Patent Application
20240299473
The skin is the largest organ in the human body and functions as the first line of defense by providing a protective barrier between the environment and inner body. The skin harbors several hundreds of resident microorganisms, which function in communities and protect the body from invasion of pathogens. Several studies have shown that shifts in the skin microbiota are associated with various skin diseases.
Acne vulgaris (commonly called acne) is the most common skin disease, affecting 80-85% of the population. It is most prevalent in adolescents and rarely occurs in people over the age of 50. Although acne is not life threatening, it can lead to severe pain and scarring on the skin, and has profoundly negative psychosocial effects. Acne is a disease of the pilosebaceous unit (commonly known as the hair follicle). While its etiology is still unclear with multiple factors involved, the Gram-positive lipophilic anaerobe Propionibacterium acnes has been thought to play a role in acne pathogenesis.
In some aspects, provided herein are methods and compositions related to treating or preventing a skin disease (e.g., inflammatory skin disease, such as acne, rosacea, or Porphyria Cutanea Tarda (PCT)), preventing and/or slowing skin aging (e.g., preventing the formation of wrinkles), and maintaining healthy skin by administering to the subject a composition disclosed herein. In some aspects, provided herein are methods of treating or preventing a skin condition in a subject and/or reducing the amount of porphyrins on the skin of a subject by administering (e.g., a subject with symptoms of skin aging, or a subject with a skin condition, such as acne, rosacea, or PCT) a composition comprising at least one strain of P. granulosum, at least one strain of P. avidum, and/or at least one strain of P. humerusii to the subject. The strain of P. granulosum may be HL078PG1 and/or HL082PG1. The strain of P. avidum may be HL063PV1, HL083PV1, and HL307PV1. The strain of P. humerusii may be HL044PA1.
In some aspects, the invention relates, at least in part, to determining whether a subject is at risk for a skin disease (e.g., inflammatory skin disease, such as acne, rosacea, or Porphyria Cutanea Tarda (PCT)), comprising obtaining a skin sample from the subject, optionally isolating microbial DNA from the skin sample and identifying the amount of P. granulosum in the microbiome of the skin sample. In some embodiments, if the microbiome is less than 5%, (e.g., less than 4%, less than 3%, less than 2%, less than 1%, or less than 0.5%) P. granulosum, the subject is considered at risk for a skin disease. In some embodiments, the methods further comprise administering to the subject a composition comprising P. granulosum if P. granulosum is less than 5% (e.g., less than 4%, less than 3%, less than 2%, less than 1%, or less than 0.5%) of the microbiome of the skin sample.
In some embodiments, provided herein are methods determining whether a subject has acne by obtaining a skin sample from the subject, sequencing DNA from the skin sample, and analyzing the DNA for the enrichment of one or more metagenomic elements in Table 1 associated with acne or the depletion of one or more metagenomic elements in Table 1 associated with health, wherein the subject is determined as having acne if one or more of the metagenomic elements in Table 1 associated with acne is enriched in the skin sample or one or more of the metagenomic elements in Table 1 associated with health is depleted in the skin sample. In some embodiments, provided herein are methods for determining whether a subject has acne by obtaining a skin sample from the subject, optionally isolating bacterial DNA from the skin sample, using one or more primer sets to amplify the DNA, using one or more probes to detect the amplified DNA; and analyzing the probe signals for the enrichment of one or more metagenomic elements in Table 1 associated with acne or the depletion of one or more metagenomic elements in Table 1 associated with health, wherein the subject is determined as having acne if one or more of the metagenomic elements in Table 1 associated with acne is enriched in the skin sample or one or more of the metagenomic elements in Table 1 associated with health is depleted in the skin sample.
In some embodiments, the microbial DNA is isolated from a skin follicle. Also provided herein are methods of determining whether a subject has acne, comprising obtaining a skin sample from a subject and analyzing (e.g., sequencing) the skin sample for enrichment or depletion of a metagenomic element disclosed herein. In some aspects, provided herein are methods of treating acne by administering a composition comprising a metabolite produced by a strain of P. granulosum.
Also provided here are methods for treating acne in a subject, comprising administering a composition comprising a metabolite produced by a strain of P. granulosum. P. avidum, and/or P. humerusii, wherein the metabolite is selected from bacterial culture supernatant, cell lysate, proteins, nucleic acids, lipids, and other bacterial molecules.
In some embodiments, the subject has a skin disease (e.g., inflammatory skin disease, such as acne, rosacea, or Porphyria Cutanea Tarda (PCT)).
Provided herein are compositions (e.g., cosmetic or pharmaceutical compositions) comprising at least one strain of P. granulosum, at least one strain of P. avidum, and/or at least one strain of P. humerusii to the subject. The strain of P. granulosum may be HL078PG1 and/or HL082PG1. The strain of P. avidum may be HL063PV1, HL083PV1, and HL307PV1. The strain of P. humerusii may be HL044PA1. The composition may further comprises one or more strains of P. acnes (e.g., an RT1, RT2, RT3, or RT6 strain of P. acnes). The composition may be enriched for any one of the genes associated with healthy skin in Table 1. In some embodiments, the composition comprises a phage against a strain of P. acnes (e.g., a strain of P. acnes selected from RT4, RT5, RT7, RT8, RT9 or RT10). In some embodiments, the composition comprises two or more phages against a strain of P. acnes. The composition may further comprise an antibiotic (preferably an antibiotic that does not target and/or kill P. granulosum. P. avidum, and/or P. humerusii). In certain embodiments, the compositions disclosed herein may be formulated for oral or topical delivery.
Also provided herein are methods of improving skin health or slowing or preventing skin aging in a subject, comprising administering a composition disclosed herein (e.g., a composition comprising P. granulosum, P. humerusii and/or P. avidum) to the subject.
In some aspects, provided herein are methods and compositions related to treating or preventing a skin disease (e.g., acne, rosacea, or PCT), preventing and/or slowing skin aging (e.g., preventing or inhibiting the formation of wrinkles), and maintaining healthy skin by administering to the subject a composition disclosed herein (e.g., a composition comprising at least one strain of P. granulosum, at least one strain of P. avidum, and/or at least one strain of P. humerusii). In some aspects, provided herein are methods of treating or preventing a skin condition in a subject and/or reducing the amount of porphyrins on the skin of a subject (e.g., a subject with symptoms of skin aging, or a subject with a skin condition, such as acne, rosacea, or PCT) by administering to the subject a composition comprising at least one strain of P. granulosum, at least one strain of P. avidum, and/or at least one strain of P. humerusii. In some aspects, the invention relates to determining whether a subject is at risk for a skin disease, comprising obtaining a skin sample from the subject, optionally isolating microbial DNA from the skin sample, and identifying the amount of P. granulosum. P. avidum, and/or P. humerusii in the microbiome of the skin sample. In some embodiments, the microbial DNA is isolated from a skin follicle. Also provided herein are methods of determining whether a subject has acne, comprising obtaining a skin sample from a subject and analyzing (e.g., sequencing) the skin sample for the enrichment or depletion of one or more metagenomic elements disclosed herein in the skin sample. In some aspects, provided herein are methods of treating acne comprising administering a composition comprising a metabolite produced by a strain of P. granulosum. P. avidum, and/or P. humerusii.
