Botulinum neurotoxin for therapeutic purposes

Abstract

The present invention provides compositions and methods for cosmetic procedures. [Solution] Disclosed herein are compositions and methods for therapeutic treatments. Use of "rapid-acting" botulinum toxin in cosmetic treatments and / or for preventing or reducing scarring. "Rapid-acting" means botulinum toxin that exerts its effect in a patient more rapidly than, for example, botulinum neurotoxin type A. "Rapid-acting" botulinum toxin, for example, type E.

Description

[Technical Field]

[0001] This specification relates to the use of neurotoxins in therapeutic purposes. [Background technology]

[0002] Cosmetic surgery or reconstructive surgery includes surgical and non-surgical procedures that improve and reshape the body's structure to enhance appearance and self-confidence. These procedures are becoming increasingly common. [Overview of the project] [Problems that the invention aims to solve]

[0003] Disclosed herein are compositions and methods for cosmetic treatments. For example, the disclosed embodiments include, for instance, the use of “fast-acting” botulinum toxin to treat frown lines. Disclosed herein are compositions and methods for use in minimizing scarring. For example, the disclosed embodiments include the use of “fast-acting” botulinum toxin to reduce muscle tension adjacent to the wound, and thus prevent or reduce scarring. [Means for solving the problem]

[0004] In this embodiment, the activity of muscles adjacent to the skin incision or laceration is reduced, thus reducing or preventing scar formation. In embodiments, the disclosed method includes additional cosmetic procedures. For example, the disclosed embodiments include the administration of rapid-acting botulinum neurotoxin in combination with, for example, dermal filler injections, eye lifts, rhinoplasty, or similar procedures.  Botulinum toxin is a "rapidly recovering" toxin. In an embodiment, the "rapid-acting" botulinum toxin is also a "fast-recovering" toxin. In some embodiments, the cosmetic treatment may include an additional administration of botulinum toxin after the initial dose. In embodiments, the disclosed method includes, for example, the administration of a rapid-acting botulinum neurotoxin in combination with a slower-acting neurotoxin. In embodiments, the disclosed method includes, for example, the administration of a rapidly regenerating botulinum neurotoxin in combination with a slowly regenerating neurotoxin. In one embodiment, the dose of neurotoxin is expressed in terms of the amount of protein. [Brief explanation of the drawing]

[0005] [Figure 1] This is a diagram illustrating the injection sites used in cosmetic surgery. [Figure 2] This graph shows the primary effectiveness of a trial treating frown lines between the eyebrows. [Figure 3] This graph shows the secondary efficacy of a trial treating frown lines. [Figure 4] This graph shows the effect of a single topical administration of the disclosed type E botulinum toxin composition in a rat model of postoperative pain. [Modes for carrying out the invention]

[0006] Embodiments disclosed herein reduce local muscle activity, thereby reducing cosmetic imperfections or irregularities in appearance, such as facial wrinkles. In embodiments, cosmetic imperfections include frown lines, forehead wrinkles, "bunny" lines, laugh lines, jawline irregularities, platysmal bands, "marionette" lines, lip wrinkles, crow's feet, brow irregularities, combinations thereof, and similar. Embodiments include methods involving dermatological surgery, such as the treatment of actinic keratosis, seborrheic keratosis, basal cell carcinoma, squamous cell carcinoma, and other lesions or subcutaneous cysts. Useful injection sites for carrying out the disclosed embodiments may include the glabellar complex, including the corrugator supercilii and procerus muscles; the orbicularis oculi muscle; the superior lateral fibers of the orbicularis oculi muscle; the frontalis muscle; the nasal muscle; the levator labii superioris alaeque nasi muscle; the orbicularis oris muscle; the masseter muscle; the depressor anguli oris muscle; and the platysma muscle. A typical injection site useful for treating glabellar wrinkles is shown in Figure 1. Embodiments include the treatment of general wrinkles.

[0007] The disclosed embodiments may include, for example, the treatment of skin disorders, such as acne and similar conditions. The disclosed embodiments may include the treatment of inflammatory skin diseases. For example, the disclosed embodiments may include the treatment of psoriasis, eczema and similar conditions. Embodiments disclosed herein can reduce local muscle activity, thereby reducing the development of scars, such as surgical scars. In embodiments, the surgical procedure may include cosmetic surgery, such as rhinoplasty, eye lifts, "tummy" tucks (abdominal wall reconstruction), or similar procedures. In embodiments, the procedure may include other types of medical procedures, such as appendectomy, organ transplantation, and similar procedures. In embodiments, the method includes administering the disclosed composition near a wound. Embodiments disclosed herein can reduce local muscle activity, thereby reducing the development of scars, such as those resulting from trauma. For example, after a traumatic injury, embodiments of the disclosed embodiment may include administering the composition disclosed to the vicinity of the injury, such as a laceration or amputation. Useful administration sites for carrying out the disclosed embodiments may include any area where muscle activity is to be reduced. For example, when used in conjunction with facial cosmetic surgery, the disclosed embodiments may include administration to the glabellar complex including the corrugator supercilii and procerus muscles; the orbicularis oculi muscle; the superior lateral fibers of the orbicularis oculi muscle; the frontalis muscle; the nasal muscle; the levator labii superioris alaeque nasi muscle; the orbicularis oris muscle; the masseter muscle; the depressor anguli oris muscle; and the platysma muscle.

[0008] When used in conjunction with other surgical procedures, the disclosed embodiments may include, for example, administration to the muscles of the arms, legs, torso, and similar bodies. The disclosed embodiments may include methods for preparing the surgical site prior to surgery in order to reduce tension in muscles adjacent to the incision. The disclosed embodiments can, for example, promote the production of elastin, collagen, and similar substances. The disclosed embodiments may include methods for increasing skin elasticity. In embodiments, the compositions disclosed herein may include a rapid-acting botulinum toxin, for example, type E. In embodiments, the compositions disclosed herein may include a rapidly recovering botulinum toxin, such as type E. In embodiments, the compositions disclosed herein may include a rapid-acting, quickly reversible botulinum toxin, such as type E.

[0009] Definition: "Administer" or "to administer" means the step of giving (i.e., administering) a pharmaceutical composition or active ingredient to a subject. The pharmaceutical compositions disclosed herein may be administered by several suitable routes, but as described in the method of disclosure, the compositions are administered topically, for example, by an intramuscular route, such as injection or implantable tablet. "Botulinum toxin" or "botulinum neurotoxin" means wild-type neurotoxin derived from Clostridium botulinum, as well as modified, genetically modified, hybrid, and chimeric botulinum toxins. Genetically modified botulinum toxin has its light and / or heavy chains produced by genetic modification from a non-Clostridium species. As used herein, "botulinum toxin" encompasses botulinum toxin serotypes A, B, C, D, E, F, G, and H. As used herein, "botulinum toxin" also includes both botulinum toxin complexes (i.e., 300, 600, and 900 kDa complexes) and pure botulinum toxin (i.e., approximately 150 kDa neurotoxin molecules), all of which are useful in the practice of the present invention. "Pure botulinum toxin" refers to botulinum toxin complexes isolated or substantially isolated from pure botulinum toxin or other proteins, as well as impurities that may be present when botulinum toxin is obtained by culture or fermentation processes. Therefore, in pure botulinum toxin, at least 95%, more preferably at least 99%, of non-botulinum toxin proteins and impurities have been removed.

[0010] "Biocompatible" means that there is little inflammatory reaction at the implantation site of the implant tablet. "Clostridium neurotoxin" means a neurotoxin produced by Clostridium bacteria such as Clostridium botulinum, Clostridium butyricum or Clostridium beratti, or a neurotoxin specific to those bacteria, as well as a Clostridium neurotoxin produced by genetic recombination by non-Clostridium species. "Completely free" (the term "consisting of") means that within the detection range of the device or process being used, a substance cannot be detected or its presence cannot be confirmed. "Substantially free" means that within the detection range of the device or process being used, only trace amounts of a substance can be detected.

[0011] As used herein, "rapid onset" refers to a botulinum toxin that exhibits an effect more rapidly in a patient than, for example, that exerted by botulinum neurotoxin type A. For example, the effect of a rapid onset botulinum toxin can be visually observed within 36 hours. As used herein, "rapid recovery" refers to a botulinum toxin whose effect decreases more rapidly in a patient than, for example, the effect exerted by botulinum neurotoxin type A. For example, the effect of an early recovery botulinum toxin decreases within, for example, 120 hours, 150 hours, 300 hours, 350 hours, 400 hours, 500 hours, 600 hours, 700 hours, 800 hours, or the like. Botulinum neurotoxin type A is known to have an effect for up to 12 months. However, the normal duration of intramuscular injection of botulinum neurotoxin type A is typically about 3 to 4 months. As used herein, "moderate acting" refers to a botulinum toxin that exhibits an effect more slowly than a rapid onset toxin. "Neurotoxin" means a biologically active molecule that has a specific affinity for neuronal cell surface receptors. Neurotoxins include both Clostridium toxins as pure toxins and as complexes with one or more non-toxins, toxin-related proteins.

