H and Ci-Ce alkoxy, and

[0093] R5is H; or

[0094] (ii) R3and R4are taken together to form a 5-6 membered heteroaryl with the carbon atoms to which R3, R4, and R5are attached, and R5is selected from H and Ci-Ce alkyl;

[0095] — is a double bond, or — is a single bond when (a) R1is NRaand R3is oxo or (b) R3and R4are taken together to form a 5-6 membered heteroaryl with the carbon atoms to which R3, R4, and R5are attached; and

[0096] Rais H or Ci-Ce alkyl; wherein at least one of R3, R4, and R5is not hydrogen.

[0097] In some embodiments, R1can be CH2. In certain embodiments, R1can be CH. In other embodiments, R1can be NRa.

[0098] In some embodiments, R2can be H.

[0099] In certain embodiments, R3can be H. In other embodiments, R3can be Ci-Ce alkyl, e.g., Ci alkyl, C2 alkyl, C3 alkyl, C4 alkyl, C5 alkyl, or Ce alkyl. In some embodiments, R3can be oxo.

[0100] In certain embodiments, R4can be H. In other embodiments, R4can be Ci-Ce alkoxy, e.g., Ci alkoxy, C2 alkoxy, C3 alkoxy, C4 alkoxy, C5 alkoxy, or Ce alkoxy.

[0101] In some embodiments, R3and R4can be taken together to form a 5-6 membered heteroaryl with the carbon atoms to which R3, R4, and R5are attached.

[0102] In other embodiments, R5can be H. In certain embodiments, R5can be Ci-Ce alkyl, e.g., Ci alkyl, C2 alkyl, C3 alkyl, C4 alkyl, C5 alkyl, or Ce alkyl.

[0103] In some embodiments, Rais H. In other embodiments, Rais Ci-Ce alkyl, e.g., Ci alkyl, C2 alkyl, C3 alkyl, C4 alkyl, C5 alkyl, or Ce alkyl.

[0104] In certain embodiments, the compound is selected from the group consisting of:

[0105] Compositions

[0106] The present disclosure also provides a pharmaceutical formulation or a pharmaceutical composition including a disclosed compound and a pharmaceutically acceptable excipient for use in the methods of the invention.

[0107] A formulation can be prepared in any of a variety of forms for use such as for administering an active agent to a patient, who may be in need thereof. For example, the pharmaceutical compositions of the present disclosure can be formulated for administration in solid or liquid form, including those adapted for the following: (1) oral administration, for example, drenches (aqueous or non-aqueous solutions or suspensions), tablets (e.g., those targeted for buccal, sublingual, and / or systemic absorption), boluses, powders, granules, and pastes for application to the tongue; (2) parenteral administration by, for example, subcutaneous, intramuscular, intraperitoneal, intravenous or epidural injection as, for example, a sterile solution or suspension, or sustained-release formulation; (3) topical administration, for example, as a cream, ointment, or a controlled-release patch or spray applied to the skin; (4) intravaginal or intrarectal administration, for example, as a pessary, cream or foam; (5) sublingual administration; (6) ocular administration; (7) transdermal administration; or (8) nasal administration.

[0108] The pharmaceutical compositions of the present disclosure can be topical formulations. The topical formulations may be formulated as a liquid, a solution, an emulsion, a cream, a lotion, a suspension, a triturate, a gel, a jelly, a foam, a paste, an ointment, a shampoo, an adhesive, a patch, a foam, an aerosol foam, or the like. In certain embodiments, the topical formulations of the present disclosure may be formulated as a liquid. In certain embodiments, the topical formulations of the present disclosure may be formulated as a solution. In certain embodiments, the topical formulations of the present disclosure may be formulated as an emulsion. In certain embodiments, the topical formulations of the present disclosure may be formulated as a cream. In certain embodiments, the topical formulations of the present disclosure may be formulated as a lotion. In certain embodiments, the topical formulations of the present disclosure may be formulated as a suspension. In certain embodiments, the topical formulations of the present disclosure may be formulated as a triturate. In certain embodiments, the topical formulations of the present disclosure may be formulated as a gel. In certain embodiments, the topical formulations of the present disclosure may be formulated as a jelly. In certain embodiments, the topical formulations of the present disclosure may be formulated as a foam. In certain embodiments, the topical formulations of the present disclosure may be formulated as a paste. In certain embodiments, the topical formulations of the present disclosure may be formulated as an ointment. In certain embodiments, the topical formulations of the present disclosure may be formulated as a shampoo. In certain embodiments, the topical formulations of the present disclosure may be formulated as an adhesive. In certain embodiments, the topical formulations of the present disclosure may be formulated as a patch. In certain embodiments, the topical formulations of the present disclosure upon application to a skin of a subject may form a patch.

[0109] Ointments, as is well known in the art of pharmaceutical formulation, are semisolid preparations that are typically based on petrolatum or other petroleum derivatives. The specific ointment base to be used, as will be appreciated by those skilled in the art, is one that will provide for optimum drug delivery, and, preferably, will provide for other desired characteristics as well, e.g., emolliency or the like. As with other carriers or vehicles, an ointment base should be inert, stable, nonirritating and nonsensitizing. As explained in Remington: The Science and Practice of Pharmacy, 19th Ed. (Easton, Pa.: Mack Publishing Co., 1995), at pages 1399-1404, ointment bases may be grouped in four classes: oleaginous bases; emulsifiable bases; emulsion bases; and water-soluble bases. Oleaginous ointment bases include, for example, vegetable oils, fats obtained from animals, and semisolid hydrocarbons obtained from petroleum. Emulsifiable ointment bases, also known as absorbent ointment bases, contain little or no water and include, for example, hydroxystearin sulfate, anhydrous lanolin, and hydrophilic petrolatum. Emulsion ointment bases are either water-in-oil (W / O) emulsions or oil-in-water (O / W) emulsions, and include, for example, cetyl alcohol, glyceryl monostearate, lanolin, and stearic acid. In embodiments, water-soluble ointment bases are prepared from polyethylene glycols of varying molecular weight; again, see Remington: The Science and Practice of Pharmacy for further information.

[0110] Creams, as also well known in the art, are viscous liquids or semisolid emulsions, either oil-in-water or water-in-oil. Cream bases are water- washable, and contain an oil phase, an emulsifier, and an aqueous phase. The oil phase, also called the “internal” phase, is generally included of petrolatum and a fatty alcohol such as cetyl or stearyl alcohol. The aqueous phase usually, although not necessarily, exceeds the oil phase in volume, and generally contains a humectant. The emulsifier in a cream formulation is generally a nonionic, anionic, cationic, or amphoteric surfactant.

[0111] Gels are semisolid, suspension-type systems. Single-phase gels contain organic macromolecules distributed substantially uniformly throughout the carrier liquid, which is typically aqueous, but also contains an alcohol and, optionally, an oil. In embodiments, “organic macromolecules,” i.e., gelling agents, are crosslinked acrylic acid polymers such as the “carbomer” family of polymers, e.g., carboxypolyalkylenes that may be obtained commercially under the CARBOPOL™ trademark. Hydrophilic polymers such as polyethylene oxides, polyoxyethylene-polyoxypropylene copolymers, and polyvinyl alcohol; cellulosic polymers such as hydroxypropyl cellulose, hydroxyethyl cellulose, hydroxypropyl methylcellulose, hydroxypropyl methylcellulose phthalate, and methyl cellulose; gums such as tragacanth and xanthan gum; sodium alginate; and gelatin may also be included. In order to prepare a uniform gel, dispersing agents such as alcohol or glycerin can be added, or the gelling agent can be dispersed by trituration, mechanical mixing, or stirring, or combinations thereof.

[0112] Lotions are preparations to be applied to the skin surface without friction, and are typically liquid or semiliquid preparations in which solid particles, including the active agent, are present in a water or alcohol base. Lotions are usually suspensions of solids, and preferably, for the present purpose, include a liquid oily emulsion of the oil-in-water type. In embodiments, lotions are used for treating large body areas, because of the ease of applying a more fluid composition. It is generally necessary that the insoluble matter in a lotion be finely divided. Lotions will typically contain suspending agents to produce better dispersions as well as compounds useful for localizing and holding the active agent in contact with the skin, e.g., methylcellulose, sodium carboxymethylcellulose, or the like.

[0113] Pastes are semisolid dosage forms in which the compound of Formula (I), or a pharmaceutically acceptable salt thereof, is suspended in a suitable base. Depending on the nature of the base, pastes are divided between fatty pastes or those made from a single-phase aqueous gels. The base in a fatty paste is generally petrolatum, hydrophilic petrolatum, or the like. The pastes made from single-phase aqueous gels generally incorporate carboxymethylcellulose or the like as a base. Formulations may also be prepared with liposomes, micelles, and microspheres. Liposomes are microscopic vesicles having a lipid wall including a lipid bilayer, and can be used as drug delivery systems herein as well. Generally, liposome formulations are poorly soluble or insoluble pharmaceutical agents. Liposomal preparations for use in the instant disclosure may include cationic (positively charged), anionic (negatively charged), and neutral preparations. Cationic liposomes are readily available. For example, N[l-2,3- dioleyloxy)propyl]-N,N,N-triethylammonium (DOTMA) liposomes are available under the trade name LIPOFECTIN™. (ThermoFisher). Similarly, anionic and neutral liposomes are readily available as well, e.g., from Avanti Polar Lipids (Birmingham, Ala.), or can be easily prepared using readily available materials. Such materials include phosphatidyl choline, cholesterol, phosphatidyl ethanolamine, dioleoylphosphatidyl choline (DOPC), dioleoylphosphatidyl glycerol (DOPG), and dioleoylphoshatidyl ethanolamine (DOPE), among others. These materials can also be mixed with DOTMA in appropriate ratios. Methods for making liposomes using these materials are well known in the art.

[0114] Micelles are known in the art as included of surfactant molecules arranged so that their polar head groups form an outer spherical shell, while their hydrophobic, hydrocarbon chains are oriented towards the center of the sphere, forming a core. Micelles form in an aqueous solution containing surfactant at a high enough concentration so that micelles naturally result. Surfactants useful for forming micelles include, but are not limited to, potassium laurate, sodium octane sulfonate, sodium decane sulfonate, sodium dodecane sulfonate, sodium lauryl sulfate, docusate sodium, decyltrimethylammonium bromide, dodecyltrimethylammonium bromide, tetradecyltrimethylammonium bromide, tetradecyltrimethylammonium chloride, dodecylammonium chloride, polyoxyl 8 dodecyl ether, polyoxyl 12 dodecyl ether, nonoxynol 10, and nonoxynol 30. Micelle formulations can be used in conjunction with the present disclosure either by incorporation into the reservoir of a topical or transdermal delivery system, or into a formulation to be applied to the body surface.

[0115] Microspheres, similarly, may be incorporated into the present formulations and drug delivery systems. Like liposomes and micelles, microspheres essentially encapsulate a drug or drug-containing formulation. Microspheres are generally, although not necessarily, formed from synthetic or naturally occurring biocompatible polymers, but may also be included of charged lipids such as phospholipids. Preparation of microspheres is well known in the art and described in the pertinent texts and literature. Various additives, known to those skilled in the art, may be included in the topical formulations. For example, solvents, including relatively small amounts of alcohol, may be used to solubilize certain formulation components. In embodiment, the formulation includes a suitable enhancer, e.g., but are not limited to, ethers such as diethylene glycol monoethyl ether (available commercially as TRANSCUTOL™) and di ethylene glycol monomethyl ether; surfactants such as sodium laurate, sodium lauryl sulfate, cetyltrimethylammonium bromide, benzalkonium chloride, Poloxamer (231, 182, 184), Tween (20, 40, 60, 80), and lecithin (U.S. Patent No. 4,783,450); alcohols such as ethanol, propanol, octanol, benzyl alcohol, and the like; polyethylene glycol and esters thereof such as polyethylene glycol monolaurate (PEGML); amides and other nitrogenous compounds such as urea, dimethylacetamide (DMA), dimethylformamide (DMF), 2-pyrrolidone, l-methyl-2-pyrrolidone, ethanolamine, diethanolamine, and triethanolamine; terpenes; alkanones; and organic acids, particularly citric acid and succinic acid. AZONETM and sulfoxides such as DMSO and CIO MSO may also be used.

