Rhamnolipids for modified gene expression during wound healing

Abstract

The technology presented herein, in general, relates to the use of biosurfactants, such as rhamnolipids, for treating and / or accelerating the healing of wounds. More particularly, the present technology relates to a method for treating chronic and acute wounds, by applying a composition comprising a mixture of rhamnolipids to the wound, wherein application of the composition facilitates coordinated gene expression associated with wound healing.

Description

Attorney Docket No. 68622WO01RHAMNOLIPIDS FOR MODIFIED GENE EXPRESSION AND ACCELERATED WOUND HEALINGCROSS-REFERENCE TO RELATED APPLICATIONSThis application claims priority to United States Provisional Application No. 63 / 724974, filed November 26, 2024, and to United States Provisional Application No. 63 / 724975, filed November 26, 2024. The entire content and specification of these provisional applications are hereby incorporated by reference.FIELD OF THE INVENTION

[0001] The present technology, in general, relates to the use of biosurfactants, such as rhamnolipids, for treating and / or accelerating the healing of wounds. More particularly, the present technology relates to a method for treating wounds, by applying a composition comprising a mixture of specific rhamnolipids to the wound, wherein application of the composition accelerates wound closure. In a further aspect, the present technology relates to a method for treating wounds, by applying a composition comprising a mixture of specific rhamnolipids to the wound, wherein application of the composition modifies the gene expression profile associated with the wound healing process.BACKGROUND OF THE INVENTION

[0002] The wound-healing process consists of a progression through a number of highly integrated and overlapping phases, including: hemostasis (e.g. vascular constriction, platelet aggregation, degranulation, and fibrin formation (thrombus));Attorney Docket No. 68622WO01 inflammation (e.g., neutrophil infiltration, monocyte infiltration and differentiation to macrophage, and lymphocyte infiltration); proliferation (e.g., re-epithelialization, angiogenesis, collagen synthesis, and ECM formation); and tissue remodeling or resolution (e.g., collagen remodeling, and vascular maturation and regression).

[0003] These phases, and their associated biophysiological functions, must occur in the proper sequence, at specific times and durations. Optimal wound healing in adult humans generally involves at least the following the events: (1 ) rapid hemostasis; (2) appropriate inflammation; (3) mesenchymal cell differentiation, proliferation, and migration to the wound site; (4) suitable angiogenesis; (5) prompt re-epithelialization (regrowth of epithelial tissue over the wound surface); and (6) proper synthesis, crosslinking, and alignment of collagen to provide strength to the healing tissue.

[0004] Wounds that exhibit both normal and impaired healing, including both acute and chronic wounds, generally have failed to progress through the normal stages of healing (i.e., enter a state of pathologic inflammation due to a postponed, incomplete, or uncoordinated healing process).

[0005] There is therefore an ongoing need for new wound healing compositions, and methods of use, that facilitate both a robust and coordinated healing process, including compositions and methods that facilitate acceleration of wound closure. Likewise, an ongoing need for new wound healing compositions, and methods of use, that facilitate the coordinated activation of genes involved in the normal stages of healing.

[0006] There has also been a recent trend to formulate products with ingredients that are based on renewable raw materials. Such ingredients are considered “green” or “natural”, since they are derived from renewable and / or sustainable sources. As a result,Attorney Docket No. 68622WO01 they are more environmentally friendly than ingredients derived from fossil fuels or other non-renewable sources. An ingredient having a high Bio-renewable Carbon Index (BCI), such as greater than 80, indicates that the ingredient contains carbons that are derived primarily from plant, animal or marine-based sources.

[0007] Rhamnolipids are interface-active glycolipids produced by various bacterial species and are an example of a “green” ingredient, since they can be prepared by means of fermentation based on renewable raw materials. It would be desirable to provide compositions that include active ingredients derived from renewable sources, such as rhamnolipids, that can be used to facilitate the treatment and healing of wounds. Providing wound healing / treatment compositions comprising rhamnolipids would satisfy sustainability goals of ensuring sustainable consumption through the use of bio-based antibacterial materials.

[0008] Applicants have determined that specific mixtures of rhamnolipid salts can meet the above objectives while also advancing UN Sustainability Goals (“SDG”). The rhamnolipid salt mixtures of the present technology contribute to better health and wellbeing by delivering equal or better efficacy in the treatment of wounds. The rhamnolipid salt mixtures are advantageously bio-based, renewably sourced actives obtained from a bacterial fermentation process that generates biodegradable waste products that are less impactful on the environment. These benefits further SDG #3 (Good Health and Wellbeing) and SDG #12 (Responsible Consumption and Production).Attorney Docket No. 68622WO01SUMMARY OF THE INVENTION

[0009] One aspect of the present technology is directed to compositions and methods for treating a wound. The methods of the present disclosure comprise applying a composition to the wound, wherein the composition comprises at least one rhamnolipid, and wherein application of the composition modifies the rate associated with wound healing and closure. In some embodiments, the wound being treated is an acute wound, including an incision, a laceration, an abrasion, an avulsion, a puncture, a penetration, or a burn wound. In other embodiments, the composition applied to the wound comprises mono-rhamnolipid, di-rhamnolipid, or a combination of both mono- and di-rhamnolipids. In further embodiments, in response to the application of the rhamnolipid compositions presented herein the rate or time to wound closure is accelerated.

[0010] In another aspect of the present technology is directed to compositions and methods for treating a wound. The methods of the present disclosure comprise applying a composition to the wound, wherein the composition comprises at least one rhamnolipid, and wherein application of the composition modifies the expression of at least one gene associated with one or more of the normal stages of healing. In some embodiments, the wound being treated is an acute wound, including an incision, a laceration, an abrasion, an avulsion, a puncture, a penetration, or a burn wound. Some embodiments include treating chronic wounds (e.g., diabetic foot ulcers (DFU), pressure ulcers (PU) or venous leg ulcers (VLU)). In other embodiments, the composition applied to the wound comprises mono-rhamnolipid, di-rhamnolipid, or a combination of both mono- and di-rhamnolipids. In further embodiments, the expression of at least one gene is increased or decreased in response to the application of the rhamnolipid compositions presented herein.Attorney Docket No. 68622WO01

[0011] In another aspect, the present technology is directed to a rhamnolipid composition for use in methods of treating wounds, wherein the composition comprises a mixture of mono-rhamnolipids and di-rhamnolipids having a mono-rhamnolipids:di- rhamnolipids weight ratio of about 40:60 to about 60:40, preferably about 40:60 to about 48:52. In addition, the rhamnolipid composition comprises, based on the total weight of rhamnolipids present in the composition: an amount of C10-C10 mono-rhamnolipid of about 29% to about 40% by weight, preferably 29% to about 37% by weight; an amount of C10-C10 di-rhamnolipid of about 35% to about 50% by weight, preferably about 35% to about 45%; an amount of C8-C10 mono-rhamnolipid of about 2% to about 5% by weight; an amount of C8-C10 di-rhamnolipid of about 2% to about 5% by weight; an amount of C10-C12 mono-rhamnolipid of about 2% to about 6% by weight; and an amount of C10-C12 di-rhamnolipid of about 8% to about 14% by weight. The composition further comprises at least one acceptable carrier, and optionally one or more additives, in an amount to total 100% by weight of the composition.BRIEF DESCRIPTION OF THE DRAWINGS

[0012] Figure 1 shows an example of the wound healing progression for the positive control (collagen hydrogel without rhamnolipid) versus 1 % SEP-RM rhamnolipid in collagen hydrogel at days 0, 3, 17, and 24.

[0013] Figure 2 shows a graph of the average wound size (normalized to day 0) for the positive control group (collagen hydrogel).

[0014] Figure 3 shows a graph of the average wound size (normalized to day 0) for the 1 % SEP-RM RL mixed with collagen hydrogel group.Attorney Docket No. 68622WO01DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0015] While the present technology will be described in connection with one or more preferred embodiments, it will be understood by those skilled in the art that the technology is not limited to only those specific embodiments. To the contrary, the presently described technology includes all alternatives, modifications, and equivalents as may be included within the spirit and scope of the appended claims.

[0016] Generally, the present disclosure is directed to compositions and methods for treating a wound. The methods disclosed herein comprise applying a composition to a wound, wherein the composition comprises at least one rhamnolipid, and wherein application of the composition to the wound accelerates wound closure. Wounds treated herein include, for example, acute wounds including incisions, lacerations, abrasions, avulsions, punctures, penetrations, or burns.

[0017] Generally, the present disclosure is directed to compositions and methods for treating a wound. The methods disclosed herein comprise applying a composition to a wound, wherein the composition comprises at least one rhamnolipid, and wherein application of the composition to the wound modifies the expression of at least one gene in the wound bed or the peri wound area. Wounds treated herein include, for example, acute wounds (incisions, lacerations, abrasions, avulsions, punctures, penetrations, or burns) and chronic wounds (DFLI, PU, and VLU).

[0018] As defined herein, a “rhamnolipid” is a glycolipid that has a lipid portion that includes one or more, typically linear, saturated or unsaturated B-hydroxy-carboxylic acid moieties and a saccharide portion of one or two units of rhamnose.Attorney Docket No. 68622WO01

[0019] The saccharide portion and the lipid portion are linked via a B-glycosidic bond between the 1 -OH group of a rhamnose moiety of the saccharide portion and the 3-OH group of a B-hydroxy-carboxylic acid of the lipid portion. Thus, the carboxylic acid of one carboxylic acid moiety defines the end of the rhamnolipid. Where more than one rhamnose-moiety is included in a rhamnolipid, each of the rhamnose moieties not linked to the lipid portion is linked to another rhamnose moiety via a 1 ,4B-glycosidic bond. In embodiments where two or more B-hydroxy-carboxylic acids are present in a rhamnolipid, the B-hydroxy-carboxylic acid moieties are selected independently from each other. B- hydroxy carboxylic acid moieties may in some embodiments be identical. In some embodiments, they are different from each other.

[0020] The present technology generally relates to a wound care / treatment composition that comprises a particular mixture of rhamnolipids in their salt form. The rhamnolipids may have the following structure (I):In this formula, R9is a hydrogen atom (H) or an aliphatic group that has a main chain of one to about 46, such as one to about 42, one to about 40, one to about 38, one to about 36, one to about 34, one to about 30, one to about 28, including e.g. 2, 3, 4, 5, 6, 7, 8, 9,Attorney Docket No. 68622WO0110, 1 1 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27 or 28 carbon atoms and one to about three, including two, oxygen atoms. In some embodiments, the main chain of the respective aliphatic group carries a terminal carboxylic acid group and / or an internal ester group. As an illustrative example in this regard, R9may be of the formula - CH(R5)— -CH2- COOR6. In these illustrative moieties, R5may be an aliphatic moiety with a main chain that has a length from 1 to about 19, such as from 1 to about 17, from 1 to about 15, from 1 to about 13, about 2 to about 13, about 3 to about 13 or about 4 to about 13, including e.g. 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 or 12 carbon atoms. R4in formula (I) is a hydrogen atom (H), or a rhamnopyranosyl moiety. R6is a hydrogen atom.

