WO2019222850A1 - Synergistic tetrodotoxin formulations and methods of treatment for neuropathic pain - Google Patents

Synergistic tetrodotoxin formulations and methods of treatment for neuropathic pain Download PDF

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WO2019222850A1
WO2019222850A1 PCT/CA2019/050702 CA2019050702W WO2019222850A1 WO 2019222850 A1 WO2019222850 A1 WO 2019222850A1 CA 2019050702 W CA2019050702 W CA 2019050702W WO 2019222850 A1 WO2019222850 A1 WO 2019222850A1
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Prior art keywords
amount
ttx
duloxetine
reboxetine
tetrodotoxin
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PCT/CA2019/050702
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French (fr)
Inventor
Christopher C. Gallen
Walter N. KORZ
Donald C. K. WONG
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Wex Pharmaceuticals Inc.
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Publication of WO2019222850A1 publication Critical patent/WO2019222850A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/38—Heterocyclic compounds having sulfur as a ring hetero atom
    • A61K31/381—Heterocyclic compounds having sulfur as a ring hetero atom having five-membered rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/529—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim forming part of bridged ring systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/535—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one oxygen as the ring hetero atoms, e.g. 1,2-oxazines
    • A61K31/5375—1,4-Oxazines, e.g. morpholine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D491/00—Heterocyclic compounds containing in the condensed ring system both one or more rings having oxygen atoms as the only ring hetero atoms and one or more rings having nitrogen atoms as the only ring hetero atoms, not provided for by groups C07D451/00 - C07D459/00, C07D463/00, C07D477/00 or C07D489/00
    • C07D491/22—Heterocyclic compounds containing in the condensed ring system both one or more rings having oxygen atoms as the only ring hetero atoms and one or more rings having nitrogen atoms as the only ring hetero atoms, not provided for by groups C07D451/00 - C07D459/00, C07D463/00, C07D477/00 or C07D489/00 in which the condensed system contains four or more hetero rings

Definitions

  • the invention described herein relates generally to the use of a sodium channel blocker, for example tetrodotoxin or its analogs and/or derivatives as well as their pharmaceutically acceptable salts, as a medicament for the treatment of neuropathic pain in combination with other compounds. More particularly, but not exclusively, to use such a medicament in synergistic doses to ameliorate pain.
  • a sodium channel blocker for example tetrodotoxin or its analogs and/or derivatives as well as their pharmaceutically acceptable salts
  • Tetrodotoxin is a marine organic toxin which is mainly found in testis, ovaries, eggs, liver, spleen, gastrointestinal tract, skin, muscle, eyes, and blood of puffer fish. Tetrodotoxin is also found in several diverse animal species, including goby fish, some newts and frogs, the blue ringed octopus, and even in some marine alga. See, e.g., Bane et al , Tetrodotoxin: Chemistry, Toxicity, Source, Distribution and Detection, Toxins 6:693-755 (2014).
  • Tetrodotoxin is an alkaloid found in puffer fish ( Tetradontidae ).
  • the chemical name of TTX is [4R-(4a,4aa,5a,7a,9a, 10a, 10a ,1 1 S*, 12S*)]-Octahydro-12- (hydroxymethyl)-2-imino-5, 9:7, 10a-dimethano- 10aH-[1 ,3]dioxocino[6,5-d]pyrimidine- 4,7, 10, 1 1 , 12-pentol with a molecular formula CiiHi 7 N 3 0 8 and a molecular weight of 319.27 Da. It is a potent non-protein, small molecule neurotoxin and an indispensable tool for the study of neurobiology and physiology.
  • the present invention is directed to compositions and methods useful to treat pain in a subject mammal.
  • An aspect of the present invention are synergistic combinations of a sodium channel blocker, for example tetrodotoxin, and modulators of synaptic signaling, for example, antidepressants, selective serotonin reuptake inhibitors, mixed serotonin and norepinephrine reuptake inhibitors, or norepinephrine reuptake inhibitors.
  • Other aspects of the present invention are methods of using such synergistic combinations to treat pain.
  • a composition for the treatment of pain includes: (a) an amount of tetrodotoxin; and (b) an amount of either a serotonin-norepinephrine reuptake inhibitor (SNRI), selective serotonin reuptake inhibitor (SSRI), or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat pain.
  • SNRI serotonin-norepinephrine reuptake inhibitor
  • SSRI selective serotonin reuptake inhibitor
  • NRI norepinephrine reuptake inhibitor
  • composition for the treatment of neuropathic pain includes: (a) an amount of tetrodotoxin; and (b) an amount of any one of duloxetine and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat neuropathic pain.
  • the present invention comprises a method of treating neuropathic pain in a mammal in need thereof, includes: (a) administering an amount of tetrodotoxin; and (b) administering an amount of either a serotonin-norepinephrine reuptake inhibitor (SNRI), a selective serotonin reuptake inhibitor (SSRI) or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and reboxetine exhibit synergism to treat neuropathic pain.
  • SNRI serotonin-norepinephrine reuptake inhibitor
  • SSRI selective serotonin reuptake inhibitor
  • NRI norepinephrine reuptake inhibitor
  • the present invention comprises a method of treating neuropathic pain in a mammal in need thereof, includes: (a) administering an amount of tetrodotoxin; and (b) administering an amount of either a serotonin-norepinephrine reuptake inhibitor (SNRI), a selective serotonin reuptake inhibitor (SSRI) or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and duloxetine exhibit synergism to treat neuropathic pain.
  • SNRI serotonin-norepinephrine reuptake inhibitor
  • SSRI selective serotonin reuptake inhibitor
  • NRI norepinephrine reuptake inhibitor
  • composition for the treatment of nociceptive pain includes:
  • component (a) an amount of tetrodotoxin; and (b) an amount of any one of duloxetine and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat nociceptive pain.
  • the present invention comprises a method of treating nociceptive pain in a mammal in need thereof, includes: (a) administering an amount of tetrodotoxin; and (b) administering an amount of either a serotonin-norepinephrine reuptake inhibitor (SNRI), a selective serotonin reuptake inhibitor (SSRI) or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and reboxetine exhibit synergism to treat nociceptive pain.
  • SNRI serotonin-norepinephrine reuptake inhibitor
  • SSRI selective serotonin reuptake inhibitor
  • NRI norepinephrine reuptake inhibitor
  • the present invention comprises a method of treating nociceptive pain in a mammal in need thereof, includes: (a) administering an amount of tetrodotoxin; and (b) administering an amount of either a serotonin-norepinephrine reuptake inhibitor (SNRI), a selective serotonin reuptake inhibitor (SSRI) or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and duloxetine exhibit synergism to treat nociceptive pain.
  • SNRI serotonin-norepinephrine reuptake inhibitor
  • SSRI selective serotonin reuptake inhibitor
  • NRI norepinephrine reuptake inhibitor
  • Figure 1 shows the effects of TTX at 1 , 2 and 3 pg/kg on baseline Paw Withdrawal Test (PWT) (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy.
  • PWT Paw Withdrawal Test
  • Figure 2 shows the effects of duloxetine at 5, 10 and 15 mg/kg on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin- induced neuropathy.
  • Figure 3 shows the effects of Reboxetine at 1 , 3 and 5 mg/kg on baseline PWT (A) and baseline PWT, PWT at 1 and 2 hours after first dosing (B) in rats with oxaliplatin- induced neuropathy. The dosing schedule lasted for 5 days.
  • Figure 4 shows the effects of the combination of TTX and Duloxetine on baseline PWT (A) and baseline PWT, PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin- induced neuropathy.
  • Figure 5 shows the effects of the combination of TTX and Reboxetine on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy.
  • Figure 6 shows the effect of 3 pg/kg TTX and 2 mg/kg Reboxetine, administered alone and in combination for 5 days, on baseline PWT (A), and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy.
  • Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0, D4-0, D5-0, D7, D10, D14: baseline value for the 2nd, 3rd, 4th, 5th, 7th, 10th, and 14th days after the start of the course of dosing; 1 h, 2 h: 1 hour and 2 hours after each dosing. N 2 for each group.
  • Figure 7 shows the effect of 3 pg/kg TTX and 10 mg/kg duloxetine, administered alone and in combination, for 14 days, on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy.