[0012] "Patient" means a human or non-human subject receiving medical or veterinary care. "Pharmaceutical composition" means a formulation in which the active ingredient may be botulinum toxin. The term "formulation" means that in the pharmaceutical composition, in addition to the botulinum neurotoxin active ingredient, there is at least one additional ingredient (such as, but not limited to, albumin [such as human serum albumin or recombinant human albumin] and / or sodium chloride, etc.). Thus, the pharmaceutical composition is a formulation suitable for diagnostic, therapeutic or cosmetic administration to a subject such as a human patient. The pharmaceutical composition may be in a lyophilized or vacuum dried state, a solution formed after re-dissolving the lyophilized or vacuum dried pharmaceutical composition in physiological saline or water, for example, or a solution that does not require re-dissolving. As already described, the pharmaceutical composition may be liquid or solid. The pharmaceutical composition may be free of animal proteins.

[0013] "Substantially free" means present at a level of less than 1 percent by mass of the mass of the substance in a medium, fermentation medium, pharmaceutical composition or other substance in which the mass percentage of the substance is calculated. As used herein, "additional administration" refers to a subsequent administration of botulinum following an initial administration of neurotoxin.

[0014] "Therapeutic formulation" means a formulation that can be used to treat and thereby alleviate a disorder or disease, such as a disorder or disease characterized by the activity of peripheral muscles, and / or the symptoms associated therewith. "Therapeutic dose" means the level, amount, or concentration of a drug (e.g., botulinum toxin or a pharmaceutical composition containing botulinum toxin) necessary to treat a disease, disorder, or condition without causing significant negative or adverse side effects. "To treat," "to treat," or "to treat" means to reduce or diminish (including slight reduction, significant reduction, near-complete reduction, and complete reduction), eliminate or prevent (temporary or permanent) a disease, disorder or condition in order to achieve a desired therapeutic or cosmetic outcome, for example, by healing damaged or impaired tissue, or by partially altering, altering, improving, refining, restoring, and / or beautifying an existing or perceived disease, disorder or condition. "Unit" or "U" refers to the amount of active BoNT standardized to have a neuromuscular blocking effect equivalent to one unit of commercially available botulinum neurotoxin type A.

[0015] Neurotoxin composition Embodiments disclosed herein include neurotoxin compositions, for example, rapid-acting neurotoxin compositions, for example, botulinum toxin type E compositions. Such neurotoxins may be formulated in any pharmacopoeia acceptable form and in any pharmacopoeia acceptable formulation. Neurotoxins may also be used in any pharmacopoeia acceptable form supplied by any manufacturer. Embodiments disclosed herein include neurotoxin compositions, for example, rapidly regenerating neurotoxins. Such neurotoxins may be formulated in any pharmacopoecitable form and in any pharmacopoecitable formulation. Neurotoxins may also be used in any pharmacopoecitable form supplied by any manufacturer. The embodiments disclosed herein may include a variety of neurotoxins. For example, in embodiments, the disclosed composition may include two neurotoxins, such as two types of botulinum neurotoxins, such as a rapid-acting and a slower-acting neurotoxin, for example, type E and type A botulinum neurotoxins. In embodiments, the disclosed composition may include a fragment of botulinum neurotoxin, for example, a 50 kDa light chain (LC). The neurotoxin may be produced by Clostridium bacteria such as Clostridium botulinum, Clostridium butyricum, or Clostridium bellattii. Furthermore, the neurotoxin may be a modified neurotoxin; a neurotoxin in which at least one amino acid is removed, modified, or substituted compared to the natural or wild-type neurotoxin. Furthermore, the neurotoxin may be a genetically modified neurotoxin or its derivatives or fragments.

[0016] In embodiments, the disclosed Type E composition has 40% amino acid homology to Type A, and both share the same basic domain structure consisting of two chains linked by disulfide bonds: a 100 kDa heavy chain (HC) and a 50 kDa light chain (LC) (Whelan 1992). The HC contains a receptor-binding domain and a translocation domain, while the LC contains synaptosome-associated protein (SNAP) enzyme activity. The domain structure is identical to that shared by all botulinum neurotoxin serotypes. In the disclosed embodiments, the neurotoxin is formulated in a unit dose form. For example, it may be provided as a sterile solution in a vial, or as a vial or sachet containing a lyophilized powder that is redissolved in an injection-suitable solvent such as physiological saline. In one embodiment, botulinum toxin is formulated in a solution containing physiological saline and pasteurized human serum albumin to stabilize the toxin and minimize loss due to nonspecific adsorption. The solution may be aseptically filtered (0.2 μm filter), filled into individual vials, and then vacuum-dried to obtain a sterile lyophilized powder. For use, the powder may be redissolved by adding sterile, preservative-free physiological saline (0.9% sodium chloride for injection).

[0017] In this embodiment, botulinum toxin type E is supplied in a 5 mL vial as a sterile solution for injection with a nominal concentration of 20 ng / mL in 0.03 M sodium phosphate, containing 0.12 M sodium chloride and 1 mg / mL human serum albumin (HSA), and at pH 6.0. In one embodiment, botulinum toxin type E is supplied in a 5 mL vial as a sterile solution for injection with a nominal concentration of 10 ng / mL in 0.03 M sodium phosphate, containing 0.12 M sodium chloride and 1 mg / mL HSA, and with a pH of 6.0. In one embodiment, botulinum toxin type E is supplied in a 5 mL vial as a sterile solution for injection with a nominal concentration of 5 ng / mL in 0.03 M sodium phosphate, containing 0.12 M sodium chloride and 1 mg / mL HSA, and with a pH of 6.0. In one embodiment, botulinum toxin type E is supplied in a 5 mL vial as a sterile solution for injection with a nominal concentration of 1 ng / mL in 0.03 M sodium phosphate, containing 0.12 M sodium chloride and 1 mg / mL HSA, and with a pH of 6.0.

[0018] The composition may contain only one type of neurotoxin, such as botulinum toxin type E, but the disclosed composition may contain two or more types of neurotoxins that can enhance the therapeutic effect against the disorder. For example, the composition administered to the patient may contain botulinum toxin types A and E. Administering a single composition containing two different neurotoxins allows for a lower effective concentration of each neurotoxin than when a single neurotoxin is administered to the patient, while still achieving the desired therapeutic effect. The composition administered to the patient may also contain other pharmaceutically active components, such as protein receptors or ion channel modulators, in combination with one or more neurotoxins. These modulators can contribute to reducing neurotransmission between various nerve cells. For example, the composition may contain GABA A It may contain a gamma-aminobutyric acid (GABA) type A receptor modulator that enhances receptor-mediated inhibitory effects. A Receptors suppress nerve activity by effectively short-circuiting electrical currents passing through the cell membrane. GABA A The receptor modulator is GABA A GABA can enhance the inhibitory effect of receptors and reduce electrical or chemical signaling from nerve cells. AExamples of receptor modulators include benzodiazepines, such as diazepam, oxazepam, lorazepam, prazepam, alprazolam, harazepam, cordiazepoxide, and chlorazepate. The composition may also include glutamate receptor modulators that reduce the excitatory effects mediated by glutamate receptors. Examples of glutamate receptor modulators include agents that suppress current flux via AMPA, NMDA, and / or kainate-type glutamate receptors. The composition may also include agents that modulate dopamine receptors, such as antipsychotics, norepinephrine receptors, and / or serotonin receptors. The composition may also include agents that affect ion flux via voltage-gated calcium channels, potassium channels, and / or sodium channels. Therefore, the composition used in the disclosed embodiments may include one or more neurotoxins, such as botulinum toxin, in addition to ion channel receptor modulators that can reduce neurotransmission.

[0019] How to use The methods disclosed herein may include administering a rapid-acting neurotoxin to a patient. In one preferred embodiment, the neurotoxin is botulinum toxin type E. The methods disclosed herein may include administering a rapidly recovering neurotoxin to a patient. In a preferred embodiment, the neurotoxin is botulinum toxin type E. Before administering any of the compositions disclosed herein, careful consideration must be given to the anatomical morphology of the treatment site. For example, in some embodiments, the therapeutic goal is to inject into the area where neuromuscular junctions are most concentrated, if known. For example, in the case of intramuscular administration, the position of the needle within the muscle can be confirmed by moving the muscle through its range of motion and observing the resulting movement of the needle tip before intramuscular injection. General anesthesia, local anesthesia, and sedation are used depending on the patient's age, the number of injection sites, and the patient's specific needs. More than one injection and / or injection sites may be required to achieve the desired results. Also, depending on the muscle to be injected, some injections may require the use of a thin, hollow, TEFLON®-coated needle guided by electromyography.