[0021] The term "aliphatic" means, unless otherwise stated, a straight or branched hydrocarbon chain, which may be saturated or mono- or poly-unsaturated and include heteroatoms. The term "heteroatom" as used herein means an atom of any element other than carbon or hydrogen. Herein, an unsaturated aliphatic group contains one or more double bonds (alkenyl moieties). The branches of the hydrocarbon chain may include linear chains as well as non-aromatic cyclic elements. The hydrocarbon chain, which may, unless otherwise stated, be of any length, and contain any number of branches. Typically, the hydrocarbon (main) chain includes 1 to about 5, to about 10, to about 15 or to about 20 carbon atoms. Examples of alkenyl moieties are straight-chain or branched hydrocarbon moieties that contain one or more double bonds. Alkenyl moieties generally contain about two to about twenty carbon atoms and one or more, for instance two, double bonds, such as about two to about ten carbon atoms, and one double bond. Examples of alkyl groups are methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, the n isomers of these radicals, isopropyl, isobutyl, isopentyl, sec-butyl, tert-butyl, neopentyl,Attorney Docket No. 68622WO013,3-dimethylbutyl. Both the main chain as well as the branches may furthermore contain heteroatoms as for instance N, O, S, Se or Si or a carbon atom may be replaced by one of these heteroatoms. An aliphatic moiety may be substituted or unsubstituted with one or more functional groups. Substituents may be any functional group, as for example, but not limited to, amino, amido, carbonyl, carboxyl, hydroxyl, nitro, thiol, and sulfonyl.

[0022] In a more particular embodiment, the rhamnolipid salts in said structure have the structure (II):wherein x is 1 or 2, y is 4, 6 or 8, z is 4, 6, or 8, and M is H, or a metal, such as alkali metals Li, Na, or K, alkali earth metals Mg or Ca, or transition metals Mn, Fe, Cu, or Zn. In the cases of the alkali earth and transition metals, multiple rhamnolipid salt moieties may associate with each metal.

[0023] The mixture of rhamnolipids preferably comprises mono (where x=1 ) and di (where x=2) rhamnolipids where y and z are 6 and M is H or Na. The mono-rhamnolipid may be referred to as Rha-C10-C10, with a formula of C26H48C>9. The IUPAC Name is 3-Attorney Docket No. 68622WO01[3-[(2R,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyloxan-2-yl]oxydecanoyloxy]decanoic acid. The di-rhamnolipid may be referred to as RhaRha-C10-C10, with a formula of C32H58O13. The IUPAC name is 3-[3-[ 4, 5-dihydroxy-6-methyl-3-(3,4,5-trihydroxy-6- methyloxan-2-yl)oxyoxan-2-yl]oxydecanoyloxy]decanoic acid. In general, the mixture of rhamnolipids disclosed herein comprises various types of mono and di rhamnolipids and the mixture specifically encompasses all possible combinations of mono and di rhamnolipids as disclosed herein. Further, unless otherwise stated, an amount of an individual mono or di-rhamnolipid as disclosed herein means that the respective mono or di-rhamnolipid can be present in the mixture of rhamnolipids in the indicated amount, and the mixture of rhamnolipids disclosed herein specifically includes all possible combinations of amounts of mono and di-rhamnolipids as disclosed herein. Generally preferred mixtures of rhamnolipids are SEP-RM rhamnolipid compositions as described below.

[0024] Rha-C10-C10 may be present in the mixture in an amount of about 29% to about 40%, alternatively about 30% to about 40%, alternatively about 29% to about 37%, alternatively about 35% to about 37% by weight based on the total weight of rhamnolipids. RhaRha-C10-C10 may be present in the mixture in an amount of about 35% to about 50%, alternatively about 35% to about 45%, alternatively about 36% to about 40%, alternatively about 36% to about 38% by weight based on the total weight of rhamnolipids.

[0025] In addition to Rha-C10-C10 and RhaRha-C10-C10, the mixture of rhamnolipids may comprise RhaRha-C10-C12 in an amount of about 8% to about 14%, alternatively about 9% to about 12%, alternatively about 10% to about 12.5% by weight based on the total weight of rhamnolipids, and Rha-C10-C12 in an amount of about 2% to about 6% byAttorney Docket No. 68622WO01 weight, alternatively about 2% to about 5%, alternatively about 3.5% to about 5% by weight based on the total weight of rhamnolipids. The mixture of rhamnolipids may also comprise RhaRha-C10-C12:1 in an amount of about 2% to about 5% by weight, alternatively about 3% to about 5% by weight, based on the total weight of rhamnolipids, an amount of RhaRha-C8-C10 in the range of about 2% to about 5% by weight, alternatively about 2% to about 4% by weight, based on the total weight of rhamnolipids, and an amount of Rha-C8-C10 in the range of about 2% to about 5% by weight, alternatively about 2% to about 4% by weight, based on the total weight of rhamnolipids. The mixture of rhamnolipids may also comprise Rha-Rha 012-012 in an amount of about 0.1 % to about 0.5% by weight, alternatively about 0.2% to about 0.4% by weight, alternatively about 0.2% to about 0.3% by weight, alternatively about 0.25% to about 0.3% by weight, based on the total weight of rhamnolipids present in the composition.

[0026] The mixture of rhamnolipids may comprise a mixture of mono-rhamnolipids and di-rhamnolipids. The mono-rhamnolipids may be present in an amount of about 40% to about 50%, preferably about 42% to about 48%, based on the total weight of rhamnolipids in the mixture. The di-rhamnolipids may be present in an amount of about 50% to about 60% by weight, preferably about 52% to about 58%, based on the total weight of rhamnolipids. The ratio of mono-rhamnolipids:di-rhamnolipids can be from about 40:60 to about 60:40, alternatively about 40:60 to about 50:50, alternatively about 40:60 to about 48:52, preferably about 42:58 to about 48:52.

[0027] The terms “active”, “% active”, and “% active weight” refer to the amount of the active ingredient without regard to the amount of water or other solvent that may be present with the ingredient.Attorney Docket No. 68622WO01

[0028] As used herein, “effective amount” refers to an amount of an active ingredient or composition that, when administered to a wound, is capable of accelerating or otherwise facilitating the healing process. The actual amount may vary depending on a number of factors, including, but not limited to, the severity of the wound, the age and health status of the subject, and the form of administration. Also, as used herein “accelerating” or accelerated” wound healing is in reference to either with or without a standard of care.

[0029] The mono-rhamnolipid may comprise one or more mono-rhamnolipid-mono- lipidic congeners, including for example: Rha-C8:2; Rha-C8; Rha-C10; Rha-C12:2; Rha- C12; Rha-C14:2; or combinations thereof. The mono-rhamnolipid may also comprise one or more mono-rhamnolipid-di-lipidic congeners, including for example: Rha-C8-C8; Rha- 08-010:1 ; Rha-C10:1 -C8; Rha-C8-C10; Rha-C10-C8; Rha-C10-C10:1 ; Rha-C10-C10; Rha-C8-C12; Rha-C12-C8; Rha-C10-C12:1 ; Rha-C12:1 -C10; Rha-C10-12; Rha-C12- C10; Rha-C10-C14:1 ; Rha-C12-C12:1 ; Rha-C10-C14; Rha-C12-C12; Rha-C12-C14; Rha-C14-C14; Rha-C14-C16; Rha-C16-C16; Rha-C10-C10-CH3; Decenoyl-Rha-C10- C10; or combinations thereof.

[0030] The di-rhamnolipid may comprise one or more di-rhamnolipid-mono-lipidic congeners, including for example: Rha-Rha-C8; Rha-Rha-C10; Rha-Rha-C12:1 ; Rha- Rha-C12; Rha-Rha-C14; or combinations thereof. The di-rhamnolipid may also comprise one or more di-rhamnolipid-di-lipidic congeners, including for example: Rha-Rha-C8-C8; Rha-Rha-C8-C10; Rha-Rha-C10-C8; Rha-Rha-C10-C10:1 ; Rha-Rha-C10-C10; Rha- Rha-C8-C12:1 ; Rha-Rha-C12:1 -C8; Rha-Rha-C10-C12:1 ; Rha-Rha-C12:1 -C10; Rha- Rha-C10-C12; Rha-Rha-C12-C10; Rha-Rha-C10-C14:1 ; Rha-Rha-C12-C12:1 ; Rha-Attorney Docket No. 68622WO01Rha-012 :1 -012 ; Rha-Rha-012-012; Rha-Rha-012-014; Rha-Rha-014-012; Rha-Rha- 014-C14; Rha-Rha-014-C16; Rha-Rha-C16-014; Rha-Rha-C16-016; Rha-Rha-014- C14-C14; Rha-Rha-010-010-CH3; Decenoyl-Rha-Rha-C10-C10; or combinations thereof.

[0031] In one aspect, the present technology provides a wound care composition, and methods of use, wherein the composition comprises a mixture of rhamnolipids in an amount of 0.1 % to 99% by weight, based on the total weight of the composition, wherein the mixture of rhamnolipids comprises mono-rhamnolipids and di-rhamnolipids in a weight ratio 40:60 to 60:40 mono-rhamnolipids:di-rhamnolipids, alternatively 40:60 to 50:50 mono-rhamnolipids:di-rhamnolipids, alternatively about 40:60 to about 48:52, alternatively 42:58 to 48:52 mono-rhamnolipids:di-rhamnolipids.

[0032] A further aspect of the present technology provides a method for treating a wound comprising administering to the wound an effective amount of a composition comprising a mixture of rhamnolipids, thereby accelerating or otherwise facilitating healing, wherein the mixture of rhamnolipids comprises mono-rhamnolipids and di- rhamnolipids in a weight ratio of 40:60 to 60:40 mono-rhamnolipids:di-rhamnolipids, alternatively 40:60 to 50:50 mono-rhamnolipids:di-rhamnolipids, alternatively 40:60 to 48:52 mono-rhamnolipids:di-rhamnolipids, alternatively 42:58 to 48:52 mono- rhamnolipids:di-rhamnolipids.

[0033] In some embodiments, the present technology provides a method for treating a wound, as described above, in which the mixture of rhamnolipids comprises mono- rhamnolipids and di-rhamnolipids in a weight ratio of 42:58 to 48:52, an amount of Rha- C10-C10 mono-rhamnolipid of about 29% to about 40% by weight, and an amount ofAttorney Docket No. 68622WO01RhaRha-C10-C10 di-rhamnolipid of about 35% to 50% by weight, based on the total weight of the rhamnolipids in the mixture of rhamnolipids.