  • Pre1 , Pre2, Pre3 1 st, 2nd and 3rd days before first oxaliplatin injection
  • W1 , W2 1 st and 2nd week after first oxaliplatin injection, respectively
  • D1-0 pre-dosing control
  • D2-0, D3-0. ... D14-0, ... , D27-0 Baseline value for the 2nd, 3rd, ... 14th, ... 27th days after the start of the course of dosing;
  • Figure 8 shows the effect of 3 pg/kg TTX and 2 mg/kg reboxetine, administered alone and in combination, for 14 days, on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy.
  • D1-0 pre-dosing control; D2-0, D3-0. ... D14-0, ... , D27-0: Baseline value for the 2nd, 3rd, ... 14th, ... 27th days after the start of the course of dosing;
  • Figure 9 shows the effect of 5 pg/kg TTX, administered alone and in combination with 5 mg/kg Reboxetine or 15 mg/kg Duloxetine for 5 days, on baseline PWT (A) and baseline PWT, PWT at 1 and 2 hours after first dosing (B) in rats with oxaliplatin-induced neuropathy. The dosing schedule lasted for 5 days.
  • Figure 10 shows the effect of 5 pg/kg TTX and 15 mg/kg duloxetine, administered alone and in combination, for 14 days, on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy.
  • Pre1 , Pre2, Pre3 1 st, 2nd and 3rd days before first oxaliplatin injection
  • W1 , W2 1 st and 2nd week after first oxaliplatin injection, respectively
  • D1-0 pre-dosing control
  • D27-0 shows the effect of 5 pg/kg TTX and 15 mg/kg duloxetine, administered alone and in combination, for 14 days, on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy.
  • Pre1 , Pre2, Pre3 1
  • Figure 1 1 shows the effect of 5 pg/kg TTX and 5 mg/kg reboxetine, administered alone and in combination, for 14 days, on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy.
  • D1-0 pre-dosing control;
  • D2-0, D3-0. ... D14-0, .... D27-0 Baseline value for the 2nd, 3rd, ... 14th, ... 27th days after the start of the course of dosing;
  • Figure 12 depicts the dose-effect relationships, as estimated using a hyperbolic model, for each of the three drugs administered alone.
  • Figure 12A shows the dose-effect relationship for TTX alone.
  • Figure 12B shows the dose-effect relationship for reboxetine alone.
  • Figure 12C shows the dose-effect relationship for duloxetine alone.
  • the hyperbolic curved lines in each panel represent the dose response curves.
  • the dots represent the mean % PE (over the post-baseline period) for each rat.
  • Figure 13 shows the performance of the reboxetine/TTX drug combination, compared with the TTX alone dose effect curve.
  • the observed effect is much higher above the curve, signifying strong synergism.
  • the interaction index of 0.469 indicates the combination took less than half of the dose (if using TTX alone) to achieve the same response. That is, using TTX alone, a dose of 25.1 pg/kg would be required to achieve the same MPE of 66.5%.
  • Figure 14 shows the performance of duloxetine/TTX drug combination, compared with the TTX alone dose effect curve.
  • Figure 15 show response surface plots produced with the methods of Tallarida.
  • Figure 16 shows the Tallarida response surface plots depicting the expected responses under additive assumption and the actual (observed) responses. This figure shows the performance of the TTX/duloxetine combination.
  • Figure 17 shows the effects of 3 pg/kg TTX and Duloxetine administered alone and in combination for 14 days on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy.
  • Pre1 , Pre2, Pre3 1 st, 2nd and 3rd days before first oxaliplatin injection
  • W1 , W2 1 st and 2nd week after first oxaliplatin injection, respectively
  • D1-0 pre-dosing control
  • D27 baseline value for the 2nd, 3rd... 14th, and 27th days after the start of dosing course
  • 1 h, 2 h 1 hour and 2 hours after each dosing.
  • N 7 for each group.
  • Figure 18 shows the effects of 3 pg/kg or 5 pg/kg TTX and varying amounts of Reboxetine administered alone and in combination for 14 days on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy.
  • Figure 19 depicts the dose response curves, as estimated using a hyperbolic model, at Day 5 Hour 1 , for each of the three drugs administered alone.
  • Figure 19A shows the dose-effect relationship for TTX alone.
  • Figure 19B shows the dose-effect relationship for reboxetine alone.
  • Figure 19C shows the dose-effect relationship for duloxetine alone.
  • the hyperbolic curved lines in each panel represent the dose response curves.
  • the dots represent the mean %MPE (over the post-baseline period) for each rat.
  • Figure 20 shows response surface plots depicting the expected responses under additive assumption and the actual (observed) responses from the drug combinations, produced with the methods of Tallarida.
  • Figure 20A shows the expected responses under additive assumption and the actual (observed) responses from the drug combinations of TTX administered in combination with Reboxetine.
  • Figure 20B shows the expected responses under additive assumption and the actual (observed) responses from the drug combinations of TTX administered in combination with Duloxetine.
  • a point above the surface represent the existence of synergism in the drug combination. The strong performance of TTX 5 pg/kg + Duloxetine 15 mg/kg is clearly noticeable.
  • Figure 21 depicts the dose response curves, as estimated using a hyperbolic model, at Day 14 Hour 1 , for each of the three drugs administered alone.
  • Figure 21 A shows the dose-effect relationship for TTX alone.
  • Figure 21 B shows the dose-effect relationship for reboxetine alone.
  • Figure 21 C shows the dose-effect relationship for duloxetine alone.
  • the hyperbolic curved lines in each panel represent the dose response curves.
  • the dots represent the mean %MPE (over the post-baseline period) for each rat.
  • Figure 22 shows the Tallarida response surface plots depicting the expected responses under additive assumption and the actual (observed) responses from the drug combinations.
  • Figure 22A shows the expected responses under additive assumption and the actual (observed) responses from the drug combinations of TTX administered in combination with Reboxetine.
  • Figure 22B shows the expected responses under additive assumption and the actual (observed) responses from the drug combinations of TTX administered in combination with Duloxetine.
  • the strong performance of TTX 5 pg/kg + Reboxetine 5 mg/kg combination, TTX 3 pg/kg + Duloxetine 10 mg/kg, and TTX 5 pg/kg + Duloxetine 15 mg/kg are clearly seen.
  • Drug synergy describes the total effect of two drugs in combination. When the combined effect of a drug combination is greater than predicted (additive effect) by their individual potencies, it is called super-additive; when the combination is said to be synergistic (enhanced effect more than mere additivity).
  • Synergy or synergism is a well- known pharmacological phenomenon with specific quantitative techniques to identify, explore, and characterize relationships between two different drugs.
  • R. J. Tallarida discusses various methods to identify and characterize synergy between two drugs in“Drug Synergism: Its Detection and Applications,” Perspectives in Pharmacology, 298: 865-872 (2001).
  • a composition for the treatment of pain comprising: (a) an amount of tetrodotoxin; and (b) an amount of either a selective serotonin reuptake inhibitor (SSRI), a serotonin-norepinephrine reuptake inhibitor (SNRI), or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat pain.
  • SSRI selective serotonin reuptake inhibitor
  • SNRI serotonin-norepinephrine reuptake inhibitor
  • NRI norepinephrine reuptake inhibitor
  • a composition for the treatment of neuropathic pain comprises: (a) an amount of tetrodotoxin; and (b) an amount of any one of paroxetine, fluoxetine, duloxetine, and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat neuropathic pain.
  • a composition for the treatment of nociceptive pain comprises: (a) an amount of tetrodotoxin; and (b) an amount of any one of paroxetine, fluoxetine, duloxetine, and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat nociceptive pain.
  • TTX is a well-known compound described for example in WO02/22129 as systemically acting as analgesic.
  • publications describing TTX are, for example, Tu, Anthony (Ed.) Handbook of Natural Toxins, Vol. 3: Marine Toxins and Venoms, 1988, 185-210, Kao (1966), Pharmacol. Rev. 18:997-1049, and Bane et al., Tetrodotoxin:
  • TTX is extracted from marine organisms (e.g. JP 270719 Goto and Takahashi), but numerous other methods of preparation or synthesis are also described (and used for the preparation of TTX in connection to this invention) in U.S. Pat. No. 6,552, 191 , U.S. Pat. No. 6,478,966, U.S. Pat. No. 6,562,968 or 2002/0086997, all of which are included here by reference.
  • Each of the preceding patents are incorporated by reference in their entirety and in particular for the methods of isolation, synthesis, and purification of TTX and TTX analogs.