[0020] The disclosed composition may be administered by injection to, for example, the following skeletal muscles or the area surrounding one or more of them, such as the occipitofrontalis muscle, nasal muscle, orbicularis oris muscle, depressor anguli oris muscle, platysma muscle, sternohyoid muscle, serratus anterior muscle, rectus abdominis muscle, external oblique muscle, tensor fasciae latae muscle, brachioradialis muscle, iliopsoas muscle, psoas major muscle, pectineus muscle, adductor longus muscle, sartorius muscle, gracilis muscle, vastus lateralis muscle, rectus femoris muscle, vastus medialis muscle, quadriceps femoris tendon, patella, gastrocnemius muscle, soleus muscle, tibia, peroneus longus muscle, tibialis anterior muscle, patellar ligament, iliotibial band, and small This may include injections into the thenar muscles, thenar muscles, flexor carpi ulnaris, flexor digitorum superficialis, palmaris longus, flexor carpi radialis, brachioradialis, pronator teres, brachialis, biceps brachii, triceps brachii, pectoralis major, deltoid, trapezius, sternocleidomastoid, masseter, orbicularis oculi, temporalis, galea aponeurotica, teres major, extensor digitorum, extensor carpi ulnaris, anconeus, abductor pollicis longus, plantaris, calcaneal tendon, soleus, adductor magnus, gluteus maximus, gluteus medius, latissimus dorsi, infraspinatus, and combinations thereof, as well as similar substances.

[0021] Administration of the disclosed composition may be by injection into, for example, the following nerves or the vicinity of one or more of them: e.g., axillary nerve, phrenic nerve, spinal ganglia, spinal cord, sympathetic ganglion chain, pudendal nerve, common palmar digital nerve, ulnar nerve, deep branch of the ulnar nerve, sciatic nerve, peroneal nerve, tibial nerve, saphenous nerve, interosseous nerve, superficial peroneal nerve, intermediate dorsal cutaneous nerve, medial plantar nerve, medial dorsal cutaneous nerve, deep peroneal nerve, muscular branch of the tibial nerve, infrapatellar branch of the saphenous nerve, common peroneal nerve, muscular branch of the femoral nerve, anterior cutaneous branch of the femoral nerve, muscular branch of the sciatic nerve, femoral nerve, ilioinguinal nerve, filum terminale, hypogastric iliac crest This may include injections into nerves, obstructive nerves, ulnar nerves, radial nerves, obstructive nerves, radial nerves, subcostal nerves, intercostal nerves, dorsal branches of intercostal nerves, medial cutaneous branches of intercostal nerves, musculocutaneous nerves, deltoid nerves, vagus nerves, brachial plexus, supraclavicular nerves, facial nerves, auricular-temporal nerves, and combinations thereof, as well as similar substances. Smooth muscles suitable for administration of the disclosed composition may include blood vessel walls, gastric walls, ureters, intestines, within the aorta (medial layer), iris of the eye, prostate, gastrointestinal tract, airways, arterioles, reproductive systems (both male and female), veins, renal glomeruli (referred to as mesangial cells), bladder, uterus, arrector pili muscles, ciliary muscles, sphincters of the skin, trachea, bile ducts, and any of the same. The frequency and amount of injections under the disclosed method may be determined based on the nature and location of the specific area being treated. However, in certain cases, repeated or additional injections may be desired to achieve optimal results. The frequency and amount of injections for each specific case may be determined by those skilled in the art.

[0022] In one embodiment, the rapid-acting neurotoxin is administered prior to the surgical procedure. In one embodiment, the administration is performed, for example, within 36 hours, 24 hours, 22 hours, 20 hours, 18 hours, 16 hours, 14 hours, 12 hours, 11 hours, 10 hours, 9 hours, 8 hours, 7 hours, 6 hours, 5 hours, 4 hours, 3 hours, 2 hours, 60 minutes, 50 minutes, 40 minutes, 30 minutes, 20 minutes, 10 minutes, 5 minutes, 2 minutes, etc. In the embodiments, the rapid-acting neurotoxin is administered prior to the surgical procedure. In the embodiments, the administration is performed, for example, at least 36 hours before surgery, at least 24 hours before surgery, at least 22 hours before surgery, at least 20 hours before surgery, at least 18 hours before surgery, at least 16 hours before surgery, at least 14 hours before surgery, at least 12 hours before surgery, at least 11 hours before surgery, at least 10 hours before surgery, at least 9 hours before surgery, at least 8 hours before surgery, at least 7 hours before surgery, at least 6 hours before surgery, at least 5 hours before surgery, at least 3 hours before surgery, at least 2 hours before surgery, at least 60 minutes before surgery, at least 50 minutes before surgery, at least 40 minutes before surgery, at least 30 minutes before surgery, at least 20 minutes before surgery, at least 10 minutes before surgery, at least 5 minutes before surgery, at least 2 minutes before surgery, etc. In one embodiment, the administration of a rapid-acting neurotoxin is performed simultaneously with the surgical procedure. In a given embodiment, the rapid-acting neurotoxin is administered after surgery. For example, administration may be performed within 1 minute, 2 minutes, 3 minutes, 4 minutes, 5 minutes, 6 minutes, 7 minutes, 8 minutes, 9 minutes, 10 minutes, 20 minutes, 30 minutes, 40 minutes, 50 minutes, 60 minutes, 90 minutes, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, etc.

[0023] The methods disclosed herein may include additional doses following an initial dose of a rapid-acting neurotoxin to a patient. Embodiments including additional doses may further include physician- or patient-to-physician evaluation of the results of the prior neurotoxin administration. Such evaluation may include, for example, the use of photographs, scans, or similar methods.

[0024] In the embodiment, the evaluation of the results of the initial neurotoxin administration is performed, for example, at 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 54 hours, 60 hours, 66 hours, 72 hours, and 78 hours after the initial administration. This may be done within 84 hours, 90 hours, 96 hours, 102 hours, 108 hours, 114 hours, 120 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, etc., after the first dose.

[0025] In embodiments including an additional dose, the additional dose may be administered within, for example, 6 hours, 8 hours, 10 hours, 12 hours, 14 hours, 16 hours, 18 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 54 hours, 60 hours, 66 hours, 72 hours, 78 hours, 84 hours, 90 hours, 96 hours, 102 hours, 108 hours, 114 hours, 120 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, etc.

[0026] In the embodiment, additional doses are, for example, 6 hours after the initial dose, 8 hours after the initial dose, 10 hours after the initial dose, 12 hours after the initial dose, 14 hours after the initial dose, 16 hours after the initial dose, 18 hours after the initial dose, 24 hours after the initial dose, 30 hours after the initial dose, 36 hours after the initial dose, 42 hours after the initial dose, 48 hours after the initial dose, 54 hours after the initial dose, 60 hours after the initial dose, 66 hours after the initial dose, 72 hours after the initial dose, 78 hours after the initial dose, and the initial dose It may be performed within 84 hours, 90 hours after the first dose, 96 hours after the first dose, 102 hours after the first dose, 108 hours after the first dose, 114 hours after the first dose, 120 hours after the first dose, 1 week after the first dose, 2 weeks after the first dose, 3 weeks after the first dose, 4 weeks after the first dose, 5 weeks after the first dose, 6 weeks after the first dose, 7 weeks after the first dose, 8 weeks after the first dose, 9 weeks after the first dose, 10 weeks after the first dose, 11 weeks after the first dose, 12 weeks after the first dose, etc.

[0027] The methods disclosed herein can provide a rapidly initiating effect (for example, using a fast-acting neurotoxin). For example, the disclosed embodiments can provide a visually noticeable cosmetic effect at, for example, 30 minutes, 45 minutes, 60 minutes, 75 minutes, 90 minutes, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, and 16 hours after administration. It can be provided within the following timeframes: 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours, 24 hours, 30 hours, 36 hours, 42 hours, 48 ​​hours, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, etc. The methods disclosed herein can provide shorter-term effects (for example, using a fast-acting neurotoxin). For example, the disclosed embodiments can provide a visually noticeable cosmetic effect that soothes within, for example, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 45, 60, 75, 90, 105 days after administration.

[0028] Side effects may be associated with botulinum injection. The disclosed embodiments may provide neurotoxin therapy that results in fewer or shorter-lasting side effects than conventional neurotoxin therapy. For example, the disclosed embodiments may result in fewer (or shorter-lasting) consequences of double vision or blurred vision, blepharoplasty (inability of the subject to fully open their eyelids), loss of facial muscle movement, hoarseness, bladder dysfunction, shortness of breath, dysphagia, difficulty speaking, death, and similar conditions.