As used herein the specification, “a” or “an” may mean one or more. As used herein in the claim(s), when used in conjunction with the word “comprising”, the words “a” or “an” may mean one or more than one. As used herein “another” may mean at least a second or more.
The term “preventing” is art-recognized, and when used in relation to a condition, such as a local recurrence, is well understood in the art, and includes administration of a composition which reduces the frequency of, or delays the onset of, symptoms of a medical condition in a subject relative to a subject which does not receive the composition. Thus, prevention of acne includes, for example, reducing the number of detectable acne lesions in a population of patients receiving a prophylactic treatment relative to an untreated control population, and/or delaying the appearance of detectable lesions in a treated population versus an untreated control population, e.g., by a statistically and/or clinically significant amount.
The term “prophylactic” or “therapeutic” treatment is art-recognized and includes administration to the host of one or more of the subject compositions. If it is administered prior to clinical manifestation of the unwanted condition (e.g., disease or other unwanted state of the host animal) then the treatment is prophylactic (i.e., it protects the host against developing the unwanted condition), whereas if it is administered after manifestation of the unwanted condition, the treatment is therapeutic (i.e., it is intended to diminish, ameliorate, or stabilize the existing unwanted condition or side effects thereof).
The term “ribotype” refers to strains of P. acnes. The ribotyped strains were characterized as in Fitz-Gibbon et al. (J. Investigative Dermatology 133:2152-60 (2013)).
The term “subject” refers to a mammal, including, but not limited to, a human or non-human mammal, such as a bovine, equine, canine, ovine, or feline.
A “therapeutically effective amount” of a compound with respect to the subject method of treatment refers to an amount of the compound(s) in a preparation which, when administered as part of a desired dosage regimen (to a mammal, preferably a human) alleviates a symptom, ameliorates a condition, or slows the onset of disease conditions according to clinically acceptable standards for the disorder or condition to be treated or the cosmetic purpose, e.g., at a reasonable benefit/risk ratio applicable to any medical treatment.
As used herein, the term “treating” or “treatment” includes reversing, reducing, or arresting the symptoms, clinical signs, and underlying pathology of a condition in a manner to improve or stabilize a subject's condition.
Provided herein are methods of treating or preventing a skin condition, preventing and/or slowing skin aging, and/or maintaining healthy skin. In some aspects, the methods relate to treating or preventing a skin disease in a subject, comprising administering a composition comprising Propionibacterium granulosum (P. granulosum), P. avidum, and/or P. humerusii to the subject. Compositions described herein may have at least one strain of P. granulosum, at least one strain of P. avidum, and/or at least one strain of P. humerusii. Compositions may contain two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more strains of P. granulosum, P. avidum, and/or P. humerusii. In some aspects, provided herein are methods related to reducing the amount of porphyrins on the skin of a subject, comprising administering a composition comprising P. granulosum. P. avidum, and/or P. humerusii. In some embodiments, the subject has a skin disease or symptoms associated with skin aging (e.g., formation of wrinkles). In some embodiments, the skin disease is an inflammatory skin disease, such as acne, rosacea, or Porphyria Cutanea Tarda (PCT). The composition may further comprise an additional strain of bacteria (e.g., a strain of P. acnes, such as RT1, RT2, RT3, or RT6 strain of P. acnes). In some aspects, provided herein are methods of slowing or preventing skin aging in a subject by administering a composition comprising P. granulosum. P. avidum, and/or P. humerusii to the subject.
The strain of P. granulosum may be HL078PG1 and/or HL082PG1. The strain of P. avidum may be HL063PV1, HL083PV1, and HL307PV1. The strain of P. humerusii may be HL044PA1.
In some embodiments, the composition further comprises a phage against a strain of P. acnes (e.g., a RT4, RT5, RT7, RT8, RT9) or RT10 strain of P. acnes). In some embodiments, the composition comprises two or more (e.g., three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more) phages against a strain of P. acnes. The type of phage that may be administered in a composition disclosed herein depends on the type of acne or skin disease, the medical history of an individual, or the symptoms of a subject with a skin disease. Non-limiting examples of phages include PHL111M01. PHL082M00, PHL060L00, PHL067M10, PHL07 IN05, PHL112N00, PHL037M02, PHL085N00, PHL115M02, PHL085M01, PHL114L00, PHL010M04, PHL066M04, PHL071N05, PHL113M01, PHL112N00, and PHL037M02. Information about P. acnes phages can be found in U.S. Patent Publication US20150086581A1, hereby incorporated in its entirety. In some embodiments, the composition may comprise an antibiotic (e.g., an antibiotic that does not target P. granulosum, P. avidum, and/or P. humerusii).
The compositions disclosed herein may be administered to a subject by any means known in the art, for example, the composition may be formulated for topical delivery. The formulation may be a liquid, gel, or cream. In some embodiments, the composition is formulated for oral delivery. The composition may be in the form of a pill, tablet, or capsule. In some embodiments, the subject may be a mammal (e.g., a human). In some embodiments, the composition is self-administered.
In some aspects, provided herein are methods of determining whether a subject is at risk for a skin disease by obtaining a skin sample from the subject, isolating bacterial DNA from the skin sample, identifying the amount of P. granulosum in the microbiome of the skin sample, and if P. granulosum comprises less than 5%, less than 4.5%, less than 4%, less than 3.5%, less than 3%, less than 2.5%, less than 2%, less than 1.9%, less than 1.8%, less than 1.7%, less than 1.6%, less than 1.5%, less than 1.4%, less than 1.3%, less than 1.2%, less than 1.1%, less than 1%, less than 0.9%, less than 0.8%, less than 0.7%, less than 0.6%, less than 0.5%, less than 0.4%, less than 0.3%, less than 0.2%, less than 0.1%, less than 0.09%, less than 0.08%, less than 0.07%, less than 0.06%, less than 0.05%, less than 0.04%, less than 0.03%, less than 0.02%, less than 0.01% of the microbiome of the skin sample, the subject is considered at risk for a skin disease. In some embodiments, the composition comprising P. granulosum is administered to the subject if P. granulosum comprises less than 5%, less than 4.5%, less than 4%, less than 3.5%, less than 3%, less than 2.5%, less than 2%, less than 1.9%, less than 1.8%, less than 1.7%, less than 1.6%, less than 1.5%, less than 1.4%, less than 1.3%, less than 1.2%, less than 1.1%, less than 1%, less than 0.9%, less than 0.8%, less than 0.7%, less than 0.6%, less than 0.5%, less than 0.4%, less than 0.3%, less than 0.2%, less than 0.1%, less than 0.09%, less than 0.08%, less than 0.07%, less than 0.06%, less than 0.05%, less than 0.04%, less than 0.03%, less than 0.02%, less than 0.01% of the microbiome of the skin sample.
