Novel phthalimide compounds or tissue / cell regeneration and compositions comprising thereof

Abstract

The present invention relates to phthalimide-based compounds and pharmaceutical compositions comprising phthalimide-based pharmacophore-containing compounds of formula (A) or a pharmaceutically acceptable salts thereof and pharmaceutically acceptable carriers, isomers, deuterium analogs for compounds of formula (A). The present invention also provides a method of preparation of the compound of formula (A). The phthalimide-based pharmaceutical composition are useful as wound healers, tissue / cell / muscle regeneration substances, and hair follicle development substances.

Description

NOVEL PHTHALIMIDE COMPOUNDS FOR TISSUE / CELLREGENERATION AND COMPOSITIONS COMPRISING THEREOFTECHNICAL FIELD OF INVENTION

[0001] The present invention relates to novel compounds of formula A, or pharmaceutically acceptable salts thereof, and pharmaceutical compositions comprising novel compounds of formula A or salts thereof. Particularly, the present invention relates to substituted phthalimidebased compounds and compositions thereof to be used for wound healing or tissue / cell regeneration and hair follicle development.BACKGROUND OF INVENTION

[0002] The pharmaceuticals are out of the grasp of the underprivileged due to their high price and limited availability. It generates a significant need for low-cost medications or novel small organic compounds to lessen the burden of tropical parasite diseases. Phthalimide (Pht)-based compounds have shown extensive potential in medicinal chemistry. Aside from many Pht-based FDA- approved drugs, recent reports have demonstrated the broad- spectrum biological properties of Pht against various diseases such as schistosomiasis, malaria, leishmaniasis, tuberculosis, and dengue. Pht also showed remarkable activity when hybridized with other significant pharmacophores, i.e., triazole, piperazine, and thiazole.

[0003] Annually, millions worldwide endure compromised wound healing / tissue or cell regeneration due to various factors like trauma, surgery, acute illness, or chronic diseases. These results from inadequately controlled aspects of the normal tissue repair process, including inflammation, angiogenesis, matrix deposition, and cell recruitment. The breakdown of these cellular processes often correlates with underlying clinical conditions like diabetes, vascular disease, or ageing, commonly associated with healing -related disorders. A comprehensive comprehension of the foundational biology of repair and regeneration is crucial for advancing treatment strategies that can augment the body's innate repair mechanisms.

[0004] A wound is defined as the beach in the living histoarchitecture regarding anatomical, functional, and cellular integrity imposed by any kind of physicochemical, thermal, environmental, and microbial deterrents. It is also described as an interruption of healthy tissue's structure and function due to contusion, abrasion, and laceration following the loss of the epithelial barrier of the skin layer. 1 % of the world’s population is affected by various kinds of wounds. Wound healing is an overlapping yet well-defined process occurring at three different stages, namely hemostasis and inflammation, proliferation, and remodelling, respectively. Several factors affect the healing process, viz. age, sex, disease, and immune status of the individual. Diabetes, venous leg ulcers, and the immunocompromised state of the individual affect the healing process. Wound healing proceeds through a series of collaborative episodes in tissue-cell lineage and cytokine-mediated reactions, resulting in the restoration of the functional integrity of damaged tissues.

[0005] In this context, we introduce the discovery of innovative compounds that serve as wound healers, revitalizing damaged skin cells / tissue or cell regeneration / hair follicle development.OBJECTIVES OF THE INVENTION

[0006] The primary objective of the present invention is to provide novel substituted phthalimidebased compounds and / or pharmaceutically acceptable salts thereof.

[0007] Another objective of the present invention is to provide novel substituted phthalimide -based compounds and / or pharmaceutically acceptable salts thereof for use in wound healing or tissue / cell regeneration, and hair follicle development.

[0008] Yet another objective of the present invention is to provide a composition comprising the novel substituted phthalimide-based compounds or pharmaceutically acceptable salts thereof; with at least one pharmaceutically acceptable excipient (such as a pharmaceutically acceptable carrier or diluent).

[0009] Yet another objective of the invention is to provide a method of preparation of novel substituted phthalimide -based compounds, which are synthesized under microwave irradiation following milder reaction conditions.