[0034] The rhamnolipids may be produced from a rhamnolipid-producing microorganism that has the capacity to synthesize / produce rhamnolipids under suitable conditions. Such microorganisms include, but are not limited to, bacteria, particularly bacteria of the phyla Pseudomonadota, Actinobacteria, Fimicutes, and Proteobacteria. The rhamnolipids are naturally derived and therefore have a BCI of 100. In a particular embodiment, the rhamnolipid-producing microorganism for producing the rhamnolipids is Pseudomonas aeruginosa. Methods of culturing the rhamnolipid-producing bacteria and the production of rhamnolipids from fermentation are known in the art from, for example U.S. Patent No 1 1 ,142,782 and U.S. Patent No. 10,144,943, incorporated herein by reference in their entirety. Methods of purifying the rhamnolipids are also known in the art from, for example, U.S. Patent No 9,884,883 and U.S. Patent No. 10,829,507, incorporated herein by reference in their entirety.

[0035] The mixture of rhamnolipid salts can be used alone, as the sole active ingredient in the wound care I treatment composition. When used alone, the mixture of rhamnolipids may be in the range of about 0.01 % to about 99% by active weight, based on the total weight of the composition, alternatively about 0.02% to about 25%, alternatively about 0.1 % to about 10%, alternatively about 0.2% to about 6% by active weight, based on the total weight of the composition. The mixture of rhamnolipid salts may be used as a coactive in combination with another active ingredient, such humectants and other skin wellness ingredients (e.g., that help promote skin wellness and improve the skin’s natural barrier properties). The mixture of rhamnolipid salts may also be used as a co-active inAttorney Docket No. 68622WO01 combination with another active ingredient, such as antibiotics, antimicrobial agents, vitamins (e.g., vitamins E, A, and C), Hyaluronic Acid, proteins, peptides, and nucleic acids. When used in combination, the mixture of rhamnolipids may be in the range of about 0.01 % to about 99% by active weight, based on the total weight of the composition, alternatively about 0.02% to about 25%, alternatively about 0.1 % to about 10%, alternatively about 0.2% to about 6% by active weight, by active weight based on the total weight of the composition. The combination of the mixture of rhamnolipid salts and another co-active ingredient may help to alleviate the irritation potential of the co-active without reducing or inhibiting its activity. Combining the mixture of rhamnolipid salts with another co-active ingredient may also allow for the reduction of the other co-active ingredients, which can also help to reduce the overall irritation potential.

[0036] The wound care / treatment compositions can be formulated into any treatment form commonly used for dermatological / topical applications. For example, the compositions can be in the form of an aqueous solution, suspension, cream, lotion, gel, paste, spray, cream, foam or emollient, or impregnated onto pads, wipes, bandages and / or dressings. Potential dressings include, for example, polyurethane foams, methylcellulose, alginates, collagen, in addition to cellular and acellular matrix-like products.

[0037] The wound care I treatment compositions of the present technology also include at least one carrier suitable for wound care I treatment to bring the total percentage of the composition to 100%. As will be appreciated by at least those skilled in the art, a variety of carriers, vehicles, diluents, and the like are suitable for use in the practice of the present technology. Thus, it will also be appreciated that the terms “carrier”, “vehicle”, andAttorney Docket No. 68622WO01“diluent” are to be considered non-exhaustive and interchangeable with respect to the present technology and in describing the various formulations, applications, uses, and compositions thereof.

[0038] Water is a suitable carrier, and can be de-ionized water, hard water, soft water, distilled water, tap water or combinations thereof. Water can be used alone as the carrier, or in combination with other carriers suitable for personal care, such as for example, alcohols such as ethanol, isopropanol, or benzyl alcohol; glycols such as propylene glycol, or polyethylene glycol. Other carriers can include, but are not limited to solvents, emulsifiers, or solubilizers.

[0039] When the treatment form is a cream, gel, or paste, the wound care I treatment compositions can include, but are not limited to, vegetable gums, starches, celluloses, waxes, silicone, silica, or clays, as carrier ingredients. When the treatment form is a spray, the composition may include a propellant, film formers, formation fibers, and / or quick drying agents to promote bandage formation.

[0040] In addition to the rhamnolipid active and carrier, the wound care I treatment compositions of the present technology can include optional ingredients as known in the art. Such other components or additives can include, but are not limited to, surfactants, pH adjustment agents, skin conditioners, antioxidants, preservatives, fragrances, pigments, dyes, and other excipients (e.g., anesthetics such as Benzocaine, and other antibiotics).

[0041] The wound care / treatment compositions of the present technology can have pH values in the range of about 4.0 to about 8.5, alternatively, about 5.0 to about 8.0, ideally 5.5 to 7.0.Attorney Docket No. 68622WO01

[0042] The wound care / treatment compositions of the present technology may be used by applying the composition to the wound of a subject in an amount effective to treat, and / or otherwise facilitate wound healing. “Applying” can refer to any commonly used method of application, such as, but not limited to, spreading a cream or gel containing the wound care / treatment composition on the surface of the wound and allowing the cream or gel to remain on the wound; spraying a liquid containing the wound care / treatment composition on the surface of the wound and, if desired, surrounding tissue; wiping the wound with a wipe impregnated with the wound care / treatment composition and allowing the composition to remain on the wound and, if desired, surrounding tissue; applying a pad impregnated with the wound care / treatment composition and allowing the pad to remain on the surface of the wound and, if desired, surrounding tissue; or an aqueous or non-aqueous liquid wash intended to treat the surface of the wound and, if desired, the surrounding tissue.

[0043] Dosage forms and treatment regimens using the wound care / treatment compositions of the present technology can vary with the type and grade or intensity of the wound (e.g., based on recommended clinical guidelines). In one or more embodiments, methods of treatment in accordance with the present technology may use a one, two, three, four, or more daily dosage regime. The daily dosage regimen can continue for 1 -6 days, alternatively one, two, three, four, five, six, or more weeks according to the condition and response of the patient. In another embodiment, methods of treatment in accordance with the present technology may use once a day, 2-3 times per week, or once per week regime.Attorney Docket No. 68622WO01

[0044] Methods of the present disclosure comprise applying a rhamnolipid containing wound care / treatment composition to a wound, wherein application of the composition accelerates wound closure.

[0045] Methods of the present disclosure comprise applying a rhamnolipid containing wound care / treatment composition to a wound, wherein application of the composition modifies the expression of at least one gene associated with one or more of the normal stages of healing, including: hemostasis (e.g. vascular constriction, platelet aggregation, degranulation, and fibrin formation (thrombus)); inflammation (e.g., neutrophil infiltration, monocyte infiltration and differentiation to macrophage, and lymphocyte infiltration); proliferation (e.g., re-epithelialization, angiogenesis, collagen synthesis, and ECM formation); and tissue remodeling or resolution (e.g., collagen remodeling, and vascular maturation and regression).

[0046] In one aspect of the present disclosure, the application to a wound of a rhamnolipid composition contemplated herein results in the activation or repression of gene expression for at least one of the following genes: Xaa-Pro aminopeptidase 1 (XPNPEP1 ); autophagy related 14 (ATG14); cannabinoid receptor 1 (CNR1 ); Jun protooncogene (JUN); TIMP metallopeptidase inhibitor 3 (TIMP3); colony stimulating factor 2 receptor subunit beta (CSF2RB); interleukin 1 beta (IL1 B); platelet factor 4 (PF4); colony stimulating factor 3 receptor (CSF3R); nuclear receptor subfamily 4, group A, member 1 (NR4A1 ); platelet derived growth factor receptor alpha (PDGFRA); platelet derived growth factor receptor beta (PDGFRB); paired immunoglobin like type 2 receptor beta (PILRB); TNF receptor superfamily member 1 B (TNFRSF1 B); tyrosine kinase with immunoglobulin-like and EGF-like domains 1 (TIE1 ); leucyl-tRNA synthetase 1 (LARS1 );Attorney Docket No. 68622WO01 pannexin 1 (PANX1 ); plexin domain containing 2 (PLXDC2); secreted frizzled related protein 1 (SFRP1 ); cathepsin K (CTSK); matrix metallopeptidase 14 (MMP14); integrin subunit beta 5 (ITGB5); discs large MAGUK scaffold protein 4 (DLG4); platelet derived growth factor subunit A (PDGFA); or combinations thereof.

[0047] In another aspect of the present disclosure, the application to a wound of a rhamnolipid composition contemplated herein results in the activation (i.e. , upregulation) of gene expression during inflammation (e.g., at day 3 after wound) for at least one of the following genes: TIE1 ; LARS1 ; PANX1 ; PLXDC2; SFRP1 ; CTSK; MMP14; ITGB5; DLG4; and PDGFA.

[0048] In a further aspect of the present disclosure, the application to a wound of a rhamnolipid composition contemplated herein results in the activation (i.e., upregulation) of gene expression during proliferation (e.g., at day 17 after wound) for at least one of the following genes: CSF2RB; IL1 B; PF4; CSF3R; NR4A1 ; PDGFRA; PDGFRB; PILRB; and TNFRSF1 B.