[0116] The present formulations may also include conventional additives such as opacifiers, antioxidants, fragrance, colorant, gelling agents, thickening agents, stabilizers, surfactants, and the like. Other agents may also be added, such as antimicrobial agents, to prevent spoilage upon storage, i.e., to inhibit growth of microbes such as yeasts and molds. Suitable antimicrobial agents are typically selected from the group consisting of the methyl and propyl esters of / ?-hydroxybenzoic acid (i.e., methyl and propyl paraben), sodium benzoate, sorbic acid, imidurea, and combinations thereof.

[0117] The formulations may also contain irritation-mitigating additives to minimize or eliminate the possibility of skin irritation or skin damage resulting from the pharmacologically active base or other components of the composition. Suitable irritation-mitigating additives include, for example: a-tocopherol; monoamine oxidase inhibitors, particularly phenyl alcohols such as 2 -phenyl- 1 -ethanol; glycerin; salicylic acids and salicylates; ascorbic acids and ascorbates; ionophores such as monensin; amphiphilic amines; ammonium chloride; N- acetylcysteine; cis-urocanic acid; capsaicin; and chloroquine. The irritant-mitigating additive, if present, may be incorporated into the present formulations at a concentration effective to mitigate irritation or skin damage, typically representing not more than about 20 wt. %, more typically not more than about 5 wt. %, of the composition.

[0118] Amounts of a disclosed compound as described herein in a formulation may vary according to factors such as the disease state, age, sex, and weight of the individual. Dosage regimens may be adjusted to provide the optimum therapeutic response. For example, a single bolus may be administered, several divided doses may be administered over time or the dose may be proportionally reduced or increased as indicated by the exigencies of the therapeutic situation. It is especially advantageous to formulate parenteral compositions in dosage unit form for ease of administration and uniformity of dosage. Dosage unit form as used herein refers to physically discrete units suited as unitary dosages for the mammalian subjects to be treated; each unit containing a predetermined quantity of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier.

[0119] The specification for the dosage unit forms are dictated by and directly dependent on (a) the unique characteristics of the compound selected and the particular therapeutic effect to be achieved, and (b) the limitations inherent in the art of compounding such an active compound for the treatment of sensitivity in individuals.

[0120] Therapeutic compositions typically must be sterile and stable under the conditions of manufacture and storage. The composition can be formulated as a solution, microemulsion, liposome, or other ordered structure suitable to high drug concentration. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof. The proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants. In many cases, it will be preferable to include isotonic agents, for example, sugars, polyalcohols such as mannitol, sorbitol, or sodium chloride in the composition.

[0121] The compounds can be administered in a time release formulation, for example in a composition which includes a slow release polymer. The compounds can be prepared with carriers that will protect the compound against rapid release, such as a controlled release formulation, including implants and microencapsulated delivery systems. Biodegradable, biocompatible polymers can be used, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, polylactic acid and polylactic, polyglycolic copolymers (PLG). Many methods for the preparation of such formulations are generally known to those skilled in the art.

[0122] In some embodiments, a compound can be formulated with one or more additional compounds that enhance the solubility of the compound. In certain embodiments, pharmaceutical compositions described herein can be administered in combination with one or more additional therapeutic agents to treat a disorder described herein. Methods of Use and Treatment

[0123] Disclosed compounds can be used in methods of treating or preventing a disease or disorder, e.g., a skin cancer, in a subject in need thereof. Also provided herein are compounds that can be used in methods of preventing or reducing the severity of a pre-cancerous skin lesion. In particular, in certain embodiments, the disclosure provides a method of treating the below medical indications comprising administering to a subject in need thereof a therapeutically effective amount of a compound described herein, such as a compound of Formula (I), or a pharmaceutically acceptable salt thereof. The disclosure also provides a compound described herein, such as a compound of Formula (I), or a pharmaceutically acceptable salt thereof, for use in treating the below medical indications.

[0124] The skin cancers or pre-cancerous skin lesions described herein may result from exposure to ultraviolet (UV) radiation. Most UV exposure in humans is due to sun exposure; however, other sources, such as tanning beds and welding torches, also provide UV exposure. There are three types of UV radiation: UVA, UVB, and UVC. UVA radiation has been linked to long-term skin damage (e.g., wrinkles) and UVC radiation reacts with ozone and does not reach the ground. UVB radiation is most commonly associated with skin cancer; UVB rays cause sunburn and can damage DNA in skin cells directly (e.g., mutations in p53). In some embodiments, the skin cancers or pre-cancerous skin lesions result from UV exposure, such as UVB exposure.

[0125] In some embodiments, the skin cancer is squamous cell skin cancer. In certain embodiments, the skin cancer is cutaneous squamous cell carcinoma. Squamous cell carcinoma (SCC) is an invasive malignant neoplasm of epidermal keratinocytes and is usually related to ultraviolet radiation exposure.

[0126] In some embodiments, the skin cancer is basal cell carcinoma. Basal cell carcinoma (BCC) is an autosomal dominant disorder that results from mutations in the p53 and / or PTCH genes. UV radiation is the most important risk factor in the development of BCC.

[0127] In some embodiments, the skin cancer is Merkel cell carcinoma. Merkel cell carcinoma (MCC) is a rare aggressive neuroendocrine carcinoma located between the dermal- epidermal junction. MCC is caused by chronic exposure to ultraviolet radiation.

[0128] In certain embodiments, the skin cancer is melanoma.

[0129] In some embodiments, the skin cancer is unresectable skin cancer. In some embodiments, the skin cancer is locally advanced cutaneous squamous cell carcinoma.

[0130] In certain embodiments, the skin cancer is metastatic skin cancer.

[0131] In some embodiments, the skin cancer is refractory skin cancer.

[0132] In some embodiments, the skin cancer is relapsed skin cancer.

[0133] In certain embodiments, the skin cancer is advanced.

[0134] In some embodiments, a pre-cancerous skin lesion is actinic keratosis (AK), which may turn into squamous cell carcinoma if left untreated. Actinic keratosis is caused by chronic exposure to ultraviolet radiation. It is also referred to as solar keratosis and senile keratosis.

[0135] Combination Therapy

[0136] The present disclosure also provides a combination therapy, which includes the administration of a compound described herein, e.g., a compound of Formula (I) or related compound described herein and a second therapeutic agent as part of a specific treatment regimen intended to provide the beneficial effect from the co-action of these therapeutic agents. The beneficial effect of the combination may include pharmacokinetic or pharmacodynamic co-action resulting from the combination of therapeutic agents.

[0137] In some embodiments, a combination comprising a compound described herein or a pharmaceutically acceptable salt thereof and octatrienoic acid is provided.

[0138] In certain embodiments, a contemplated method of treating a skin cancer or a pre- cancerous skin lesion in a patient in need thereof may further comprise administering to the subject a therapeutically effective amount of a second therapeutic agent. In other embodiments, the second therapeutic agent is octatrienoic acid.

[0139] The compound described herein and a second therapeutic agent can be administered simultaneously (e.g., at the same time), in the same or in separate compositions, or sequentially. Sequential administration refers to administration of one compound or therapy before administration of another compound or therapy.

[0140] EXAMPLES

[0141] The compounds of Formula (I) as disclosed herein, as well as their pharmaceutically acceptable salts, can be prepared by methods known from the literature.

[0142] The following abbreviations may be used herein and have the indicated definitions: A. SYNTHESIS OF COMPOUNDS

[0143] EXAMPLE 1: Synthesis of Compounds 1-7

[0144] 1. Synthesis of Compound 1

[0145] Synthesis of Compound 1 starts from compound A-l. Compound A-3 was prepared by treating A-2 with KHMDS. Compound A-3 on reaction with KOH afforded compound Compound 1.

[0146] Reagents were purchased from commercial sources and were used as received.

[0147] Preparation of compound A-3

[0148] To a stirred solution of A-2 (0.485 g, 2.288 mmol), 18-crown-6 (2.749 g, 10.41mmol) in THF (5 mL) was charged KHMDS (1 M in THF, 2.082 mL, 2.082 mmol) at 0 °C. The reaction mixture was stirred for 20 min at 0 °C and was cooled to -20 °C. Compound A-l (0.200 g, 2.082 mmol) was added slowly to the reaction mixture at -20 °C. The reaction mixture was stirred for 18 h at room temperature. Upon reaction completion, the reaction mixture was quenched with HC1 (2 N, 10 mL) and extracted with EtOAc (2x25 mL). Combined organic layer was washed with water (2x 10 mL) followed by brine solution (10 mL). The organic layer was dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure.

[0149] The crude residue was purified initially by preparative TLC using CH2Q2: CH3OH (95:5) as eluent. The resulted impure compound was further purified by preparative HPLC to afford compound A-3 (0.010 g, 2.6%) as yellow liquid.

[0150] Preparation of Compound 1

[0151] To a stirred solution of A-3 (0.010 g, 0.059 mmol) in a mixture of EtOH / H2O (1 : 1 / 2 mL) was charged KOH (0.003 g, 0.054 mmol) at room temperature. The reaction mixture was stirred for 3 h at room temperature. Upon reaction completion, the reaction mixture was concentrated under reduced pressure and the residue was acidified with 2 N HC1 (pH ~ 2) and extracted with EtOAc (3x 10 mL). Combined organic layer was dried with anhydrous Na2SO4, filtered and concentrated under reduced pressure. Crude material, consisting of two isomers (ratio 7:3), were purified by preparative HPLC to afford single isomer of Compound 1 (0.003 g, 33%) as a light yellow solid.

[0152] Analytical Report: Purification of Compound 1 by preparative HPLC Analytical Method development conditions for the purification of Compound 1

[0153] Method conditions for the purification of Compound 1

[0154] }HNMR (400 MHz, CDChy. 5 6.88 (d, J= 10.4 Hz, 1H), 6.51-6.48 (m, 2H), 6.23-6.17 (m, 1H), 5.97-5.91 (m, 1H), 3.76 (s, 3H), 1.85-1.82 (m, 3H); ESI (m / z) [C9H12O3 - H]’ 167

[0155] 2. Synthesis of Compound 2

[0156] B-l in tetrahydrofuran (THF) was reacted with freshly prepared Lithium diisopropylamide (LDA), TMSC1 and ethyl formate at -78°C to afford B-2. B-2 in tetrahydrofuran (THF) was reacted with freshly prepared Lithium diisopropylamide (LDA) and acetaldehyde at -78°C to afford B-3. B-3 in tetrahydrofuran (THF) treated with a solution of triethyl phosphonoacetate (B-4) and sodium hydride (NaH) to afford B-5. Ester hydrolysis of B-5 was done using potassium hydroxide (KOH) to afford Compound 2.

[0157] All reagents and solvents were received from commercial suppliers.