[0010] Other objectives and advantages of the present invention will become apparent from the following description taken in connection with the accompanying drawings, wherein, by way of illustration and example, the aspects of the present invention are disclosed.SUMMARY OF INVENTION

[0011] An embodiment of the present invention discloses compounds of formula A, or pharmaceutically acceptable salts thereof, and pharmaceutical compositions thereof, for wound healing or revitalizing damaged skin cells or tissue or cell regeneration, and hair follicle development. Particularly, the present invention discloses substituted phthalimide -based compounds or pharmaceutically salts thereof, and compositions comprising the substituted phthalimide-based compounds of formula A.whereinX is selected from the group consisting of naphthalene, 5-7 membered heterocyclo alkyl, aniline optionally substituted with halogen, or an aminophenyl group, wherein the phenyl group is optionally substituted with halogen(s), aryl optionally substituted with one or more groups selected from hydrogen, methyl or halogen;Ri is selected from the group consisting of Hydrogen, fluoro, chloro, bromo, (C1-5) alkyl, nitro, trifluoromethyl, tert-butyl, phenyl, methoxy, cyano, naphthyl;R2 is selected from the group consisting of hydrogen, hydroxyl, (C1-5) amino alkyl, cyano, alkyl, nitro, alkoxy, halogen; optionally substituted oxalate, malonate, succinate that may be substituted with hydrogen, deuterium, hydroxyl, amino;R3 is Hydrogen, (C1-C5) alkyl, (C1-C5) cycloalkyl, aryl, and heteroaryl rings optionally substituted with halogen, tert-butyl, methyl, (C1-C5) hydroxyalkyl, phenyl, cycloalkyl, methoxy, aryls, sulphur dioxide, trifluoromethyl, and trifluoromethoxy.

[0012] In another embodiment, the present invention provides a pharmaceutical composition comprising a compound of formula (A) or a possible isomer or a pharmaceutically acceptable salt thereof and at least one pharmaceutically acceptable excipient (such as a pharmaceutically acceptable carrier or diluent).BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 illustrates a reaction scheme for the synthesis of compound of formula A.

[0014] Figure 2 illustrates the1H NMR spectrum of compound 1 in DMSO-D6.

[0015] Figure 3 illustrates the13C NMR spectrum of compound 1 in DMSO-D6.

[0016] Figure 4 illustrates the ESI (HR-MS) spectrum of compound 1.

[0017] Figure 5 illustrates the wound healing contraction studies.DETAILED DESCRIPTION OF THE INVENTION

[0018] The present invention will be better understood after reading the following detailed description of the presently preferred aspects thereof, in which the features, other aspects, and advantages of the invention shall be more apparent from certain exemplary embodiments of the invention.

[0019] Accordingly, those of ordinary skill in the art will recognize that various changes and modifications of the embodiments described herein can be made without departing from the scope of the invention. In addition, descriptions of well-known terms and functions are omitted for clarity and conciseness.

[0020] The terms and words used in the following description are not limited to the bibliographical meanings, but are merely used to enable a clear and consistent understanding of the invention. Accordingly, it should be apparent to those skilled in the art that the following description of exemplary embodiments of the present invention are provided for illustrative purpose only and not for the purpose of limiting the invention.

[0021] It is to be understood that the singular forms “a”, “an” and “the” include plural referents unless the context clearly dictates otherwise.

[0022] The term “phthalimide -based compounds” herein includes derivatives, pharmaceutically acceptable salts, all optically active forms including, isomers, tautomers, stereoisomers, enantiomers, diastereomers, racemic mixtures, and their pharmaceutically acceptable salts.

[0023] The term “pharmacore” herein includes triazole, piperazine, and thiazole that may be hybridized with “phthalimide-based compounds” to increase the effectivity of the composition.

[0024] The term “tissue / cell regeneration” herein refers to wound healing, cell repairing, Reepithelialisation, and hair follicle development.

[0025] It should be emphasized that the term “comprises / comprising” when used in this specification is taken to specify the presence of stated features, integers, steps, or components but does not preclude the presence or addition of one or more other features, integers, steps, components, or groups thereof.

[0026] Accordingly, the present invention relates to novel compounds of formula (A) and compositions thereof. Notably, the present invention relates to novel substituted phthalimide-based compounds or pharmaceutically acceptable salts thereof, and pharmaceutical compositions thereof that are used as wound-repairing agents, damaged skin cells revitalizing agents, tissue / cell regeneration substances, and hair follicle development agents.

[0027] In yet another exemplary embodiment, the present invention discloses the chemical structures of phthalimide -based pharmacophore-containing compounds of formula (A) and their chemical synthesis.

[0028] In an embodiment, the present invention provides substituted phthalimide -based compounds of formula (A):X is selected from the group consisting of naphthalene, 5-7 membered heterocycloalkyl, aniline optionally substituted with halogen, or an aminophenyl group, wherein the phenyl group is optionally substituted with halogen(s) and aryl optionally substituted with hydrogen, methyl, halogen;Ri is selected from the group consisting of hydrogen, fluoro, chloro, bromo, (C1-5) alkyl, nitro, trifluoromethyl, tert-butyl, phenyl, methoxy, cyano, and naphthyl;R2 is selected from the group consisting of hydrogen, hydroxyl, (C1-5) amino alkyl, cyano, alkyl, nitro, alkoxy, halogen; optionally substituted oxalate, malonate, succinate substituted with hydrogen, deuterium, hydroxyl, amino;R3 is Hydrogen, (C1-C5) alkyl, (C1-C5) cycloalkyl, aryl and heteroaryl rings optionally substituted with halogen, tert-butyl, methyl, (C1-C5) hydroxy alkyl, phenyl, cycloalkyl, methoxy, aryl, sulphur dioxide, trifluoromethyl, and trifluoromethoxy.