[0049] In a still further aspect of the present disclosure, the application to a wound of a rhamnolipid composition contemplated herein results in the deactivation or suppression (i.e., downregulation) of gene expression during proliferation (e.g., at day 17 after wound) for at least one of the following genes: CNR1 ; TIMP3; ATG14; JUN; and XPNPEP1 .ITEMS OF THE DISCLOSUREThe current disclosure further relates to any one or more of the following items:1 A rhamnolipid composition for use in methods of accelerating wound closure, wherein the composition comprises a mixture of mono-rhamnolipids andAttorney Docket No. 68622WO01 di-rhamnolipids having a mono-rhamnolipids:di-rhamnolipids weight ratio of about 40:60 to about 60:40; and wherein application of the composition to a wound accelerates wound closure. The rhamnolipid composition also comprises, based on the total weight of rhamnolipids present in the composition, an amount of 010- C10 mono-rhamnolipid of about 29% to about 40% by weight, an amount of C10- C10 di-rhamnolipid of about 35% to about 50% by weight, an amount of C8-C10 mono-rhamnolipid of about 2% to about 5% by weight; an amount of C8-C10 di- rhamnolipid of about 2% to about 5% by weight; an amount of C10-C12 mono- rhamnolipid of about 2% to about 6% by weight; and an amount of C10-C12 di- rhamnolipid of about 8% to about 14% by weight. The composition also further comprises at least one acceptable carrier, and optionally one or more additives, in an amount to total 100% by weight of the composition.2. A rhamnolipid composition for use in methods of modifying gene expression in a target tissue, wherein the composition comprises a mixture of monorhamnolipids and di-rhamnolipids having a mono-rhamnolipids:di-rhamnolipids weight ratio of about 40:60 to about 60:40. The rhamnolipid composition also comprises, based on the total weight of rhamnolipids present in the composition, an amount of 010-010 mono-rhamnolipid of about 29% to about 40% by weight, an amount of 010-010 di-rhamnolipid of about 35% to about 50% by weight, an amount of 08-010 mono-rhamnolipid of about 2% to about 5% by weight; an amount of 08-010 di-rhamnolipid of about 2% to about 5% by weight; an amount of 010-012 mono-rhamnolipid of about 2% to about 6% by weight; and an amount of C10-C12 di-rhamnolipid of about 8% to about 14% by weight. The compositionAttorney Docket No. 68622WO01 also further comprises at least one acceptable carrier, and optionally one or more additives, in an amount to total 100% by weight of the composition. The target tissue is preferably derived from a wound, or an area surrounding a wound.3. The composition of the preceding items, wherein the mixture of rhamnolipids comprises an amount of total mono rhamnolipid of about 40% to about 50% by weight, preferably about 40% to about 48% by weight, such as about 42 % to about 48% by weight, or about 43% to about 47% by weight, or about 43% to about 45% by weight, based on the total weight of rhamnolipids present in the composition.4. The composition of any one of the preceding items, wherein the mixture of rhamnolipids comprises an amount of total di-rhamnolipid of about 50% to about 60% by weight, preferably about 52% to about 60% by weight, such as about 52% to about 58% by weight, or about 53% to about 57% by weight, or about 55% to about 57% by weight, based on the total weight of rhamnolipids present in the composition.5. The composition of any one of the preceding items, wherein the mixture of rhamnolipids comprises an amount of the mono-rhamnolipid C8-C10 congener in the range of about 2% to about 4% by weight, based on the total weight of rhamnolipids present in the composition.6. The composition of any one of the preceding items, wherein the mixture of rhamnolipids comprises an amount of the mono-rhamnolipid C10-C10 congener in the range of about 29% to about 37% by weight, preferably about 32% to about 37% by weight, such as about 34% to about 37% by weight, or about 35% to aboutAttorney Docket No. 68622WO0137% by weight, based on the total weight of rhamnolipids present in the composition.7. The composition of any one of the preceding items, wherein the mixture of rhamnolipids comprises an amount of the mono-rhamnolipid C10-C12 congener in the range of about 2% to about 5% by weight, preferably about 3.5% to about 5% by weight, based on the total weight of rhamnolipids present in the composition.8. The composition of any one of the preceding items, wherein the mixture of rhamnolipids comprises an amount of the di-rhamnolipid C8-C10 congener in the range of about 2% to about 4% by weight, based on the total weight of rhamnolipids present in the composition.9. The composition of any one of the preceding items, wherein the mixture of rhamnolipids comprises an amount of the di-rhamnolipid C10-010 congener in the range of about 35% to about 45% by weight, preferably about 36% to about 40% by weight, such as about 36% to about 38% by weight, based on the total weight of rhamnolipids present in the composition.10. The composition of any one of the preceding items, wherein the mixture of rhamnolipids comprises an amount of the di-rhamnolipid C10-C12.1 congener in the range of about 2% to about 5% by weight, preferably about 3% to about 5% by weight, such as about 3% to about 4% by weight, based on the total weight of rhamnolipids present in the composition.1 1 . The composition of any one of the preceding items, wherein the mixture of rhamnolipids comprises an amount of the di-rhamnolipid C10-C12 congener in theAttorney Docket No. 68622WO01 range of about 9% to about 12% by weight, preferably in the range of about 10% to about 12% by weight, or about 10% to about 12.5% by weight, based on the total weight of rhamnolipids present in the composition.12. The composition of any one of the preceding items, wherein the mixture of rhamnolipids comprises an amount of the di-rhamnolipid C12-C12 congener in the range of about 0.1 % to about 0.5% by weight, preferably about 0.2% to about 0.4% by weight, such as about 0.2% to about 0.3% by weight, or about 0.25% to about 0.3% by weight, based on the total weight of rhamnolipids present in the composition.13. The composition of any one of the preceding items, wherein the mixture of rhamnolipids is a SEP-RM Rhamnolipid composition.14. The composition of any one of the preceding items, wherein the mixture of rhamnolipids is a SEP-RM Rhamnolipid composition obtained by a solvent extraction process including a bleaching step after acidulation and before the solvent extraction.15. The composition of any one of the preceding items, wherein modification of gene expression in a target tissue includes the activation (i.e., upregulation) of gene expression during inflammation (e.g., at day 3) for at least one of the following genes: TIE1 ; LARS1 ; PANX1 ; PLXDC2; SFRP1 ; CTSK; MMP14; ITGB5; DLG4; or PDGFA.16. The composition of any one of the preceding items, wherein modification of gene expression in a target tissue includes the activation (i.e., upregulation) ofAttorney Docket No. 68622WO01 gene expression during proliferation (e.g., at day 17) for at least one of the following genes: CSF2RB; IL1 B; PF4; CSF3R; NR4A1 ; PDGFRA; PDGFRB; PILRB1 ; or TNFRSF1 B.17. The composition of any one of the preceding items, wherein modification of gene expression in a target tissue includes the deactivation or suppression (i.e. , downregulation) of gene expression during proliferation (e.g., at day 17) for at least one of the following genes: CNR1 ; TIMP3; ATG14; JUN; or XPNPEP1 .EXAMPLES

[0050] The presently described technology and its advantages will be better understood by reference to the following examples. These examples are provided to describe specific embodiments of the present technology. By providing these examples, the inventors do not limit the scope and spirit of the present technology.Experimental Procedures***Test MaterialsRhamnolipid Compositions

[0051] Rhamnolipids are produced, as understood in the art, through the fermentation of rhamnolipid producing bacteria such as Pseudomonas aeruginosa. The rhamnolipids may be produced from a rhamnolipid-producing microorganism that has the capacity to synthesize / produce rhamnolipids under suitable conditions. Such microorganisms include, but are not limited to, bacteria, particularly bacteria of the phyla Pseudomonadota, Actinobacteria, Fimicutes, and Proteobacteria. The rhamnolipids areAttorney Docket No. 68622WO01 naturally derived and therefore have a BCI of 100. In a particular embodiment, the rhamnolipid-producing microorganism for producing the rhamnolipids is Pseudomonas aeruginosa. Methods of culturing the rhamnolipid-producing bacteria and the production of rhamnolipids from fermentation are known in the art from, for example U.S. Patent No 1 1 ,142,782 and U.S. Patent No. 10,144,943, incorporated herein by reference in their entirety. Methods of purifying the rhamnolipids are also known in the art from, for example, U.S. Patent No 9,884,883 and U.S. Patent No. 10,829,507, incorporated herein by reference in their entirety.

[0052] Fermentation whole broth then undergoes multiple purification steps. The specific combination of steps depends on the rhamnolipid actives and purity requirements of the target application. Certain rhamnolipid mixtures used herein are purified by a solvent extraction process. As used herein, the acronym SEP-RM refers to Solvent Extraction Purified Rhamnolipid Mixture (or composition), in accordance with the current specification.

[0053] For example, the SEP-RM Rhamnolipid compositions used herein are prepared in accordance with at least the following processing steps: (1 ) fermentation of appropriate Rhamnolipid producing bacterium; (2) biomass separation; (3) sterilization; (4) clarification (e.g. filtration); (5) acidulation; (6) bleaching; (7) washing; (8) solvent extraction; (9) neutralization and dilution; and (10) final polishing step(s). As discussed below, the solvent extraction process / step is intended to yield a higher purity rhamnolipid mixture / composition for personal care applications. The additional solvent extraction steps provide a rhamnolipid composition having a lighter color and milder odor profile that are preferred for the personal care market.Attorney Docket No. 68622WO01

[0054] At the end of fermentation, the whole broth typically contains rhamnolipids along with biomass and other by-products of fermentation. To separate the biomass solids, the broth can be centrifuged. The resulting centrifuged broth is then subjected to sterilization (e.g., high temperature sterilization), after which, the centrifuged sterilized broth is clarified by filtration to remove suspended solids.

[0055] Further purification is achieved by treating the clarified broth with acid, which converts the rhamnolipid to a water-insoluble form that settles to the bottom and separates from the bulk aqueous phase. This dense acidulated rhamnolipid is then isolated (and referred to as Acidulated, Concentrated Clarified Broth (ACCB). ACCB is then treated with bleaching agent, then water washed to remove residual bleaching agent and other water-soluble impurities. At this point, the decolorized washed ACCB is concentrated (e.g., > about 45% actives, or between about 35% to about 55% actives) and purer (e.g., > about 75% purity, or between about 65% to about 85% purity).

[0056] Solvent Extraction Process - To achieve better purity, color, and odor, the decolorized Acidulated, Concentrated Clarified Broth (ACCB) undergoes a solvent extraction process. This is generally performed by dissolving the decolorized, washed ACCB in organic solvent, preferably ethyl acetate. The rag layer that typically forms is separated from the bulk solution. Activated carbon is then added to the rhamnolipid solution in ethyl acetate for further decolorization and deodorization. The slurry is filtered and the resulting solution is stripped under vacuum to remove ethyl acetate. The highly concentrated ACCB obtained as residue is then neutralized and diluted (e.g., from between about 20% to about 30% actives, preferably about 25% actives) to give crude SEP-RM. As a final polishing step, crude SEP-RM is then washed with ethyl acetate toAttorney Docket No. 68622WO01 extract the antifoam and yield the final product as optically clear SEP-RM at about 25% active and about 85% purity.Mono-Rhamnolipid:Di-Rhamnolipid Ratio and Congener Distribution

[0057] The mono-rhamnolipid:di-rhamnolipid ratio, and congener distribution for SEP- RM rhamnolipid compositions of the present disclosure can be determined using HPLC (High Performance Liquid Chromatography). As understood by the person of ordinary skill in the art, HPLC will separate rhamnolipid congeners by chain length and number of rhamnose moieties (with the more polar congeners eluting first). Fractions are collected and analyzed by mass spec to identify each peak. The congener distribution is reported as area percent (with actives being calculated relative to known standards).