In some aspects, provided herein are methods of determining whether a subject is at risk for a skin disease by obtaining a skin sample from the subject, isolating bacterial DNA from the skin sample, identifying the amount of P. avidum in the microbiome of the skin sample, and if P. avidum comprises less than 5%, less than 4.5%, less than 4%, less than 3.5%, less than 3%, less than 2.5%, less than 2%, less than 1.9%, less than 1.8%, less than 1.7%, less than 1.6%, less than 1.5%, less than 1.4%, less than 1.3%, less than 1.2%, less than 1.1%, less than 1%, less than 0.9%, less than 0.8%, less than 0.7%, less than 0.6%, less than 0.5%, less than 0.4%, less than 0.3%, less than 0.2%, less than 0.1%, less than 0.09%, less than 0.08%, less than 0.07%, less than 0.06%, less than 0.05%, less than 0.04%, less than 0.03%, less than 0.02%, less than 0.01% of the microbiome of the skin sample, the subject is considered at risk for a skin disease. In some embodiments, the composition comprising P. avidum is administered to the subject if P. avidum comprises less than 5%, less than 4.5%, less than 4%, less than 3.5%, less than 3%, less than 2.5%, less than 2%, less than 1.9%, less than 1.8%, less than 1.7%, less than 1.6%, less than 1.5%, less than 1.4%, less than 1.3%, less than 1.2%, less than 1.1%, less than 1%, less than 0.9%, less than 0.8%, less than 0.7%, less than 0.6%, less than 0.5%, less than 0.4%, less than 0.3%, less than 0.2%, less than 0.1%, less than 0.09%, less than 0.08%, less than 0.07%, less than 0.06%, less than 0.05%, less than 0.04%, less than 0.03%, less than 0.02%, less than 0.01% of the microbiome of the skin sample.
In some aspects, provided herein are methods of determining whether a subject is at risk for a skin disease by obtaining a skin sample from the subject, isolating bacterial DNA from the skin sample, identifying the amount of P. humerusii in the microbiome of the skin sample, and if P. humerusii comprises less than 5%, less than 4.5%, less than 4%, less than 3.5%, less than 3%, less than 2.5%, less than 2%, less than 1.9%, less than 1.8%, less than 1.7%, less than 1.6%, less than 1.5%, less than 1.4%, less than 1.3%, less than 1.2%, less than 1.1%, less than 1%, less than 0.9%, less than 0.8%, less than 0.7%, less than 0.6%, less than 0.5%, less than 0.4%, less than 0.3%, less than 0.2%, less than 0.1%, less than 0.09%, less than 0.08%, less than 0.07%, less than 0.06%, less than 0.05%, less than 0.04%, less than 0.03%, less than 0.02%, less than 0.01% of the microbiome of the skin sample, the subject is considered at risk for a skin disease. In some embodiments, the composition comprising P. humerusii is administered to the subject if P. humerusii comprises less than 5%, less than 4.5%, less than 4%, less than 3.5%, less than 3%, less than 2.5%, less than 2%, less than 1.9%, less than 1.8%, less than 1.7%, less than 1.6%, less than 1.5%, less than 1.4%, less than 1.3%, less than 1.2%, less than 1.1%, less than 1%, less than 0.9%, less than 0.8%, less than 0.7%, less than 0.6%, less than 0.5%, less than 0.4%, less than 0.3%, less than 0.2%, less than 0.1%, less than 0.09%, less than 0.08%, less than 0.07%, less than 0.06%, less than 0.05%, less than 0.04%, less than 0.03%, less than 0.02%, less than 0.01% of the microbiome of the skin sample. In some aspects, provided herein are methods of determining whether a subject has a skin disease (e.g., acne), by obtaining a skin sample from the subject, sequencing DNA from the skin sample, analyzing the DNA for the enrichment of a metagenomic element in Table 1 associated with acne or the depletion of a metagenomic element in Table 1 associated with healthy skin, and determining the subject to have a skin disease (e.g., acne) if the metagenomic element in Table 1 associated with acne is enriched in the skin sample or if one or more of the metagenomic elements in Table 1 associated with health (e.g., healthy skin, or skin not afflicted with a skin disease, such as acne) is depleted in the skin sample. The DNA in the skin sample may be sequenced by any known technique in the art, including, but not limited to. Maxam Gilbert sequencing. Sanger sequencing, shotgun sequencing, bridge PCR, or next generation sequencing methods, such as massively parallel signature sequencing (MPSS), polony sequencing. 454 pyrosequencing. Illumina (Solexa) sequencing. SOLID sequencing. Ion torrent semiconductor sequencing. DNA nanoball sequencing, heliscope single molecule sequencing, single molecule real time (SMRT) sequencing, or nanopore DNA sequencing. In some embodiments, the skin sample is considered enriched for a metagenomic elements in Table 1 associated with acne afflicted skin if at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40)%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 95%, or at least 100% of the bacteria in the sample show enrichment of the metagenomic element in Table 1 associated with acne, or if the level of a metagenomic element in Table 1 across the sample is at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 95%, or at least 100% higher than a level associated with health. Similarly, the skin sample may be considered depleted for a metagenomic element in Table 1 associated with healthy skin if less than 100%, less than 95%, less than 90%, less than 85%, less than 80%, less than 75%, less than 70%, less than 65%, less than 60%, less than 55%, less than 50%, less than 45%, less than 40%, less than 35%, less than 30%, less than 25%, less than 20%, less than 15%, less than 10%, less than 5% of the bacteria in the sample show depletion of the metagenomic element in Table 1 associated with healthy skin, or if the level of a metagenomic element in Table 1 across the sample is at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 95% lower than a level associated with health.