[0029] This method is particularly suitable for treating cosmetic imperfections that are typically the result of aging, environmental exposure, weight loss, childbirth, trauma, surgery, or a combination thereof. Aging and environmental exposure often cause wrinkles in various locations on the skin. On the other hand, weight loss and childbirth often cause stretch marks in various locations on the skin, particularly on the abdomen, lower body, and legs. Trauma and surgery frequently result in scars in the traumatic and surgical areas. Specific contour defects suitable for treatment by the disclosed method include, but are not limited to, wrinkles caused by frowning, forehead lines, wrinkles, crow's feet, marionette lines, stretch marks, wounds, accidents, bites, surgery, or similar conditions. Contour defects in areas such as the eyes, cheeks, nose, lips, forehead, and neck are particularly suitable for treatment by the present invention. Furthermore, the disclosed embodiments can provide patients with more reliable and longer-lasting cosmetic results. For example, a 20% variation in the duration of effect due to a longer-acting neurotoxin can result in a one-month difference in the duration of effectiveness. With the disclosed early-recovering neurotoxin, this 20% variation does not result in such an extreme difference in the duration of effectiveness.

[0030] Additional administration of a rapid-acting neurotoxin can effectively modify or increase the prior administration of a cosmetic neurotoxin. For example, the methods disclosed herein may include additional administration to correct uneven cosmetic outcomes from the prior administration, to enhance the cosmetic outcomes of the prior administration, or to accelerate the onset of results compared to results achieved with a non-rapid-acting neurotoxin. The disclosed rapid-acting neurotoxin compositions may be administered using a needle or a needle-free device. In certain embodiments, the method involves injecting the composition subcutaneously into an individual. For example, administration involves injecting the composition with a needle of about 30 gauge or less. In certain embodiments, the method involves administering a composition comprising botulinum toxin type E.

[0031] The composition may be injected by syringe, catheter, needle, and other means of injection. The injection may be administered to any area of ​​the mammalian body in need of treatment, including but not limited to the face, neck, torso, arms, hands, legs, and feet. The injection may be administered to any location in a particular area, such as the epidermis, dermis, fat, muscle, or subcutaneous layer. The frequency and amount of injections under the disclosed method may be determined based on the nature and location of the specific cosmetic imperfection being treated. However, in certain cases, repeated injections may be desired to achieve optimal results. The frequency and amount of injections for each specific case may be determined by those skilled in the art. While examples of administration routes and dosages are provided, the appropriate administration route and dosage are generally determined on a case-by-case basis by the attending physician. Such decisions are commonplace for those skilled in the art. For example, the administration routes and dosages of Clostridium neurotoxins disclosed in the present invention can be selected based on criteria such as the solubility properties of the selected neurotoxin and the intensity and extent of the cosmetic condition being treated.

[0032] Fast-acting neurotoxins are approximately 10 -3 It may be administered in an amount ranging from U / kg body weight to approximately 35 U / kg body weight. In one embodiment, the neurotoxin is approximately 10 -2 It is administered in amounts ranging from U / kg to approximately 25 U / kg. In another embodiment, the neurotoxin is approximately 10 -1 The neurotoxin is administered in amounts ranging from U / kg to approximately 15 U / kg. In another embodiment, the neurotoxin is administered in amounts ranging from approximately 1 U / kg to approximately 10 U / kg. In many cases, administration of a neurotoxin such as botulinum toxin type E in amounts ranging from approximately 1 unit to approximately 500 units provides effective therapeutic relief. In one embodiment, a neurotoxin such as botulinum toxin type E can be used in amounts ranging from approximately 5 units to approximately 200 units, and in another embodiment, a neurotoxin such as botulinum toxin type E can be administered topically to target tissues such as muscle in amounts ranging from approximately 10 units to approximately 100 units.

[0033] In an embodiment, the administration is approximately 4 units of neurotoxin, or approximately 5 units of neurotoxin, or approximately 6 units of neurotoxin, or approximately 7 units of neurotoxin, or approximately 8 units of neurotoxin, or approximately 10 units of neurotoxin, or approximately 15 units of neurotoxin, or approximately 20 units of neurotoxin, or approximately 30 units of neurotoxin, or approximately 40 units of neurotoxin, or approximately 50 units of neurotoxin, or approximately 60 units of neurotoxin, or approximately 70 units of neurotoxin, or approximately 80 units of neurotoxin, or approximately 90 units of neurotoxin. , or about 100 units of neurotoxin, or about 110 units of neurotoxin, or about 120 units of neurotoxin, or about 130 units of neurotoxin, or about 140 units of neurotoxin, or about 150 units of neurotoxin, or about 160 units of neurotoxin, or about 170 units of neurotoxin, or about 180 units of neurotoxin, or about 190 units of neurotoxin, or about 200 units of neurotoxin, or about 210 units of neurotoxin, or about 220 units of neurotoxin, or about 230 units of neurotoxin, or Approximately 240 units of neurotoxin, or approximately 250 units of neurotoxin, or approximately 260 units of neurotoxin, or approximately 270 units of neurotoxin, or approximately 280 units of neurotoxin, or approximately 290 units of neurotoxin, or approximately 290 units of neurotoxin, or approximately 300 units of neurotoxin, or approximately 310 units of neurotoxin, or approximately 320 units of neurotoxin, or approximately 330 units of neurotoxin, or approximately 340 units of neurotoxin, or approximately 350 units of neurotoxin, or approximately 360 units of neurotoxin, or approximately 370 The dosage may include units of neurotoxin, or approximately 380 units of neurotoxin, or approximately 390 units of neurotoxin, or approximately 400 units of neurotoxin, or approximately 410 units of neurotoxin, or approximately 420 units of neurotoxin, or approximately 430 units of neurotoxin, or approximately 440 units of neurotoxin, or approximately 450 units of neurotoxin, or approximately 460 units of neurotoxin, or approximately 470 units of neurotoxin, or approximately 480 units of neurotoxin, or approximately 490 units of neurotoxin, or approximately 500 units of neurotoxin, and so on.

[0034] In embodiments, the administration is approximately 4 units of botulinum toxin type E, or approximately 5 units of botulinum toxin type E, or approximately 6 units of botulinum toxin type E, or approximately 7 units of botulinum toxin type E, or approximately 8 units of botulinum toxin type E, or approximately 10 units of botulinum toxin type E, or approximately 15 units of botulinum toxin type E, or approximately 20 units of botulinum toxin type E, or approximately 30 units of botulinum toxin type E, or approximately 40 units of botulinum toxin type E, or approximately 50 units of botulinum toxin type E, or approximately 60 units of botulinum toxin Botulinum toxin type E neurotoxin, or approximately 70 units of botulinum toxin type E neurotoxin, or approximately 80 units of botulinum toxin type E neurotoxin, or approximately 90 units of botulinum toxin type E neurotoxin, or approximately 100 units of botulinum toxin type E neurotoxin, or approximately 110 units of botulinum toxin type E neurotoxin, or approximately 120 units of botulinum toxin type E neurotoxin, or approximately 130 units of botulinum toxin type E neurotoxin, or approximately 140 units of botulinum toxin type E neurotoxin, or approximately 150 units of botulinum toxin type E neurotoxin, or approximately 160 units of botulinum toxin type E neurotoxin, or approximately 170 units of botulinum toxin type E neurotoxin, Or approximately 180 units of botulinum toxin type E, or approximately 190 units of botulinum toxin type E, or approximately 200 units of botulinum toxin type E, or approximately 210 units of botulinum toxin type E, or approximately 220 units of botulinum toxin type E, or approximately 230 units of botulinum toxin type E, or approximately 240 units of botulinum toxin type E, or approximately 250 units of botulinum toxin type E, or approximately 260 units of botulinum toxin type E, or approximately 270 units of botulinum toxin type E, or approximately 280 units of botulinum toxin type E, or approximately 290 units 1 unit of botulinum toxin type E, or approximately 290 units of botulinum toxin type E, or approximately 300 units of botulinum toxin type E, or approximately 310 units of botulinum toxin type E, or approximately 320 units of botulinum toxin type E, or approximately 330 units of botulinum toxin type E, or approximately 340 units of botulinum toxin type E, or approximately 350 units of neurotoxin, or approximately 360 units of botulinum toxin type E, or approximately 370 units of botulinum toxin type E, or approximately 380 units of botulinum toxin type E, or approximately 390 units of botulinum toxin type E,Alternatively, it may contain doses such as approximately 400 units of botulinum toxin type E, or approximately 410 units of botulinum toxin type E, or approximately 420 units of botulinum toxin type E, or approximately 430 units of botulinum toxin type E, or approximately 440 units of botulinum toxin type E, or approximately 450 units of botulinum toxin type E, or approximately 460 units of botulinum toxin type E, or approximately 470 units of botulinum toxin type E, or approximately 480 units of botulinum toxin type E, or approximately 490 units of botulinum toxin type E, or approximately 500 units of botulinum toxin type E.