[0158] HPLC Method for Compound 2

[0159] Preparation of B-2

[0160] To a stirred solution of diisopropyl amine (2.4 mL, 17.2 mmol) in tetrahydrofuran (10 mL) at -78 °C, w-BuLi (6.24 mL, 15.6 mmol) was added dropwise and stirred at same temperature (-78 °C) for 1 hour. After 1 h a solution of B-1 (1.00 g, 5.20 mmol) in tetrahydrofuran (5 mL) at -78 °C was added to freshly prepared LDA solution, stirred for 10 min., followed by drop wise addition of TMSCI (0.7 mL, 5.46 mmol) in tetrahydrofuran (5.0 mL), the reaction mixture turned yellow. After 15 min., a solution of ethyl formate (0.8 mL, 10.4 mmol) in tetrahydrofuran (5 mL) was added dropwise and the resulting reaction mixture stirred for 1 h at -78°C. Progress of reaction was monitored by TLC, after Ih; the reaction mixture was allowed to come at 0 °C, quenched with saturated ammonium chloride (10 mL). The reaction mixture was again diluted with H2O (20.0 mL), extracted with ethyl acetate (EtOAc) (20.0 mL x 2); combined organic layers were washed with brine (20.0 mL), dried over anhydrous sodium sulfate (ISfeSCh) and concentrated under reduced pressure to get crude material. The obtained crude material was purified by combi flash chromatography eluted with 5% methanol in dichloromethane to afford B-2 (0.42 g, 37%) as a colourless liquid.

[0161] Preparation of B-3

[0162] To a stirred solution of diisopropyl amine (4.4 mL, 31.5 mmol) in tetrahydrofuran (20 mL) at -78°C, w-BuLi (11.5 mL, 28.7 mmol) was added dropwise and stirred at same temperature (-78 °C) for 1 hour. A solution of B-2 (2.10 g, 9.55 mmol) in tetrahydrofuran (10 mL) at -78°C was charged to the freshly prepared LDA solution and stirred for 30 min. After 30 min, a solution of acetaldehyde (0.84 g, 19.1 mmol) in tetrahydrofuran (5.0 mL) was added to above reaction mixture and stirred for same temperature for 2 hours. The progress of reaction was monitored by TLC. After 2 hour, reaction mixture was allowed to come at 0 °C, quenched with saturated ammonium chloride (10 mL), followed by dilution with H2O (20.0 mL). An aqueous layer extracted with methyl tert-butyl ether (MTBE) (20.0 mL x 2); combined organic layers were washed with brine (10.0 mL), dried over anhydrous sodium sulfate (ISfeSCh) and concentrated under reduced pressure at 0°C to afford B-3 (1.08 g), crude as a yellow liquid. The obtained crude material was taken for next step without further purification.

[0163] Preparation of B-5

[0164] To a suspension of NaH (0.57 g, 14.2 mmol) in THF (10.0 mL), under argon atmosphere triethylphophonoacetate B-4 (3.18 g, 14.2 mmol) was added at 0 °C and stirred for 20 min. After 20 min. reaction mixture turned to clear solution, then, a solution of B-3 (1.04 g, 9.45 mmol) in THF (10.0 mL) were added drop wise to above reaction mixture over a period of 10 min. After addition completed, resulting reaction mixture was stir for another 30 min at 0 °C. The progress of reaction was monitored by TLC. After 30 min. reaction mixture was warmed to room temperature for 2 h, quenched with ice cold saturated solution of ammonium chloride (NH4CI) (10.0 mL), diluted with H2O (10.0 mL). An aqueous layer extracted with methyl tert-butyl ether (MTBE) (10.0 mL x 2), combined organic layers were washed with brine (10.0 mL), dried over anhydrous sodium sulfate (ISfeSCh) and concentrated under reduced pressure to obtain crude material. The obtained crude material was purified by combi flash chromatography eluted with 20% MTBE in hexanes to afford B-5 [0.12 g, 7% (over 2 steps)] as colourless liquid. Preparation of Compound 2

[0165] To a solution of B-5 [2.80 g, 15.6 mmol] in mixture THF: MeOH: H2O (20.0 mL: 20.0 mL: 10.0 mL) was added KOH (1.74 g, 31.2 mmol) and resulting reaction mixture was stirred at room temperature for 18 h. The progress of reaction was monitored by TLC. Upon the completion of reaction, excess of solvent was evaporated under reduced pressure left behind viscous mass, which was diluted with H2O (25.0 mL), pH was adjusted to ~3 (using 2 N aqueous HC1) and extracted with methyl tert-butyl ether (MTBE) (50.0 mL x 2). The combined organic layers were washed with brine (25.0 mL), dried over anhydrous Na2SO4, concentrated under reduced pressure to obtain crude material, which was recrystallized using 80% MTBE in hexanes to afford Compound 2 (0.130 g, 38%) as a pale-yellow solid.

[0166] !HNMR (400 MHz, DMSO-d65 12.5 (s, 1H), 7.21 (d, J= 15.5 Hz, 1H), 6.54-6.43 (m, 2H), 6.05-5.94 (m, 1H), 5.78 (d, J= 15.5 Hz, 1H), 1.86-1.80 (m, 6H);13CNMR (100 MHz, DMSO- d6)-. 5 167.67, 148.44, 138.33, 135.06, 130.71, 127.94, 116.96, 18.46, 12.05; ESI(m / z) [C9H12O2 + H]+153.

[0167] 3. Synthesis of Compound 3

[0168] Synthesis of Compound 3 was achieved from commercially available C-l. C-l undergoes Suzuki coupling with C-2 in presence of potassium carbonate (K2CO3) and XPhos- Pd-G2 to afford C-3. Ester hydrolysis of C-3 was done using lithium hydroxide monohydrate (LiOH»H2O) to afford Compound 3.

[0169] All reagents and solvents were used as received from commercial suppliers.

[0170] HPLC Method for Compound 3

[0171] Preparation of C-3

[0172] To a stirred solution of C-l (0.2 g, 0.975 mmol) in tetrahydrofuran: water (THF / H2O) was added potassium carbonate (0.269 g, 1.95mmol) and resulting reaction mixture was degassed with argon for 10 min followed by addition of XPhos-Pd-G2 (0.076g, 0.09 mmol) and C-2 (0.21 g, 1.17 mmol) at room temperature. After addition completed, resulting reaction was sealed in argon atmosphere and heated at 100 °C for 16 h. The reaction mixture was filtered through celite pad and filtrate was evaporated to obtain crude material. The obtained crude material was purified by combi flash chromatography, eluted with 50% EtOAc in hexanes to afford C-3 (0.03 g, 17%) as a pale-yellow liquid.

[0173] Preparation of Compound 3

[0174] To a stirred solution of C-3 (0.12 g, 0.6 mmol) in mixture of THF / H2O (3.0 mL: 0.5 mL), LiOH»H2O (0.036 g, 0.86 mmol) was added and stirred for 16 h at room temperature. Upon completion of reaction, excess of organic solvent was evaporated under reduced pressure left behind viscous mass, which was diluted with H2O (10.0 mL). pH of an aqueous layer was adjusted to ~4 using HC1 (2 N aqueous), extracted with EtOAc (20.0 mL x 2). The combined organic layers were washed with brine (20.0 mL), dried over anhydrous Na2SO4 and concentrated under reduced pressure to obtain crude material. The obtained crude material was purified by prep-HPLC eluted with CH3CN: Water to afford Compound 3 (0.040 g, 33%) as an off white solid.

[0175] }HNMR (400 MHz, DMSO-d6. 5 12.9 (bars, 1H), 7.65 (d, J= 0.8 Hz, 1H), 7.04 (s, 1H), 5.58-

[0176] 5.46 (m, 2H), 3.10 (d, J= 5.2 Hz, 2H), 1.64 (d, J= 4.8 Hz, 3H); ESI (m / z) [C9H10O3 + H]+167.2.

[0177] 4. Synthesis of Compound 4

[0178] Synthesis of Compound 4 was achieved from commercially available D-l and D-2. D- 1 coupled with amine (D-2) using EDC HC1 and HOBt condition to afford D-3. D-3 in dry toluene was treated with Ru(CO)HCl(PPh3)3 to afford D-4. Ester hydrolysis of D-4 was done using lithium hydroxide monohydrate (LiOH»H2O) to afford Compound 4.

[0179] D-4 Compound 4

[0180] All reagents and solvents were used as received from commercial suppliers.

[0181] HPLC Method for Compound 4

[0182] Preparation of D-3

[0183] To a stirred solution of D-l (0.5 g, 3.84 mmol) in THF were added triethyl amine (0.5ml, 3.84 mmol) followed by HOBt (1.03 g, 7.68 mmol), EDC HC1 (0.75 g, 3.95 mmol) and D-2 (0.27 g, 3.84 mmol) at 0 °C. After addition completed, resulting reaction mixture was stirred at room temperature for 1 h. After 1 h, reaction mixture was diluted with water (EEO) (50.0 mL) and extracted with ethyl acetate (EtOAc) (50.0 mL x 3). The combined organic layers were washed with brine (25.0 mL), dried over anhydrous sodium sulfate (ISfeSCL) and concentrated under reduced pressure to obtain crude material. The obtained crude material was purified by combi flash chromatography eluted with 50% EtOAc in hexanes to afford impure D-3 (0.59g, 71%) as a colourless liquid, which was taken to next step without any further purification.

[0184] Preparation of D-4

[0185] To a stirred solution of compound, D-3 (1.5g, 8.19 mmol) in dry Toluene (15 mL), Ru(CO)HCl(PPh3)3 (0.78 g, 0.81 mmol) was charged at room temperature, sealed and resulting reaction mixture was heated at 120 °C for 2 h. The reaction mixture was filtered through celite pad; filtrate was evaporated under reduced pressure to obtain crude material. The obtained crude material was purified by combi flash chromatography eluted with 50% EtOAc in hexanes to afford D-4 (0.18 g, 12%) as a colourless liquid.

[0186] Preparation of Compound 4 To a stirred solution of D-4 (0.4 g, 2.17 mmol) in mixture of THF / H2O (3.0 mL:0.5 mL), lithium monohydrate LiOH»H2O (0.118 g, 2.82 mmol) was added and resulting reaction mixture was stirred at room temperature for 3 h. The progress of reaction was monitored by TLC. Upon completion of reaction, excess of organic solvent was evaporated under reduced pressure left behind viscous mass, which was diluted with H2O (10.0 mL). pH of an aqueous layer was adjusted to ~4 using HC1 (2 N aqueous), extracted with EtOAc (20.0 mL x 2). The combined organic layers were washed with brine (20.0 mL), dried over anhydrous ISfeSCh and concentrated under reduced pressure to obtain crude material. The obtained crude material was purified by combi flash chromatography eluted with 20% EtOAc in hexanes to afford Compound 4 (0.20 g, 54%) as an off white solid.

[0187] 1H NMR (400 MHz, DMSO-d6)'. 5 12.9 (brs, 1H), 7.13 (d, J= 15.2 Hz, 1H), 6.55 (d, J= 15.6 Hz, 1H), 6.28 (dd, J = 7.6 Hz, 1H), 5.65-5.63 (m, 1H), 2.97 (s, 3H), 1.48 (dd, J= 7.2 Hz, 3H); ESI (m / z) [C8HnNO3+ H]+170.1.

[0188] 5. Synthesis of Compound 5

[0189] Synthesis of Compound 5 was achieved from commercially available E-l. E-l in chloroform (CHCI3) was reacted with AICI3 and bromine (E ) to afford E-2. E-2 in tetrahydrofuran (THF) was reacted with w-BuLi (2.5M in hexane) to afford E-3. E-3 in tetrahydrofuran; Water (THF / H2O) was reacted with potassium carbonate (K2CO3) and X-Phos-

[0190] Pd-G2 was charged to E-4 to afford E-5. Ester hydrolysis of E-5 was done using lithium hydroxide monohydrate (LiOH»H2O) to afford Compound 5.