In some aspects, provided herein are methods for determining whether a subject has a skin disease (e.g., acne) comprising obtaining a skin sample from the subject, isolating microbial DNA from the skin sample, using one or more primer sets to amplify the DNA, using one or more probes to detect the amplified DNA, and analyzing the probe signals for the enrichment of one or more metagenomic elements in Table 1 associated with acne or the depletion of one or more metagenomic elements in Table 1 associated with health (e.g., healthy skin, or skin not afflicted with a skin disease, such as acne), wherein the subject is determined as having acne if one or more of the metagenomic elements in Table 1 associated with acne is enriched in the skin sample or if one or more of the metagenomic elements in Table 1 associated with health is depleted in the skin sample.
In some embodiments, the skin sample is enriched if at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 95%, or at least 100% of the bacteria in the sample has the metagenomic element in Table 1 associated with acne, or if the level of a metagenomic element in Table 1 across the sample is at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 95%, or at least 100% higher than a level associated with health. In some embodiments, the skin sample is depleted of a metagenomic element if less than 100%, less than 95%, less than 90%, less than 85%, less than 80%, less than 75%, less than 70%, less than 65%, less than 60%, less than 55%, less than 50%, less than 45%, less than 40%, less than 35%, less than 30%, less than 25%, less than 20%, less than 15%, less than 10%, less than 5% of the bacteria in the sample has a metagenomic element in Table 1 associated with healthy skin, or if the level of a metagenomic element in Table 1 across the sample is at least 1%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 95% lower than a level associated with health.
P.
acnes Gene ID
The skin sample may be any sample taken from the skin of a subject (e.g., a human). The subject may have skin afflicted with an inflammatory skin disease (e.g., acne, PCT, or rosacea) or may have skin not afflicted with a skin disease. A skin sample may be obtained by any means known in the art including, but not limited to, swabbing the skin with a tool able to collect skin cells (e.g., a Q-tip or cotton swab), placing an adhesive or tape on the surface of the skin and removing the adhesive or tape, thereby yielding a skin sample on the adhesive or tape, or through a biopsy (e.g., a shave biopsy, a punch biopsy, an incisional biopsy, a saucerization biopsy, or an excisional biopsy).
In some embodiments, the skin sample includes epithelial cells, epidermal cells, dermal cells, skin flora (e.g., skin microbes such as P. granulosum, P. humerusii. P. avidum, P. acnes, Staphylococcus epidermidis, Staphylococcus aureus, Streptococcus pyogenes, Streptococcus mitis, Propionibacterium acnes, Corynebacterium spp., Acinetobacter johnsonii, Pseudomonas aeruginosa), fungus (e.g., Candida albicans, Rhodotorula rubra, Torulopsis and Trichosporon cutaneum, dermatophytes (skin living fungi) such as Microsporum gypseum, and Trichophyton rubrum and nondermatophyte fungi (opportunistic fungi that can live in skin) such as Rhizopus stolonifer, Trichosporon cutaneum, Fusarium, Scopulariopsisbrevicaulis, Curvularia, Alternaria alternata, Paecilomyces, Aspergillus flavus and Penicillium species), hair/skin follicles, adipose tissue (i.e., subcutaneous fat), and/or connective tissue. Preferably the skin sample includes at least some skin cells of the patient (whether living or dead) and skin flora.
In some aspects, provided herein are methods for treating acne in a subject, comprising administering a composition comprising a metabolite produced by a strain of P. granulosum, P. humerusii, and/or P. avidum wherein the metabolite is selected from bacterial culture supernatant, cell lysate, proteins, nucleic acids, lipids, and other bacterial molecules.
In general, the above methods directly act to reduce the amount of pathogenic bacteria in a subject. In some embodiments, this includes any such therapy that achieves the same goal of reducing the number of pathogenic organisms, when used in combination with the composition described herein, would lead to replacement of the pathogenic microflora involved in the diseased state with natural microflora enriched in skin not afflicted with a skin disease, or less pathogenic species occupying the same ecological niche as the type causing a disease state. For example, a subject may undergo treatment with antibiotics or a composition comprising antibiotics to target and decrease the prevalence of pathogenic organisms, and subsequently be treated with a composition described herein.
Suitable antimicrobial compounds include capreomycins, including capreomycin IA, capreomycin IB, capreomycin IIA and capreomycin IIB; carbomycins, including carbomycin A; carumonam; cefaclor, cefadroxil, cefamandole, cefatrizine, cefazedone, cefazolin, cefbuperazone, cefcapene pivoxil, cefclidin, cefdinir, cefditoren, cefime, ceftamet, cefmenoxime, cefmetzole, cefminox, cefodizime, cefonicid, cefoperazone, ceforanide, cefotaxime, cefotetan, cefotiam, cefoxitin, cefpimizole, cefpiramide, cefpirome, cefprozil, cefroxadine, cefsulodin, ceftazidime, cefteram, ceftezole, ceftibuten, ceftiofur, ceftizoxime, ceftriaxone, cefuroxime, cefuzonam, cephalexin, cephalogycin, cephaloridine, cephalosporin C, cephalothin, cephapirin, cephamycins, such as cephamycin C, cephradine, chlortetracycline; chlarithromycin, clindamycin, clometocillin, clomocycline, cloxacillin, cyclacillin, danofloxacin, demeclocyclin, destomycin A, dicloxacillin, dirithromycin, doxycyclinepicillin, erythromycin A, ethanbutol, fenbenicillin, flomoxef, florfenicol, floxacillin, flumequine, fortimicin A, fortimicin B, forfomycin, foraltadone, fusidic acid, gentamycin, glyconiazide, guamecycline, hetacillin, idarubicin, imipenem, isepamicin, josamycin, kanamycin, leumycins such as leumycin A1, lincomycin, lomefloxacin, loracarbef, lymecycline, meropenam, metampicillin, methacycline, methicillin, mezlocillin, micronomicin, midecamycins such as midecamycin A1, mikamycin, minocycline, mitomycins such as mitomycin C, moxalactam, mupirocin, nafcillin, netilicin, norcardians such as norcardian A, oleandomycin, oxytetracycline, panipenam, pazufloxacin, penamecillin, penicillins such as penicillin G, penicillin N and penicillin O, penillic acid, pentylpenicillin, peplomycin, phenethicillin, pipacyclin, piperacilin, pirlimycin, pivampicillin, pivcefalexin, porfiromycin, propiallin, quinacillin, ribostamycin, rifabutin, rifamide, rifampin, rifamycin SV, rifapentine. rifaximin. ritipenem, rekitamycin, rolitetracycline, rosaramicin, roxithromycin, sancycline, sisomicin, sparfloxacin, spectinomycin, streptozocin, sulbenicillin, sultamicillin, talampicillin, teicoplanin, temocillin, tetracyclin, thostrepton, tiamulin, ticarcillin, tigemonam, tilmicosin, tobramycin, tropospectromycin, trovafloxacin, tylosin, and vancomycin, and analogs, derivatives, pharmaceutically acceptable salts, esters, prodrugs, and protected forms thereof.