[0058] The HPLC congener distribution for SEP-RM rhamnolipid compositions of the present technology may comprise mono-rhamnolipid C8-C10 congener in an amount of about 2% to about 5% by weight, di-rhamnolipid C8-C10 congener in an amount of about 2% to about 5% by weight, mono-rhamnolipid C10-C10 congener in an amount of about 29% to about 40% by weight, preferably about 35% to about 40% by weight, di- rhamnolipid C10-C10 congener in an amount of about 35% to about 50 weight, preferably about 35% to about 45% by weight, mono-rhamnolipid C10-C12 congener in an amount of about 2% to about 6% by weight, di-rhamnolipid C10-C12 congener in an amount of about 8% to about 14% by weight, with total mono-rhamnolipid congeners in an amount of about 40% to about 48% by weight, and total di-rhamnolipid congeners in an amount of about 52% to about 60% by weight, based on the total weight of the congeners in the composition.Attorney Docket No. 68622WO01

[0059] The mixture of rhamnolipids comprises a mixture of mono-rhamnolipids and dirhamnolipids. In some embodiments, the mono-rhamnolipids may be present in an amount of about 10% to about 90%, alternatively about 15% to about 90%, alternatively about 20% to about 90%, alternatively about 30% to about 90%, alternatively about 40% to about 90%, alternatively about 10% to about 80%, alternatively about 20% to about 80%, alternatively about 30% to about 80%, alternatively about 40% to about 80%, alternatively about 10% to about 70%, alternatively about 15% to about 70%, alternatively about 20% to about 70%, alternatively about 30% to about 70% by weight, based on the total weight of rhamnolipids in the mixture and as measured by high-performance liquid chromatography (HPLC).

[0060] In some embodiments, the mono-rhamnolipids may be present in an amount of about 10% to about 60%, alternatively about 15% to about 60%, alternatively about 20% to about 60%, alternatively about 25% to about 60%, alternatively about 30% to about 60%, alternatively about 25% to about 50%, alternatively about 10% to about 48%, alternatively about 15% to about 48%, alternatively about 20% to about 48%, alternatively about 25% to about 48%, alternatively about 30% to about 48% by weight, based on the total weight of rhamnolipids in the mixture, as determined by HPLC.

[0061] In some embodiments, the di-rhamnolipids may be present in an amount of about 10% to about 90%, alternatively about 15% to about 90%, alternatively about 20% to about 90%, alternatively about 30% to about 90%, alternatively about 40% to about 90%, alternatively about 50% to about 90%, alternatively about 10% to about 80%, alternatively about 20% to about 80%, alternatively about 30% to about 80%, alternatively about 40% to about 80%, alternatively about 50% to about 80%, alternatively about 10%Attorney Docket No. 68622WO01 to about 75%, alternatively about 15% to about 75%, alternatively about 20% to about 75%, alternatively about 30% to about 75%, alternatively about 40% to about 75%, alternatively about 50% to about 75% by weight, based on the total weight of rhamnolipids in the mixture, as determined by HPLC.

[0062] In some embodiments, the di-rhamnolipids may be present in an amount of about 40% to about 70%, alternatively about 50% to about 70%, alternatively about 52% to about 90%, alternatively about 52% to about 85%, alternatively about 52% to about 80%, alternatively about 52% to about 75%, alternatively about 52% to about 70%, based on the total weight of rhamnolipids in the mixture, as determined by HPLC.

[0063] The ratio of mono-rhamnolipids:di-rhamnolipids can be from about 10:90 to about 90:10, alternatively 10:90 to about 60:40, alternatively about 15:85 to about 60:40, alternatively about 20:80 to about 60:40, alternatively about 25:75 to about 60:40, alternatively about 30:70 to about 60:40, alternatively about 10:90 to about 48:52, alternatively about 15:85 to about 48:52, alternatively about 20:80 to about 48:52, alternatively about 25:75 to about 48:52, alternatively about 30:70 to about 48:52. In preferred embodiments, the ratio of mono-rhamnolipids:di-rhamnolipids is about 25:75 to about 60:40, alternatively about 25:75 to about 50:50, or about 25:72 to about 48:52.

[0064] The mixture of rhamnolipids preferably comprises mono (where x=1 ) and di (where x=2) rhamnolipids where y and z are 6 and M is H or Na. The mono-rhamnolipid may be referred to as Rha-C10-C10, with a formula of C26H48O9. The IUPAC Name is 3-[3-[(2R,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyloxan-2-yl]oxydecanoyloxy]decanoic acid. The di-rhamnolipid may be referred to as RhaRha-C10-C10, with a formula ofAttorney Docket No. 68622WO01C32H58O13. The IUPAC name is 3-[3-[ 4, 5-dihydroxy-6-methyl-3-(3,4,5-trihydroxy-6- methyloxan-2-yl)oxyoxan-2-yl]oxydecanoyloxy]decanoic acid.

[0065] Rha-C10-010 may be present in the mixture in an amount of about 5% to about 85%, alternatively about 5% to about 50%, alternatively about 5% to about 38%, alternatively about 10% to about 38%, alternatively about 15% to about 38%, alternatively about 20% to about 38%, alternatively about 25% to about 37.5%, alternatively about 29% to about 37.5%, by weight based on the total weight of rhamnolipids. RhaRha-C10- C10 may be present in the mixture in an amount of about 5% to about 85%, alternatively about 10% to about 80%, alternatively about 20% to about 80%, alternatively about 30% to about 80%, alternatively about 35% to about 75%, alternatively about 35% to about 65%, alternatively about 35% to about 60%, alternatively about 35% to about 55%, alternatively about 35.5% to about 50%, alternatively about 36% to about 45%, alternatively about 40% to about 60% by weight based on the total weight of rhamnolipids.

[0066] In addition to Rha-C10-C10 and RhaRha-C10-C10, the mixture of rhamnolipids may comprise RhaRha-C10-C12 in an amount of about 3% to about 15%, alternatively about 3% to about 12%, alternatively about 3% to about 12.5% by weight based on the total weight of rhamnolipids, and Rha-C10-C12 in an amount of about 0.2% to about 6% by weight, alternatively about 0.4% to about 5% by weight based on the total weight of rhamnolipids. The mixture of rhamnolipids may also comprise RhaRha-C10-C12:1 in an amount of about 0.04% to about 4% by weight, alternatively about 0.05% to about 4% by weight, alternatively about 0.05% to about 3% by weight, based on the total weight of rhamnolipids, an amount of RhaRha-C8-C10 in the range of about 0.2% to about 5% by weight, alternatively about 0.6% to about 4% by weight, based on the total weight ofAttorney Docket No. 68622WO01 rhamnolipids, and an amount of Rha-C8-C10 in the range of about 0.2% to about 5% by weight, alternatively about 0.6% to about 4% by weight, based on the total weight of rhamnolipids. In some embodiments, the mixture of rhamnolipids may also comprise an amount of RhaRha-C12-C12 in the range of about 3% to about 10% by weight, based on the total weight of rhamnolipids in the mixture.

[0067] In some embodiments, the mixture of rhamnolipids comprises a mixture of mono-rhamnolipids and di-rhamnolipids in a weight ratio of mono-rhamnolipids:di- rhamnolipids in the range of about 25:75 to about 60:40, alternatively about 25:75 to about 50:50, or alternatively about 25:75 to about 45:55.

[0068] The mono-rhamnolipid may comprise one or more mono-rhamnolipid-mono- lipidic congeners, including for example: Rha-C8-:2; Rha-C8; Rha-C10; Rha-C12:2; Rha- C12; Rha-C14:2; or combinations thereof. The mono-rhamnolipid may also comprise one or more mono-rhamnolipid-di-lipidic congeners, including for example: Rha-C8-C8; Rha- C8-C10:1 ; Rha-C10:1 -C8; Rha-C8-C10; Rha-C10-C8; Rha-C10-C10:1 ; Rha-C10-C10; Rha-C8-C12; Rha-C12-C8; Rha-C10-C12:1 ; Rha-C12:1 -C10; Rha-C10-12; Rha-C12- C10; Rha-C10-014:1 ; Rha-C12-C12:1 ; Rha-C10-C14; Rha-C12-C12; Rha-C12-C14; Rha-C14-C14; Rha-C14-C16; Rha-C16-C16; Rha-C10-C10-CH3; Decenoyl-Rha-C10- C10; or combinations thereof.

[0069] The di-rhamnolipid may comprise one or more di-rhamnolipid-mono-lipidic congeners, including for example: Rha-Rha-C8; Rha-Rha-C10; Rha-Rha-C12:1 ; Rha- Rha-C12; Rha-Rha-C14; or combinations thereof. The di-rhamnolipid may also comprise one or more di-rhamnolipid-di-lipidic congeners, including for example: Rha-Rha-C8-C8; Rha-Rha-C8-C10; Rha-Rha-C10-C8; Rha-Rha-C10-C10:1 ; Rha-Rha-C10-C10; Rha-Attorney Docket No. 68622WO01Rha-C8-C12:1 ; Rha-Rha-012:1 -C8; Rha-Rha-010-012:1 ; Rha-Rha-C12:1 -C10; Rha- Rha-C10-C12; Rha-Rha-C12-C10; Rha-Rha-C10-C14:1 ; Rha-Rha-C12-C12:1 ; Rha- Rha-C12:1 -C12; Rha-Rha-C12-C12; Rha-Rha-C12-C14; Rha-Rha-014-012; Rha-Rha- 014-014; Rha-Rha-014-016; Rha-Rha-016-014; Rha-Rha-016-016; Rha-Rha-014- 014-014; Rha-Rha-010-010-CH3; Decenoyl-Rha-Rha-010-010; or combinations thereof.

[0070] The HPLC congener distribution for an exemplary SEP-RM rhamnolipid composition of the present disclosure, is presented below:Optional Bleaching Step

[0071] The SEP-RM rhamnolipid composition provided herein can be processed either with, or without a bleaching step (occurring after acidulation, and before the solventAttorney Docket No. 68622WO01 extraction step(s)). For the SEP-RM rhamnolipid compositions of the present disclosure, the mono-rhamnolipid:di-rhamnolipid ratio, and the associated congener distribution, are unaffected by whether a bleaching step is included, or not included, in the purification process.Wound Closure Analysis

[0072] Wound closure, such as that shown in Figure 1 , can be quantified by measuring the open area of the wound through the duration of healing. Percent wound closure can be analyzed using NIH Imaged (Bethesda, MD) by standardizing each image using the millimeter ruler captured in each photo. The open wound margin can be traced with a freehand tool, generating an area in mm2. The wound bed for each day is then normalized to time 0 using the following equation: 100 = Percent Wound Closurewhere DO = Initial wound area measurement and Dn = Day of wound area measurement.Example 1

[0073] This study determined how certain rhamnolipid compositions impacted the progression of wound healing, and in particular the rate of wound healing determined as set forth above, and in this Example.