[0035] Disclosed herein are methods for representing and communicating neurotoxin doses. In embodiments, the dose is expressed in terms of protein quantity, for example, nanograms (ng). In embodiments, the neurotoxin may include botulinum toxin. The methods disclosed herein may include administering a neurotoxin, for example, a rapid-acting neurotoxin, to a patient, wherein the dose of the neurotoxin is expressed as the amount of protein, for example, the amount of protein per dose. In one embodiment, the rapid-acting neurotoxin is botulinum toxin, for example, botulinum toxin type E.

[0036] In embodiments, the dose of neurotoxin is expressed as the amount or concentration of protein. For example, in embodiments, the neurotoxin may be administered in amounts of about 0.2 ng to 20 ng. In one embodiment, the neurotoxin is administered to a target tissue such as muscle in amounts such as about 0.3 ng to 19 ng, about 0.4 ng to 18 ng, about 0.5 ng to 17 ng, about 0.6 ng to 16 ng, about 0.7 ng to 15 ng, about 0.8 ng to 14 ng, about 0.9 ng to 13 ng, about 0.0 ng to 12 ng, about 0.5 ng to 11 ng, about 0.2 ng to 10 ng, about 0.5 ng to 7 ng, etc.

[0037] In embodiments, the administration may include total doses such as 5 to 7 ng, 7 to 9 ng, 9 to 11 ng, 11 to 13 ng, 13 to 15 ng, 15 to 17 ng, and 17 to 19 ng. In one embodiment, the administration may include total doses of 5 ng or less, 6 ng or less, 7 ng or less, 8 ng or less, 9 ng or less, 10 ng or less, 11 ng or less, 12 ng or less, 13 ng or less, 14 ng or less, 15 ng or less, 16 ng or less, 17 ng or less, 18 ng or less, 19 ng or less, 20 ng or less, etc. In one embodiment, the administration may include total doses of 5 ng or more, 6 ng or more, 7 ng or more, 8 ng or more, 9 ng or more, 10 ng or more, 11 ng or more, 12 ng or more, 13 ng or more, 14 ng or more, 15 ng or more, 16 ng or more, 17 ng or more, 18 ng or more, 19 ng or more, 20 ng or more, etc.

[0038] In embodiments, the administration is approximately 0.1 ng of neurotoxin, 0.2 ng of neurotoxin, 0.3 ng of neurotoxin, 0.4 ng of neurotoxin, 0.5 ng of neurotoxin, 0.6 ng of neurotoxin, 0.7 ng of neurotoxin, 0.8 ng of neurotoxin, 0.9 ng of neurotoxin, 1.0 ng of neurotoxin, 1.1 ng of neurotoxin, 1.2 ng of neurotoxin, 1.3 ng of neurotoxin, 1.4 ng of neurotoxin, 1.5 ng of neurotoxin, 1.6 ng of neurotoxin, 1.7 ng of neurotoxin, 1.8 ng of neurotoxin, 1.9 ng of neurotoxin, 2.0 ng of neurotoxin, 2.1 ng of neurotoxin, 2.2 ng of neurotoxin, 2.3 ng of neurotoxin, 2.4 ng of neurotoxin, 2.5 ng of neurotoxin, 2.6 ng of neurotoxin, 2.7 ng of neurotoxin, 2.8 ng of neurotoxin, 2.9 ng of neurotoxin, and 3.0 ng The total dose may include 1.5 ng of neurotoxin, 3.1 ng of neurotoxin, 3.2 ng of neurotoxin, 3.3 ng of neurotoxin, 3.4 ng of neurotoxin, 3.5 ng of neurotoxin, 3.6 ng of neurotoxin, 3.7 ng of neurotoxin, 3.8 ng of neurotoxin, 3.9 ng of neurotoxin, 4.0 ng of neurotoxin, 4.1 ng of neurotoxin, 4.2 ng of neurotoxin, 4.3 ng of neurotoxin, 4.4 ng of neurotoxin, 4.5 ng of neurotoxin, 5 ng of neurotoxin, 6 ng of neurotoxin, 7 ng of neurotoxin, 8 ng of neurotoxin, 9 ng of neurotoxin, 10 ng of neurotoxin, 11 ng of neurotoxin, 12 ng of neurotoxin, 13 ng of neurotoxin, 14 ng of neurotoxin, 15 ng of neurotoxin, 16 ng of neurotoxin, 17 ng of neurotoxin, 18 ng of neurotoxin, 19 ng of neurotoxin, 20 ng of neurotoxin, and so on.

[0039] In embodiments, the administration is, for example, per injection, about 0.1 ng of botulinum toxin type E, 0.2 ng of botulinum toxin type E, 0.3 ng of botulinum toxin type E, 0.4 ng of botulinum toxin type E, 0.5 ng of botulinum toxin type E, 0.6 ng of botulinum toxin type E, 0.7 ng of botulinum toxin type E, 0.8 ng of botulinum toxin type E, 0.9 ng of botulinum toxin type E, 1.0 ng of botulinum toxin type E, 1.1 ng of botulinum toxin type E, and 1.2 ng of botulinum toxin. Botulinum toxin type E, 1.3 ng botulinum toxin type E, 1.4 ng botulinum toxin type E, 1.5 ng botulinum toxin type E, 1.6 ng botulinum toxin type E, 1.7 ng botulinum toxin type E, 1.8 ng botulinum toxin type E, 1.9 ng botulinum toxin type E, 2.0 ng botulinum toxin type E, 2.1 ng botulinum toxin type E, 2.2 ng botulinum toxin type E, 2.3 ng botulinum toxin type E, 2.4 ng neurotoxin, 2.5 ng neurotoxin, 2.6 ng g of botulinum toxin type E, 2.7 ng of botulinum toxin type E, 2.8 ng of botulinum toxin type E, 2.9 ng of botulinum toxin type E, 3.0 ng of botulinum toxin type E, 3.1 ng of botulinum toxin type E, 3.2 ng of botulinum toxin type E, 3.3 ng of botulinum toxin type E, 3.4 ng of botulinum toxin type E, 3.5 ng of botulinum toxin type E, 3.6 ng of botulinum toxin type E, 3.7 ng of botulinum toxin type E, 3.8 ng of botulinum toxin type E, 3. The dosage may include 9 ng of botulinum toxin type E, 4.0 ng of botulinum toxin type E, 4.1 ng of botulinum toxin type E, 4.2 ng of botulinum toxin type E, 4.3 ng of botulinum toxin type E, 4.4 ng of botulinum toxin type E, 4.5 ng of botulinum toxin type E, 5 ng of botulinum toxin type E, 6 ng of botulinum toxin type E, 7 ng of botulinum toxin type E, 8 ng of botulinum toxin type E, 9 ng of botulinum toxin type E, 10 ng of botulinum toxin type E, and so on. Ultimately, however, both the amount of toxin administered and the frequency of its administration will be left to the discretion of the treating physician, balancing safety with the effects of the toxin.

[0040] Controlled-release systems may be used in the embodiments described herein to deliver neurotoxins in vivo at a predetermined rate over a specific period of time. Generally, the release rate is determined by the design of the system and may be largely independent of environmental conditions such as pH. Controlled-release systems are known that can deliver drugs over periods of several years. In contrast, sustained-release systems typically deliver drugs within 24 hours, and environmental factors may affect the release rate. Therefore, the release rate of neurotoxins from an implanted controlled-release system ("implanted tablet") is a function of the physiochemical properties of the implanted carrier material and the drug itself. Typically, implanted tablets are made of inert materials that induce little to no host response. The release control system may contain a neurotoxin incorporated into a carrier. The carrier may be a polymer or a bioceramic material. The release control system may be injected, inserted, or implanted in a selected site on the patient's body and remain there for an extended period while the neurotoxin is released by the implanted tablet in a manner and concentration that provides the desired therapeutic effect. Polymeric substances can release neurotoxins by diffusion, chemical reaction, or solvent activation, as well as under the influence of magnetic fields, ultrasound, or temperature-changing factors. Diffusion may occur from a reservoir or matrix. Chemical control may result from the degradation of the polymer or cleavage of the drug from the polymer. Solvent activation may include swelling of the polymer or osmotic effects.

[0041] The implant tablets may be prepared by mixing a suitable polymer solution in which a desired amount of stabilized neurotoxin is dissolved in methylene chloride. The solution may be prepared at room temperature. The solution may then be transferred to a Petri dish and the methylene chloride evaporated in a vacuum desiccator. Depending on the desired dimensions of the implant tablets, and thus the amount of neurotoxin incorporated, an appropriate amount of the dried implant tablets containing the neurotoxin formulation is compressed in a mold at about 8000 p.s.i. for 5 seconds or at 3000 p.s.i. for 17 seconds to form an implant disk encapsulating the neurotoxin.