Suitable anti-fungal compounds include ketoconazole, miconazole, fluconazole, clotrimazole, undecylenic acid, sertaconazole, terbinafine, butenafine, clioquinol, haloprogin, nystatin, naftifine, tolnaftate, ciclopirox, amphotericin B. or tea tree oil and analogs, derivatives, pharmaceutically acceptable salts, esters, prodrugs, and protected forms thereof.
Suitable antiviral agents include acyclovir, azidouridine, anismoycin, amantadine, bromovinyldeoxusidine, chlorovinyldeoxusidine, cytarabine, delavirdine, didanosine, deoxynojirimycin, dideoxycytidine, dideoxyinosine, dideoxynucleoside, desciclovir, deoxyacyclovir, efavirenz, enviroxime, fiacitabine, foscamet, fialuridine, fluorothymidine, floxuridine, ganciclovir, hypericin, idoxuridine, interferon, interleukin, isethionate, nevirapine, pentamidine, ribavirin, rimantadine, stavudine, sargramostin. suramin. trichosanthin, tribromothymidine, trichlorothymidine, trifluorothymidine, trisodium phosphomonoformate, vidarabine, zidoviridine, zalcitabine and 3-azido-3-deoxy thymidine and analogs, derivatives, pharmaceutically acceptable salts, esters, prodrugs, and protected forms thereof.
Other suitable antiviral agents include 2′,3′-dideoxyadenosine (ddA), 2′,3′-dideoxyguanosine (ddG), 2′,3′-dideoxycytidine (ddC), 2′,3′-dideoxy thymidine (ddT), 2′3′-dideoxy-dideoxy thymidine (d4T), 2′-deoxy-3′-thia-cytosine (3TC or lamivudime), 2′,3′-dideoxy-2′-fluoroadenosine, 2′,3′-dideoxy-2′-fluoroinosine, 2′,3′-dideoxy-2′-fluorothymidine, 2′,3′-dideoxy-2′-fluorocytosine, 2′3′-dideoxy-2′,3′-didehydro-2′-fluorothymidine (Fd4T), 2′3′-dideoxy-2′-beta-fluoroadenosine (F-ddA), 2′3′-dideoxy-2′-beta-fluoro-inosine (F-ddI), and 2′,3′-dideoxy-2′-beta-flurocytosine (F-ddC). In some embodiments, the antiviral agent is selected from trisodium phosphomonoformate, ganciclovir, trifluorothymidine, acyclovir, 3′-azido-3′-thymidine (AZT), dideoxyinosine (ddI), and idoxuridine and analogs, derivatives, pharmaceutically acceptable salts, esters, prodrugs, and protected forms thereof.
In some aspects, the invention relates to a composition (e.g., a pharmaceutical composition) comprising a strain of P. granulosum, a strain of P. humerusii, or a strain of P. avidum. A composition may have two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more strains of P. granulosum. P. humerusii, and/or P. avidum. A composition may comprise multiple strains from one species of bacteria, or a composition may have multiple strains from different species of bacteria. For example, a composition disclosed herein may have multiple strains of P. granulosum, or may have a combination of strains from P. granulosum. P. avidum, and/or P. humerusii. The pharmaceutical composition may be formulated for topical administration. The pharmaceutical composition may be a probiotic. The pharmaceutical compositions disclosed herein may be delivered by any suitable route of administration, including orally, buccally, sublingually, parenterally, and topically, as by powders, ointments, drops, liquids, gels, or creams. In certain embodiments, the pharmaceutical compositions are delivered generally (e.g., via oral or parenteral administration). In certain other embodiments, the pharmaceutical compositions are delivered locally through injection.
Actual dosage levels of the active ingredients in the pharmaceutical compositions may be varied so as to obtain an amount of the active ingredient which is effective to achieve the desired therapeutic response for a particular patient, composition, and mode of administration, without being toxic to the patient.
The selected dosage level will depend upon a variety of factors including the activity of the particular agent employed, the route of administration, the time of administration, the rate of excretion or metabolism of the particular compound being employed, the duration of the treatment, other drugs, compounds and/or materials used in combination with the particular compound employed, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and like factors well known in the medical arts.
A physician or veterinarian having ordinary skill in the art can readily determine and prescribe the effective amount of the pharmaceutical composition required. For example, the physician or veterinarian could prescribe and/or administer doses of the compounds employed in the pharmaceutical composition at levels lower than that required in order to achieve the desired therapeutic effect and gradually increase the dosage until the desired effect is achieved.
Exemplary identities of various constituents of the topical formulations of some embodiments of the present invention are described below.
Suitable topical vehicles and vehicle components for use with the formulations of the invention are well known in the cosmetic and pharmaceutical arts, and include such vehicles (or vehicle components) as water; organic solvents such as alcohols (particularly lower alcohols readily capable of evaporating from the skin such as ethanol), glycols (such as propylene glycol, butylene glycol, and glycerol (glycerin)), aliphatic alcohols (such as lanolin); mixtures of water and organic solvents (such as water and alcohol), and mixtures of organic solvents such as alcohol and glycerol (optionally also with water); lipid-based materials such as fatty acids, acylglycerols (including oils, such as mineral oil, and fats of natural or synthetic origin), phosphoglycerides, sphingolipids and waxes; protein-based materials such as collagen and gelatin; silicone-based materials (both non-volatile and volatile) such as cyclomethicone, dimethiconol, dimethicone, and dimethicone copolyol; hydrocarbon-based materials such as petrolatum and squalane; and other vehicles and vehicle components that are suitable for administration to the skin, as well as mixtures of topical vehicle components as identified above or otherwise known to the art.
In one embodiment, the compositions of the present invention are oil-in-water emulsions. Liquids suitable for use in formulating compositions of the present invention include water, and water-miscible solvents such as glycols (e.g., ethylene glycol, butylene glycol, isoprene glycol, propylene glycol), glycerol, liquid polyols, dimethyl sulfoxide, and isopropyl alcohol. One or more aqueous vehicles may be present.
In some embodiments, formulations do not have methanol, ethanol, propanols, or butanol.