  • tetrodotoxin a sodium channel blocker
  • administration of tetrodotoxin, a sodium channel blocker is highly effective for the treatment for pain, including neuropathic pain/allodynia/hyperalgesia.
  • the present invention relates to the use of sodium channel blockers.
  • the sodium channel blocker is tetrodotoxin, its analogs/derivatives, optionally in the form of its racemate, pure stereoisomers, especially enantiomers or diastereomers or in the form of mixtures of stereoisomers, in any suitable mixing ratio; in neutral form, in the form of an acid or base or in the form of a salt, in particular a physiologically acceptable salt, or in the form of a solvate, especially a hydrate for the production of a medicament for the treatment of pain.
  • the sodium channel blocker is an analog of TTX.
  • an analog may be one disclosed in Lin, U.S. Pat. No. 8,486,901 , which is incorporated by reference in its entirety, particularly for the identity, composition of matter, purification, and characterization of TTX analogs.
  • Further embodiments may include a TTX analog as disclosed in Buschmann et al., U.S. Pat. No. 9,018,222, which is incorporated by reference in its entirety, particularly for the identity and composition of matter of various TTX analogs.
  • any serotonin-norepinephrine reuptake inhibitor can be used in the present compositions and methods.
  • the SNRI is Duloxetine.
  • Duloxetine molecular formula C-isH-igNOS, is a commercially available central nervous system penetrant serotonin-norepinephrine reuptake inhibitor. Duloxetine also has some effect on the reuptake of norepinephrine. Duloxetine is (+)-N-methyl-3-(1- naphthalenyloxy)-3-(2-thienyl)propanamine hydrochloride. Duloxetine was first taught by U.S. Patent 4,956,388, which discloses its high potency serotonin reuptake inhibition as well as its inhibition of norepinephrine uptake. U.S.
  • Patent 4,956,388 is incorporated by reference in its entirety, in particular for the method of synthesis and purification of duloxetine.
  • “duloxetine” is used to refer to the free base of the molecule as well as any salt, solvate, crystalline or amorphous form of the compound as well as commercially formulated and commercially sold unit doses of the molecule.
  • the U.S. Food and Drug Administration marketing label for Duloxetine Delayed-Release Capsules, marketed in the U.S. under the trade name Cymbalta®, is incorporated by reference in its entirety, and particularly for the formulation and dosing information therein.
  • NRI norepinephrine reuptake inhibitor
  • Reboxetine is a commercially available central nervous system penetrant selective norepinephrine reuptake inhibitor (NRI). Reboxetine has the molecular formula C19H23NO3. Reboxetine is also known as 2-[(2-ethoxyphenoxy)phenylmethyl]-, (R*,R*)-Morpholine and (S*,S*) form, and mixtures thereof.
  • “reboxetine” is used to refer to the free base of the molecule as well as any salt, solvate, crystalline or amorphous form of the compound, whether sterochemically pure or a mixture, and commercially formulated and commercially sold unit doses of the molecules.
  • the European Medicines Agency Summary of Product Characteristics for reboxetine, marketed as Edronax® in Europe, is incorporated by reference in its entirety, in particular for its formulation and dosing information.
  • the composition for the treatment of pain includes: (a) an amount of tetrodotoxin; and (b) an amount of either a selective serotonin reuptake inhibitor (SSRI), serotonin-norepinephrine reuptake inhibitor (SNRI), or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat pain.
  • the composition includes a singly therapeutically effective amount of tetrodotoxin or a singly subtherapeutic amount of tetrodotoxin.
  • the amount of component (b) is a singly therapeutically effective amount of component (b) or a singly subtherapeutic amount of component (b).
  • component (b) is an SSRI.
  • component (b) is a mixed SSRI/NRI.
  • component (b) is an NRI.
  • component (b) is one or more of a SSRI, mixed SSRI/NRI, and NRI.
  • component (b) is an SNRI.
  • component (b) is a mixed SNRI/NRI.
  • component (a) and component (b) are formulated in the same pharmaceutically acceptable carrier. In an embodiment, component (a) and component (b) are formulated in the same pharmaceutically acceptable carrier and administered simultaneously via the same route of administration. In an embodiment, component (a) and component (b) are formulated in the same pharmaceutically acceptable carrier and form a unit dose.
  • component (a) and component (b) are formulated in different pharmaceutically acceptable carriers.
  • component (a) and component (b) are formulated in different pharmaceutically acceptable carriers and administered via different routes of administration.
  • component (a) and component (b) are formulated in different pharmaceutically acceptable carriers and component (a) is administered subcutaneously and component (b) is administered orally.
  • component (a) and component (b) are formulated in different pharmaceutically acceptable carriers and component (a) is administered subcutaneously, intramuscularly, or
  • component (a) and component (b) are formulated in different pharmaceutically acceptable carriers and component (a) is administered subcutaneously, intramuscularly, or intravenously; and component (b) is separately administered subcutaneously, intramuscularly, or intravenously.
  • the formulation may include a second active pharmaceutical ingredient.
  • the second active pharmaceutical ingredient may be an analgesic present in a therapeutically effective amount to ameliorate pain in conjunction with an embodiment of the present compositions or used in an embodiment of the present methods.
  • the embodied compositions may be co-administered with an analgesic; the analgesic being present at a therapeutically effective amount.
  • a composition for the treatment of pain includes: (a) an amount of tetrodotoxin; and (b) an amount of either a selective serotonin reuptake inhibitor (SSRI) , serotonin-norepinephrine reuptake inhibitor (SNRI), or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat pain.
  • SSRI selective serotonin reuptake inhibitor
  • SNRI serotonin-norepinephrine reuptake inhibitor
  • NRI norepinephrine reuptake inhibitor
  • TTX administered will be dependent on the human or mammal being treated, the severity of the disorder or condition, the rate of administration, the disposition of the compounds and the discretion of the prescribing physician.
  • an effective dosage of each is in the range of 3.0 pg to about 150 pg of TTX per day, in single or divided doses.
  • dosage levels below the lower limit of the aforesaid range may be more than adequate, while in other cases still larger doses may be employed without causing any harmful side effect, e.g. , by dividing such larger doses into several small doses for administration throughout the day.
  • TTX is administered in multiple doses. Dosing may be at least once, twice, three times, four times, five times, six times, or more than six times per day. Dosing may be every day for one, two, three, four, or more consecutive days with this regimen repeating.
  • a daily dosage of TTX or an analog thereof is in the range of 3.0 pg to about 150 pg, about 144 pg, in the range of about 3.0 pg to about 50 pg, about 44 pg, in the range of about 3.0 pg to about 30 pg, in the range of about 3.0 pg to about 15 pg, in the range of about 3.0 pg to 7.5 pg, or about 3.0 pg.
  • a daily dosage of TTX or an analog thereof is a mammal equivalent of a rat dosage, such that a daily dosage of TTX or an analog thereof is 3.0 pg/kg or 5.0 pg/kg.
  • the daily dosage in an adult human patient may be between about 0.5 to about 1 .5 of the mammal equivalent of a rat dosage, between about 0.75 to about 1.25 of the mammal equivalent of a rat dosage, or between about 0.9 to about 1 .1 of the mammal equivalent of a rat dosage.
  • the daily dosage in an adult human patient may be about 0.5, about 0.55, about 0.6, about 0.65, about 0.7, about 0.75, about 0.8, about 0.85, about 0.9, about 0.95, about 1 .0, about 1 .05, about 1.1 , about 1 .15, about 1.2, about 1 .25, about 1.3, about 1.35, about 1 .4, about 1 .45, or about 1.5 of the mammal equivalent of a rat dosage.
  • a daily dosage of TTX or an analog thereof is in the range of 148.5 pg to about 181.5 pg, 123.75 pg to about 206.25 pg, 82.5 pg to about 247.5 pg, about 82.5 pg, about 90.75 pg, about 99.0 pg, about 107.25 pg, about 1 15.5 pg, about 123.75 pg, about 132.0 pg, about 140.25 pg, about 148.5 pg, about 156.75 pg, about 165 pg, about 173.25 pg, about 181.5 pg, about 189.75 pg, about 198.0 pg, about 206.25 pg, about 214.5 pg, about 222.75 pg, about 231.0 pg, about 239.25 pg
  • a daily dosage of TTX or an analog thereof is in the range of 150 pg to about 180 pg, 125 pg to about 205 mg, 85 mg to about 250 mg, about 85 mg, about 90 mg, about 100 mg, about 105 mg, about 1 15 mg, about 125 mg, about 130 mg, about 140 mg, about 150 mg, about 155 mg, about 175 mg, about 180 mg, about 190 mg, about 200 mg, about 205 mg, about 215 mg, about 225 mg, about 230 mg, about 240 mg, or about 250 mg.