[0191] All reagents and solvents were used as received from commercial suppliers. HPLC Method for Compound 5

[0192] Preparation of E-2

[0193] To a solution of E-l (5 g, 35.6 mmol) in chloroform (CHCI3) (50 mL) at 0 °C, AICI3 (10.2 g, 35.6 mmol) followed by Bromine (Br2) (8.5 g, 53.5 mmol) were added. After addition completed, resulting reaction mixture was stirred at room temperature for 16 h. The progress of reaction was monitored by TLC. After 16 h, resulting reaction mixture was quenched with saturated hypo solution (50.0 mL), an aqueous layer extracted with ethyl acetate (EtOAc) (50.0 mL x 3). The combined organic layers were washed with brine (25.0 mL), and dried over anhydrous sodium sulfate (ISfeSCh) and concentrated under reduced pressure to obtain crude material. The obtained crude material was purified by combi flash chromatography (by using gold column), eluted with 10% EtOAc in hexanes to afford E-2 (2.0 g, 20%) as an off white solid.

[0194] Preparation of E-3 To a solution of E-2 (0.6 g, 0.2 mmol) in tetrahydrofuran (THF) (60.0 mL) at -78 °C under argon atmosphere, w-BuLi (2.5M in hexane) (0.8 ml, 0.22 mmol) was added drop-wise (over 5 min) and stirred for 10 min at same temperature. After 10 min. resulting reaction mixture was allowed to room temperature and quenched with ice cold solution of ammonium chloride (NH4CI) (25.0 mL). The reaction mixture was diluted with water (H2O) (20.0 mL), an aqueous layer extracted with ethyl acetate (EtOAc) (20.0 mL x 3). The combined organic layers were washed with brine (25.0 mL), dried over anhydrous sodium sulfate (ISfeSCU) and concentrated under reduced pressure to obtain crude material. The obtained crude material was purified by combi flash chromatography eluted with 50% EtOAc in hexanes to afford compound E-3. (0.12g, 27%) as an off white solid.

[0195] Preparation of E-5

[0196] To a solution of E-3 (0.34 g, 1.56 mmol) in mixture of tetrahydrofuran: water (THF / H2O) (5 mL: 1 mL), potassium carbonate (K2CO3) (0.432 g, 3.132 mmol) was added and resulting reaction mixture was degassed with argon for 10 min, followed by addition of XPhos- Pd-G2 (0.123g, 0.156 mmol) and E-4 (0.313 g, 1.72 mmol) at room temperature. After addition completed, resulting reaction mixture was sealed and heated at 100 °C, stirred for 16 h. The reaction mixture was filtered through celite pad; filtrate was evaporated under reduced pressure to obtain crude material. The obtained crude material was purified by combi flash chromatography, eluted with 50% EtOAc in hexanes to afford E-5 (0.13 g, 43%) as a pale-yellow liquid.

[0197] Preparation of Compound 5

[0198] Compound E-5 (0.150 g, 0.77 mmol) in mixture of THF / H2O (3.0 mL: 0.5 mL) was added lithium monohydrate LiOH»H2O (0.081g, 1.93 mmol) and resulting reaction mixture was stirred at room temperature for 16 h. Upon completion of reaction, excess of organic solvent was evaporated under reduced pressure left behind viscous mass, which was diluted with H2O (10.0 mL), pH of an aqueous layer was adjusted to ~4 using HC1 (2 N aqueous), extracted with EtOAc (20.0 mL x 2). The combined organic layers were washed with brine (20.0 mL), dried over anhydrous Na2SO4 and concentrated under reduced pressure to obtain crude material. The obtained crude material was purified by preparative HPLC, eluted with CH3CN: water to afford Compound 5 (0.040 g, 28.7%) as an off white solid.

[0199] !HNMR (400 MHz, DMSO-d6)'. 5 12.8 (brs, 1H), 7.53 (s, 1H), 5.53-5.43 (m, 2H), 3.05 (d, J = 4.4 Hz, 2H), 2.18 (s, 3H), 1.63 (d, J= 4.8 Hz, 3H); ESI (m / z) [C10H12O3 + H]+181.

[0200] 6. Synthesis of Compound 6

[0201] Synthesis of Compound 6 was achieved from commercially available F-l and F-2. F-l was coupled with amine (F-2) using HATU and DIPEA to afford F-3. Isomerization of F-3 with Ru(CO)HCl(PPh3)3 in dry toluene afforded F-4. Ester hydrolysis of F-4 was achieved using lithium hydroxide monohydrate (LiOH»H2O) to afford Compound 6. F-2

[0202] HATU, DIPEA

[0203] HOOC-^-'C°°MeDMF, rt, 6 h

[0204] Ru(CO)HCI(PPh3)3

[0205] Toluene

[0206] 120 °C, 2 h Compound 6

[0207] All reagents and solvents were used as received from commercial suppliers.

[0208] HPLC Analytical Method for Compound 6

[0209] Column Eclipse plus C18, 100 x 4.6 mm, 3.5 pm

[0210] Mobile Phase A 0.05%TFA in WATER: ACN (95:5) (%v / v)

[0211] Mobile Phase B 0.05%TFA in ACN

[0212] Gradient

[0213] Preparation of F-3

[0214] To a stirred solution of F-l (1.50 g, 11.53 mmol) and F-2 (0.79 g, 13.84 mmol) in dimethylformamide (DMF, 20.0 mL) was added diisopropylethylamine (DIPEA, 3.96 mL, 23.06 mmol) followed by l-[Bis(dimethylamino)methylene]-lH-l,2,3-triazolo[4,5-b]pyridinium 3- oxide hexafluorophosphate (HATU, 6.57 g, 17.29 mmol) at room temperature. The resulting reaction mixture was stirred at room temperature for 6 h, then the reaction was monitored by TLC. After completion of the reaction, the mixture was diluted with water (50.0 mL) and extracted with ethyl acetate (50.0 mL x 3). The combined organic layers were washed with brine (50.0 mL), dried over anhydrous sodium sulfate, filtered the drying agent. The filtrate was evaporated under reduced pressure to obtain crude material. The crude material was purified by silica gel combi flash chromatography by eluting with 80% EtOAc in hexane. The pure column fractions were evaporated to afford F-3 (1.60 g, 82%) as an off-white solid.

[0215] !HNMR (400 MHz, DMS0-d6): 5 8.68 (s, 1H), 7.05 (d, J= 15.2 Hz, 1H), 6.60 (d, J= 15.60 Hz, 1H), 5.87-5.78 (m, 1H), 5.19-5.08 (m, 2H), 3.83-3.80 (m, 2H), 3.73 (s, 3H); S E57) m / z 170.09 [CsHnNCh + H]+.

[0216] Preparation of F-4

[0217] To a stirred solution of F-3 (1.60 g, 9.46 mmol) in dry toluene (20.0 mL) was added Ru(CO)HCl(PPh3)3 (0.90 g, 0.94 mmol) at room temperature, sealed and resulting reaction mixture was heated at 120 °C for 2 h. The reaction mixture was filtered through celite pad; the filtrate was evaporated under reduced pressure to obtain crude material. The crude material was purified by silica gel combi flash chromatography eluted with 20% EtOAc in hexane. The pure column fractions were evaporated to afford F-4 (0.45 g, 28%) as an off-white solid.

[0218] !HNMR (400 MHz, DMSO-d6): 5 10.31 (d, J= 10.00 Hz, 1H), 7.01 (d, J= 15.60 Hz, 1H), 6.72- 6.68 (m, 1H), 6.64 (d, J = 15.20 Hz, 1H), 5.37-5.28 (m, 1H), 3.73 (s, 3H), 1.66 (dd, J = 1.60, 6.80 Hz, 3H). MS (ESI) m / z 170.12 [C8HnNO3+ H]+.

[0219] Preparation of Compound 6

[0220] To a stirred solution of F-4 (0.45 g, 2.66 mmol) in mixture of THF / H2O (10.0 mL / 2.0 mL) was added lithium hydroxide monohydrate (0.13 g, 3.19 mmol) at 0 °C. The resulting reaction mixture was stirred at room temperature for 3 h. The progress of the reaction was monitored by TLC. After completion of the reaction, concentrated under reduced pressure to give crude material. The crude was diluted with H2O (10.0 mL) and washed with EtOAc (50.0 mL). The aqueous layer was adjusted to pH ~2 using IN aq. HC1. The solid was thronged out, filtered and the solid was dissolved in EtOAc (100.0 mL). The organic layers were dried over anhydrous Na2SO4 and evaporated under reduced pressure to obtain Compound 6 (0.30 g, 73%) as an off white solid.

[0221] !HNMR (400 MHz, DMSO-d6): 5 12.93 (brs, 1H), 10.27 (d, J= 10.0 Hz, 1H), 6.92 (d, J= 15.2 Hz, 1H), 6.72-6.66 (m, 1H), 6.57 (d, J= 16.40 Hz, 1H), 5.36-5.27 (m, 1H), 1.66 (dd, J =1.60, 6.80 Hz, 3H). MS (ESI) m / z 156.09 [C7H9NO3 + H]+. 7. Synthesis of Compound 7

[0222] Synthesis of Compound 7 was achieved from commercially available G-l. G-l was treated with isobutyl chloroformate (G-2) using EtsN to afford crude G-3 which was treated with NaBH4 to afford G-4. Oxidation of G-4 was done using Dess-Martin periodinane (DMP) to afford G-5. G-5 was treated with tert-Butyl propiolate using LDA to afford G-6. G-6 was treated with EtsN to afford G-7. G-7 was hydrolysed using TFA to afford Compound 7.

[0223] All reagents and solvents were used as received from commercial suppliers. HPLC analytical method for Compound 7 is identical to that for Compound 6. Note: Butylated hydroxytoluene (BHT) was added in small amounts during the reactions, purifications, even after isolation in each step for Compound 7 for stability purpose and to arrest polymerization.

[0224] Preparation of G-4

[0225] To a stirred solution of G-l (5.00 g, 51.02 mmol) in diethyl ether (500 mL) was added isobutyl chloroformate (G-2) (7.26 mL, 56.12 mmol) followed by EtsN (7.87 mL, 56.12 mmol) at 0 °C. The resulting reaction mixture was stirred at 0 °C for 30 min. The progress of the reaction monitored by TLC. The reaction mixture was diluted with water (100 mL). The organic layer was separated and dried over anhydrous sodium sulfate, decanted the organic layer and this solution (containing G-3) was added dropwise to a stirred solution of NaBEL (3.87 g, 102.4 mmol) in methanol (20.0 mL) at 0 °C. The reaction mixture was stirred at room temperature for 1 h, the progress of reaction monitored by TLC. After completion of reaction, the reaction mixture was quenched with aqueous NH4CI solution (200 mL) and extracted with diethyl ether (3 x 100 mL). The combined organic layer was washed with brine (50.0 mL), dried over anhydrous sodium sulfate, evaporated under reduced pressure at 0 °C to obtain G-4 (2.80 g, crude) as a yellow colour liquid.

[0226] !HNMR (400 MHz, CDCI3): 5 6.41-6.24 (m, 2H), 5.90-5.83 (m, 1H), 5.11 (dd, J= 0.80, 9.60 Hz, 1H), 4.79 (t, J= 17.20 Hz, 1H), 4.21 (d, J= 5.60 Hz, 2H).