[0042] Preferably, the implant material used is substantially non-toxic, non-carcinogenic, and non-immunogenic. Suitable implant materials include poly(2-hydroxyethyl methacrylate) (p-HEMA), poly(N-vinylpyrrolidone) (p-NVP) + , poly(vinyl alcohol) (PVA), poly(acrylic acid) (PM), polydimethylsiloxane (PDMS), ethylene-vinyl acetate (EVAc) copolymer, polyvinylpyrrolidone / methyl acrylate copolymer, polymethyl methacrylate (PMMA), polylactic acid (PLA), polyglycolic acid (PGA), polyanhydrides, polyorthoesters, collagen and cellulose derivatives and bioceramics such as hydroxyapatite (HPA), tricalcium phosphate (TCP), and aluminum calcium phosphate (ALCAP), and other polymers. Lactic acid, glycolic acid, and collagen may be used to make biodegradable implant tablets. The implant material may be biodegradable or bioerodible. The advantage of bioerodible implant tablets is that they do not need to be removed from the patient. Bioerodible implant tablets may be based on either membrane or matrix release of the bioactive substance. Biodegradable microspheres prepared from PLA-PGA are known for subcutaneous or intramuscular administration.

[0043] Kits for carrying out the disclosed embodiments are also encompassed by this disclosure. The kit may include a needle of 30 gauge or less and a corresponding syringe. The kit may also include a Clostridium neurotoxin composition, such as a botulinum toxin type E composition. The neurotoxin composition may be provided in a syringe. The composition is injectable via the needle. The kit may be designed in various forms based on the dimensions of the syringe and needle and the volume of the injectable composition contained therein, and then based on the specific cosmetic defect for which the kit is designed to treat. [Examples]

[0044] The following non-limiting embodiments are provided solely for illustrative purposes to facilitate a more complete understanding of the representative embodiments. These embodiments should not be construed as limiting any of the embodiments described in the specification of the present invention.

[0045] (Example 1) Use of botulinum toxin type E to treat frown lines (GL) This first-in-human, randomized, double-blind, placebo-controlled, ascending-dose cohort study enrolled 42 subjects who received either EB-001 (botulinum toxin type E composition disclosed herein) (N=35) or placebo (N=7). The primary efficacy outcome was the proportion of subjects showing an IR-2 improvement as assessed by the investigator in GL severity at maximum grimacing. Safety evaluations included adverse events (AEs), laboratory studies, and physical examinations. IR-2 responses were observed starting in the third cohort (EB-001) and were more frequent at higher doses. Onset of clinical effect occurred within 24 hours and lasted for a duration ranging from 14 to 30 days with respect to the highest dose. The incidence of AEs was low, with mild to moderate headache being the most common. There were no serious AEs or ptosis, and no clinically significant changes were observed in other safety evaluations. In this clinical trial at GL, EB-001 demonstrated favorable safety and tolerability, as well as dose-dependent efficacy with an 80% response rate at the highest dose. The maximum clinical effect of EB-001 was observed within 24 hours and lasted for 14 to 30 days. This distinctive EB-001 profile, with its rapid onset and short duration of effect, supports development toward desirable aesthetic and therapeutic applications.

[0046] Botulinum neurotoxin inhibits the presynaptic release of acetylcholine and is among the most potent molecules in nature. When injected into muscles, botulinum neurotoxin inhibits neuromuscular transmission, producing dose-dependent local muscle relaxation. Purified botulinum neurotoxin, including serotypes A and B, has been developed as an injectable drug and is widely used to treat various neuromuscular conditions. Serotype E of botulinum neurotoxin is a novel serotype that has not been developed for clinical use until now. Of all botulinum neurotoxins, serotype E has the fastest onset of action and the shortest duration of action. Serotype E has a domain structure similar to serotype A, consisting of two protein chains linked by disulfide bonds: a 100kDa heavy chain and a 50kDa light chain. Serotype E inhibits neuromuscular transmission by cleaving the same presynaptic vesicle protein (synaptosome-associated protein 25) as serotype A, but at a different cleavage site than serotype A. Two junction sites on motor axons are mediated by botulinum neurotoxin to facilitate high-affinity recognition of nerve cells. Binding is first mediated by cell surface gangliosides and then by specific protein receptors. These receptors are found at the neuromuscular junction and at the motor axon terminals. Both botulinum toxin types A and E have been shown to bind to the specific receptor synaptic vesicle protein 2, and only these two serotypes share this receptor. This study was the first clinical trial to evaluate the safety and efficacy of ascending doses of botulinum toxin type E in a target population under the guidelines.

[0047] This study was a first-in-human evaluation of the safety and efficacy of EB-001, focusing on the treatment of moderate to severe GL. EB-001 is a proprietary purified form of botulinum toxin type E formulated as an injectable solution (Bonti, Inc., Newport Beach, California, USA). This study was a randomized, double-blind, placebo-controlled, ascending-dose cohort study conducted at two specialized clinical centers: Steve Yoelin, MD Medical Associates, Newport Beach, California, USA; and the Center for Dermatology Clinical Research, Fremont, California, USA. This study was accredited by the Aspire Institutional Review Board (Santee, California, USA) and conducted in accordance with the guidelines set forth in the Declaration of Helsinki. Written informed consent was received from all participants prior to participation in the study.

[0048] A total of 42 healthy, toxin-free men and women aged 18 to 60 years were enrolled in the study. Each participant was scheduled to participate for approximately 6 weeks. The primary inclusion criteria were: the presence of symmetrical glabella (GL) rated as moderate to severe when grimacing to the maximum extent; sufficient visual acuity without glasses to accurately assess one's own facial wrinkles (contact lens use was permitted); and the ability to comply with the requirements of the study. The primary exclusion criteria were: any uncontrolled systemic disease or other medical condition; any medical condition that may increase the risk of exposure to botulinum neurotoxin (including diagnosed myasthenia gravis, Eaton-Lambert syndrome, amyotrophic lateral sclerosis, or any other medical condition that impairs neuromuscular function); current or previous botulinum neurotoxin treatment; known immunization or hypersensitivity to botulinum neurotoxin; pre-defined dermatological procedures within 3 to 12 months prior to the study (such as non-invasive surface repairs, facial cosmetic procedures, or topical / oral retinoid treatments); and a history of surgery or treatment around the orbit. Women who were pregnant, breastfeeding, or planning to become pregnant were not enrolled. Men with a fertile female partner were enrolled only if they agreed to use double contraception for 3 months after administration.

[0049] During screening, the demographic characteristics, medical history, and prior and concomitant medications of the subjects were recorded, and alcohol / drug screening was performed. Standardized facial photographs were taken at baseline before treatment, at each follow-up visit, and at the end of the study, but the photographs were not used to evaluate efficacy.

[0050] Seven cohorts (six subjects per cohort) were enrolled and administered EB-001 or placebo in ascending doses in a 5:1 ratio. The best recommended starting dose (safety factor 10-fold) for this first-in-human trial was determined based on the no-observed-adverse-effect level (NOAEL) from preclinical safety and toxicity studies (data not yet published). From this, the baseline dose (cohort 1) was calculated, and the sub-efficacious dose was determined. Cohorts 2 through 7 received the baseline dose 3, 9, 12, 16, 21, and 28 times, respectively. This corresponded to the sub-efficacious to maximum effective dose of EB-001. The total dose was administered in a standardized manner to five injection sites with equal volumes (0.1 mL per site to the procerus, left and right medial corrugator supercilii muscles, and left and right lateral corrugator supercilii muscles) (see Figure 1). The injection into the lateral corrugator supercilii muscle was administered approximately 1 cm above the supraorbital ridge. EB-001 was supplied in a 5 mL vial as a sterile solution for injection. The placebo was supplied in an identical vial without EB-001. Each participant completed visits for screening (-30 to -1 day), baseline / injection (day 0), days 1, 2, 7, 14, day 30 (end of study), and day 42 (final safety follow-up).

[0051] Safety was assessed through adverse events (AEs), laboratory studies, electrocardiograms (ECGs), physical examinations, vital signs (pulse rate, respiratory rate, and blood pressure), urinary pregnancy tests (in women of childbearing potential), and intensive neurological examinations to assess the potential for botulinum neurotoxin diffusion. Treatment-induced adverse events (TEAEs) were defined as any AE that commenced or worsened in severity after exposure to the study treatment. AEs and TEAEs were aggregated by organ-specific major classifications and basic terminology using the Medical Dictionary for Regulatory Activities (MedDRA, version 19.0). Serious AEs (SAEs, or AEs that meet regulatory criteria for drug severity) and discontinuation due to AEs were also assessed. The severity of AEs was recorded as mild, moderate, severe, or life-threatening. Prior to enrollment in each dose cohort, a Safety Data Review Board (SAR) was convened to analyze all safety data from previous cohorts.