Many topical formulations contain chemical emulsions which use surface active ingredients (emulsifiers and surfactants) to disperse dissimilar chemicals in a particular solvent system. For example, most lipid-like (oily or fatty) or lipophilic ingredients do not uniformly disperse in aqueous solvents unless they are first combined with emulsifiers, which form microscopic aqueous soluble structures (droplets) that contain a lipophilic interior and a hydrophilic exterior, resulting in an oil-in-water emulsion. In order to be soluble in aqueous media, a molecule must be polar or charged so as to favorably interact with water molecules, which are also polar. Similarly, to dissolve an aqueous-soluble polar or charged ingredient in a largely lipid or oil-based solvent, an emulsifier is typically used which forms stable structures that contain the hydrophilic components in the interior of the structure while the exterior is lipophilic so that it can dissolve in the lipophilic solvent to form a water-in-oil emulsion. It is well known that such emulsions can be destabilized by the addition of salts or other charged ingredients which can interact with the polar or charged portions of the emulsifier within an emulsion droplet. Emulsion destabilization results in the aqueous and lipophilic ingredients separating into two layers, potentially destroying the commercial value of a topical product.
Surfactants suitable for use in the present invention may be ionic or non-ionic. These include, but are not limited to: cetyl alcohol, polysorbates (Polysorbate 20, Polysorbate 40, Polysorbate 60, Polysorbate 80), steareth-10 (Brij 76), sodium dodecyl sulfate (sodium lauryl sulfate), lauryl dimethyl amine oxide, cetyltrimethylammonium bromide (CTAB), polyethoxylated alcohols, polyoxyethylene sorbitan, octoxynol, N,N-dimethyldodecylamine-N-oxide, hexadecyltrimethylammonium bromide (HTAB), polyoxyl 10 lauryl ether, bile salts (such as sodium deoxycholate or sodium cholate), polvoxyl castor oil, nonylphenol ethoxylate, cyclodextrins, lecithin, dimethicone copolyol, lauramide DEA, cocamide DEA, cocamide MEA, oleyl betaine, cocamidopropyl betaine, cocamidopropyl phosphatidyl PG-dimonium chloride, dicetyl phosphate (dihexadecyl phosphate), ceteareth-10) phosphate, methylbenzethonium chloride, dicetyl phosphate, ceteth-10 phosphate (ceteth-10 is the polyethylene glycol ether of cetyl alcohol where n has an average value of 10; ceteth-10 phosphate is a mixture of phosphoric acid esters of ceteth-10), ceteth-20, Brij S10 (polyethylene glycol octadecyl ether, average Mn˜711), and Poloxamers (including, but not limited to, Poloxamer 188 (HO(C2H4O)a(CH(CH3)CH2O)b(C2H4O)aH, average molecular weight 8400) and Poloxamer 407 (HO(C2H4O)a(CH(CH3)CH2O)b(C2H4O)aH, wherein a is about 101 and b is about 56)). Appropriate combinations or mixtures of such surfactants may also be used according to the present invention. Many of these surfactants may also serve as emulsifiers in formulations of the present invention.
Other suitable emulsifiers for use in the formulations of the present invention include, but are not limited to, behentrimonium methosulfate-cetearyl alcohol, non-ionic emulsifiers like emulsifying wax, polyoxyethylene oleyl ether. PEG-40 stearate, cetostearyl alcohol (cetearyl alcohol), ceteareth-12, ceteareth-20, ceteareth-30), ceteareth alcohol. Ceteth-20) (Ceteth-20 is the polyethylene glycol ether of cetyl alcohol where n has an average value of 20), oleic acid, oleyl alcohol, glyceryl stearate, PEG-75 stearate. PEG-100) stearate, and PEG-100 stearate, ceramide 2, ceramide 3, stearic acid, cholesterol, steareth-2, and steareth-20, or combinations/mixtures thereof, as well as cationic emulsifiers like stearamidopropyl dimethylamine and behentrimonium methosulfate, or combinations/mixtures thereof.
Moisturizers. Emollients, and Humectants
One of the most important aspects of topical products in general, and cosmetic products in particular, is the consumer's perception of the aesthetic qualities of a product. For example, while white petrolatum is an excellent moisturizer and skin protectant, it is rarely used alone, especially on the face, because it is greasy, sticky, does not rub easily into the skin and may soil clothing. Consumers highly value products which are aesthetically elegant and have an acceptable tactile feel and performance on their skin. Suitable moisturizers for use in the formulations of the present invention include, but are not limited to, lactic acid and other hydroxy acids and their salts, glycerol, propylene glycol, butylene glycol, sodium PCA, sodium hyaluronate, Carbowax 200, Carbowax 400, and Carbowax 800. Suitable emollients or humectants for use in the formulations of the present invention include, but are not limited to, panthenol, cetyl palmitate, glycerol (glycerin). PPG-15 stearyl ether, lanolin alcohol, lanolin, lanolin derivatives, cholesterol, petrolatum, isostearyl neopentanoate, octyl stearate, mineral oil, isocetyl stearate, myristyl myristate, octyl dodecanol. 2-ethylhexyl palmitate (octyl palmitate), dimethicone, phenyl trimethicone, cyclomethicone. C12-C15 alkyl benzoates, dimethiconol, propylene glycol. Theobroma grandiflorum seed butter, ceramides (e.g., ceramide 2 or ceramide 3), hydroxypropyl bispalmitamide MEA, hydroxypropyl bislauramide MEA, hydroxypropyl bisisostearamide MEA. 1,3-bis(N-2-(hydroxyethyl)stearoylamino)-2-hydroxy propane, bis-hydroxyethyl tocopherylsuccinoylamido hydroxypropane, urea, aloe, allantoin, glycyrrhetinic acid, safflower oil, oleyl alcohol, oleic acid, stearic acid, dicaprylate/dicaprate, diethyl sebacate, isostearyl alcohol, pentylene glycol, isononyl isononanoate, and 1,3-bis(N-2-(hydroxyethyl)palmitoylamino)-2-hydroxypropane. In addition, appropriate combinations and mixtures of any of these moisturizing agents and emollients may be used in accordance with the present invention.
The composition may further include components adapted to improve the stability or effectiveness of the applied formulation.
Suitable preservatives for use in the present invention include, but are not limited to: ureas, such as imidazolidinyl urea and diazolidinyl urea; phenoxyethanol; sodium methyl paraben, methylparaben, ethylparaben, and propylparaben; potassium sorbate; sodium benzoate; sorbic acid; benzoic acid; formaldehyde; citric acid; sodium citrate; chlorine dioxide; quaternary ammonium compounds, such as benzalkonium chloride, benzethonium chloride, cetrimide, dequalinium chloride, and cetylpyridinium chloride; mercurial agents, such as phenylmercuric nitrate, phenylmercuric acetate, and thimerosal; piroctone olamine; Vitis vinifera seed oil; and alcoholic agents, for example, chlorobutanol, dichlorobenzyl alcohol, phenylethyl alcohol, and benzyl alcohol.