[0227] Preparation of G-5

[0228] To a stirred solution of G-4 (2.30 g, 27.38 mmol) in dichloromethane (DCM, 50.0 mL) was added portionwise Dess-Martin periodinane (11.60 g, 27.38 mmol) at 0 °C. The reaction mixture was stirred at room temperature for 2 h, the progress of reaction monitored by TLC. After completion of reaction, the mixture was filtered through celite pad, washed with DCM (100 mL). The filtrate was washed with aqueous NaHCCL solution (100 mL), brine (50.0 mL), dried over anhydrous sodium sulfate, filtered. The filtrate was evaporated under reduced pressure at 0 °C to obtain crude material. The crude material was purified by silica gel column chromatography eluted with 50% n-pentane in DCM to afford G-5 (2.14 g, with DCM) as a yellow colour liquid.

[0229] !HNMR (400 MHz, CDCI3): 5 9.60 (d, J= 7.60 Hz, 1H), 7.13-7.07 (m, 1H), 6.55-6.65 (m, 1H), 6.15-6.21 (m, 1H), 5.77 (d, = 0.4 Hz, 1H), 5.73 (d, J = 0.4 Hz, 1H).

[0230] Preparation of G-6

[0231] To a stirred solution of tert-butyl propiolate (3.00 g, 23.80 mmol) in THF (50.0 mL) was added dropwise 2.0 M Lithium diisopropylamine (LDA) solution in THF (13.09 mL, 26.19 mmol) at -78 °C. The resulting reaction mixture was stirred at -78 °C for 10 min, then G-5 (2.14 g, 26.19 mmol) in THF (10.0 mL) was added dropwise and continued stirring at same temperature for 1 h, the progress of reaction monitored by TLC. After completion of reaction was quenched with aqueous NH4CI solution (100 mL) at same temperature, stirred for 10 min and extracted with EtOAc (3 x 50 mL), combined organic layer was washed with brine (50.0 mL), dried over anhydrous Na2SO4, filtered and evaporated under reduced pressure to obtain crude material. The crude material was purified by silica gel column chromatography eluted with 10% EtOAc in hexane. The pure column fractions were evaporated to afford G-6 (0.80 g,16%) as a yellow colour liquid.

[0232] !HNMR (400 MHz, CDCI3): 5 6.43-6.30 (m, 2H), 5.78 (dd, J= 6.00, 14.80 Hz, 1H), 5.35-5.31 (m, 1H) ,5.23-5.21 (m, 1H), 5.03 (t, J= 6.80 Hz, 1H), 2.16 (dd, J=1.20, 6.60 Hz, 1H), 1.50 (s, 9H).

[0233] Preparation of G-7 To a stirred solution of G-6 (0.40 g, 1.92 mmol) in acetonitrile (ACN, 10.0 mL) was added Et3N (0.26 mL, 1.92 mmol) at 0 °C, the resulting reaction mixture was stirred at room temperature for 1.5 h, the progress of reaction monitored by TLC. After completion of reaction, the reaction mixture was diluted with ice water (10.0 mL), extracted with EtOAc (3 x 50 mL). The combined organic layer was washed with IN aq. HC1 solution (10.0 mL), brine (50.0 mL), dried over anhydrous sodium sulfate, filtered and evaporated under reduced pressure to obtain crude material (G-7 and G-7A). The crude material was purified by silica gel column chromatography eluted with 7% EtOAc in hexane. The pure column fractions were evaporated to afford G-7 (0.15 g, 37%) as a yellow colour liquid.

[0234] !HNMR (400 MHz, CDCh): 5 7.33-7.29 (m, 1H), 7.24-7.20 (m, 1H), 6.68 (d, J= 15.60 Hz, 1H), 6.57-6.48 (m, 1H), 6.43 (d, J= 15.60 Hz, 1H), 5.76-5.72 (m, 1H), 5.63 (d, J= 9.60 Hz, 1H), 1.52 (s, 9H).

[0235] Preparation of Compound 7

[0236] To a stirred solution of G-7 (0.20 g, 0.96 mmol) in DCM (12.0 mL), was added dropwise trifluoroacetic acid (TFA, 1.00 mL) at 0 °C. The resulting reaction mixture was stirred at room temperature for 5 h and progress of reaction was monitored by TLC. After completion of reaction, excess of organic solvent was evaporated under reduced pressure to obtain crude material. The crude material was purified by prep-TLC by eluting with 40% EtOAc in hexane to afford Compound 7 (80.0 mg, 54%) as a yellow gummy solid.

[0237] !HNMR (400 MHz, DMSO-d6): 5 13.05 (brs, 1H), 7.43-7.36 (m, 1H), 7.30 (d, J = 15.60 Hz, 1H), 6.69 (d, J= 6.40 Hz, 1H), 6.65 (d, J= 7.20 Hz, 1H ), 6.62-6.57 (m, 1H), 5.89 (d, J= 16.8 Hz, 1H), 5.68 (d, J= 10.0 Hz, 1H); MS (ESI) m / z 151.17 [C8H8O3- H]’.

[0238] B. BIOLOGICAL ACTIVITY OF COMPOUNDS

[0239] Materials

[0240] Ml 54 defined medium, human keratinocyte growth supplements (HKGS), fetal bovine serum (FBS), L-glutamine, penicillin / streptomycin, trypsin / EDTA, and D-PBS were purchased from Invitrogen Technologies (Monza, Italy). DMEM basal medium was purchased from Euroclone (Milan, Italy). P-actin antibody (A5441) (1 : 10000), GAPDH antibody (G9545) (1 :5000), Catalase antibody (C0979) (1 : 1000), propidium iodide solution and 2’,7’- dichlorofluorescein diacetate (DCFH2-DA) were from Sigma-Aldrich (Milan, Italy). AurumTM Total RNA Mini kit, SYBR Green PCR Master Mix, Bradford reagent were from Bio-Rad (Milan, Italy). RevertAidTM First Strand cDNA synthesis kit was from Thermo Fisher Scientific (Monza, Italy). IL-6 and IL-8 ELISA kits were from Diaclone SAS (Besancon Cedex, France). The antibodies for p21 Wafl / Cipl (#2947) (1 : 1000), phospho-p44 / 42 MAPK (ERK1 / 2) (Thr202 / Tyr204) (#4370) (1 :2000), phospho-p38 MAP kinase (Thrl80 / Tyrl82) (#4511) (1 : 1000), p38 MAPK (#9212) (1 : 1000), PPARy (#2430) (1 : 1000), phospho-AKT (Ser473) (#4060) (1 :1000), AKT (#2920) (1 : 1000), phospho-histone H2A.X (#9718) (1 : 1000), cyclin DI (#2978) (1 : 1000), secondary anti-mouse IgG HRP-conjugated antibody (#7076) (1 :3000) and anti-rabbit IgG HRP-conjugated antibody (#7074) (1 :8000) were purchased from Cell Signaling (Danvers, MA, USA); anti-ERK2 (SC-1647) (1 : 1000), anti-Fibronectin (SC-8422) (1 : 1000) and anti-Filaggrin (SC-66192) (1 :250) were from Santa Cruz Biotechnology (CA, USA); anti-p53 (M7001) (1 : 1000) and E-cadherin (M3612) (1 : 1000) were purchased from DakoCytomation (Glostrup, Denmark); the antibodies for Cytokeratin 10 (ab76318) (1 :2000), Involucrin (ab53112) (1 :500), Loricrin (ab85679) (1 :500), and Vimentin (ab92547) (1 : 1000), were purchased from Abeam (Cambridge, UK). Amersharm ECL Western Blotting Detection Reagent was from GE Healthcare (Buckinghamshine, UK). RIPA lysis buffer, broad spectrum protease inhibitor cocktail, and broad spectrum phosphatase inhibitor cocktail were from Boster Biological Technology Co. (Pleasanton, CA, USA). Recombinant Human TGF-pi was purchased from PeproTech (Cranbury, NJ, USA). RNASE A was from Biobasic Canada Inc. (Ontario, Canada). The Amaxa® human keratinocyte Nucleofector kit was from Lonza (Basel, Switzerland).

[0241] Cell Cultures

[0242] Normal human keratinocytes (NHKs) were isolated from the neonatal foreskin following a previously described procedure (323 Kovacs, D. 2012). NHKs were maintained at 37 °C under 5% CO2 in the defined medium Ml 54 with HKGS, L-glutamine (2mM), and penicillin / streptomycin (100 pg / mL). NHKs were sub-cultivated once a week and the experiments were carried out in cells between passages 2 to 4. The A431 squamous carcinoma cell line was purchased from ATCC (USA) and was maintained in basal medium DMEM, supplemented with 10% FBS, L-glutamine (2mM), penicillin / streptomycin (100 pg / mL), in a humidified atmosphere containing 5% CO2 at 37 °C. Cell cultures were routinely tested for Mycoplasma infection. The Institute’s Research Ethics Committee (IFO) approval was obtained to collect samples of human material for research (Prot CE / 286 / 06, approved on 21 April 2006). The study was conducted according to the Declaration of Helsinki Principles. Patients gave written informed consent. For each experiment, at least three different donors were used. Cells were plated and 24 h later were stimulated with chemicals in fresh medium, in accordance with the experimental design. UVB irradiation of NHKs

[0243] NHKs were incubated in a medium without phenol-red and irradiated with UVB at the dose of 25 mJ / cm2. Control cells were treated identically but without UVB exposure. The BioSun irradiation apparatus (Vilbert Lourmat, Mame-la-Vallee, France) was employed. The UVB lamps emit ultraviolet rays between 280 and 320nm, with peak luminosity at 312nm. UVB lamps do have not UVC emission. UVB was supplied by a closely spaced array of two UVB lamps which delivered uniform irradiation at a distance of 10cm. Based on a programmable microprocessor, the Bio-Sun system constantly monitors UV light emission. The irradiation stops automatically when the received energy matches the programmed energy (range of measure: 0 to 9,999 J / cm2).

[0244] RNA Extraction and Quantitative Real-Time RT-PCR

[0245] Total RNA was isolated using the AurumTM Total RNA Mini kit, according to the manufacturer’s instructions. Total RNA samples were stored at -80 °C until use. Following DNAse I treatment, cDNA was synthesized using a mix of oligo-dT and random primers and RevertAidTM First Strand cDNA synthesis kit according to the manufacturer’s instructions. Quantitative real-time RT-PCR was performed in a total volume of 10 pL with SYBR Green PCR Master Mix and 200 nM concentration of each primer. Reactions were carried out in triplicates using a CFx96TM Real-Time System (Bio-Rad). Melt curve analysis was performed to confirm the specificity of the amplified products. Expression of mRNA (relative) was normalized to the expression of GAPDH mRNA by the change in the A cycle threshold (ACt) method and calculated based on 2Cl.

[0246] Western Blot Analysis

[0247] Cells were lysed in RIPA lysis buffer supplemented with a protease / phosphatase inhibitor cocktail, then sonicated. Total cell lysates were clarified by centrifugation at 12.000 rpm for 10 minutes at 4°C and then stored at -80°C until analysis. Following spectrophotometric protein measurement, equal amounts of protein were resolved on acrylamide SDS-PAGE and transferred onto nitrocellulose membrane (Amersham Biosciences, Milan, Italy). Protein transfer efficiency was checked with Ponceau S staining (Sigma-Aldrich, St Louis, MO, USA). Membranes were first washed with water, blocked with EveryBlot Blocking Buffer (Bio-Rad Laboratories Sri, Milan, Italy) for 10 min at room temperature, and then treated overnight at 4°C with primary antibodies (according to data sheet instructions). A secondary anti-mouse IgG HRP-conjugated antibody or anti-rabbit IgG HRP-conjugated antibody was used. Antibody complexes were visualized using enhanced chemiluminescence (ECL) substrate. A subsequent hybridization with anti-0 actin or anti-GAPDH was used as a loading control. Protein levels were quantified by measuring the optical densities of specific bands using the UVI-TEC Imaging System (Cambridge, UK). The control value was taken as one-fold in each case.