  • the daily dosage of TTX or an analog thereof is about 165 pg.
  • a daily dosage of TTX or an analog thereof is in the range of 137.5 pg to about 412.5 pg, 206.25 pg to about 343.75 pg, 247.5 pg to about 302.5 pg, about 137.5 pg, about 151.25 pg, about 165.0 pg, about 178.75 pg, about 192.5 pg, about 206.25 pg, about 220.0 pg, about 233.75 pg, about 247.5 pg, about 261.25 pg, about 275.0 pg, about 288.75 pg, about 302.5 pg, about 316.25 pg, about 330.0 pg, about 343.75 pg, about 357.5 pg, about 371 .25 pg, about 385.0 pg, about 398.75 pg
  • a daily dosage of TTX or an analog thereof is in the range of 140 pg to about 415 pg, 205 pg to about 345 pg, 250 pg to about 305 pg, about 140 pg, about 150 pg, about 180 pg, about 195 pg, about 205 pg, about 235 pg, about 250 pg, about 260 pg, about 290 pg, about 305 pg, about 315 pg, about 345 pg, about 360 pg, about 370 pg, about 400 pg, or about 415 pg.
  • the daily dosage of TTX or an analog thereof is about 275 pg.
  • the amount of TTX is less than an individually therapeutic amount. In an embodiment, the amount of TTX is less than an individually effective amount. In an embodiment, the amount of TTX is a subtherapeutic amount. The doses above are understood to be therapeutically effective amounts of TTX.
  • the total daily dose of TTX is a therapeutically effective amount. In a further embodiment, the total daily dose of TTX is a therapeutically effective amount administered subcutaneously, orally, intramuscularly, intravenously, intranasally, or transdermally.
  • the total daily dose of TTX is a subtherapeutic amount.
  • the total daily dose is 95% of a therapeutically effective amount, 90% of a therapeutically effective amount, 85% of a therapeutically effective amount, 80% of a therapeutically effective amount, 75% of a therapeutically effective amount, 70% of a therapeutically effective amount, 65% of a therapeutically effective amount, 60% of a therapeutically effective amount, 55% of a therapeutically effective amount, 50% of a therapeutically effective amount, 45% of a therapeutically effective amount, 40% of a therapeutically effective amount, 35% of a therapeutically effective amount, 30% of a therapeutically effective amount, 25% of a therapeutically effective amount, 20% of a therapeutically effective amount, 18% of a therapeutically effective amount, 15% of a therapeutically effective amount, 10% of a therapeutically effective amount, or 5% of a therapeutically effective amount.
  • dosage levels below the lower limit of the aforesaid ranges may be more than adequate, while in other cases still larger doses may be employed without causing any harmful side effect - for example, by dividing such larger doses into several small doses for administration throughout the day.
  • the SNRI is duloxetine.
  • duloxetine is administered orally.
  • duloxetine is administered parenterally.
  • duloxetine is administered transdermally.
  • duloxetine is administered subcutaneously.
  • duloxetine is administered subcutaneously in the same pharmaceutically acceptable carrier as TTX.
  • duloxetine is administered once per day.
  • the once daily dose is 20 mg.
  • the once daily dose is 30 mg.
  • the once daily dose is 60 mg.
  • duloxetine is administered in divided doses throughout a single day.
  • the total daily dose is 20 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg, 80 mg, 90 mg, 100 mg, 1 10 mg, or 120 mg.
  • the mammal is given the mammal equivalent of a rat dosage of about 1 mg/kg to 50 mg/kg.
  • a daily dosage of duloxetine is a mammal equivalent of a rat dosage, such that a daily dosage of duloxetine is 10 mg/kg or 15 mg/kg.
  • the daily dosage in an adult human patient may be between about 0.5 to about 1.5 of the mammal equivalent of a rat dosage, between about 0.75 to about 1 .25 of the mammal equivalent of a rat dosage, or between about 0.9 to about 1 .1 of the mammal equivalent of a rat dosage.
  • the daily dosage in an adult human patient may be about 0.5, about 0.55, about 0.6, about 0.65, about 0.7, about 0.75, about 0.8, about 0.85, about 0.9, about 0.95, about 1.0, about 1.05, about 1 .1 , about 1.15, about 1 .2, about 1.25, about 1 .3, about 1 .35, about 1.4, about 1.45, or about 1.5 of the mammal equivalent of a rat dosage.
  • a daily dosage of duloxetine is in the range of 275 g to about 825 mg, 412.5 mg to about 687.5 mg, 495 mg to about 605 mg, about 275 mg, about 302.5 mg, about 330 mg, about 357.5 mg, about 385 mg, about 412.5 mg, about 440 mg, about 467.5 mg, about 495 mg, about 522.5 mg, about 550 mg, about 577.5 mg, about 605 mg, about 632.5 mg, about 660 mg, about 687.5 mg, about 715 mg, about 742.5 mg, about 770 mg, about 797.5 mg, or about 825 mg.
  • a daily dosage of duloxetine is in the range of 415 mg to about 690 mg, about 305 mg, about 360 mg, about 415 mg, about 470 mg, about 525 mg, about 580 mg, about 635 mg, about 690 mg, about 745 mg, or about 800 mg. In preferred embodiments, the daily dosage of duloxetine is about 550 mg.
  • a daily dosage of duloxetine is in the range of 412.5 mg to about 1 .2375 g, 618.75 mg to about 1.03125 g, 742.5 mg to about 907.5 mg, about 412.5 mg, about 453.75 mg, about 495 mg, about 536.25 mg, about 577.5 mg, about 618.75 mg, about 660 mg, about 701.25 mg, about 742.5 mg, about 783.75 mg, about 825 mg, about 866.25 mg, about 907.5 mg, about 948.75 mg, about 990 mg, about 1.03125 g, about 1.0725 g, about 1.1 1375 g, about 1.155 g, about 1 .19625 g, or about 1 .2375 g.
  • a daily dosage of duloxetine is in the range of 415 mg to about 1.240 g,
  • the daily dosage of duloxetine is about 825 mg.
  • duloxetine is given once per day. In some embodiments, reboxetine is given twice per day. In some embodiments the dose is oral. In other embodiments the dose may be administered via the routes of administration as described below. The doses above are understood to be therapeutically effective amounts of duloxetine.
  • the total daily dose of duloxetine is from about 5 mg to 120 mg.
  • the total daily doses of duloxetine are selected from the group consisting of about 5 mg, about 10 mg, about 20 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 1 10 mg, and about 120 mg.
  • the total daily dose of duloxetine is in the range of about 1 mg to 120 mg. The doses above are understood to be therapeutically effective amounts of duloxetine.
  • An amount of TTX and an amount of a serotonin-norepinephrine reuptake inhibitor may be given by the same route of administration or a different route of administration.
  • An amount of TTX and an amount of a serotonin-norepinephrine reuptake inhibitor may be given by the same route of administration or a different route of administration.
  • the amount of TTX administered is not a therapeutically effective amount of TTX when administered alone.
  • the amount of TTX administered is a therapeutically effective amount of TTX when administered alone.
  • the amount of an SNRI administered is not a therapeutically effective amount of the SNRI when administered alone. In an embodiment, the amount of an SNRI administered is a therapeutically effective amount of the SNRI when administered alone.
  • the total daily dose is subtherapeutic for duloxetine alone.
  • the total daily dose is 95% of a therapeutically effective amount, 90% of a therapeutically effective amount, 85% of a therapeutically effective amount, 80% of a therapeutically effective amount, 75% of a therapeutically effective amount, 70% of a therapeutically effective amount, 65% of a therapeutically effective amount, 60% of a therapeutically effective amount, 55% of a therapeutically effective amount, 50% of a therapeutically effective amount, 45% of a therapeutically effective amount, 40% of a therapeutically effective amount, 35% of a therapeutically effective amount, 30% of a therapeutically effective amount, 25% of a therapeutically effective amount, 20% of a therapeutically effective amount, 18% of a therapeutically effective amount, 15% of a therapeutically effective amount, 10% of a therapeutically effective amount, or 5% of a therapeutically effective amount.