[0029] In yet another embodiment, the present invention provides methods for treating a disease or a disorder comprising administering a therapeutically effective amount of a compound represented by a compound of formula (A) or a pharmaceutically acceptable salt thereof to a subject, e.g., a human, in need thereof.

[0030] In an exemplary embodiment, the compounds of Formula A are selected from:

[0031] In an exemplary embodiment, the present invention provides substituted phthalimide-based compounds of formula (A)or a pharmaceutically acceptable salt thereof, or a stereoisomer, or a tautomer thereof, for wound repairing, revitalizing damaged skin cells, tissue / cell regeneration, and hair follicle development, wherein:X is selected form the group consisting of 6 membered heterocycloalkyl ring, or an aminophenyl group, wherein the phenyl group is optionally substituted with halogens;Ri is hydrogen;R2 is selected from the group consisting of hydrogen, hydroxyl;Ra is selected from the group consisting of halogen, tert-butyl, methyl, (C1-C5) alkyl, hydroxy, (Ci-C5)hydroxyalkyl, trifluoromethyl, and trifluoromethoxy.

[0032] In a preferred embodiment, the present invention provides substituted phthalimide-based compounds of formula A, or a pharmaceutically acceptable salt thereof,Wherein, X is piperazine, Ri is hydrogen, R2 is hydroxyl; R3 is (C1-C5) hydroxyalkyl group.

[0033] In an embodiment, the present invention also includes stereoisomers of substituted phthalimide-based compounds of formula A (such as compounds of Formula I, II, and III) having molecular formula 2-(2-hydroxy-3 -(4-(2-hydroxyethyl)piperazin- 1 -yl)propyl)isoindoline- 1,3- dione, (7?)-2-(2-hydroxy-3-(4-(2-hydroxyethyl)piperazin- l-yl)propyl)isoindoline- 1 ,3-dione and (S)-2-(2-hydroxy-3-(4-(2-hydroxyethyl)piperazin-l-yl)propyl)isoindoline- 1,3-dione, respectively. The said compounds of formula I, II, and III can be synthesized and studied for wound repairing, development of hair follicles, revitalizing damaged skin cells / tissues, and cell regeneration.

[0034] In an embodiment, the present invention provides a pharmaceutical composition comprising the novel substituted phthalimide -based compounds of formula A and at least one pharmaceutically acceptable excipient. Preferably, the pharmaceutical composition comprises a therapeutically effective amount of at least one compound of formula A that may be associated with a pharmaceutically acceptable excipient.

[0035] In a preferred embodiment, the pharmaceutical composition comprises at least one substituted phthalimide-based compound of formula A in a quantitative amount ranging from 20% to 90% of the total pharmaceutical composition; and a pharmaceutically acceptable excipient in a quantitative amount ranging from 10% to 30% of the total pharmaceutical composition.

[0036] In an exemplary embodiment, the pharmaceutically acceptable excipients are selected from, such as, but not limited to, carriers, isomers, deuterium analogs, diluents, or their combination. The pharmaceutically acceptable excipients may be diluted by a carrier or enclosed within a carrier, which can be in the form of a capsule, sachet, paper, or other container.

[0037] In another exemplary embodiment, the pharmaceutically acceptable excipients are selected from metal phosphate, metal carbonate, metal sulfate, hallites, metal oxides, sugar, alchohol, artificial sweeteners, starch, cellulose ester, cellulose ethers, polyethylene glycol, polypropylene glycol, glycerin, fatty acid esters, and their combination thereof.

[0038] In an embodiment, the present invention provides a pharmaceutical composition comprising a compound of formula (A) or possible isomers or a pharmaceutically acceptable salt thereof and at least one pharmaceutically acceptable excipient.

[0039] In another embodiment, the present invention provides a pharmaceutical composition comprising a compound of formula (A) or a pharmaceutically acceptable salt thereof for treating diseases.

[0040] In an embodiment, the present invention provides a pharmaceutical composition comprising the said compound of formula A, or a stereoisomer, or a tautomer thereof as therapeutic agents.

[0041] In one embodiment, the present invention provides a compound of formula (A) or a pharmaceutical-acceptable salt or a stereoisomer or a tautomer thereof for use in the treatment of wound healing or repair.