[0052] At screening, baseline, and on days 1, 2, 7, 14, and 30, participants' GL (Greater Lumpiness) was assessed using the Facial Wrinkle Scale (FWS) at maximum frown and at rest. Assessments were performed by both the investigator and the participants. The FWS is a widely accepted method used to assess the severity of facial wrinkles. In this study, the 4-point scale for GL severity was as follows: 0 = none, 1 = mild, 2 = moderate, 3 = severe. Participants were considered to have responded if they achieved at least a 2-grade improvement (reduction) (IR-2) based on the investigator's FWS assessment. The primary efficacy variable was the proportion of IR-2 responders at maximum frown at any post-baseline visit up to day 30. An additional efficacy endpoint for participants was the proportion of responders who achieved none or mild (analyzed per visit) in the investigator's assessed FWS grade on days 1, 2, 7, 14, or 30.

[0053] Two analysis populations were pre-specified: a safety population and an efficacy population. Subjects who received placebo were pooled for all analyses. The safety population included all subjects who received the study treatment and subsequently underwent at least one safety assessment. All TEAEs and SAEs were aggregated by treatment group. All safety parameters, including laboratory tests, ECG, physical examination, vital signs, urine pregnancy tests, and intensive neurological examinations, were reviewed by the investigators and assessed for clinical significance. The efficacy population was a modified intent-to-treat (mITT) population, defined as all randomized subjects who received at least one dose of the study treatment and had at least one post-baseline efficacy evaluation. Demographic and baseline characteristics analyses were performed for the mITT population. Medical history was based on the safety population, coded using MedDRA, and aggregated by organ system subcategory and basic terminology. Prior and concomitant medications were based on the safety population, coded using the WHO (World Health Organization) Anatomical Therapeutic Chemical Classification index, and aggregated by drug class and treatment group. Efficacy analyses were performed using the mITT population. FWS grades were aggregated by treatment and trial day using frequency counts and response rates (%). Analysis comparing the proportion of IR-2 responders in each EB-001 cohort against (pooled) placebo was performed using Fisher's exact test with a significance level of 0.05.

[0054] Of the 59 participants selected for the trial, 43 were enrolled in one of seven cohorts. One participant did not receive treatment, resulting in 42 participants being included in the mITT and safety populations (35 receiving EB-001 and 7 receiving placebo). One participant dropped out of follow-up, and 41 participants completed the trial. The demographic and baseline characteristics of the mITT population are shown in Table 1. The mean (range) age of the participants was 47.9 (22–60) years for the EB-001-treated (pooled) group and 50.4 (32–57) years for the (pooled) placebo group, respectively. The majority of participants were female (EB-001 = 91.4%; placebo = 85.7%) and white (71.4% in both groups). The baseline mean (standard deviation [SD]) of the maximum facial grimacing GL, as assessed by the investigator, was 2.6 (0.50) and 2.9 (0.38) in the EB-001 group and the placebo group, respectively. There was no substantial difference between the EB-001 group and the placebo group, indicating a good balance.

[0055] Figure 2 shows the proportion of subjects in the mITT population who achieved an IR-2 response to maximum facial grimacing GL severity at any post-baseline visit up to day 30, broken down by dose cohort. In Cohort 3, 40% of subjects were IR-2 responders. This response rate was comparable to or higher in all higher-dose cohorts, with 80% IR-2 responders in Cohorts 6 and 7. Cohorts 6 and 7 showed a significantly higher percentage of IR-2 responders compared to placebo (P=0.046). Figure 3 summarizes the proportion of subjects in each cohort whose FWS grade, as assessed by the investigator, was no GL or mild GL at any post-baseline visit up to day 30. Cohorts 2 through 7 (including the ends) had a higher percentage of responders compared to placebo, achieving rates of 60% to 100% in Cohorts 3 and above. In cohorts 3 through 7, no response or mild response was most frequently observed on days 1, 2, and / or 7. One response (20%) was observed on day 14 in cohorts 3, 5, 6, and 7, and on day 30 in cohorts 3 and 5. The safety results support the safety of all evaluated doses of EB-001 administered as IM injection in this population. No clinically significant changes from baseline were observed in any subject in neurological examination, ECG, physical examination, or laboratory tests.

[0056] Five subjects who received EB-001 treatment reported TEAEs, while none in the placebo group reported any. No SAEs were reported, and no TEAEs led to discontinuation of the study. All TEAEs were mild or moderate in severity. Pharyngeal soreness and flu-like symptoms were considered unrelated to treatment. Three subjects reported headache as a TEAE, one of which was considered treatment-related. There was no dose-related increase in headache incidence. No other TEAEs were observed that appeared to be related to ptosis or toxin diffusion.

[0057] To our knowledge, this is the first controlled clinical trial of a botulinum toxin type E product for any aesthetic or therapeutic use. This first-in-human trial of EB-001, a novel purified form of botulinum toxin type E administered via IM, served the objective of evaluating the safety, tolerability, and effective dose range of EB-001. A higher proportion of therapeutic responses and dose-response were observed in cohorts receiving higher doses of EB-001. IR-2 responses were observed starting in cohort 3 and increased in higher-dose cohorts, suggesting that the effective dose range of EB-001 may be the doses used in cohorts 4 through 7. Cohorts 6 and 7 had an 80% IR-2 response rate, similar to the response rates of approved botulinum toxin type A products. Subjects achieving none or mild FWS grade were observed starting in cohort 2. Regarding the onset of effect, therapeutic response was observed as early as 24 hours after administration, supporting previous reports suggesting that botulinum toxin type E has a faster onset than type A.

[0058] Regarding the duration of effect, defined as the proportion of subjects with no or mild response ratings, by day 14, one subject was observed in most of the five cohorts at the higher dose, and by day 30, one subject was observed in two of the five cohorts at the higher dose. All doses of EB-001 were well-tolerated without causing local injection site reactions. No SAEs or serious TEAEs were reported, and there were no discontinuations due to TEAEs. The most common TEAE, headache, was mild to moderate in severity, and there were no other treatment-related AEs. There were no cases of ptosis at any dose level, and no events that appeared to be related to toxin diffusion. Therefore, the clinical safety and tolerability profile in this study appears favorable. The efficacy and safety profile of EB-001 is promising and supports its potential as a unique treatment option for the treatment of GL and other aesthetic facial applications. Rapid initiation can satisfy the unmet needs of people seeking rapid treatment of facial wrinkles before unexpected social or occupational events. The limited duration of effect may be beneficial for people considering botulinum toxin therapy for the first time, and long-term commitments are undesirable. EB-001 treatment will allow people to evaluate aesthetic effects over a shorter duration compared to the 12-week duration of effect of botulinum toxin type A products. In this first clinical trial in GL subjects, EB-001 demonstrated favorable safety and tolerability in all cohorts. Five out of seven cohorts showed numerically higher response rates compared to placebo. This supports the efficacy of EB-001 in reducing the severity of GL. The two highest doses resulted in an 80% response rate, similar to that of approved botulinum toxin type A products. In contrast to the known time course of type A products, the clinical effect of EB-001 was observed within 24 hours (initiation) and lasted for 14–30 days (duration). This distinct clinical profile supports future advancements of EB-001 for aesthetic and critical therapeutic uses on the face, where rapid onset and short duration of effect are desirable.

[0059] [Table 1] (Example 2) Use of botulinum toxin type E to treat crow footprints A 57-year-old man with crow's feet caused by years of sun exposure seeks treatment from his doctor. The doctor recommends the composition disclosed herein and administers it subcutaneously to both sides of the patient's eyes. Approximately three units of botulinum toxin type E are administered to each injection site, with several injections on each side of the eye. The crow's feet disappear approximately two days after treatment and remain reduced for two months.

[0060] (Example 3) Use of botulinum toxin type E for eyebrow lifts A 60-year-old woman has eyebrows that extend below the brow bone. The physician recommends the composition disclosed herein and administers it subcutaneously over each eye. Approximately 10 units of botulinum toxin type E are administered to each injection site, with several injections on either side of the eye. The drooping of the eyebrows decreases within approximately two days and is substantially reduced for three months after administration.

[0061] (Example 4) Use of botulinum toxin type E for breast augmentation A 30-year-old woman chooses breast augmentation surgery. Four hours before the surgery, botulinum toxin type E is administered near the site where the surgical incision will be made. The administration reduces muscle tension in the incision area, resulting in minimal scarring. Two weeks after the surgery, an additional dose is administered.

[0062] (Example 5) Use of botulinum toxin type E for breast reconstruction A 30-year-old woman chooses breast reconstruction surgery. Fourteen hours before the surgery, botulinum toxin type E is administered near the site where the surgical incision will be made. The administration reduces muscle tension in the incision area, resulting in minimal scarring. Two weeks after the surgery, an additional dose is administered.

[0063] (Example 6) Use of botulinum toxin type E to treat episiotomy A 44-year-old woman undergoes an episiotomy. Immediately after the surgery, 4 ng of type E botulinum toxin is administered to the tissue surrounding the surgical site. Within 20 hours, muscle and nerve activity around the wound is significantly reduced.