  • the total daily dose of duloxetine is a subtherapeutic amount in the range of about 5% of a therapeutically effective amount to about 98% of a therapeutically effective amount.
  • the NRI is reboxetine.
  • reboxetine is administered orally.
  • reboxetine is administered parenterally.
  • reboxetine is administered transdermally.
  • reboxetine is administered subcutaneously.
  • reboxetine is administered subcutaneously in the same pharmaceutically acceptable carrier as TTX.
  • the method further comprises the administration of an analgesic.
  • the composition further comprises an analgesic.
  • reboxetine is administered once or twice per day (b.i.d.); for example, each dose 4 mg, for a total daily dose of 8 mg. In other embodiments, reboxetine is administered twice per day in evenly divided doses for a total daily dose of 10 mg. In other embodiments, reboxetine is administered twice per day in evenly divided doses for a total daily dose of 12 mg. In some embodiments, reboxetine is administered in total daily doses of 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 1 1 mg, or 12 mg. In an embodiment, the mammal is given the mammal equivalent of a rat dosage of about 1 mg/kg to 6 mg/kg.
  • the daily dose of reboxetine is the mammal equivalent of a rat dosage, such that the daily dosage of reboxetine is 2 mg/kg or 5 mg/kg.
  • the daily dosage in an adult human patient may be between about 0.5 to about 1.5 of the mammal equivalent of a rat dosage, between about 0.75 to about 1 .25 of the mammal equivalent of a rat dosage, or between about 0.9 to about 1 .1 of the mammal equivalent of a rat dosage.
  • the daily dosage in an adult human patient may be about 0.5, about 0.55, about 0.6, about 0.65, about 0.7, about 0.75, about 0.8, about 0.85, about 0.9, about 0.95, about 1.0, about 1.05, about 1 .1 , about 1.15, about 1 .2, about 1.25, about 1 .3, about 1 .35, about 1.4, about 1.45, or about 1.5 of the mammal equivalent of a rat dosage.
  • a daily dosage of reboxetine is in the range of 55 mg to about 165 mg, 82.5 mg to about 137.5 mg, 99 mg to about 121 mg, about 55 mg, about 60.5 mg, about 66 mg, about 71.5 mg, about 77 mg, about 82.5 mg, about 88 mg, about 93.5 mg, about 99 mg, about 104.5 mg, about 1 10 mg, about 1 15.5 mg, about 121 mg, about 126.5 mg, about 132 mg, about 137.5 mg, about 143 mg, about 148.5 mg, about 154 mg, about 159.5 mg, or about 165 mg.
  • a daily dosage of reboxetine is in the range of 80 mg to about 140 mg, 100 mg to about 120 mg, about 60 mg, about 65 mg, about 70 mg, about 75 mg, about 80 mg, about 85 mg, about 90 mg, about 95 mg, about 100 mg, about 105 mg, about 1 15 mg, about 120 mg, about 125 mg, about 130 mg, about 135 mg, about 140 mg, about 145 mg, about 150 mg, about 155 mg, or about 160 mg.
  • the daily dosage of reboxetine is about 1 10 mg.
  • a daily dosage of reboxetine is in the range of 137.5 mg to about 412.5 mg, 206.25 mg to about 343.75 mg, 247.5 mg to about 302.5 mg, about 137.5 mg, about 151.25 mg, about 165.0 mg, about 178.75 mg, about 192.5 mg, about 206.25 mg, about 220.0 mg, about 233.75 mg, about 247.5 mg, about 261.25 mg, about 275.0 mg, about 288.75 mg, about 302.5 mg, about 316.25 mg, about 330.0 mg, about 343.75 mg, about 357.5 mg, about 371.25 mg, about 385.0 mg, about 398.75 mg, or about 412.5 mg.
  • a daily dosage of reboxetine is in the range of 140 mg to about 415 mg, 205 mg to about 345 mg, 250 mg to about 300 mg, about 140 mg, about 150 mg, about 180 mg, about 195 mg, about 205 mg, about 235 mg, about 250 mg, about 260 mg, about 290 mg, about 300 mg, about 315 mg, about 345 mg, about 360 mg, about 370 mg, about 400 mg, or about 415 mg.
  • the daily dosage of reboxetine is about 275 mg.
  • reboxetine is given once per day. In some embodiments, reboxetine is given twice per day. In some embodiments reboxetine is administered orally.
  • reboxetine is administered parenterally.
  • the doses above are understood to be therapeutically effective amounts of reboxetine.
  • reboxetine is administered at a total daily dose selected from about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 6 mg, about 7 mg, about 8 mg, about 9 mg, about 10 mg, about 1 1 mg, about 12 mg, about 13 mg, about 14 mg, about 15 mg.
  • the doses above are understood to be therapeutically effective amounts of reboxetine.
  • reboxetine is administered orally and the total daily dose is subtherapeutic for reboxetine alone.
  • the total daily dose is 95% of a therapeutically effective amount, 90% of a therapeutically effective amount, 85% of a therapeutically effective amount, 80% of a therapeutically effective amount, 75% of a therapeutically effective amount, 70% of a therapeutically effective amount, 65% of a therapeutically effective amount, 60% of a therapeutically effective amount, 55% of a therapeutically effective amount, 50% of a therapeutically effective amount, 45% of a therapeutically effective amount, 40% of a therapeutically effective amount, 35% of a therapeutically effective amount, 30% of a therapeutically effective amount, 25% of a therapeutically effective amount, 20% of a therapeutically effective amount, 18% of a therapeutically effective amount, 15% of a therapeutically effective amount, 10% of a therapeutically effective amount, or 5% of a therapeutically effective amount.
  • the total daily dose of reboxetine is a subtherapeutic amount in the range of about 5% of a therapeutically effective amount to about 98% of a therapeutically effective amount.
  • the amount of a NRI administered is not a therapeutically effective amount of the NRI for analgesia when administered alone. In an embodiment, the amount of a NRI administered is a therapeutically effective amount for analgesia of the NRI when administered alone.
  • the invention relates generally to methods for treating pain by administering one or more doses of a sodium channel blocker in combination with one or more selective serotonin reuptake inhibitors. In certain aspects, the invention relates generally to methods for treating pain by administering a sodium channel blocker in combination with one or more mixed serotonin and/or norepinephrine reuptake inhibitors. In certain aspects, the invention relates generally to methods for treating pain by administering a sodium channel blocker in combination with one or more serotonin-norepinephrine reuptake inhibitors. In certain aspects, the invention relates generally to methods for treating pain by administering one or more amounts of a sodium channel blocker in combination with one or more norepinephrine reuptake inhibitors. In preferred embodiments, the sodium channel blocker is TTX.
  • the method is used to treat a subject undergoing treatment for chemotherapy induced neuropathic pain.
  • a method for the treatment of neuropathic pain comprises: (a) an amount of tetrodotoxin; and (b) an amount of any one of paroxetine, fluoxetine, duloxetine, and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat neuropathic pain.
  • the method for the treatment of neuropathic pain wherein the neuropathic pain is selected from the group consisting of pain arising from diabetic peripheral neuropathy, fibromyalgia, and chemotherapy induced neuropathic pain.
  • the method for the treatment of neuropathic pain wherein the mammal is a human patient undergoing treatment for chemotherapy induced neuropathic pain.
  • the composition or method is used to treat a subject undergoing treatment for cancer pain and wherein said subject also receives at least one low dose of an analgesic or an opioid.
  • the subject is further treated with about 500 mg morphine equivalent dose of an opioid.
  • the method is used to treat a subject undergoing treatment for nociceptive pain.
  • a method for the treatment of nociceptive pain comprises: (a) an amount of tetrodotoxin; and (b) an amount of any one of paroxetine, fluoxetine, duloxetine, and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat nociceptive pain.
  • the method for the treatment of nociceptive pain wherein the nociceptive pain is selected from the group consisting of radicular pain, somatic pain, and visceral pain.
  • the method for the treatment of nociceptive pain wherein the mammal is a human patient undergoing treatment for nociceptive pain.
  • compositions disclosed herein may be administered
  • the methods of the present invention may include administration subcutaneously, orally, or any other route know to the art.
  • the composition for the treatment of neuropathic pain comprising: (a) an amount of tetrodotoxin; and (b) an amount of any one of duloxetine and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat neuropathic pain.
  • compositions disclosed herein include a combination of (a) a dosage amount of tetrodotoxin and (b) a dosage amount of duloxetine or reboxetine, wherein component (a) and component (b) exhibit synergism to treat neuropathic and/or nociceptive pain.