[0042] In one embodiment, the present invention provides a pharmaceutical composition comprising a compound of formula (A) or a pharmaceutical-acceptable salt or a stereoisomer or a tautomer thereof for use in the treatment of regeneration applications or revitalizing damaged skin cells or disorders.

[0043] In one embodiment, the present invention provides a pharmaceutical composition comprising a compound of formula (A) or a pharmaceutical-acceptable salt or a stereoisomer or a tautomer thereof for use in treating tissue or cell regeneration or disorders.

[0044] In one embodiment, the present invention provides a pharmaceutical composition comprising a compound of formula (A) or a pharmaceutical-acceptable salt or a stereoisomer or a tautomer thereof for use in hair follicle development.

[0045] In one embodiment, the present invention provides a method of treating rare diseases comprising administering to a cell, in need thereof, a therapeutically effective amount of a compound of formula (A) or a pharmaceutically acceptable salt or a stereoisomer or a tautomer thereof.

[0046] In one embodiment, the present invention provides a pharmaceutical composition comprising a compound of formula (A), for use in the manufacture of a medicament.

[0047] In an embodiment, the present invention provides a method of preparation of the substituted phthalimide-based compounds of formula (A), said process comprising the following steps:

[0048] A method of preparing the pharmaceutical compound of formula A, wherein the method comprises the steps of: a) preparing a mixture of compounds optionally substituted isoindoline- 1,3-dione, substituted heterocycloalkyl or aryl in a molar ratio (1:1), in ethanol solvent in a pressure seal tube; b) heating the mixture of step (a) under microwave conditions at 200 °C for 1 minute; c) removing the ethanol solvent under vacuum on a rotary evaporator to obtain crude product; d) extracting the crude product with ethyl acetate and brine solution; e) separating the organic layer of the crude product and drying over anhydrous sodium sulphate; f) removing the excess ethyl acetate solution using a rotary evaporator to obtain the product; g) purifying the product obtained in step (f) to prepare the compound of formula A.

[0049] In an exemplary embodiment, the optionally substituted isoindoline- 1,3 -dione of step (a) is 2-(oxiran-2-ylmethyl)isoindoline- 1,3-dione, and the substituted heterocycloalkyl of step (a) is 2-(piperazin-l-yl)ethan-l-ol .EXAMPLE:

[0050] The present invention will be described in detail with reference to specific examples provided below. However, such examples should not be construed to limit the scope of the present invention.

[0051] EXAMPLE 1:

[0052] Synthesis of compound 1:Compound 1The compound 1 represent the compounds of formula I, II, and III. As shown in Figure 1, the method of synthesis of the compound 1 is discussed herein below. To prepare these compounds,2-(oxiran-2-ylmethyl)isoindoline-l, 3-dione (1.0 mmol), 2-(piperazin-l-yl)ethan-l-ol (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.JH NMR,13C NMR, and HR-MS techniques confirmed the chemical composition of 1.

[0053] EXAMPLE 2:

[0054] Synthesis of compound 2 (2-(2-hydroxy-3-(p-tolylamino)propyl)isoindoline-l,3- dione):Compound 22-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), p-aminotoluene (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under microwave at 200 °C for 5-10 minutes. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0055] EXAMPLE 3:

[0056] Synthesis of compound 3 (2-(3-((4-(tert-butyl)phenyl)amino)-2- hydroxypropyl)isoindoline-l,3-dionedione):Compound 32-(oxiran-2-ylmethyl)isoindoline-l, 3-dione (1.0 mmol), 4-(tert-butyl)phenylamine (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °Cfor 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0057] EXAMPLE 4:

[0058] Synthesis of compound 4 (2-(2-hydroxy-3-((4- methoxyphenyl)amino)propyl)isoindoline-l, 3-dione):Compound 42-(oxiran-2-ylmethyl)isoindoline-l, 3-dione (1.0 mmol), 4-(methoxyphenylamine) (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0059] EXAMPLE 5:

[0060] Synthesis of compound 5 (2-(3-((4-fluorophenyl)amino)-2- hydroxypropyl)isoindoline- 1,3-dione) :Compound 52-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 4-(fluorophenylamine) (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate(20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0061] EXAMPLE 6:

[0062] Synthesis of compound 6 (2-(3-((4-bromophenyl) amino)-2-hydroxypropyl) isoindoline- 1 ,3-dione) :Compound 62-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 4-(bromophenylamine) (1.0 mmol), ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0063] EXAMPLE 7:

[0064] Synthesis of compound 7 (2-(2-hydroxy-3-((4-(trifluoromethoxy)phenyl)amino)propyl)isoindoline-l, 3-dione):Compound 72-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 4-(trifluoromethoxyphenylamine) (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried overanhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0065] EXAMPLE 8:

[0066] Synthesis of compound 8 (2-(2-hydroxy-3-((4-(trifluoromethyl)phenyl)amino)propyl)isoindoline-l, 3-dione):Compound 82-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 4-(trifluoromethoxyphenylamine) (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0067] EXAMPLE 9:

[0068] Synthesis of compound 9 (2-(2-hydroxy-3-((2- methoxyphenyl)amino)propyl)isoindoline-l, 3-dione):Compound 92-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 3 -methoxy aniline (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brinesolution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.EXAMPLE 10:

[0069] Synthesis of compound 10 (2-(3-((3-fhiorophenyl)amino)-2- hydroxypropyl)isoindoline-l, 3-dione:Compound 102-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 3-fluorophenylamine (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0070] EXAMPLE 11:

[0071] Synthesis of compound 11 (2-(3-((3-bromophenyl)amino)-2- hydroxypropyl)isoindoline- 1,3-dione) :Compound 112-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 3 -bromophenylamine (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydroussodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0072] EXAMPLE 12:

[0073] Synthesis of compound 12 (2-(2-hydroxy-3-((3-(trifluoromethyl)phenyl)amino)propyl)isoindoline-l, 3-dione):Compond 122-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 3 -trifluoromethylphenylamine (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.EXAMPLE 13:

[0074] Synthesis of compound 13 (2-(2-hydroxy-3-((2- methoxyphenyl)amino)propyl)isoindoline-l, 3-dione ) :Compound 132-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 2-methoxylphenylamine (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate(20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0075] EXAMPLE 14:

[0076] Synthesis of compound 14 (2-(3-((2-fluorophenyl)amino)-2- hydroxypropyl)isoindoline- 1,3-dione) :Compound 142-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 2-fluorophenylamine (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.EXAMPLE 15:

[0077] Synthesis of compound(2-(3-((2-bromophenyl)amino)-2- hydroxypropyl)isoindoline- 1,3-dione) :Compound 152-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 2-bromophenylamine (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum ona rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.

[0078] EXAMPLE 16:

[0079] Synthesis of compound 16 (2-(3-((2,4-difhiorophenyl)amino)-2- hydroxypropyl)isoindoline- 1,3-dione) :Compound 162-(oxiran-2-ylmethyl)isoindoline- 1,3-dione (1.0 mmol), 2,4-difluorophenylamine (1.0 mmol), and ethanol (2 mL) were taken in a pressure seal tube and heated under the microwave at 200 °C for 1 minute. After completion of the reaction, the reaction the reaction contents were transferred into a round bottom flask, and the solvent was removed under vacuum on a rotary evaporator. The crude product was extracted with ethyl acetate (20 mL x 3) and brine solution (15 mL x 3). The organic layer separated and was dried over anhydrous sodium sulphate. Excess ethyl acetate was removed on a rotary evaporator, and the product was purified either by washing with hexane or recrystallizing with ethyl acetate and hexane in 1:9 ratios.EXPERIMENTAL STUDY:

[0080] The following illustrates the experimental data and should not be construed to limit the scope of the present invention.

[0081] Solvents and reagents were received and utilized for the experiments without additional purification. Thin-layer chromatography (TLC) on alumina-coated Merck plates was used to determine the purity of all the necessary products. TLC plates were loaded with compounds dissolved in chloroform (CHCL) and developed in Ethyl acetate / Hexane (1:1, v / v). The compounds were purified by Flash column chromatography (Yamazen, Japan) using alumina gel columns (100-200 mesh size, CDH) for minor impurities found by iodine vapor / UV light visualization. Microwave reactions were carried out in a sealed vial using the “Start Synth Microwave Synthesis Labstation (Milestone microwave laboratory systems)” microwave apparatus utilizing a power supply of 300 W at 80° C by controlled temperature programming with a 2-minute ramp and holding for 20 minutes. BUCHI Labortechnik AG CH-9230 was used to measure melting points. Under ambient temperature, an Anton Paar Polarimeter was used to record specific rotation using a 1 percent chemical concentration (lg / 100 mL) in ethanol at 589nm wavelength. Nuclear Magnetic Resonance (NMR) spectra (JH &13C) were recorded in CDCI3, DMSO, and D2O medium on a JEOL ECX-400P NMR at 400 MHz and 100 MHz, respectively. The NMR spectroscopy study utilized Trimethylsilane (TMS) as an internal standard. Chemical shifts (-scale) and coupling constants were measured in parts per million (ppm) and hertz (Hz). Singlet (s), doublet (d), doublet of doublet, triplet (t), quartet (q), and multiplet (m) were the letters written to the splitting patterns (m). A Biosystems Q-Star mass spectrometer also confirmed the product's chemical structures. A High-Performance Liquid Chromatography (HPLC) system equipped with an analytical column (Cl 8) and a Thermo Separation Spectra SERIES UV100 detector paired with software was used to test the purity of the synthesized compounds. The mobile phase was comprised of acetonitrile and water (v / v), and the compounds were >95% pure.