[0248] MTT assay

[0249] Cells treated with Octa (90 pM) or compound 1 (90 pM) for 72 h were then incubated with 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) (1 mg / mL) for 2 h at 37 °C, and lysed in dimethyl sulfoxide (DMSO). The absorbance at 570 nm was measured by a spectrophotometer DTX880 Multimode Detector (Beckman Coulter srl., Milano, Italy). The measurement was performed in triplicate for each sample.

[0250] Cell number analysis

[0251] Cells were treated with 2,4,6-octatrienoic acid (“Octa”) (90 pM) or compound 1 (90 pM) for 72 h and then detached by trypsinization. Cell number was evaluated by cell counting using the phase-contrast microscope Axiovert 40C (Zeiss, Milan, Italy). None of the employed chemicals determined positivity to the Trypan Blue exclusion assay test.

[0252] Flow cytometric analysis of cell cycle

[0253] After the treatment, cells were detached by trypsinization and fixed with ice-cold 70% ethanol solution overnight at 4°C for maximum resolution of cellular DNA. The samples were stained in PBS containing propidium iodide (50 pg / ml) and RNAse A (40 pg / ml). The DNA content of the cells was measured by MACSQuant Analyzer (Miltenyi Biotec GmbH; Bergisch Gladbach, Germany). A total of 104events for each sample were acquired. The measurement was performed in duplicate for each sample. Flow cytometric histograms were analysed by defining borders between pre-Gi, Gi, S, and G2 + M phase populations. MACSQuantify™ Software was used to analyse data.

[0254] Immunofluorescence Analysis

[0255] A431 cells were fixed either with 4% paraformaldehyde followed by 0.1% Triton X-100 to allow permeabilisation or with cold methanol at -20°C. Cells were then incubated with the following primary antibodies: anti-vimentin rabbit polyclonal antibody (1 :400) (Abeam), anti- fibronectin mouse monoclonal antibody (1 :400) (Santa Cruz Biotechnology), anti-E-cadherin monoclonal antibody (1 :200) (Dako). The primary antibodies were visualised using goat antirabbit Alexa Fluor 546 conjugate, goat anti-mouse Alexa Fluor 546, and goat anti-mouse Alexa Fluor 488 conjugate antibodies (1 :800) (ThermoFisher Scientific, Italy). Coverslips were mounted using ProLong Gold antifade reagent with DAPI (Invitrogen). The fluorescence signal was evaluated by recording stained images, using a CCD camera (Zeiss, Oberkochen, Germany). For vimentin and fibronectin stainings, the number of positive cells was measured by counting at least 150 cells randomly taken from 10 different microscopic fields and results are expressed as fold change respect with control. Quantitative analysis of E-cadherin fluorescence intensity was performed using the Zen 2.6 (blue edition) software (Zeiss) and the results are expressed as mean fluorescence intensity / cell ±SD.

[0256] Lipid extraction from A431 cells

[0257] Lipids were extracted from A431 cells accordingly with the Bligh & Dryer procedure with slight modifications (410 BLIGH, E G 1959). Briefly, lipids were extracted with chloroform: methanol (2: 1) (2 x 1 ml) after the addition of butylhydroxytoluene to prevent oxidation of oxygen-sensitive compounds. 75 pl of a mixture of deuterated standards (d6CH 4pM; dl7C16:0 8pM; d98TG48:0 2.4pM; d4PGD2 IpM; d4PGE2 IpM; d85HETE IpM) were added to control the analytical performance and to calculate the relative abundance of the lipid species detected. The organic layers were collected and evaporated under nitrogen. The dried lipid extract was dissolved in 75 pl of acetonitrile and stored at -80 °C until the analysis.

[0258] GC-MS lipid analysis of FFAs and CH

[0259] Free fatty acids (FFAs) and cholesterol (CH) were analyzed after direct silylation with N, O-bis-(trimethylsilyl)-trifluoroacetamide containing 1% trimethylchlorosilane as catalyst (Sigma-Aldrich, Milan, Italy). After 30 min at 60°C, the samples were analyzed with GC 7890A coupled to MS 5975 VL analyzer (Agilent Technologies, Santa Clara, CA, USA). The chromatographic separation was performed on an HP-5MS (Agilent Technologies, Santa Clara, CA, USA) capillary column (30m x 250 pm x 0,25 pm), using helium as the carrier gas. An oven temperature gradient from 80 °C to 200 °C at 8 °C / min and then to 250°C at 10°C / min was used. The injector and the GC-MS transfer lines were kept at 260°C and 280°C, respectively. Samples were analyzed in scan mode utilizing electron impact (El) mass spectrometry. The identity of the detected FFAs and CH was verified by the comparison with authentic standards and the match with library spectral data. The area of the peaks corresponding to C16:0, C16: l, C18:0, Cl 8: 1, and CH was integrated with the qualitative analysis software. The pmole amount of FFAs and CH were calculated against the added pmoles of d6CH and dl7C16:0, respectively.

[0260] In vivo study

[0261] Mice were cared for in accordance with the Principles of Laboratory Animal Care (National Institutes of Health publ. no. 85-23, revised 1985), and with national laws. The experimental protocols comply with the principles of (https: / / www.nc3rs.org.uk / arrive- guidelines) and were approved by the National Ethics Committee for Animal Experimentation of the Italian Ministry of Health. The mice were housed in single cages with wood-derived bedding material in a specific pathogen-free facility with a 12-h light / dark cycle under controlled temperatures (20-22 °C) and received water and food ad libitum. All experimental procedures conformed to protocols approved by the Regina Elena National Cancer Institute Animal Care and Use Committee and were performed in accordance with the Guide for the Care and Use of Laboratory Animals and the guidelines of the National Institutes of Health, according to the current National Legislation (Art. 31 D.lgs 26 / 2014, 4 March 2014). The animals used in the study were 6-8 weeks old MITO-Luc mice (349 Goeman, F. 2012) of both sexes, maintained on an FVB background.

[0262] Carcinogenesis mouse model and In Vivo Bioluminescence imaging (BLI) analysis

[0263] Skin papillomas were generated by local administration of 7,12- dimethylbenz(a)anthracene and the forbolic ester 12-O-tetradecanoylphorbol-13 acetate (DMBA-TPA) on the shaved ventral skin of MITO-Luc mice (348 Vantaggiato,C. 2016). Mice were treated with a single topical application of 860 pM DMBA (Sigma-Aldrich) in 0.2 ml acetone or with the solvent alone. One week after, 100 pM of TPA (Alexis Biochemicals, San Diego, CA) in 0.2 ml acetone or the solvent alone was topically applied twice weekly. After the appearance of papillomas, topical treatments with compound 1 (20 mg dissolved in 1 ml of DMSO and 4 ml glycerol) were performed five days weekly. Solvent alone was applied as placebo.

[0264] In vivo bioluminescence BLI represents a new approach to molecular imaging. It allows us to visualise internally generated light linked to specific physiological and / or pathological cellular processes in small living animals (407 Signore, A. 2010). In the MITO-Luc transgenic reporter mouse animal model, luciferase activity is under the control of the cyclin B2-dependent nuclear factor-Y (NF-Y) promoter and is consequently restricted to proliferating cells (403 Manni,I. 2016; 308 Manni,I. 2019; 405 Rizzi, N. 2015). Bioluminescence imaging (BLI) analysis was performed using the IVIS Lumina II equipped with the Living Image 2.20 software package for data quantification (PerkinElmer, Waltham, MA, USA) (349 Goeman, F. 2012). For in vivo imaging, mice were anesthetised, and D-luciferin (75 mg / kg body weight) (PerkinElmer) dissolved in phosphate-buffered saline (PBS) was administered i.p. Ten minutes later, quantification of light emission was performed in photons per second per square centimeter per steradian (photons / s / cm2 / sr). and visualised in a pseudocolor scaling. Time exposure ranged from 1 to 5 min, depending on light intensity.

[0265] Statistical Analysis

[0266] Data were represented as mean ± SD of three independent experiments using at least three different donors for primary cells. Statistical significance was assessed using paired Student’s t-test or ANOVA followed by Tukey’s multiple comparisons test using GraphPad Prism (GraphPad Software). The minimal level of significance was p < 0.05. Example 2: Effects of compound 1 on cellular antioxidant defence, DNA damage and inflammatory response in UVB-irradiated NHKs.

[0267] The treatment with compound 1 (10, 50, 90 pM) for 6 h significantly increased the mRNA expression level of CATALASE (CAT), HEME-OXIGENASE 1 (HO-1), and NAD(P)H QUINONE DEHYDROGENASE 1 (NQO1) (Fig. 1 A), three key enzymes involved in cellular defence against oxidative stress. The western blot analysis of catalase confirmed the gradual induction of the protein after 24 h of treatment (Fig. IB).

[0268] Then, whether compound 1 may exert a protective effect against oxidative stress and DNA damage induced by UVB exposure in keratinocytes was investigated. To this aim, NHKs were pre-treated with compound 1 (10, 50, 90 pM) for 24 h, exposed to UVB (25 mJ / cm2), and then allowed to recover. As expected, after 6 h, UVB alone caused a significant decrease in the mRNA level of CAT, HO-1, and NQO1 compared to the control, whereas compound 1 pre-treatment counteracted this effect (Fig. 1C). The western blot analysis of catalase protein expression at 24 h confirmed the mRNA data (Fig. ID).

[0269] Furthermore, the protein expression level of two DNA damage markers, p53 and phosphorylated histone H2A.X (y-H2AX), the expression of which is associated with the amount of doublestrand breaks after UV-radiation, were analyzed. NHKs pre-treated with compound 1 showed a reduced level of these UV-related genotoxicity markers compared to those stimulated with UVB alone, suggesting that also compound 1 exerted a protective effect on UV-induced damage (Fig. ID). The compound 1 dose of 90 pM proved to be the most effective and hence was selected for the subsequent experiments.

[0270] Example 3: Effects of compound 1 and Octa on viability, proliferation, and differentiation of A431 cells.

[0271] The effect on cell viability and the anti-proliferative / pro-differentiative potential of the two compounds on the human cutaneous squamous cell carcinoma (cSCC) A431 cells was investigated. The MTT assay indicated no significant effect of Octa and compound 1 after 72 h of treatment on the NAD(P)H-dependent cellular mitochondrial activity, an indicator of cell viability (Fig. 2F). Notably, a significant reduction in the number of A431 cells only after compound 1 treatment was observed (Fig. 2A). In compliance with these results, 48 h of treatment with compound 1 determined an increase of cells in the G0 / G1 phase of the cell cycle (Fig. 2B). To corroborate the compound 1 effect on proliferation slow down, the expression level of the cell cycle regulators p21WAFl / CIPl (p21) and cyclin DI was analyzed. Compound 1 significantly induced the expression of the cell cycle progression inhibitor p21 and down- regulated the level of the cell proliferation-associated protein cyclin DI (Fig. 2C). Moreover, as forNHKs (Fig. 3 A and 3B), compound 1 significantly increased both mRNA and protein levels of the different markers filaggrin (FLG), involucrin (IVL), and loricrin (LOR), whereas Octa was not able to modulate their expression (Fig. 2D and 2E, respectively).

[0272] Example 4: Compound 1 and Octa inhibit TGF-pi-induced EMT in A431 cells.