  • a composition includes a combination of (a) 5 pg per kg TTX and (b) 5 mg per kg Reboxetine, wherein the 5 pg/kg TTX and 5 mg/kg Reboxetine exhibit synergism to treat neuropathic and/or nociceptive pain.
  • a composition in another embodiment, includes a combination of (a) 3 pg per kg TTX and (b) 2 mg per kg Reboxetine, wherein the 3 pg/kg TTX and 2 mg/kg Reboxetine exhibit synergism to treat neuropathic and/or nociceptive pain.
  • a composition includes a combination of (a) 3 pg per kg TTX and (b) 10 mg per kg Duloxetine, wherein the 3 pg/kg TTX and 10 mg/kg Duloxetine exhibit synergism to treat neuropathic and/or nociceptive pain.
  • a composition includes a combination of (a) 5 pg per kg TTX and (b)
  • the composition is a single formulation with both an amount of TTX and an amount of any one of paroxetine, fluoxetine, duloxetine, and reboxetine.
  • a single formulation is administered subcutaneously.
  • such a single formulation is administered intramuscularly.
  • such a single formulation is administered intravenously.
  • such a single formulation is administered transdermally.
  • the TTX is administered via the same route of administration as the SSRI. In one embodiment, the TTX is administered via the same route of administration as the SNRI. In one embodiment, the TTX is administered via the same route of administration as the NRI.
  • the TTX is administered via a different route of administration as the SSRI. In one embodiment, the TTX is administered via a different route of administration as the SNRI. In one embodiment, the TTX is administered via a different route of administration as the NRI.
  • At least one analgesic is administered via the same route of administration as the TTX. In some embodiments, at least one analgesic is administered via a different route of administration than the TTX. In some embodiments, at least one analgesic is administered via the same route of administration as the SSRI. In some embodiments, at least one analgesic is administered via the same route of administration as the SNRI. In some embodiments, at least one analgesic is administered via the same route of administration as the NRI. In some embodiments, the combination, an embodiment of the present compositions and an analgesic, is administered in a single dose. Such
  • administration may be by injection, e.g., intravenous injection, in order to introduce pain control quickly.
  • routes including the preferred oral route, may be used as appropriate.
  • a single dose of the combination of an embodiment of the present compositions and an analgesic may also be used for treatment of acute pain.
  • the amount of TTX and the amount of either a SSRI, SNRI or NRI may be given simultaneously or at differing times.
  • An amount of TTX in a first pharmaceutically acceptable formulation, the second pharmaceutically acceptable formulation including a neurotransmitter reuptake inhibitor, or a combination of TTX and a neurotransmitter reuptake inhibitor may be administered in either single or multiple doses by any of the accepted modes of administration of agents having similar utilities, including rectal, buccal, intranasal and transdermal routes, by intra-arterial injection, intravenously, intraperitoneally, parenterally, intramuscularly, subcutaneously, orally, topically, or as an inhalant. Kits
  • An embodiment may be a kit comprising a unit dose of an amount of TTX and an amount of an SNRI.
  • An embodiment may be a kit comprising a daily dose of an amount of TTX, which may be more than one unit dose, and a daily dose of an amount of an SNRI, which may be more than one unit dose.
  • the kit comprises one or more unit doses of TTX and one or more oral doses of the SNRI duloxetine.
  • the one or more unit doses of TTX may be for subcutaneous injection, or for administration orally, intramuscularly, intravenously, intranasally, or transdermally.
  • An embodiment may be a kit comprising a unit dose of an amount of TTX and an amount of an NRI.
  • An embodiment may be a kit comprising a daily dose of an amount of TTX, which may be more than one unit dose, and a daily dose of an NRI, which may be more than one unit dose.
  • the kit comprises one or more unit doses of TTX and one or more oral doses of the NRI reboxetine.
  • the one or more unit doses of TTX may be for subcutaneous injection, or for administration orally, intramuscularly, intravenously, intranasally, or transdermally.
  • An embodiment may be a kit comprising a unit dose of an amount of TTX and an amount of an SSRI.
  • An embodiment may be a kit comprising a daily dose of an amount of TTX, which may be more than one unit dose, and a daily dose of an amount of an SSRI, which may be more than one unit dose.
  • the kit comprises one or more unit doses of TTX and one or more oral doses of an SSRI (e.g., paroxetine or fluoxetine).
  • the one or more unit doses of TTX may be for subcutaneous injection, or for administration orally, intramuscularly, intravenously, intranasally, or transdermally.
  • the kit comprises one or more unit doses of TTX and one or more SNRIs. In one embodiment, the kit comprises one or more unit doses of TTX, one or more SNRIs, and one or more analgesics. In one embodiment, the kit comprises one or more unit doses of TTX, an amount of at least one SNRI, and an amount of at least one NRI. In one embodiment, the kit comprises one or more unit doses of TTX, at least one SNRI, at least one NRI, and at least one analgesic.
  • the kit comprises one or more unit doses of TTX, an amount of at least one SNRI, an amount of at least one NRI , and an amount of one or more analgesics. In one embodiment, the kit comprises one or more unit doses of TTX and one or more NRIs. In one embodiment, the kit comprises one or more unit doses of TTX, one or more NRIs, and one or more analgesics. In any of the preceding embodiments, the one or more unit doses of TTX may be for subcutaneous injection, or for administration orally, intramuscularly, intravenously, intranasally, or transdermally. In preferred embodiments, the one or more unit doses of TTX is for subcutaneous injection.
  • tetrodotoxin refers collectively to tetrodotoxin, its analogs, derivatives, metabolites, degradation products, hydrates thereof, salts thereof, and combinations thereof.
  • Synergism or drug synergy as discussed above in paragraph [0024] means that the combined effect of two drugs in combination is greater than predicted (additive effect) by their individual potencies, it is called super-additive, and the combination is said to be synergistic (i.e. enhanced effect).
  • analogs as used in this application is defined here as meaning a chemical compound that is a derivative of a compound which has similar biochemical activity to that compound.
  • derivatives as used in this application is defined here as meaning a chemical compound having undergone a chemical derivatization such as substitution or addition of a further chemical group to change (for pharmaceutical use) any of its physicochemical properties, such as solubility or bioavailability.
  • Derivatives include so-called prodrugs, e.g. ester and ether derivatives of an active compound that yield the active compound per se after administration to a subject.
  • analgesic refers to a substance that acts to relieve pain.
  • Analgesics may be drawn from any of the following: Cyclo-oxygenase-2 inhibitors (e.g. valdecoxib, rofecoxib, or celecoxib); antimigraine agents (e.g. sumatriptan, methylsergide maleate, frovatriptan, naratriptan, almotriptan, ergotamine, rizatriptan, zolmitriptan, or dihydroergotamine); non-steroidal anti-inflammitory agents (e.g.
  • ibuprofen e.g. aspirin or magnesium salicylate
  • narcotic agents e.g.
  • morphine oxycodone, fentanyl, oxymorphone, hydromorphone, meperidine, buprenorphine, methadone, tramadol, butorphanol, tapentadol, propoxyphene, alfentanil, liposomal morphene, sufentanil, remifentanil, or pentazocine).
  • an effective amount or“therapeutically effective amount” refers to that amount of a compound or combination of compounds as described herein that is sufficient to effect the intended application including, but not limited to, disease treatment.
  • therapeutically effective amount may vary depending upon the intended application (in vitro or in vivo), or the subject and disease condition being treated (e.g., the weight, age and gender of the subject), the severity of the disease condition, the manner of administration, etc. which can readily be determined by one of ordinary skill in the art.
  • the term also applies to a dose that will induce a particular response in target cells (e.g., the reduction of platelet adhesion and/or cell migration).
  • the specific dose will vary depending on the particular compounds chosen, the dosing regimen to be followed, whether the compound is administered in combination with other compounds, timing of administration, the tissue to which it is administered, and the physical delivery system in which the compound is carried.
  • “Subtherapeutic” doses are defined as doses that are less than doses that would be an individually effective amount or an individually therapeutically effective amount of the compound, composition, or combinations of compounds.
  • A“therapeutic effect” as that term is used herein, encompasses a therapeutic benefit and/or a prophylactic benefit.
  • a prophylactic effect includes delaying or eliminating the appearance of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing, halting, or reversing the progression of a disease or condition, or any combination thereof.