Suitable antioxidants include, but are not limited to, ascorbic acid and its esters, sodium bisulfite, butylated hydroxytoluene, butylated hydroxyanisole, tocopherols, tocopheryl acetate, sodium ascorbate/ascorbic acid, ascorbyl palmitate, propyl gallate, and chelating agents like EDTA (e.g., disodium EDTA), citric acid, and sodium citrate.
In some embodiments, the antioxidant or preservative comprises (3-(4-chlorophenoxy)-2-hydroxy propyl)carbamate.
In some embodiments, antioxidants or preservatives of the present invention may also function as a moisturizer or emollient, for example.
In addition, combinations or mixtures of these preservatives or anti-oxidants may also be used in the formulations of the present invention.
The composition can also contain any other agent that has a desired effect when applied topically to a subject. Suitable classes of active agents include, but are not limited to antibiotic agents (i.e., antibiotic agents that dot not target P. granulosum), antimicrobial agents, anti-acne agents, antibacterial agents, antifungal agents, antiviral agents, steroidal anti-inflammatory agents, non-steroidal anti-inflammatory agents, anesthetic agents, antipruriginous agents, antiprotozoal agents, anti-oxidants, antihistamines, vitamins, and hormones. Mixtures of any of these active agents may also be employed. Additionally, dermatologically-acceptable salts and esters of any of these agents may be employed.
Suitable viscosity adjusting agents (i.e., thickening and thinning agents or viscosity modifying agents) for use in the formulations of the present invention include, but are not limited to, protective colloids or non-ionic gums such as hydroxyethylcellulose, xanthan gum, and sclerotium gum, as well as magnesium aluminum silicate, silica, microcrystalline wax, beeswax, paraffin, and cetyl palmitate. In addition, appropriate combinations or mixtures of these viscosity adjusters may be utilized according to the present invention.
Additional constituents suitable for incorporation into the emulsions of the present invention include, but are not limited to: skin protectants, adsorbents, demulcents, emollients, moisturizers, sustained release materials, solubilizing agents, skin-penetration agents, skin soothing agents, deodorant agents, antiperspirants, sun screening agents, sunless tanning agents, vitamins, hair conditioning agents, anti-irritants, anti-aging agents, abrasives, absorbents, anti-caking agents, anti-static agents, astringents (e.g., witch hazel, alcohol, and herbal extracts such as chamomile extract), binders/excipients, buffering agents, chelating agents, film forming agents, conditioning agents, opacifying agents, lipids, immunomodulators, and pH adjusters (e.g., citric acid, sodium hydroxide, and sodium phosphate). For example, lipids normally found in healthy skin (or their functional equivalents) may be incorporated into the emulsions of the present invention. In certain embodiments, the lipid is selected from the group consisting of ceramides, cholesterol, and free fatty acids. Examples of lipids include, but are not limited to, ceramide 1, ceramide 2, ceramide 3, ceramide 4, ceramide 5, ceramide 6, hydroxypropyl bispalmitamide MEA, and hydroxypropyl bislauramide MEA, and combinations thereof.
Examples of peptides that interact with protein structures of the dermal-epidermal junction include palmitoyl dipeptide-5 diaminobutyloyl hydroxythreonine and palmitoyl dipeptide-6 diaminohydroxy butyrate.
Examples of skin soothing agents include, but are not limited to algae extract, mugwort extract, stearyl glycyrrhetinate, bisabolol, allantoin, aloe, avocado oil, green tea extract, hops extract, chamomile extract, colloidal oatmeal, calamine, cucumber extract, and combinations thereof.
In certain embodiments, the compositions comprise bergamot or bergamot oil. Bergamot oil is a natural skin toner and detoxifier. In certain embodiments, it may prevent premature aging of skin and may have excellent effects on oily skin conditions and acne.
In some embodiments, the composition comprises a vitamin. Examples of vitamins include, but are not limited to, vitamins A, D, E, K, and combinations thereof. Vitamin analogues are also contemplated; for example, the vitamin D analogues calcipotriene or calcipotriol. In some embodiments, the vitamin may be present as tetrahexyldecyl ascorbate. This compound exhibits anti-oxidant activity, inhibiting lipid peroxidation. In certain embodiments, use can mitigate the damaging effects of UV exposure. Studies have shown it to stimulate collagen production as well as clarifying and brightening the skin by inhibiting melanogenesis (the production of pigment) thereby promoting a more even skin tone.
In some embodiments, the composition comprises a sunscreen. Examples of sunscreens include, but are not limited to, p-aminobenzoic acid, avobenzone, cinoxate, dioxybenzone, homosalate, menthyl anthranilate, octocrylene, octyl methoxycinnamate, octyl salicylate, oxybenzone, padimate O, phenylbenzimidazole sulfonic acid, sulisobenzone, titanium dioxide, trolamine salicylate, zinc oxide, 4-methylbenzylidene camphor, methylene bis-benzotriazolyl tetramethylbutylphenol, bis-ethylhexyloxyphenol methoxyphenyl triazine, terephthalylidene dicamphor sulfonic acid, drometrizole trisiloxane, disodium phenyl dibenzimidazole tetrasulfonate, diethylamino hydroxybenzoyl hexyl benzoate, octyl triazone, diethylhexyl butamido triazone, polysilicone-15, and combinations thereof.
Suitable fragrances and colors may be used in the formulations of the present invention. Examples of fragrances and colors suitable for use in topical products are known in the art.
To identify the disease- and health-associated microbial elements in the follicular microbiota, ultra-deep metagenomic shotgun sequencing was performed of the samples collected from 38 acne patients and 30 age-matched healthy individuals (
At the species level, P. acnes was the most prevalent and abundant. It was found in all 68 individuals with an average relative abundance of 91.0% (
P.
acnes
P.
humerusii
P.
granulosum
P.
avidum
S.
epidermidis
Other cutaneous Propionibacterium species, including P. humerusii (2.7%), P. granulosum (1.6%), and P. avidum (0.4%), were also detected (Table 2). Among them, P. granulosum, a resident bacterium of sebaceous skin sites, was significantly more abundant in the healthy individuals than in the acne patients (P=0.002). This is the same trend as seen in P. acnes.
Other bacterial species frequently identified in the follicle microbiota include Staphylococcus epidermidis (0.9%), Staphylococcus capitis (0.4%), Escherichia coli (0.7%), and Clostridium sp. (0.5%). Minor bacterial taxa were found more prevalent and abundant in acne patients than in healthy individuals with varying presence and abundance. On average, acne patients had a higher diversity at the species level than healthy individuals, but not significantly different (Shannon index=0.63 and 0.44, respectively; P=0.072). Reduced P. acnes and P. granulosum and increased minor taxa observed in acne patients suggest disruptions of the commensal skin flora in acne development.