[0082] Spectroscopic Data:Color: White solid;Yield: 88%;Melting Point: 107-109 °C;1H-NMR (400 MHz, DMSO-d6) 8 7.84 - 7.78 (m, 4H, H-l and 2) 4.93 (s, 1H, OH), 4.28 (s, 1H, OH), 3.91 (s, 1H, H-3), 3.63 - 3.49 (m, 2H, H-4), 3.34 (t, J = 6.4 Hz, 2H, H-5), 2.43 - 1.92 (m, 12H, H-6 to 9);13C-NMR (100 MHz, DMSO-d6) 8 168.6 (C=O), 134.7 (C-l), 132.4 (C-la), 123.4 (C-2), 64.0 (C-3), 63.6 (H-8), 60.7 (H-5 and 9), 58.8 (H-7), 53.4 (H-6), 43.6 (H-4);HRMS (calculated): (M+H) for C17H23N3O4: 333.1689; (found): 333.1672;

[0083] Wound healing / tissue regeneration studies:

[0084] Animals of each group were examined routinely, and the gross observational changes in the healed area (mm2) of the wound were digitally photographed and calculated with the help of vernier calipers at weekly intervals for a period of 21 days. Initially, the excisional wound of ~ 2x2 cm2(400 mm2) was created on the back of the animals with the help of square transparency marking to make the accuracy in the created lesioned site.

[0085] On day 3 of the study, the planimetric analysis revealed significant (P<0.05) reduction in the wound area in animals of the Compound 1 treated group (7.245-12.18 %) and silver sulphadiazine treatment group (37.10-40.41 %) as compared to animals of the control group (47.89-52.88%). However, the animals of the silver sulphadiazine-treated group showed lesser reduction as compared to the Compound 1-treated group.

[0086] On day 7 of the study, the animals of the Compound 1 treated group (65.22 -68.23 %) and silver sulphadiazine treated group (42.21-48.31%) showed a significant reduction in wound contraction as compared to the animals of control / sham operated group (14.05-48.05%). Notably, the group of animals treated with Compound 1 showed significant (P<0.05) improvement in wound contraction percentage compared to silver sulphadiazine treated animals on day 7.

[0087] Day 14thof the study revealed significant wound contraction in the animals of the Compound 1 treated group (96.02-97.11%) as compared to the sham operated (70.01- 81.00%) and silver sulphadiazine treated group (72.00-92.96%). In contrast, sham operated and silver sulphadiazine showed no significant reduction in wound contraction percentage. Interestingly, the Compound 1 treated group of animals showed hair follicle development on the 14thday of treatment.

[0088] A perusal of data on day 21stof the study revealed a significant improvement in the wound contraction in the ethanol-soluble group, showing 100 percent contraction as compared to the sham-operated (81.00-94.93%) and silver sulphadiazine treated group (90.43-97.97%).

[0089] Table 1 illustrates using the percentage of wound contraction in the animals of Compound 1 treated group and silver sulphadiazine treated groups.Table 1Reepithelialisation and hair follicle development studies:Histopathological studies on day 3 revealed zone of inflammatory cells underneath the deeply eosinophilic exudative mass. On day 7 of the study, compound treated group of animals showed the formation of granulation tissue with new capillaries and fibroblasts was observed. Next, the 14thof the study showed reepithelialisation of the surface and the development of hair follicles. On day 21 of the study, animals treated with the compound showed the formation of hair follicles and sebaceous glands beneath a complete epidermis.Further, on day 21, the Masson’s trichome study was done to delineate the collagen deposition in the underlying skin surfaces to undermine the progression of healing in the lesioned site. The compound showed the formation of hair follicles, sebaceous glands, and collagen bundles stained blue.

[0090] Biological testsThe present study was planned to develop and evaluate the wound-healing efficacy of compounds in a full-thickness excisional rat model. The animal research was performed in the Department of Veterinary Pharmacology and Toxicology, DUVASU, Mathura, U.P., as a contract project for HeteroChem InnoTech Private Limited, New Delhi.

[0091] Procurement and Maintenance of Experimental AnimalsFor the research work, 20 male Wistar rats of 150-200 grams were procured from Disease Free Small Animal House (DFSAH), LUVAS, Hisar, and kept for acclimatization for two weeks in the Laboratory Animal House of the Department. Rats were housed under proper housing and managemental conditions under 12-12 hours light-dark cycle with ad lib balanced pelleted feed diet purchased from M / s. Ashirwad Industries, Mohali, Punjab. Clean, potable distilled water was provided to these sentinel animals throughout the experimental trial. All the experimental protocols were conducted following the ethical approval of the experiment from the Institutional Animal Ethics Committee (IAEC), DUVASU, Mathura, with the approval grant number IAEC / 24 / 1 / 60 dated 05-03-2024. Following acclimatization, rats were randomly and equally divided into four groups comprising five animals in each group at the start of the experiment, i.e., on day 0 of the study. Group I served as Sham-operated (Healthy Control), and group II as the Excisional group treated with silver sulfadiazine topically. Groups III and IV were comprised of excisional groups treated with topical compound application.