  • the terms“QD,”“qd,” or“q.d.” mean quaque die, once a day, or once daily.
  • the terms“BID,”“bid,” or“b.i.d.” mean bis in die, twice a day, or twice daily.
  • tid or“t.i.d.” mean ter in die, three times a day, or three times daily.
  • compositions that contain pharmaceutically acceptable salts of TTX and optionally pharmaceutically acceptable carriers and/or pharmaceutically acceptable excipients, and/or non-active ingredients known generally to the art to serve non-therapeutic purposes in medicaments.
  • mammal equivalent dose is used to indicate the conversion of a dose determined in one mammal species for another subject mammal species. For example, and without limitation, converting a dose determined in a rat to the equivalent human dose.
  • the methods for the determination of such equivalent doses are known to those skilled in the art and are summarized in the U.S. Food and Drug Administration Guidance to Industry, available at https://www.fda.gov/downloads/drugs/guidances/ucm078932.pdf, entitled “Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers.”
  • the document is incorporated by reference in its entirety, with particular attention to the method summarized in Table I.
  • compositions may include formulations wherein at least one component is a composition disclosed by Zhang et al. in U.S. Pat. No. 8, 124,608, which is incorporated by reference in its entirety.
  • Pharmaceutically acceptable formulations may include formulations wherein at least one component is a composition disclosed by Lin et al. in U.S. Pat. No. 8,222,258, which is incorporated by reference in its entirety.
  • compositions may include formulations wherein at least one component is a composition disclosed by Lin et al. in U.S. Pat. No. 8,530,481 , which is incorporated by reference in its entirety.
  • salt refers to salts derived from a variety of organic and inorganic counterions known in the art.
  • Pharmaceutically acceptable acid addition salts can be formed with inorganic acids and organic acids.
  • Preferred inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid and phosphoric acid.
  • Preferred organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p- toluenesulfonic acid and salicylic acid.
  • Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases.
  • Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese and aluminum.
  • Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins. Specific examples include isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. In some embodiments, the
  • cocrystal refers to a molecular complex derived from a number of cocrystal formers known in the art. Unlike a salt, a cocrystal typically does not involve hydrogen transfer between the cocrystal and the drug, and instead involves intermolecular interactions, such as hydrogen bonding, aromatic ring stacking, or dispersive forces, between the cocrystal former and the drug in the crystal structure.
  • ‘‘Pharmaceutically acceptable carrier” or“pharmaceutically acceptable excipient” is intended to include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and inert ingredients.
  • the use of such pharmaceutically acceptable carriers or pharmaceutically acceptable excipients for active pharmaceutical ingredients is well known in the art. Except insofar as any
  • a medicament according to the method includes the manufacture of a medicament wherein the medicament contains from 3.0 pg to about 150 pg of TTX or a pharmaceutically acceptable analog thereof per dosage unit.
  • mice male Sprague-Dawley Rattus norvegicus, 7-8 weeks old, were used, two animals per experimental group. Where needed, additional rats were added to the protocol in appropriate numbers to compensate for rats not developing oxaliplatin-induced peripheral neuropathy. All rats were housed in groups of 4 in an air- conditioned room on a 12-hour light/dark cycle. Food and water was available ad libitum.
  • Oxaliplatin was dissolved in 5% dextrose in distilled water to 4 mg/ml_; then further diluted the stock solution to 2 mg/ml_ with normal saline before use to induce neuropathy.
  • TTX for injection doses were prepared under aseptic conditions in the morning of administration. One or more vials, as required, was removed from the storage condition and allowed to come to ambient temperature (20°C to 25°C). Each vial of lyophilized product was reconstituted with sterile water for injection under aseptic conditions.
  • Duloxetine was dissolved in normal saline; capsules were crushed in a pestle, then normal saline was added. The material was further ground to produce an even suspension.
  • Reboxetine tablets were crushed in a pestle, then normal saline was added. The material was further ground to produce an even suspension.
  • Rat model for chemotherapy induced neuropathic pain Rats were anesthetized with 3% isoflurane mixed with medical oxygen (2 L/min). Oxaliplatin was intravenously injected through the tail vein. The rats were monitored for the development of neuropathic pain, characterized by significant mechanical allodynia. A series of graduated von Frey hairs were applied to the hind-paw to trigger a withdrawal response (Paw Withdrawal Threshold, PWT). Rats with significant mechanical allodynia (PWT ⁇ 4.0 g) were then selected for use in the drug testing experimental groups. The study groups are shown in Table 1 below.
  • TTX was administered to each rat by subcutaneous injection.
  • rats receiving a SSRI or NRI either duloxetine or reboxetine was administered orally.
  • TTX for injection and the saline control were administered subcutaneously once daily for 14 days.
  • duloxetine or reboxetine was administered orally once daily for 14 days.
  • TTX was administered by subcutaneous injection first, then immediately followed by an oral dose of either duloxetine or reboxetine, depending on the particular animal.
  • Paw Withdrawal Threshold Rats were placed in individual perspex boxes on a raised metal mesh for at least 40 min before the PWT. The test was conducted by starting from the filament of lowest force (1 g), and applying each filament perpendicularly to the center of the ventral surface of the paw until the filament slightly bends, and held for 6 seconds. If the animal withdraws or lifts the paw upon stimulation, then the von Frey hair with the immediately next lower force than the one presently generating the withdrawal response is tested. If no response is observed, then test a filament with a force immediately higher is applied as described. The lowest amount of force required to induce reliable responses (positive in 2 out of 3 trials) was recorded as the PWT result.
  • TTX at 1 , 2, and 3 pg/kg did not significantly change the baseline PWT over the 5 days of dosing undertaken in this pilot study, whilst TTX at 3 pg/kg slightly increased the baseline PWT on Day 5 of the dosing course, compared to the pre-dosing value. After the cessation of dosing, the baseline PWT declined to the pre-dosing level on Day 7.
  • TTX increased PWT at 1-hour post-dosing.
  • the PWT declined after 2 hours post-dosing.
  • Duloxetine administered at 5, 10 and 15 mg/kg, increased the baseline PWT and PWT post-dosing in rats with oxaliplatin-induced neuropathy in a dose-dependent manner.
  • Reboxetine administered at 1 , 3 and 5 mg/kg, increased the baseline PWT and PWT post-dosing in rats with oxaliplatin-induced neuropathy, in a dose-dependent manner.
  • TTX at 3 pg/kg with duloxetine at 10 mg/kg produced an increase in baseline PWT and PWT post-dosing, with a trend towards showing an effect greater than that induced by TTX or duloxetine administered alone.
  • the combination of TTX at 3 pg/kg and reboxetine at 3 mg/kg increased the baseline PWT and PWT post-dosing, with a trend towards showing an effect greater than that induced by TTX or reboxetine administered alone.
  • This study is a further pilot study to determine the most effective dose ranges for the combinations of tetrodotoxin (TTX) with duloxetine or reboxetine on mechanical allodynia in rats with oxaliplatin-induced neuropathy.
  • the study purpose was to determine the most effective dose ranges for the combinations of TTX with duloxetine (Dul) or reboxetine (Reb) on mechanical allodynia in rats with oxaliplatin-induced neuropathy.
  • Oxaliplatin Description: White powder; Vehicle: 5% dextrose, sterile; Preparation: Oxaliplatin was dissolved in 5% dextrose in distilled water, then diluted to 2 mg/ml_ with normal saline before use.
  • TTX for injection Description: Lyophilized white powder; Preparation: The dose was prepared under aseptic conditions in the morning of the administration. One or more vials, as required, was removed from storage condition and allowed to equilibrate to ambient temperature (20°C to 25°C). Each vial of lyophilized product was reconstituted with sterile water for injection under aseptic conditions.
  • Duloxetine Description: Capsules; Preparation: Duloxetine was dissolved in normal saline. Capsules were opened into a mortar, with normal saline added, and the content was ground to form an even suspension.
  • Reboxetine Description: Tablets; Preparation: Reboxetine was dissolved in normal saline. Other solutions, for example, 5% dextrose and normal saline, were prepared using standard aseptic research methods.
  • Animal study design Model system for neuropathic pain— Rats were anaesthetized with 3% isoflurane mixed with oxygen (2 L per min). Oxaliplatin was administered intravenously through the tail vein. The development of neuropathic pain, characterized by significant mechanical allodynia, was monitored using a series of graduated von Frey hairs applied to the hind-paw to trigger a withdrawal response (Paw Withdrawal Threshold, PWT). Only those rats with significant mechanical allodynia (PWT ⁇ 4 g) were selected for further drug testing.