[0074] The objective of the study was to evaluate two concentrations, 1 % and 3%, of a SEP-RM Rhamnolipid gel in an advanced wound care prototype. The animal model used was the male BKS.Cg-Dock7m + / + Lepr db / J (Jax Stock # 000642; Genotype: Homozygous genotype, Wildtype for Dock7<m>, Homozygous for Lepr<db>) diabeticAttorney Docket No. 68622WO01 mouse delayed wound healing model. The study design, using SEP-RM Rhamnolipid, is summarized below in Table 1 .Table 1 : Diabetic Mouse Wound Model Study Designmg = milligram; mL = milliliter*n=3 (sacrificed during inflammatory phase, Day 3-5)** n=3 sacrificed during proliferative phase, Day 1 1 -18

[0075] As indicated in Table 1 , wound creation, on Day 0, included a skin biopsy (10 mm disc) removed from the dorsal right side of the mouse model. The study included 6 treatment groups, including: 2 x control groups (without rhamnolipid); 2 x 1 % SEP-RM rhamnolipid (dose level = 0.1 mg; dose concentration = 10 mg / mL, 1 %; and dose volumeAttorney Docket No. 68622WO01= 100 uL); and 2 x 3% SEP-RM rhamnolipid (dose level = 0.2 mg; dose concentration = 20 mg / mL, 20%; and dose volume = 100 uL). The dose route was topical (to the area of the skin biopsy), at days 0, 3, 7, 10, 14, 17, and 21 . The study end was set at 24 days.

[0076] The Control group included Control Gel Collagen, with ambient storage. Rhamnolipid treatment group 1 comprised 1 % SEP-RM rhamnolipid in Collagen (Coloplast 7690 Woun'dres Collagen Hydrogel), with ambient storage. Rhamnolipid treatment group 2 comprised 3% SEP-RM rhamnolipid in Collagen, with ambient storage. The wound dressing comprised 3M Tegaderm™ dressing, with ambient storage.

[0077] As indicated above, the test system included species Mus muscuus (strain = BKS.Cg-Dock7m + / + Leprdb / J). The source was Jackson Laboratories (Stock# 000642; Genotype: Homozygous genotype, Wildtype for Dock7<m>, Homozygous for Lepr<db>). The age of the animals upon arrival was 8 weeks, and before use the animals were held for a minimum of 3 days for environmental acclimation.

[0078] With respect to the environment, animal holding rooms’ environmental controls were set to maintain temperatures between 20 - 26QC (68 - 799F) with a relative humidity range of 30% - 70%. A 12-hour light / 12-hour dark cycle was maintained, except when interrupted to accommodate study procedures. Ten or greater air changes per hour were also maintained in the animal holding and procedure rooms.

[0079] With respect to diet, the test animals were provided Teklad Global 18% Protein Rodent Diet 2918 ad libitum daily. The test animals were also provided filtered tap water during the acclimation period and Innovive bottled water throughout the study period, ad libitum.Attorney Docket No. 68622WO01

[0080] Light anesthesia inhalant isoflurane on O2 (1 - 4%, to effect) was used to facilitate wound creation, and for treatments, wound assessments, wound measurements, photography, and reapplication of dressings. Isoflurane overdose or carbon dioxide asphyxiation was used, to effect, for terminal tissue collections.

[0081] All animals had a wound created on Day 0 while anesthetized, and after received analgesics. With respect to pre-wound care, all test animals had their dorsum shaved prior to study start. All animals received a subcutaneous injection of Buprenorphine-SR analgesia (1.0 mg / kg) on Day 0, ~ 1 hour prior to wounding, and an additional dose 48-72 hours later, if needed.

[0082] With respect to wound creation, sterile techniques were used. The test animals were lightly anesthetized using isoflurane on O2 (1 - 4%, to effect) and placed on their abdomen. Using sterile, disposable skin biopsy punches a 10 mm disc skin biopsy was removed from the dorsal right side of each test animal. A 3D Printed biofilm (S. aureus) was placed in the wounds of animals in groups 2A and 3A only. The wound was covered with Tegademn™ and the biofilm was allowed to incorporate into the wound until treatment initiation (Day 3). From Day 0 until study conclusion all wounds were covered with a Tegaderm™ dressing as outlined above. Tegaderm™ was left on for the duration of the study.

[0083] With respect to the dose regimen, topical administration of SEP-RM rhamnolipid treatment was performed twice per week, on Study Days 0 (non-infected groups only), 3, 7, 10, 14, 17, and 21 while animals were under light anesthesia inhalant isoflurane on O2 (1-4%, to effect). Treatment administration for the infected groups began on Study Day 3 and continued twice per week outlined above. The test animals were observed for anyAttorney Docket No. 68622WO01 adverse reactions to each dose, at each session, prior to being placed back into their respective home cages.

[0084] With respect to wound measurements and assessments, on Days 0 (pretreatment, post-wound creation), 3, 7, 10, 14, 17, 21 , 24, and on each animals scheduled termination day, all animals, while lightly anesthetized via isoflurane on O2 (1 - 4%, to effect), had wounds measured by digital caliper by measuring width x length. Measurements occurred after careful removal of the Tegaderm™ dressing. These measurements were reconfirmed by Imaged analysis of the digital photographs.

[0085] At the time of wound measurements and assessments, on Days 0, 3, 7, 10, 14, 17, 21 , and on each test animal’s scheduled termination day, all test animals, while lightly anesthetized via isoflurane on O2 (1 - 4%, to effect), will have a photographic image of the wound. Images will be taken after careful removal of the Tegaderm™ dressing. Wound closure (contraction & re-epithelialization) will be determined from digital photographs.

[0086] Figure 1 shows a representative example of wound healing / closure for the positive control (collagen hydrogel without rhamnolipid) versus 1 % SEP-RM rhamnolipid in collagen hydrogel, with images taken at days 0, 3, 17, and 24.

[0087] Wound closure, such as that shown in Figure 1 , was quantified by measuring the open area of the wound through the duration of healing. Percent wound closure was analyzed using NIH Imaged (Bethesda, MD) by standardizing each image using the millimeter ruler captured in each photo. The open wound margin was traced with a freehand tool, generating an area in mm2. The wound bed for each day was normalized to time 0 using the following equation:Attorney Docket No. 68622WO01where DO = Initial wound area measurement and Dn = Day of wound area measurement.

[0088] The following table set forth a representative number of examples performed in accordance with the study design and methods set forth above:Table 2A: Representative examples performed in accordance with the study design

[0089] Figure 2 shows a graph of the average wound size (normalized to day 0) for the positive control group (collagen hydrogel), performed in accordance with the study design set forth above.Attorney Docket No. 68622WO01

[0090] Figure 3 shows a graph of the average wound for the 1 % SEP-RM RL mixed with collagen hydrogel group, performed in accordance with the study design set forth above.

[0091] Table 2B shows the change in daily wound size (i.e. , a measurement of how the wound healing is progressing overall). As the wound continues to heal, the impact of epigenetics has an influence over how the rate of wound healing changes overall as week as on a daily basis. This can also be measured from day to day or as a cumulative total result. The third and eighth column of Table 2B shows the daily wound size change as compared to the previous day. The fourth and ninth column of Table 2B shows the % average daily wound healing rate taken as compared to the previous day. The fifth and tenth column shows the % total cumulative wound healing compared to the initial wound size. As set forth in Table 2B: the % Daily Closure rate = [(previous day wound size - current day wound size) / initial wound size] x 100; and the % Total Closure = [(initial wound size - current day wound size) I initial wound size] x 100.Table 2B - Wound Healing RateAttorney Docket No. 68622WO01Attorney Docket No. 68622WO01

[0092] The present disclosure provides for accelerated wound healing and / or wound closure rates with the application of rhamnolipid composition to the wound, in accordance with the disclosure.

[0093] In some aspects, the application of rhamnolipids provides a daily closure rate, by day 3 after administration of the rhamnolipids, of from 3% to 5%, from 2% to 6%, from 2% to 7%, from 2% to 8%, from 2% to 10%, 3% to 15%, or 2% to 10% (vs control at 0% for both measures).

[0094] In another aspect, the application of rhamnolipids provides a % total closure on day 3 (after administration of the rhamnolipid) of from 3% to 5%, from 2% to 6%, from 2% to 7%, from 2% to 8%, from 2% to 10%, 3% to 15%, or 2% to 10% (vs control at 0% for both measures).

[0095] In yet another aspect, the application of rhamnolipids provides a daily closure rate, by day 7 after administration of the rhamnolipids, of from 15-20%, from 15-25%, from 15-30%, from 10-15%, from 10-20%, from 10-25%, from 10-30%, or from 5-40% (vs control at closure rate of 14%).

[0096] In another aspect, the application of rhamnolipids provides a % total closure on day 7 (after administration of the rhamnolipid) of from 40-50%, from 35-55%%, or from 30-60% (vs control at total closure of 20%).

[0097] In a further aspect, the application of rhamnolipids provides a % total closure on day 7 (after administration of the rhamnolipid) of from 85-95%, from 80-100%, or from 65-100% (vs control at 74%).Attorney Docket No. 68622WO01Example 2

[0098] This study determined how certain rhamnolipid compositions impacted gene expression during wound healing.

[0099] As outlined above, the objective of this study was to evaluate two concentrations, 1 % and 3%, of a SEP-RM Rhamnolipid gel in an advanced wound care prototype. The animal model used was the male BKS.Cg-Dock7m + / + Lepr db / J (Jax Stock # 000642; Genotype: Homozygous genotype, Wildtype for Dock7<m>, Homozygous for Lepr<db>) diabetic mouse delayed wound healing model. The study design, using SEP-RM Rhamnolipid, is summarized above for Example 1 at Table 1 .

[0100] As indicated in Table 1 , wound creation, on Day 0, included a skin biopsy (10 mm disc) removed from the dorsal right side of the mouse model. The study included 6 treatment groups, including: 2 x control groups (without rhamnolipid); 2 x 1 % SEP-RM rhamnolipid (dose level = 0.1 mg; dose concentration = 10 mg / mL, 1 %; and dose volume = 100 uL); and 2 x 3% SEP-RM rhamnolipid (dose level = 0.2 mg; dose concentration = 20 mg / mL, 20%; and dose volume = 100 uL). The douse route was topical (to the area of the skin biopsy), at days 0, 3, 7, 10, 14, 17, and 21 . The study end was set at 24 days.

[0101] The Control group included Control Gel Collagen, with ambient storage. Rhamnolipid treatment group 1 comprised 1 % SEP-RM rhamnolipid in a collagen hydrogel, with ambient storage. Rhamnolipid treatment group 2 comprised 3% SEP-RM rhamnolipid in a collagen hydrogel, with ambient storage. The wound dressing comprised 3M Tegaderm™ dressing, with ambient storage.Attorney Docket No. 68622WO01

[0102] As indicated above, the test system included species Mus muscuus (strain = BKS.Cg-Dock7m + / + Leprdb / J). The source was Jackson Laboratories (Stock# 000642; Genotype: Homozygous genotype, Wildtype for Dock7<m>, Homozygous for Lepr<db>). The age of the animals upon arrival was 8 weeks, and before use the animals were held for a minimum of 3 days for environmental acclimation.