Acne rarely occurs in individuals over the age of 50. The skin microbiome profiles of older individuals with clear skin can be used as independent references for healthy, acne-free skin. However, the skin metagenome of older healthy individuals has not been characterized. Metagenomic shotgun sequencing was performed of the samples collected from four healthy individuals of age 55-79 (
When comparing all 34 healthy individuals, including the four subjects over the age of 55, with the 38 acne patients, were found an increased prevalence and abundance of P. acnes phage in the healthy group (P 0.05) (
To better understand the role of the follicular microbiota in acne pathogenesis at the molecular level, the differences in metagenomic composition were investigated of the microbiome between acne patients and healthy individuals. Among the sequencing reads mapped to P. acnes. 2,707 operational gene units (OGUs) were identified in the samples, among which 1,943 OGUs (72%) were present in every sample. Rarefaction analysis indicated that the sequencing depth was sufficient to quantitatively compare the P. acnes functional profiles between acne patients and healthy individuals (
To determine whether specific metagenomic elements are associated with acne or healthy skin, the relative abundances of P. acnes OGUs were compared between acne patients and healthy individuals (
Among the 62 P. acnes OGUs. 25 were significantly more abundant in acne patients. Two of them are involved in thiopeptide bacteriocin precursor synthesis and transport (PAGK2104 and PAGK2105, as annotated in P. acnes HL096PA1 genome). Thiopeptide bacteriocins belong to a family of microcins, which are produced by Gram-positive bacteria and inhibit the growth of other Gram-positive species by blocking protein translation. Additionally. 19 of the 25 acne-associated OGUs were from locus 2 (PAGK0160-PAGK0178), a genomic island previously identified mainly in RT4 and RT5 strains and highly associated with acne. Locus 2 spans over 20 Kb and encodes 23 ORFs (
A higher average relative abundance and prevalence of all three loci was found in acne patients compared to healthy individuals (
Of the 62 differentially abundant P. acnes OGUs. 37 OGUs were more abundant in healthy individuals. In contrast to the enrichment of virulence-related genes observed in the acne metagenome, genes involved in microbial metabolism and nutrient biosynthesis were significantly more abundant in the healthy metagenome (
Observations of differentially abundant species, strains, and metagenomic elements between the skin microbiome of acne patients and healthy individuals led to the hypothesis that the balance between acne- and health-associated metagenomic elements determines the virulence and health properties of the microbiota in skin disease and health. The balance is defined as the ratio between the relative abundances of acne- and health-associated metagenomic elements. When the relative abundances of the 62 P. acnes OGUs and P. granulosum from all individuals were compared, the ratio in acne patients was significantly higher than in healthy individuals (P=2.9×10−5).
In addition to P. acnes OGUs, it was found that the relative abundances of P. acnes and P. granulosum were also important factors influencing the clinical status of the skin. As an example, an acne patient in the cohort (labeled with † on
It was investigated whether the relative abundance profiles of the metagenomic elements are sufficiently robust to correctly classify the clinical status of skin samples. A supervised class performed prediction analysis based on a modified weighted gene voting algorithm. One thousand permutations were generated. In each permutation, samples were randomly assigned to either acne or healthy status, with 38 samples in the acne group and 34 samples in the healthy group. Differentially abundant metagenomic elements were identified between the two groups among all but one sample, and predicted the clinical state of the withheld sample based on the abundance profiles of the metagenomic element, known as the leave-one-out cross-validation (LOOCV). With the use of the clinically defined acne-healthy grouping, the algorithm predicted the clinical states of 49 out of 72 samples (68%) with a prediction strength threshold of 0.25. Thirty-four of the 49 samples were assigned correctly (69% accuracy). The number of correctly assigned samples based on the clinical grouping was higher than most of the permutated groupings (P=0.064). The clinical grouping also had a significantly higher prediction accuracy (69%) than all but one of the permutated groupings (P 0.001) (
The classifier was improved by using the 63 metagenomic elements, which are the most robust set based on the 100 random samplings as described earlier. Since the 19 OGUs from locus 2 had similar abundance profiles across all samples, to avoid overweighing locus 2 in the classification. 19 OGUs were combined and used their average relative abundance in the classifier. The refined classifier therefore consisted of the abundance profiles of 45 metagenomic elements: P. acnes OGUs/loci, and P. granulosum. Based on this refined set, the clinical states of 31 of the 38 acne patients (82%, prediction strength threshold of 0.25) were assigned. Twenty-four acne patients were correctly assigned (accuracy of 77%). Among the healthy individuals. 21 of the 34 subjects (62%) were assigned with 20 correctly assigned (accuracy of 95%). Overall, the classifier was able to assign 72% of the subjects with an accuracy of 85% (
Furthermore, to validate that these metagenomic elements can be used as markers for clinical classification, an “independent sample set” was collected from ten additional subjects, including 4 acne patients and 6 healthy individuals, one of which was over 50 years old. The refined 45 metagenomic elements were used to predict the clinical state of each independent sample (
A performed supervised class prediction analysis. Differentially abundant metagenomic elements, including P. acnes OGUs and bacterial species, were determined between two groups of samples (healthy and acne). A method similar to that described by Golub et al. Science 286, 531-537 (1999) and Bleharski et al. Science 301, 1527-1530, hereby incorporated by reference the their entirety, was employed, using the formula:
All publications, patents, and patent applications mentioned herein are hereby incorporated by reference in their entirety as if each individual publication, patent or patent application was specifically and individually indicated to be incorporated by reference. In case of conflict, the present application, including any definitions herein, will control.
Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments are described herein. Such equivalents are intended to be encompassed by the following claims.
This application is a Continuation of application Ser. No. 16/347,706 filed on May 6, 2019, which claims the benefit of priority to 371 national-stage application based on International Application No. PCT/US17/60319 filed Nov. 7, 2017, which claims the benefit of priority to U.S. Provisional Patent Application Ser. No. 62/419,182, filed Nov. 8, 2016, and U.S. Provisional Patent Application Ser. No. 62/471,578, filed on Mar. 15, 2017, each of which are herein incorporated by reference in their entireties.
This invention was made with Government support under Grant Number GM099530, awarded by the National Institutes of Health. The Government has certain rights in the invention.
| Number | Date | Country | |
|---|---|---|---|
| 62471578 | Mar 2017 | US | |
| 62419182 | Nov 2016 | US |
| Number | Date | Country | |
|---|---|---|---|
| Parent | 16347706 | May 2019 | US |
| Child | 18381795 | US |