[0092] Sample collection and parameters studiedTissue biopsy samples from the animals of different groups were collected at various time intervals, i.e., on days 3, 7, 14, and 21, to assess the progression of multiple stages of healing from all the animals of all the groups. The following parameters were studied:1. The wound contraction area was used to determine the reduction in wound size vis-a-vis other morphological changes. The area of wound contraction (mm2) was measured at weekly intervals till the 28thday post-lesion by Wilson's formula as follows:Wound contraction (%) = Initial wound area- final wound area / Initial wound area xl00%2. Histopathological study on the skin samples and Molecular gene expression studies.

[0093] Histopathological examination

[0094] Hematoxylin and Eosin stainingSkin tissue samples from animals of each group at different time intervals, i.e., on days 3, 7, 14, and 21, were collected and fixed in 10% neutral buffered formalin. Following fixation, samples were dehydrated using a graded alcohol series and embedded within paraffin wax at 58-60 °C. Tissue blocks were sectioned 5 pm thick sections using a microtome and mounted on albumin- coated slides. The slides were deparaffinized using xylene for 30 minutes, and then slides were rehydrated successively with IPA (100%, 90%, and 80%) for 2-3 minutes each and lastly in distilled water for 3 minutes. Following blotting the excess water, slides were stained using hematoxylin for 1-2 minutes. Again, the slides were washed with distilled water, and following this step, the slides were rinsed with 95% alcohol to ensure complete dehydration and counterstained with eosin Y solution for 1 minute. Following staining, slides were dehydrated again using 95% alcohol. Finally, slides were cleaned with xylene washing and mounted with a DPX mounting medium. The obtained slides were examined and imaged under a light microscope using different magnifications.

[0095] Masson Trichome stainingOne slide from each group at a weekly time interval, i.e., on days 7, 14, 21, and 28, was processed for collagen deposition using the Masson Trichome staining method. Briefly, the slides were dipped in Bouin’s solution for 1 hr at 56°C and then washed in running tap water to remove the yellow colour, then stained with Weigert’s haematoxylin for 10 min and Biebrich scarlet-acid fuschin for 10 min with intermittent distilled water washing. Later, the slides were differentiated with the phosphomolybdic-phosphotungstic acid solution for 10-15 min to enhance collagen staining. Then, the slides were transferred to an aniline blue solution for 5- 10 min. After washing, slides were treated with 1% acetic acid solution for 2 min for stain differentiation, washed, and dehydrated using gradient alcohol concentrations. Finally, cleared using xylene. Then, the slides were mounted with a coverslip using DPX and observed under a light microscope.

[0096] The present invention provides the following advantages:• The composition of the present invention can be used as wound healers, tissue / cell / muscle regeneration substances, and hair follicle development substances.• An method of preparing phthalimide-based compounds environment friendly solvent system under microwave irradiation, and reduces the use of other hazardous chemicals.• The reaction conditions of the present invention eliminate the need for purification of the products via column chromatography, which is a time-consuming and expensive procedure.

[0097] While this invention has been described in connection with what is presently considered to be the most practical and preferred embodiment, it is to be understood that the invention is not limited to the disclosed embodiments, but, on the contrary, is intended to cover various modifications and equivalent arrangements included within the scope of the appended claims.

Claims

AMENDED CLAIMS received by the International Bureau on 18 June 2025 (18.06.2025)CLAIMS:

1. Novel phthalimide compounds of formula (A)or a pharmaceutically acceptable salt thereof, for tissue / cell regeneration, wound healing, revitalizing damaged skin cells, and hair follicle development, wherein:-X is selected from the group consisting of naphthalene, piperazine, aniline or an aminophenyl group, wherein the phenyl group is optionally substituted with halogen(s);-Ri is selected from the group consisting of Hydrogen, fluoro, chloro, bromo, (C1-5) alkyl, nitro, trifluoromethyl, tert-butyl, phenyl, methoxy, cyano, and naphthyl;-R2 is selected from the group consisting of hydrogen, hydroxyl, (C1-5) aminoalkyl, cyano, alkyl, nitro, alkoxy, halogen; and optionally substituted oxalate, malonate, succinate substituted with hydrogen, deuterium, hydroxyl, amino;-R3 is Hydrogen, (C1-C5) alkyl, (C1-C5) cycloalkyl, tert-butyl, methyl, (C1-C5) hydroxyalkyl, phenyl, cycloalkyl, methoxy, aryl, sulphur dioxide, trifluoromethyl, and trifluoromethoxy.