[0273] EMT plays a crucial role in promoting carcinoma progression and it is characterized by cancer cell transition from an epithelial differentiated state to a mesenchymal-like phenotype, more prone to invade the surrounding tissues.

[0274] To verify whether Octa and / or compound 1 were able to interfere with the EMT process in cutaneous squamous cell carcinoma, A431 cells were pre-treated with the compounds for 1 h and then exposed to the EMT inductor TGF-pi (15ng / ml) for 48 h. The mRNA expression level of N-CADHERIN (NCAD), E-CADHERIN (ECAD), SLUG, FIBRONECTIN, VIMENTIN, and MMP-2, as markers of EMT was evaluated. Pre-treatment with Octa, or compound 1, counteracted the TGF-p i -mediated upregulation of NCAD, FIBRONECTIN, VIMENTIN, and MMP2 but was ineffective in preventing the up-modulation of SLUG (Fig. 4A). The western blot analysis of vimentin and fibronectin confirmed the ability of Octa and even more of compound 1 to reduce the increase of protein expression promoted by the stimulation with TGF- i for 72h (Fig. 4B). E-cadherin transcript and protein levels were not affected by any of the treatments. In line with these results, qualitative and quantitative analyses of immunofluorescence staining for vimentin and fibronectin demonstrated that both compounds were effective in counteracting the increased expression of the EMT markers induced by TGF-pi stimulation (Figure 4C). No significant modification of E-cadherin expression was observed following the different treatments. However, the immunofluorescence analysis revealed the delocalization of the protein from the plasma membrane to the cytoplasm associated with a reduced cell-cell adhesion following TGF-pi, which was partially recovered by the treatment with compound 1 and Octa (Fig. 4C).

[0275] Example 5: Compound 1 and Octa counteract lipid metabolic reprogramming associated with TGF-pi-induced EMT in A431 cells

[0276] To confirm the ability of compound 1 and Octa to counteract the acquisition of invasive and metastatic potential of tumor cells, the effects of these compounds concerning the significant changes in lipid metabolism that characterize the EMT process were evaluated. To this end, A431 squamous carcinoma cells were pre-treated with Octa and compound 1 for 1 h and then exposed to TGF-pi (15ng / ml) for 24 h and 72 h to promote the EMT transition. The lipidomic profile of the A431 cell membranes was acquired and the monounsaturated / saturated fatty acids ratio (MUFAs / SFAs), which is positively correlated to the greater aggressiveness of cancer cells was determined. The data obtained demonstrated that Octa and compound 1 significantly reduce the A431 cell baseline level of the MUFAs / SFAs ratio after 72 h of treatment (Fig. 5 A), proving a beneficial effect of the molecules per se against the tumour. Moreover, Octa and compound 1 pre-treatment was effective in counteracting MUFA accumulation observed after 72 h of TGF- pi exposure. In particular, compound 1 reduced the desaturation index even below the A431 basal level, suggesting its higher efficacy compared to Octa in preventing EMT transition (Fig. 5A).

[0277] Considering that tumor cells can use free FAs (FFAs) as an energy supply by lipolysis for membrane biosynthesis or signalling processes, FFAs release was analysed in the cell medium. The ratio between C16: l / C16:0 and C18: l / C18:0 was reported as an index of SCD-1 activity and MUFA enrichment in the medium. TGF-pi treatment determined a significant release of palmitoleic acid (Cl 6: 1) and oleic acid (Cl 8: 1) and Octa and compound 1 effectively counteracted this effect (Fig. 5B and Fig. 5C).

[0278] The alteration in cholesterol homeostasis is another lipid metabolic reprogramming feature related to tumour initiation and progression. The higher cell cholesterol content associated with the EMT process contributes to lipid rafts domains and membrane biophysical properties modulation, fueling the maintenance of the mesenchymal state of tumoral cells. The determination of cholesterol levels demonstrated that compound 1 was able to significantly reduce the basal cholesterol content in A431 cells 72 h after the treatment, while Octa did not exert the same effect (Fig. 5D). On the other hand, both molecules successfully lowered the TGF-pi -induced increase in cholesterol content. In particular, compound 1 was more powerful than Octa in significantly counteracting the TGF-pi effect observed after 24 h and even more after 72 h of treatment (Fig. 5D). Otherwise, Octa pre-treatment almost neutralized A431 cholesterol increase, reaching statistical significance only after 72 h (Fig. 5D).

[0279] Altogether these data demonstrate a key role of the reprogramming of lipid metabolism in the EMT induced by TGF-pi in A431 cells and suggest that Octa or compound 1 protective role is associated with their ability to preserve the lipid composition of the cell membranes and counteract the release of bioactive lipids involved in the activation of EMT program. Example 6: Compound 1 counteracted DMBA-TPA effects in vivo

[0280] To demonstrate the in vivo relevance of the in vitro results, cutaneous papillomas was induced by DMBA-TPA treatments in the MITO-Luc mouse model. In this transgenic reporter mouse model, luciferase activity is under the control of a cyclin B2 promoter fragment whose activity is strictly dependent on cell proliferation. Since aberrant proliferation represents an early pre- neoplastic event and considering the superiority of compound 1 in several in vitro endpoints, the efficacy of this compound was investigated in reducing tumour mass in this murine model. Fig. 6A shows representative images of a MITO-luc mouse before induction of papillomas (preimaging), at the end of DMBA-TPA treatment (tO), and after 15 treatments (tl) with placebo or compound 1. As expected, induction of bioluminescence was detected in the treated sites after DMBA-TPA application and, of note, a significant decrease of photon emission after the treatment with the compound. On the contrary, an increase in bioluminescence was observed in placebo-treated lesions (Fig. 6B). Furthermore, the mRNA expression levels of the EMT markers NCAD, ECAD, SLUG, FIBRONECTIN, VIMENTIN, a-SMA, and 7 2 were evaluated in samples of mouse skin collected at the end of the treatments. Compound 1 was able to significantly reduce the expression of all these transcripts in comparison with the placebo-treated mice (Fig. 6C).

[0281] Summary / Analysis of Biological Data

[0282] Based on the above results, compound 1, and those fatty acid compounds closely resembling compound 1 as described herein, may represent potential pharmacological targets in the prevention or treatment of skin cancer. The present findings support the utility of compound 1 and Octa to prevent and treat keratinocyte-derived skin cancer. This is the first study, to the applicant’s knowledge, examining the ability of Octa as well as compound 1, a novel compound, to induce differentiation markers and to antagonise the TGF-01 -mediated changes associated with the EMT process in a human squamous carcinoma cell line. The in vitro data with compound 1 are in agreement with the literature reports on several malignant cell types, suggesting that compound 1 inhibits human carcinoma cell growth by regulating the expression of the cell cycle-associated proteins and inducing terminal differentiation. Interestingly, among Octa and compound 1, only compound 1 is able to inhibit cell growth and promote pro- differentiative effects. In compliance with data obtained in thyroid cancer cells and lung adenocarcinoma cells, the results on the A431 squamous cell carcinoma cells treated with Octa or compound 1, indicate the reversion of the TGF-01 -induced EMT markers. The analysis of the epithelial phenotype marker E-cadherin in A431 cells after treatment with TGF-pi did not show a decreased expression of the protein, but clearly revealed its delocalization from the plasma membrane to the cytoplasm associated with a reduced cell-cell adhesion. Literature data show a correlation between the membranous E-cadherin expression and the degree of tumour differentiation, with upregulation in well-differentiated cSCC and attenuated or missing staining in poorly differentiated tumours, which are characterized by a high cytoplasmic expression of E-cadherin. This translocation from the membrane to the intracytoplasmic region is considered a functional loss of this adhesion molecule, linked to the promotion of the EMT process.

[0283] Reprogramming of lipid metabolism is a hallmark of cancer cells because dysregulation of lipids and alteration of related enzyme profiles are correlated with oncogenic signals and malignant phenotypes, such as metastasis and EMT. Heightened de novo lipogenesis is required for cellular transformation and cancer progression. Transformed cells and cancerous tissues are characterized by a high amount of MUFAs, and up-modulation of SCD1. The data disclosed herein showed that TGF-pi induced a significant increase of MUFA / SFA ratio in A431 cells, indicating the accumulation in cellular membranes of lipids generated by de novo synthesis and over-expression of SCD-1. Compound 1 and Octa significantly counteracted this effect, suggesting that these compounds interfere with SCD-1 activity and FA metabolism dysregulation associated with the EMT program. Moreover, FA uptake from the microenvironment is altered in cancer cells and contributes to proliferation and dissemination to distant organs. TGF-pi caused a significant release of free MUFAs, in particular palmitoleic and oleic acids, in the culture media of A431 cells. This effect suggests the upregulation of lipolytic pathways that mobilize free fatty acids generating oncogenic lipid signals that, in turn, fuel aggressive features of cancer. The treatment with Octa or compound 1 significantly counteracted the release of free MUFAs, reducing the C16: l / C16:0 and C18: l / C18:0 ratios and limiting the incorporation and the utilization of exogenous MUFAs, thus dampening the pro-oncogenic lipid signalling.

[0284] Cholesterol dyshomeostasis has emerged as a key requirement for cancer initiation and progression. The increased amount of cholesterol inside cell membranes corresponds to an increase in lipid raft presence, leading to an enhanced response to cell signalling proteins involved in the neoplastic transformation and the activation of the EMT program The treatment with TGF-pi caused the enrichment of cholesterol in cellular membranes of A431 cells and Octa or compound 1 reduced significantly this accumulation, suggesting that these molecules might interfere with EMT and also counteract the alteration of lipid raft domains. References

[0285] • Nehal, K.S.; Bkjian, C.K. Update on Keratinocyte Carcinomas. N Engl J Med 2018, 379, 363-374.

[0286] • Venables, Z.C.; Autier, P.; Nijsten, T.; Wong, K.F.; Langan, S.M.; Rous, B.; Broggio, I; Harwood, C.; Henson, K.; Proby, C.M.; Rashbass, J.; Leigh, I.M. Nationwide Incidence of Metastatic Cutaneous Squamous Cell Carcinoma in England. JAMA Dermatol 2019, 155, 298-306.

[0287] • Pihl, C.; Togsverd-Bo, K.; Andersen, F.; Haedersdal, M.; Bjerring, P.; Lerche, C.M.

[0288] KeratinocyteCarcinoma and Photoprevention: The Protective Actions of Repurposed Pharmaceuticals, Phytochemicals and Vitamins. Cancers (Basel) 2021, 13,

[0289] 10.3390 / cancersl3153684.

[0290] • Piipponen, M.; Riihila, P.; Nissinen, L.; Kahari, V.M. The Role of p53 in Progression of CutaneousSquamous Cell Carcinoma. Cancers (Basel) 2021, 13, 10.3390 / cancersl3184507.

[0291] • Ciuciulete, A.R.; Stepan, A.E.; Badiu, A.M.; Andreiana, B.C.; Florescu, M.M.; Simionescu,

[0292] C.E.;Vilcea, A.M. E-cadherin, fibronectin and Slug immunoexpression in non-melanoma skincancers. Rom J Morphol Embryol 2021, 62, 705-712.

[0293] • Hodorogea, A.; Calinescu, A.; Antohe, M.; Balaban, M.; Nedelcu, R.I.; Turcu, G.; Ion,

[0294] D.A.;Badarau, I.A.; Popescu, C.M.; Popescu, R.; Popp, C.; Cioplea, M.; Nichita, L.; Hulea, I.; Brinzea,A.Epithelial-Mesenchymal Transition in Skin Cancers: A Review. Anal Cell Pathol (Amst)2019, 2019, 3851576.