[0103] With respect to the environment, animal holding rooms’ environmental controls were set to maintain temperatures between 20 - 26QC (68 - 799F) with a relative humidity range of 30% - 70%. A 12-hour light / 12-hour dark cycle was maintained, except when interrupted to accommodate study procedures. Ten or greater air changes per hour were also maintained in the animal holding and procedure rooms.

[0104] With respect to diet, the test animals were provided Teklad Global 18% Protein Rodent Diet 2918 ad libitum daily. The test animals were also provided filtered tap water during the acclimation period and Innovive bottled water throughout the study period, ad libitum.

[0105] Light anesthesia inhalant isoflurane on O2 (1 - 4%, to effect) was used to facilitate wound creation, and for treatments, wound assessments, wound measurements, photography, and reapplication of dressings. Isoflurane overdose or carbon dioxide asphyxiation was used, to effect, for terminal tissue collections.

[0106] All animals had a wound created on Day 0 while anesthetized, and after received analgesics. With respect to pre-wound care, all test animals had their dorsum shaved prior to study start. All animals recieved a subcutaneous injection of Buprenorphine-SR analgesia (1.0 mg / kg) on Day 0, ~ 1 hour prior to wounding, and an additional dose 48-72 hours later, if needed.Attorney Docket No. 68622WO01

[0107] With respect to wound creation, sterile techniques were used. The test animals were lightly anesthetized using isoflurane on O2 (1 - 4%, to effect) and placed on their abdomen. Using sterile, disposable skin biopsy punches a 10 mm disc skin biopsy was removed from the dorsal right side of each test animal. A 3D Printed biofilm (S. aureus) was placed in the wounds of animals in groups 2A and 3A only. The wound was covered with Tegaderm™ and the biofilm was allowed to incorporate into the wound until treatment initiation (Day 3). From Day 0 until study conclusion all wounds were covered with a Tegaderm™ dressing as outlined above. Tegaderm™ was left on for the duration of the study.

[0108] With respect to the dose regimen, topical administration of SEP-RM rhamnolipid treatment was performed twice per week, on Study Days 0 (non-infected groups only), 3, 7, 10, 14, 17, and 21 while animals were under light anesthesia inhalant isoflurane on O2 (1-4%, to effect). Treatment administration for the infected groups began on Study Day 3 and continued twice per week outlined above. The test animals were observed for any adverse reactions to each dose, at each session, prior to being placed back into their respective home cages.

[0109] The wound tissue and the skin region surrounding the wound, as appropriate, will be harvested and processed for NanoString quantitative gene expression analysis.

[0110] With respect to the NanoString gene expression analysis, on study Day 3-5, during the inflammatory phase, and between Study Days 11 -18, during the proliferation phase, three (3) animals from groups 1 , 1A, 2, and 3 (12 total) were designated to have wound tissue analyzed by NanoString. An 8 - 12mm punch biopsy was used to harvest wound / tissue samples, which were then placed in sample vials containing ~1 -3mL ofAttorney Docket No. 68622WO01RNAprotect Tissue Stabilization Reagent, and stored at -80°C for NanoString Analysis utilizing a custom wound healing panel (282 genes). RNA was purified and quantified using standard techniques in the art.[01 1 1] For the rhamnolipid treatment group 1 (having 1 % SEP-RM rhamnolipid in a collagen hydrogel), the following genes were activated (i.e. upregulated) at day 3 (inflammatory phase), versus control (collagen hydrogel alone): TIE1 ; LARS1 ; PANX1 ; PLXDC2; SFRP1 ; CTSK; MMP14; ITGB5; DLG4; and PDGFA. On Study Day 3, the 1% RL Gel group had 10 wound healing genes upregulated. In particular, there were two proteases (MMP14 + CTSK), six genes involved in keratinocyte and fibroblast migration, proliferation, and re-epithelialization (LARS1 , PLXDC2, TIE1 , ITGB5, PANX1 and DLG4)

[0112] The data is presented below in Table 3, which shows the genes upregulated at day 3 (inflammatory phase) for 1 % SEP-RM RL in Collagen Hydrogel group versus Collagen Hydrogel control group, in the diabetic mouse wound model study described herein (Example 2).Table 3: genes upregulated at day 3 (inflammatory phase) for 1 % SEP-RM RL in CollagenHydrogel group versus Collagen Hydrogel control group:Attorney Docket No. 68622WO01

[0113] For the rhamnolipid treatment group 1 (having 1 % SEP-RM rhamnolipid in collagen hydrogel), the following genes were activated (i.e. upregulated) at day 17 (proliferation), versus control (collagen hydrogel alone): CSF2RB; IL1 B; PF4; CSF3R; NR4A1 ; PDGFRA; PDGFRB; PILRB1 ; and TNFRSF1 B. The data is presented below in Table 4, which shows the genes upregulated and down regulated (shaded rows) at day 17 (proliferative phase) for 1 % SEP-RM RL in Collagen Hydrogel group versus Collagen Hydrogel control group, in the diabetic mouse wound model study described herein.Table 4: genes upregulated and down regulated (e.g., shaded rows) at day 17 (proliferative phase) for 1 % SEP-RM RL in Collagen Hydrogel group versus Collagen Hydrogel control groupAttorney Docket No. 68622WO01Attorney Docket No. 68622WO01

[0114] For the rhamnolipid treatment group 1 (having 1 % SEP-RM rhamnolipid in collagen hydrogel), the following genes were suppressed (i.e. downregulated) at day 17 (proliferative phase), versus control (collagen hydrogel alone): CNR1 ; TIMP3; ATG14;JUN; and XPNPEP1.

[0115] The present technology is now described in such full, clear and concise terms as to enable a person skilled in the art to which it pertains, to practice the same. It is to be understood that the foregoing describes preferred embodiments of the present technology and that modifications may be made therein without departing from the spirit or scope of the present technology as set forth in the appended claims. Further, the examples are provided to not be exhaustive but illustrative of several embodiments that fall within the scope of the claims.

Claims

Attorney Docket No. 68622WO01Claims1. A composition for use in a method of accelerating wound closure, the composition comprising:(a) a mixture of rhamnolipids, wherein the mixture of rhamnolipids comprises: mono-rhamnolipids and di-rhamnolipids in a weight ratio of about 40:60 to about 60:40 mono-rhamnolipids:di-rhamnolipids; an amount of C10-C10 mono-rhamnolipid of about 29% to about 40% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C10-C10 di-rhamnolipid of about 35% to about 50% by weight, based on the total weight of rhamnolipids present in the composition; an amount of 08-010 mono-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition; an amount of 08-010 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C10-C12 mono-rhamnolipid of about 2% to about 6% by weight, based on the total weight of rhamnolipids present in the composition; and an amount of 010-012 di-rhamnolipid of about 8% to about 14% by weight, based on the total weight of rhamnolipids present in the composition;Attorney Docket No. 68622WO01(b) at least one acceptable carrier, and optionally one or more additives, in an amount to total 100% by weight of the composition.

2. The composition of claim 1 , wherein the mixture of rhamnolipids further comprises an amount of C10-C12:1 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition.

3. The composition of claim 1 , wherein the mixture of rhamnolipids further comprises an amount of C12-C12 di-rhamnolipid of about 0.2% to about 0.4% by weight, based on the total weight of rhamnolipids present in the composition.

4. The composition of claim 1 , wherein the mixture of rhamnolipids comprises an amount of total mono-rhamnolipid of about 40% to about 50% by weight, based on the total weight of rhamnolipids present in the composition.

5. The composition of claim 1 , wherein the mixture of rhamnolipids comprises an amount of total di-rhamnolipid of about 50% to about 60% by weight, based on the total weight of rhamnolipids present in the composition.

6. The composition of any one of claims 1 to 5, wherein the mixture of rhamnolipids is in an amount of about 0.1 % to about 10%, based on the total weight of the composition.

7. The composition of any one of claims 1 to 6, wherein the wound being treated is an incision, laceration, abrasion, or burn.

8. The composition of any one of claims 1 to 6, wherein the chronic wound being treated is diabetic foot ulcer (DFU), pressure ulcer (PU), or venous leg ulcer (VLU).Attorney Docket No. 68622WO019. The composition of any one of claims 1 to 8, wherein the at least one acceptable carrier is water alone or in combination with an alcohol or glycol.

10. The composition of claim 9, wherein the alcohol is ethanol, isopropanol, or benzyl alcohol.1 1. The composition of claim 9, wherein the glycol is propylene glycol or polyethylene glycol.

12. The composition of any one of claims 1 to 1 1 , wherein the method comprises topical administration of the composition.

13. A wound care composition for use in accelerating wound closure, comprising:(a) a mixture of rhamnolipids, wherein the mixture of rhamnolipids comprises: mono-rhamnolipids and di-rhamnolipids in a weight ratio of about 40:60 to about 50:50 mono-rhamnolipids:di-rhamnolipids; an amount of C10-C10 mono-rhamnolipid of about 29% to about 37% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C10-C10 di-rhamnolipid of about 35% to about 50% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C8-C10 mono-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition;Attorney Docket No. 68622WO01 an amount of 08-010 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition; an amount of 010-012 mono-rhamnolipid of about 2% to about 6% by weight, based on the total weight of rhamnolipids present in the composition; and an amount of 010-012 di-rhamnolipid of about 8% to about 14% by weight, based on the total weight of rhamnolipids present in the composition;(b) at least one acceptable carrier, and optionally one or more additives, in an amount to total 100% by weight of the composition.

14. The wound care composition of claim 13, wherein the mixture of rhamnolipids further comprises an amount of C10-C12:1 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition.

15. The wound care composition of claim 13 or 14, wherein the mixture of rhamnolipids further comprises an amount of C12-C12 di-rhamnolipid of about 0.2% to about 0.4% by weight, based on the total weight of rhamnolipids present in the composition.

16. The wound care composition of any one of claims 13-15, wherein the mixture of rhamnolipids comprises an amount of total mono-rhamnolipid of about 40% to about 48% by weight, based on the total weight of rhamnolipids present in the composition.

17. The wound care composition of any one of claims 13-16, wherein the mixture of rhamnolipids comprises an amount of total di-rhamnolipid of about 52% to about 60% by weight, based on the total weight of rhamnolipids present in the composition.Attorney Docket No. 68622WO0118. The wound care composition of any one of claims 13-17, wherein the mixture of rhamnolipids is in an amount of about 0.1 % to about 10%, based on the total weight of the composition.

19. The wound care composition of any one of claims 13-18, wherein the at least one acceptable carrier is water alone or in combination with an alcohol or glycol.