2. The phthalimide compounds of formula (A)as claimed in claim 1, wherein:-X is selected form the group consisting of 6 membered heterocycloalkyl ring, and aniline optionally substituted with halogen or; an aminophenyl group, wherein the phenyl group is optionally substituted with halogen(s);-Ri is hydrogen;-R2 is selected from the group consisting of hydrogen, hydroxyl, tert-butyl;-R3 is selected from the group consisting of Hydrogen, (C1-C5) alkyl, (C1-C5) hydroxyalkyl, halogen, tert-butyl, methoxy, trifluoromethyl, and trifluoromethoxy.

3. The phthalimide compounds as claimed in claim 1, wherein the compounds of formula (A) is selected from the group of: i. 2-(2-hydroxy-3-(4-(2-hydroxyethyl)piperazin-l-yl)propyl)isoindoline-l, 3-dione; ii. 2-(2-hydroxy-3-(p-tolylamino)propyl)isoindoline-l,3- dione; iii. 2-(3-((4-(tert-butyl)phenyl)amino)-2- hydroxypropyl)isoindoline-l,3-dionedione); iv. 2-(2-hydroxy-3-((4- methoxyphenyl)amino)propyl)isoindoline-l ,3-dione);26v. 2-(3-((4-fluorophenyl)amino)-2- hydroxypropyl)isoindoline-l, 3-dione); vi. 2-(3-((4-bromophenyl) amino)-2-hydroxypropyl) isoindoline-1, 3-dione); vii. 2-(2-hydroxy-3-((4- (trifluoromethoxy )phenyl)amino)propyl)isoindoline- 1,3-dione); viii. 2-(2-hydroxy-3-((4- (trifluoromethyl)phenyl)amino)propyl)isoindoline-l, 3-dione); ix. 2-(2-hydroxy-3-((2- methoxyphenyl)amino)propyl)isoindoline-l ,3-dione); x. 2-(3-((3-fluorophenyl)amino)-2- hydroxypropyl)isoindoline-l ,3-dione; xi. 2-(3-((3-bromophenyl)amino)-2- hydroxypropyl)isoindoline- 1,3 -dione); xii. 2-(2-hydroxy-3-((3- (trifluoromethyl)phenyl)amino)propyl)isoindoline-l, 3-dione); xiii. 2-(2-hydroxy-3-((2- methoxyphenyl)amino)propyl)isoindoline-l, 3-dione ); xiv. 2-(3-((2-fluorophenyl)amino)-2- hydroxypropyl)isoindoline-l, 3-dione); xv. 2-(3-((2-bromophenyl)amino)-2- hydroxypropyl)isoindoline- 1,3 -dione): xvi. 2-(3-((2,4-difluorophenyl)amino)-2- hydroxypropyl)isoindoline-l ,3-dione). and the stereoisomers thereof.

4. A pharmaceutical composition comprising an effective amount of the phthalimide compound of formula A, as claimed in claim 1 to 2, and a pharmaceutically acceptable excipient, wherein:-X is piperazine;-Ri is Hydrogen;-R2 is hydroxyl;-R3 is (C1-C5) hydroxyalkyl.

5. The pharmaceutical composition as claimed in claim 4, wherein the pharmaceutically excipient is selected from the group of metal phosphate, metal carbonate, metal sulfate, hallites, metal oxides, sugar, alchohol, artificial sweeteners, starch, cellulose ester, cellulose ethers, polyethylene glycol, polypropylene glycol, glycerin, fatty acid esters, and their combination thereof.

6. A method of preparing the phthalimide compounds of formula A as claimed in claim 1 to 4, wherein the method comprises the steps of: a) preparing a mixture of compounds substituted isoindoline-1, 3-dione, and substituted heterocycloalkyl or aryl in a molar ratio (1:1), in ethanol solvent in a pressure seal tube; b) heating the mixture of step (a) under microwave; c) removing the ethanol solvent under vacuum on a rotary evaporator to obtain crude product; d) extracting the crude product with ethyl acetate and brine solution; e) separating the organic layer of the crude product and drying over anhydrous sodium sulphate; f) removing the excess ethyl acetate solution using a rotary evaporator to obtain the product; g) purifying the product obtained in step (f) to prepare the compound of formula A.

7. The method as claimed in claim 6, wherein the substituted isoindoline-1 , 3-dione is 2-(oxiran- 2-ylmethyl)isoindoline-l, 3-dione; and the substituted heterocycloalkyl is 2-(piperazin-l- yl)ethan-l-ol .

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