[0295] • Nieto, M.A. Context-specific roles of EMT programmes in cancer cell dissemination. Nat CellBiol 2017, 79, 416-418.

[0296] • Saenz-Sarda, X.; Carrato, C.; Perez-Roca, L.; Puig, L.; Ferrandiz, C.; Ariza, A.; Femandez- Figuera, M.T.s. Epithelial-to-mesenchymal transition contributes to invasion in squamous cellcarcinomas originated from actinic keratosis through the differentiated pathway, whereasproliferation plays a more significant role in the classical pathway. JEADV 2018, 32, 581-586.

[0297] • Thiery, J.P.; Acloque, H.; Huang, R.Y.; Nieto, M.A. Epithelial-mesenchymal transitions indevelopment and disease. Cell 2009, 139, 871-890.

[0298] • Usman, S.; Waseem, N.H.; Nguyen, T.K.N.; Mohsin, S.; Jamal, A.; Teh, M.T.; Waseem, A.Vimentin Is at the Heart of Epithelial Mesenchymal Transition (EMT) Mediated Metastasis. Cancers (Basel) 2021, 13, 10.3390 / cancersl3194985.

[0299] • Morandi, A.; Taddei, M.L.; Chiarugi, P.; Giannoni, E. Targeting the Metabolic ReprogrammingThat Controls Epithelial-to-Mesenchymal Transition in Aggressive Tumors. Front Oncol 2017, 7, 40. • Sciacovelli, M.; Frezza, C. Metabolic reprogramming and epithelial -to-mesenchymal transitionin cancer. FEBS J . 2017 Oct;284(19):3132-3144. doi: 10.1111 / febs.l4090. Epub 2017 May 272017, 284, 3132-3144.

[0300] • Liberti, M.V.; Locasale, J.W. The Warburg Effect: How Does it Benefit Cancer Cells? TrendsBiochem Sci 2016, 41, 211-218.

[0301] • Kang, H.; Kim, H.; Lee, S.; Youn, H.; Youn, B. Role of Metabolic Reprogramming inEpithelial Mesenchymal Transition (EMT). IntJMol Sci . 2019 Apr 25; 20(8) .2042 2019, 20,2042.

[0302] • Luo, X.; Cheng, C.; Tan, Z.; Li, N.; Tang, M.; Yang, L.; Cao, Y. Emerging roles of lipidmetabolism in cancer metastasis. Mol Cancer 2017, 16, 76-017-0646-3.

[0303] • Hao, Y.; Li, D.; Xu, Y.; Ouyang, J.; Wang, Y.; Zhang, Y.; Li, B.; Xie, L.; Qin, G. Investigationof lipid metabolism dysregulation and the effects on immune microenvironments in pan-cancerusing multiple omics data. BMC Bioinformatics 2019, 20, 195-019-2734-4.

[0304] • Raeisi, M.; Hassanbeigi, L.; Khalili, F.; Kharrati-Shishavan, H.; Yousefi, M.; Mehdizadeh, A.Stearoyl-CoA desaturase 1 as a therapeutic target for cancer: a focus on hepatocellularcarcinoma. Mol Biol Rep 2022, 49, 8871-8882.

[0305] • Che, D.; Zhang, S.; Jing, Z.; Shang, L.; Jin, S.; Liu, F.; Shen, J.; Li, Y.; Hu, J.; Meng, Q.; Yu, Y. Macrophages induce EMT to promote invasion of lung cancer cells through the IL- 6-mediated COX-2 / PGE2 / beta-catenin signalling pathway. Mol Immunol 2017, 90, 197-210.

[0306] • Li, Z.L.; Ye, S.B.; OuYang, L.Y.; Zhang, H.; Chen, Y.S.; He, J.; Chen, Q.Y.; Qian, C.N.; Zhang, X.S.; Cui, J.; Zeng, Y.X.; Li, J. COX-2 promotes metastasis in nasopharyngeal carcinoma by mediating interactions between cancer cells and myeloid-derived suppressor cells. Oncoimmunology 2015, 4, el044712.

[0307] • Goeman, F.; Manni, I.; Artuso, S.; Ramachandran, B.; Toietta, G; Bossi, G; Rando, G; Cencioni,C.; Germoni, S.; Straino, S.; Capogrossi, M.C.; Bacchetti, S.; Maggi, A.; Sacchi, A.; Ciana, P.;Piaggio, G. Molecular imaging of nuclear factor-Y transcriptional activity maps proliferationsites in live animals. Mol Biol Cell 2012, 23, 1467-1474.

[0308] • Kovacs, D.; Fiori, E.; Maresca, V.; Ottaviani, M.; Aspite, N.; Dell' Anna, M.L.; Panzella, L.;Napolitano, A.; Picardo, M.; d'Ischia, M. The eumelanin intermediate 5,6- dihydroxyindole-2-carboxylic acid is a messenger in the cross-talk among epidermal cells. J Invest Dermatol 2012, 132, 1196-1205.

[0309] • BLIGH, E.G; DYER, W.J. A rapid method of total lipid extraction and purification. Can JBiochem Physiol 1959, 37, 911-917. • Vantaggiato, C.; Dell'Omo, G.; Ramachandran, B.; Manni, I.; Radaelli, E.; Scanziani, E.; Piaggio,G.; Maggi, A.; Ciana, P. Bioluminescence imaging of estrogen receptor activity during breastcancer progression. Am J Nucl Med Mol Imaging 2016, 6, 32-41.

[0310] • Signore, A.; Mather, S.J.; Piaggio, G; Malviya, G; Dierckx, R.A. Molecular imaging ofinflammation / infection: nuclear medicine and optical imaging agents and methods. Chem Rev2010, 110, 3112-3145.

[0311] • Manni, I.; Di Rocco, G; Fusco, S.; Leone, L.; Barbati, S.A.; Carapella, C.M.; Grassi, C.; Piaggio, G; Toietta, G. Monitoring the Response of Hyperbilirubinemia in the Mouse Brain by In VivoBioluminescence Imaging. IntJMol Sci 2016, 18, 10.3390 / ijmsl8010050.

[0312] • Manni, I.; de Latouliere, L.; Gurtner, A.; Piaggio, G. Transgenic Animal Models to VisualizeCancer-Related Cellular Processes by Bioluminescence Imaging. Front Pharmacol 2019, 10,235.

[0313] • Rizzi, N.; Manni, I.; Vantaggiato, C.; Delledonne, G.A.; Gentileschi, M.P.; Maggi, A.; Piaggio, G; Ciana, P. In vivo imaging of cell proliferation for a dynamic, whole body, analysis ofundesired drug effects. Toxicol Sci 2015, 145, 296-306.

[0314] INCORPORATION BY REFERENCE

[0315] This application refers to various issued patents, published patent applications, journal articles, and other publications, all of which are incorporated herein by reference. If there is a conflict between any of the incorporated references and the instant specification, the specification shall control. In addition, any particular embodiment of the present disclosure that falls within the prior art may be explicitly excluded from any one or more of the claims. Because such embodiments are deemed to be known to one of ordinary skill in the art, they may be excluded even if the exclusion is not set forth explicitly herein. Any particular embodiment of the disclosure can be excluded from any claim, for any reason, whether or not related to the existence of prior art.

[0316] EQUIVALENTS

[0317] The invention may be embodied in other specific forms without departing from the spirit or essential characteristics thereof. The foregoing embodiments are therefore to be considered in all respects illustrative rather than limiting the invention described herein. Scope of the invention is thus indicated by the appended claims rather than by the foregoing description, and all changes that come within the meaning and range of equivalency of the claims are intended to be embraced therein.

Claims

CLAIMSWhat is claimed is:

1. A compound of Formula (I):or a pharmaceutically acceptable salt thereof, wherein:R1is selected from the group consisting of: CH2, CH, and NRa;R2is selected from the group consisting of: H, Ci-Ce alkyl, and (Ci-Ce alkylene)phenyl, wherein the phenyl may be optionally substituted with one, two or three substituents selected from the group consisting of Ci-Ce alkyl, halogen, Ci-Ce haloalkyl, Ci-Ce alkoxy, and cyano;(i) R3is selected from the group consisting of H, Ci-Ce alkyl, and oxo,R4is selected from H and Ci-Ce alkoxy, andR5is H; or(ii) R3and R4are taken together to form a 5-6 membered heteroaryl with the carbon atoms to which R3, R4, and R5are attached, andR5is selected from H and Ci-Ce alkyl;— is a double bond, or — is a single bond when (a) R1is NRaand R3is oxo or (b) R3and R4are taken together to form a 5-6 membered heteroaryl with the carbon atoms to which R3, R4, and R5are attached; andRais H or Ci-Ce alkyl; wherein at least one of R3, R4, and R5is not hydrogen.

2. The compound of claim 1, wherein R1is CH2.

3. The compound of claim 1, wherein R1is CH.

4. The compound of claim 1, wherein R1is NRa.

5. The compound of any one of claims 1-4, wherein R2is H.

6. The compound of any one of claims 1-5, wherein R3is H.

7. The compound of any one of claims 1-5, wherein R3is Ci-Ce alkyl.

8. The compound of any one of claims 1-5, wherein R3is oxo.

9. The compound of any one of claims 1-8, wherein R4is H.

10. The compound of any one of claims 1-8, wherein R4is Ci-Ce alkoxy.

11. The compound of any one of claims 1-5, wherein R3and R4are taken together to form a 5-6 membered heteroaryl with the carbon atoms to which R3, R4, and R5are attached.

12. The compound of any one of claims 1-11, wherein R5is H.

13. The compound of claim 11, wherein R5is Ci-Ce alky.

14. The compound of any one of claims 4-13, wherein Rais H.

15. The compound of any one of claims 4-13, wherein Rais Ci-Ce alkyl.

16. The compound of claim 1, wherein the compound is selected from the group consisting of:or a pharmaceutically acceptable salt thereof.

17. A compound having the formula:or a pharmaceutically acceptable salt thereof.

18. A pharmaceutical composition comprising the compound of any one of claims 1-17 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

19. A combination comprising the compound of any one of claims 1-17 or a pharmaceutically acceptable salt thereof, and octatrienoic acid.

20. A method of treating or preventing a skin cancer or a pre-cancerous skin lesion in a patient in need thereof comprising administering to the patient a therapeutically effective amount of the compound of any one of claims 1-17 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 18, or the combination of claim 19.

21. The method of claim 20, wherein the skin cancer is keratinocyte-derived skin cancer.

22. The method of claim 20 or 21, wherein the skin cancer is selected from the group consisting of cutaneous squamous cell carcinoma, basal cell carcinoma, Merkel cell carcinoma, and melanoma.

23. The method of any one of claims 20-22, wherein the skin cancer is an unresectable skin cancer.

24. The method of any one of claims 20-23, wherein the skin cancer is locally advanced cutaneous squamous cell carcinoma.

25. The method of any one of claims 20-23, wherein the skin cancer is metastatic skin cancer.

26. The method of any one of claims 20-25, wherein the skin cancer is refractory skin cancer.

27. The method of any one of claims 20-26, wherein the skin cancer is relapsed skin cancer.

28. The method of any one of claims 20-27, wherein the skin cancer is advanced.

29. The method of claim 20, wherein the pre-cancerous skin lesion is actinic keratosis.

30. The method of any one of claims 20-29, wherein the skin cancer or pre-cancerous skin lesion is caused by exposure to ultraviolet (UV) radiation.

31. The method of any one of claims 20-30, wherein the method further comprising administering to the subject a therapeutically effective amount of a second therapeutic agent.

32. The method of claim 31, wherein the second therapeutic agent is octatrienoic acid.

Images