[0064] (Example 7) Use of botulinum toxin type E to treat sports hernias A 28-year-old man suffers from a sports hernia. His physician administers 4 ng of type E botulinum toxin to the tissues surrounding both hernias. Within 10 hours, muscle and nerve activity around the affected areas is significantly reduced.

[0065] (Example 8) Use of botulinum toxin type E to treat spondylolysis A 48-year-old man suffers from spondylolysis. His physician administers 4 ng of type E botulinum toxin to the tissues surrounding both herniated discs. Within 16 hours, muscle and nerve activity around the affected areas is significantly reduced.

[0066] (Example 9) Use of botulinum toxin type E to treat anterior cruciate ligament rupture A 23-year-old woman suffered an anterior cruciate ligament (ACL) rupture. Six hours after the injury, her physician administered 7 ng of type E botulinum toxin to the muscles surrounding the ruptured ligament. Thirty hours later, muscle and nerve activity around the wound was significantly reduced. In conclusion, while aspects of this specification are emphasized by reference to specific embodiments, it should be understood that those skilled in the art will readily recognize that these disclosed embodiments serve only as illustrations of the principles of the subject matter disclosed herein. Therefore, it should be understood that the disclosed subject matter is not in any way limited to the specific methods, protocols, and / or reagents described herein. Accordingly, various modifications or changes, or alternative configurations, to the disclosed subject matter may be made in accordance with the teachings herein without deviating from the spirit of this specification. Finally, the terminology used herein is intended solely to describe specific embodiments and is not intended to limit the scope of this disclosure as defined solely by the claims. Therefore, the embodiments of this disclosure are not limited to the exact embodiments shown and described. Certain embodiments are described herein and include the best known modes for carrying out the methods and apparatus described herein. Naturally, variations of these described embodiments will be apparent to those skilled in the art by reading the foregoing description. Accordingly, this disclosure includes all modifications and equivalents of the subject matter enumerated in the claims appended herein, as permitted by applicable law. Furthermore, any combination of all possible variations of the embodiments described herein is encompassed by the disclosure unless otherwise specifically indicated herein or particularly obviously rejected by the context. Any group of alternative embodiments, elements, or steps of this disclosure should not be construed as limiting. Each group member may be referenced or claimed, either alone or in any combination with other group members disclosed herein. For convenience and / or patentability reasons, it is anticipated that one or more members of a group may be included in or removed from a group. If such inclusion or removal occurs, this specification is considered to include groups modified to satisfy the described descriptions of all Markush groups used in the appended claims.

[0067] Unless otherwise specified, all figures used herein and in the claims to represent features, items, quantities, parameters, characteristics, periods, etc., should be understood in all examples to be modified by the term “approximately.” When used herein, “approximately” means that the feature, item, quantity, parameter, characteristic, or period thus modified includes a range of plus or minus 10 percent above and below the stated value of the feature, item, quantity, parameter, characteristic, or period. Therefore, unless otherwise specified, numerical parameters expressed herein and in the appended claims are approximations that may vary. At a minimum, and not as an attempt to limit the application of the principle of equivalents to the scope of the claims, each numerical expression should be interpreted at least in light of the number of significant figures reported and by applying the usual methods of estimation. Although numerical ranges and values ​​that specify a broad scope of disclosure are approximations, numerical ranges and values ​​specified in specific examples are reported as accurately as possible. However, any numerical range and value inherently contains some degree of error that inevitably arises from the standard deviation observed in those individual test measurements. The numerical ranges of values ​​described herein are intended solely as a concise way to indicate that each distinct value falls within a range. Unless otherwise specified herein, the individual values ​​within a numerical range are incorporated herein as if they were individually described herein.

[0068] In the context describing this disclosure (particularly in the context of the following claims), the terms “a,” “an,” “the,” and similar reference subjects should be construed to include both singular and plural unless otherwise specifically indicated herein or explicitly refuted by the context. All methods described herein may be performed in any suitable order unless otherwise specifically indicated herein or explicitly refuted by the context. The use of any and all examples, or illustrative terms shown herein (e.g., “such as”), is intended merely to better illustrate the disclosure and does not present any limitation of scope as otherwise claimed. There are no terms herein that should be construed to indicate any non-claimed element essential to the implementation of the embodiments disclosed herein.

[0069] The specific embodiments of the disclosure herein may be further limited in claims using the terms "consisting of" or "essentially consisting of" the terms. When used in a claim, whether as an application or as an addition by amendment, the transitional term "consisting of" excludes any elements, steps, or components not expressed in the claim. The transitional term "consisting essentially of" limits the scope of the claim to those that do not substantially affect the expressed substance or step as well as the basic and novel features. Embodiments of the disclosure as asserted herein are essentially or expressly described and enabled herein.

Claims

1. A method for performing a cosmetic procedure, comprising administering a therapeutically effective amount of a fast-acting neurotoxin to a patient in an area where treatment is desired.

2. The method according to claim 1, wherein the rapid-acting neurotoxin comprises botulinum neurotoxin serotype E.

3. The method according to claim 2, wherein the therapeutically effective amount comprises an amount of about 0.1 to 2.8 nanograms (ng).

4. The method according to claim 2, wherein the therapeutically effective dose comprises an amount of about 0.1 to 10 ng.

5. The method according to claim 2, wherein the administration includes injection.

6. The method according to claim 2, further comprising administering a moderately active neurotoxin to a patient.

7. The method according to claim 2, further comprising administering a long-acting neurotoxin to a patient.

8. A method for correcting a previous cosmetic procedure, comprising administering a therapeutically effective amount of a fast-acting neurotoxin to a patient in the vicinity of the area previously treated with the cosmetic procedure.

9. The method according to claim 8, wherein the rapid-acting neurotoxin comprises botulinum neurotoxin serotype E.

10. The method according to claim 9, wherein the therapeutically effective dose comprises an amount of about 4 to 40 picograms / kg.

11. The method according to claim 10, wherein the therapeutically effective dose comprises an amount of about 8 to 20 picograms / kg.

12. The method according to claim 9, wherein the administration includes injection.

13. The method according to claim 9, further comprising administering a moderately active neurotoxin to a patient.

14. The method according to claim 9, further comprising administering a long-acting neurotoxin to a patient.

15. A method for administering a therapeutically effective dose of neurotoxin to a patient, wherein the dose of the neurotoxin is expressed in nanograms (ng).

16. The method according to claim 15, wherein the neurotoxin comprises botulinum neurotoxin serotype E.

17. The method according to claim 16, wherein the therapeutically effective dose comprises an amount of about 2 to 20 ng.

18. The method according to claim 17, wherein the therapeutically effective dose comprises an amount of about 8 to 20 ng.

19. The method according to claim 16, wherein the administration includes injection.

20. The method according to claim 16, further comprising administering a moderately active neurotoxin to a patient.

21. The method according to claim 16, further comprising administering a long-acting neurotoxin to a patient.

22. A method for modifying previous neurotoxin therapy, comprising administering 10 ng or less of a rapid-acting neurotoxin to a patient in proximity to an area previously treated with neurotoxin therapy.

23. The method according to claim 22, wherein the rapid-acting neurotoxin comprises botulinum neurotoxin serotype E.

24. The method according to claim 23, wherein the therapeutically effective dose comprises an amount of about 2 to 20 ng.

25. The method according to claim 24, wherein the therapeutically effective amount comprises an amount of about 8 to 20 ng.

26. The method according to claim 23, wherein the administration includes injection.

27. The method according to claim 23, further comprising administering a moderately active neurotoxin to a patient.

28. The method according to claim 23, further comprising administering a long-acting neurotoxin to a patient.

29. A method for minimizing scarring, comprising administering a therapeutically effective amount of a short-acting neurotoxin to a patient in the vicinity of a wound.

30. The method according to claim 29, wherein the short-acting neurotoxin comprises botulinum neurotoxin serotype E.

31. The aforementioned effective therapeutic dose is approximately 10 -3 The method according to claim 30, comprising an amount from U / kg to about 35 U / kg.

32. The method according to claim 31, wherein the therapeutically effective dose includes an amount of about 1 U / kg to about 25 U / kg.

33. The method according to claim 32, wherein the therapeutically effective dose includes an amount of about 5 U / kg to about 15 U / kg.

34. The method according to claim 30, wherein the therapeutically effective amount comprises an amount of about 0.2 nanograms to about 2 nanograms.

35. The method according to claim 34, wherein the therapeutically effective amount comprises an amount of about 0.5 nanograms to about 1 nanogram.

36. The method according to claim 30, wherein the administration includes injection.

37. The method according to claim 30, further comprising administering a moderately active neurotoxin to a patient.

38. The method according to claim 30, further comprising administering a long-acting neurotoxin to a patient.

39. The method according to claim 30, wherein the wound includes a surgical incision.

40. The method according to claim 39, wherein the surgical incision is the result of cosmetic surgery.

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