20. The wound care composition of claim 19, wherein the alcohol is ethanol, isopropanol, or benzyl alcohol.

21. The wound care composition of claim 19, wherein the glycol is propylene glycol or polyethylene glycol.

22. A method for accelerating closure of a wound comprising: applying to the wound a composition comprising:(a) a mixture of rhamnolipids, wherein the mixture of rhamnolipids comprises: mono-rhamnolipids and di-rhamnolipids in a weight ratio of about 40:60 to about 50:50 mono-rhamnolipids:di-rhamnolipids; an amount of C10-C10 mono-rhamnolipid of about 29% to about 37% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C10-C10 di-rhamnolipid of about 35% to about 50% by weight, based on the total weight of rhamnolipids present in the composition;Attorney Docket No. 68622WO01 an amount of C8-C10 mono-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C8-C10 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C10-C12 mono-rhamnolipid of about 2% to about 6% by weight, based on the total weight of rhamnolipids present in the composition; and an amount of C10-C12 di-rhamnolipid of about 8% to about 14% by weight, based on the total weight of rhamnolipids present in the composition;(b) at least one acceptable carrier, and optionally one or more additives, in an amount to total 100% by weight of the composition.

23. The method of claim 22, wherein the mixture of rhamnolipids further comprises an amount of 010-012:1 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition.

24. The method of claim 22 or 23, wherein the mixture of rhamnolipids further comprises an amount of 012-012 di-rhamnolipid of about 0.2% to about 0.4% by weight, based on the total weight of rhamnolipids present in the composition.

25. The method of any one of claims 22-24, wherein the mixture of rhamnolipids comprises an amount of total mono-rhamnolipid of about 40% to about 48% by weight, based on the total weight of rhamnolipids present in the composition.Attorney Docket No. 68622WO0126. The method of any one of claims 22-25, wherein the mixture of rhamnolipids comprises an amount of total di-rhamnolipid of about 52% to about 60% by weight, based on the total weight of rhamnolipids present in the composition.

27. The method of any one of claims 22-26, wherein the mixture of rhamnolipids is in an amount of about 0.1 % to about 10%, based on the total weight of the composition.

28. The method of any one of claims 22-27, wherein the at least one acceptable carrier is water alone or in combination with an alcohol or glycol.

29. The method of claim 28, wherein the alcohol is ethanol, isopropanol, or benzyl alcohol.

30. The method of claim 28, wherein the glycol is propylene glycol, hexylene glycol, 1 ,2- propanediol, 1 ,3-propanediol, or polyethylene glycol.

31. A composition for use in a method of modifying gene expression in a target tissue during treatment of a wound, the composition comprising:(a) a mixture of rhamnolipids, wherein the mixture of rhamnolipids comprises: mono-rhamnolipids and di-rhamnolipids in a weight ratio of about 40:60 to about 60:40 mono-rhamnolipids:di-rhamnolipids; an amount of C10-C10 mono-rhamnolipid of about 29% to about 40% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C10-C10 di-rhamnolipid of about 35% to about 50% by weight, based on the total weight of rhamnolipids present in the composition;Attorney Docket No. 68622WO01 an amount of C8-C10 mono-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C8-C10 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C10-C12 mono-rhamnolipid of about 2% to about 6% by weight, based on the total weight of rhamnolipids present in the composition; and an amount of C10-C12 di-rhamnolipid of about 8% to about 14% by weight, based on the total weight of rhamnolipids present in the composition;(b) at least one acceptable carrier, and optionally one or more additives, in an amount to total 100% by weight of the composition.

32. The composition of claim 31 , wherein the mixture of rhamnolipids further comprises an amount of C10-C12:1 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition.

33. The composition of claim 31 , wherein the mixture of rhamnolipids further comprises an amount of C12-C12 di-rhamnolipid of about 0.2% to about 0.4% by weight, based on the total weight of rhamnolipids present in the composition.

34. The composition of claim 31 , wherein the mixture of rhamnolipids comprises an amount of total mono-rhamnolipid of about 40% to about 50% by weight, based on the total weight of rhamnolipids present in the composition.Attorney Docket No. 68622WO0135. The composition of claim 31 , wherein the mixture of rhamnolipids comprises an amount of total di-rhamnolipid of about 50% to about 60% by weight, based on the total weight of rhamnolipids present in the composition.

36. The composition of any one of claims 31 to 35, wherein the mixture of rhamnolipids is in an amount of about 0.1 % to about 10%, based on the total weight of the composition.

37. The composition of any one of claims 31 to 36, wherein the wound being treated is an incision, laceration, abrasion, or burn.

38. The composition of any one of claims 31 to 36, wherein the wound being treated is diabetic foot ulcer (DFU), pressure ulcer (PU), or venous leg ulcer (VLU).

39. The composition of any one of claims 31 to 38, wherein the method comprises topical administration of the composition.

40. The composition of any one of claims 31 to 39, wherein the method comprises the upregulation of at least one of the following genes during the inflammatory phase of the wound healing process: TIE1 ; LARS1 ; PANX1 ; PLXDC2; SFRP1 ; CTSK; MMP14; ITGB5; DLG4; and PDGFA.

41. The composition of any one of claims 31 to 39, wherein the method comprises the upregulation of at least one of the following genes during the proliferative phase of the wound healing process: CSF2RB; IL1 B; PF4; CSF3R; NR4A1 ; PDGFRA; PDGFRB; PILRB; and TNFRSF1 B.Attorney Docket No. 68622WO0142. The composition of any one of claims 31 to 39, wherein the method comprises the downregulation of at least one of the following genes during the proliferative phase of the wound healing process: CNR1 ; TIMP3; ATG14; JUN; and XPNPEP1 .

43. A wound care composition, comprising:(a) a mixture of rhamnolipids, wherein the mixture of rhamnolipids comprises: mono-rhamnolipids and di-rhamnolipids in a weight ratio of about 40:60 to about 50:50 mono-rhamnolipids:di-rhamnolipids; an amount of C10-C10 mono-rhamnolipid of about 29% to about 37% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C10-C10 di-rhamnolipid of about 35% to about 50% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C8-C10 mono-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C8-C10 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C10-C12 mono-rhamnolipid of about 2% to about 6% by weight, based on the total weight of rhamnolipids present in the composition; and an amount of C10-C12 di-rhamnolipid of about 8% to about 14% by weight, based on the total weight of rhamnolipids present in the composition;Attorney Docket No. 68622WO01(b) at least one acceptable carrier, and optionally one or more additives, in an amount to total 100% by weight of the composition.

44. The wound care composition of claim 43, wherein the mixture of rhamnolipids further comprises an amount of C10-C12:1 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition.

45. The wound care composition of claim 43 or 44, wherein the mixture of rhamnolipids further comprises an amount of C12-C12 di-rhamnolipid of about 0.2% to about 0.4% by weight, based on the total weight of rhamnolipids present in the composition.

46. The wound care composition of any one of claims 43-45, wherein the mixture of rhamnolipids comprises an amount of total mono-rhamnolipid of about 40% to about 48% by weight, based on the total weight of rhamnolipids present in the composition.

47. The wound care composition of any one of claims 43-46, wherein the mixture of rhamnolipids comprises an amount of total di-rhamnolipid of about 52% to about 60% by weight, based on the total weight of rhamnolipids present in the composition.

48. The wound care composition of any one of claims 43-47, wherein the mixture of rhamnolipids is in an amount of about 0.1 % to about 10%, based on the total weight of the composition.

49. The wound care composition of any one of claims 43 to 48, wherein the method comprises the upregulation of at least one of the following genes during the inflammatory phase of the wound healing process: TIE1 ; LARS1 ; PANX1 ; PLXDC2; SFRP1 ; CTSK; MMP14; ITGB5; DLG4; and PDGFA.Attorney Docket No. 68622WO0150. The wound care composition of any one of claims 43 to 48, wherein the method comprises the upregulation of at least one of the following genes during the proliferative phase of the wound healing process: CSF2RB; IL1 B; PF4; CSF3R; NR4A1 ; PDGFRA; PDGFRB; PILRB; and TNFRSF1 B.

51. The wound care composition of any one of claims 43 to 48, wherein the method comprises the downregulation of at least one of the following genes during the proliferative phase of the wound healing process: CNR1 ; TIMP3; ATG14; JUN; and XPNPEP1.

52. A method of modifying gene expression in a target tissue during treatment of a wound, the method comprising applying to the wound a composition, comprising:(a) a mixture of rhamnolipids, wherein the mixture of rhamnolipids comprises: mono-rhamnolipids and di-rhamnolipids in a weight ratio of about 40:60 to about 50:50 mono-rhamnolipids:di-rhamnolipids; an amount of C10-C10 mono-rhamnolipid of about 29% to about 37% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C10-C10 di-rhamnolipid of about 35% to about 50% by weight, based on the total weight of rhamnolipids present in the composition; an amount of C8-C10 mono-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition;Attorney Docket No. 68622WO01 an amount of 08-010 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition; an amount of 010-012 mono-rhamnolipid of about 2% to about 6% by weight, based on the total weight of rhamnolipids present in the composition; and an amount of 010-012 di-rhamnolipid of about 8% to about 14% by weight, based on the total weight of rhamnolipids present in the composition;(b) at least one acceptable carrier, and optionally one or more additives, in an amount to total 100% by weight of the composition.

53. The method of claim 52, wherein the mixture of rhamnolipids further comprises an amount of C10-C12:1 di-rhamnolipid of about 2% to about 5% by weight, based on the total weight of rhamnolipids present in the composition.

54. The method of claim 52 or 53, wherein the mixture of rhamnolipids further comprises an amount of C12-C12 di-rhamnolipid of about 0.2% to about 0.4% by weight, based on the total weight of rhamnolipids present in the composition.

55. The method of any one of claims 52-54, wherein the mixture of rhamnolipids comprises an amount of total mono-rhamnolipid of about 40% to about 48% by weight, based on the total weight of rhamnolipids present in the composition.

56. The method of any one of claims 52-55, wherein the mixture of rhamnolipids comprises an amount of total di-rhamnolipid of about 52% to about 60% by weight, based on the total weight of rhamnolipids present in the composition.Attorney Docket No. 68622WO0157. The method of any one of claims 52-56, wherein the mixture of rhamnolipids is in an amount of about 0.1 % to about 10%, based on the total weight of the composition.

58. The method of any one of claims 52 to 57, wherein the method upregulates at least one of the following genes during the inflammatory phase of the wound healing process: TIE1 ; LARS1 ; PANX1 ; PLXDC2; SFRP1 ; CTSK; MMP14; ITGB5; DLG4; and PDGFA.

59. The method of any one of claims 52 to 57, wherein the method upregulates at least one of the following genes during the proliferative phase of the wound healing process: CSF2RB; IL1 B; PF4; CSF3R; NR4A1 ; PDGFRA; PDGFRB; PILRB; and TNFRSF1 B.

60. The method of any one of claims 52 to 57, wherein the method downregulates at least one of the following genes during the proliferative phase of the wound healing process: CNR1 ; TIMP3; ATG14; JUN; and XPNPEP1 .

Images