  • Paw Withdrawal Threshold Rats were placed in individual perspex boxes on a raised metal mesh for at least 40 min before the PWT. The test was conducted by starting from the filament of lowest force (1 g), and applying each filament perpendicularly to the center of the ventral surface of the paw until the filament slightly bends, and held for 6 seconds. If the animal withdraws or lifts the paw upon stimulation, then the von Frey hair with the immediately next lower force than the one presently generating the withdrawal response is tested. If no response is observed, then test a filament with a force immediately higher is applied as described. The lowest amount of force required to induce reliable responses (positive in 2 out of 3 trials) was recorded as the PWT result.
  • baseline PWT appeared to follow a similar trend as that observed in the duloxetine-alone and TTX-alone treated groups, although on day 4, PWT increased more markedly than that observed in the duloxetine-alone and TTX-alone treated groups. PWT then steadily increased between 6-8 g up to day 14, when a marked increase in PWT to 10 g was observed, before declining on day 16 and remaining at a level of 7-8 g between days 18-27. Baseline PWT in this group appeared to be higher than that of the TTX-alone treated group from day 4 to day 20, but lower than that observed in the duloxetine-alone treated group.
  • Post-dosing PWT slightly increased in the duloxetine-alone treated and the combination groups, but not in the TTX-alone treated group on day 1. From day 5 to day 14 a marked increase in post-dosing PWT at 1 and 2 hours was observed in the combination group, which was more notable than that observed in the TTX-alone treated group, where a marked increase in post-dosing PWT was only apparent on days 5, 8, 10, and 14. A marked increase in post-dosing PWT was also apparent in the duloxetine-alone treated groups on days 7, 8 and 9, although this effect was not as abrupt in occurrence as those noted in the TTX alone or the combination-treated groups. The data for this group is plotted in Figure 7. No significant adverse effects observed during the dosing/observation period.
  • baseline PWT increased sharply compared to that observed in the TTX-alone treated group from day 2 to day 5, before declining sharply on day 7 to similar levels of PWT to that observed in the TTX-alone treated group. PWT remained around this new level until day 14.
  • baseline PWT increased even more sharply from day 2 than that observed in the group administered a combination of TTX and duloxetine group. This effect reached a peak value on day 4, remained high on day 5, and then gradually declined towards day 14 to PWT levels similar to those observed in the TTX and duloxetine group.
  • PWT post-dosing increased sharply on each dosing day in the 5 pg/kg TTX-alone treated group, the combination group of 5 pg/kg TTX and duloxetine, and in the combination group of 5 pg/kg TTX and reboxetine.
  • the data for this group is plotted in Figure 9. No significant adverse effects observed during the dosing/observation period.
  • PWT post-dosing at 1 and 2 hours increased sharply from day 4 to day 14 in the 5 pg/kg TTX-alone treated group.
  • a sharp increase in PWT was observed and more notable in the combination group of TTX and duloxetine than that observed in the TTX-alone treated group, from day 2 to day 6.
  • PWT post-dosing also increased from day 1 on each dosing day, but it was less marked than the PWT observed in the combination treated group from day 2 to day 1 1 .
  • the data for this group is plotted in Figure 10. No significant adverse effects observed during the dosing/observation period.
  • the baseline PWT in the 5 mg/kg reboxetine- alone treated group also remained higher after cessation of dosing than that observed during the pre-dosing period.
  • Baseline PWT in the combination group of TTX and reboxetine appeared to be similar, between days 2 to 12, to those recorded in the TTX-alone and reboxetine-alone treated groups, except on days 13, 14, 16, 18 and 27 where it appeared to be higher than the PWT of the TTX- and reboxetine-alone treated groups.
  • PWT post-dosing in the reboxetine-alone treated group increased from day 1 and on each dosing day.
  • the post-dosing PWT in the combination group increased from day 1 and was notable from day 2 to day 14, the extent of the increase in PWT and peak value being clearly more marked than those of the TTX- and reboxetine-alone treated groups over the same period.
  • the data for this group is plotted in Figure 1 1.
  • TTX at 5 pg/kg increased the baseline PWT and post-dosing PWT in rats with oxaliplatin-induced neuropathy.
  • this dose of TTX was combined with 15 mg/kg duloxetine or 5 mg/kg reboxetine, the baseline PWT and post-dosing PWT increased to a level higher than that observed when TTX at 5 pg/kg was administered alone.
  • the combination treatment comprising 5 pg/kg TTX and 15 mg/kg duloxetine increased the post-dosing PWT, an effect more prominent than that observed with 5 pg/kg TTX or 15 mg/kg duloxetine administered alone from day 1 to day 6, in rats with oxaliplatin- induced neuropathy.
  • the combination treatment comprising of 5 pg/kg TTX and 5 mg/kg reboxetine increased the baseline PWT on days 13, 14,16, 18 and 27, and PWT post-dosing more prominently than dosing with 5 pg/kg TTX or 5 mg/kg reboxetine-alone, over the 14 day dosing course and 2 week post-treatment observation period in rats with oxaliplatin-induced neuropathy.
  • Example 3 Synergism analysis of data from pilot studies [00199] The data from Examples 1 and 2 described above were analyzed for synergism. In both of these studies there were two rats were selected for each drug dose, and for each drug combination.
  • Baseline is defined as the time point of measurement immediately prior to the treatment, that is, Day 1 Hour 0.
  • the cut-off value of 15 is the maximum PWT that can be achieved.
  • %MPE measures the improvement on PWT, with 0 indicating no improvement and 100 indicating complete recovery.
  • PWT data were collected at various time points for each rat.
  • the first step is to assess the dose effect relationship, i.e. the response from a rat under a given dose level.
  • %MPE mean response
  • the response was estimated for each rat by averaging the %MPE over all measurements, resulting in a single data point representing the response for each rat under each drug and dose (individual drug alone), and for the combinations.
  • a hyperbolic model was then constructed for the dose-effect relations:
  • MPEmax the maximum effect
  • C is the constant that represents the dose corresponding to the 50% of the MPE (or D50).
  • the dose-effect data for each drug alone (TTX, duloxetine, and reboxetine) were fitted to the above model to estimate C, using the SAS non-linear procedure PROC NLIN.
  • the relative potency measures the relative effect (strength) of two drugs.
  • the relative potency of Drug B is 0.5 (or 50%) of Drug A, meaning that either Drug B needs to double the dose to achieve the same effect as Drug A, or Drug B is a diluted dose (to 50%) of Drug A.
  • any combination of two drugs (A and B) can be converted to an equivalent dose of one drug:
  • Dose Equivalance Drug A Dose of Drug A + R X ⁇ Dose of Drug B )
  • the equivalent dose of a combination was then calculated in the hyperbolic model to obtain the expected effect (%MPE) - the effect that is expected if the two drugs are additive.
  • the expected effect was compared with the actual (observed) effect to determine if the combination drug is super-additive (synergistic, which can be an observed effect that is greater than the expected), additive (observed effect is the same as the expected), or subadditive (observed effect is lower than the expected).
  • the expected effect of the two drug combinations was calculated for the dose combinations of two drugs to construct a response surface.
  • This response surface presented as a 3-D plot, allows for a visual comparison of the observed effect of the drug combination against the expected effect under the assumption of additivity from the two drugs.
  • An interaction index defined as the ratio of the equivalent dose of the drug combination to the dose of the single drug that results in the same effect. An index of less than 1 indicates that the effect of a combination is super-additive (exhibiting synergism), 1 indicates additive, and greater than 1 indicates sub-additive (Tallarida, 2001).
  • Figure 12 shows the dose-effect relations, as estimated using a hyperbolic model, for each of the three drugs administered alone.
  • Figure 12A shows the dose-effect relationship for TTX alone.
  • Figure 12B shows the dose-effect relationship for reboxetine alone.
  • Figure 12C shows the dose-effect relationship for duloxetine alone.
  • the hyperbolic curved lines in each panel represent the dose response curves.
  • the dots represent the mean % PE (over the post-baseline period) for each rat.
  • the parameter C (or D50) in the hyperbolic function is estimated from the non-linear model; the results are shown in T able 4 below.
  • TTX For TTX, it was estimated that a dose of 12.65 pg/kg is required to achieve 50% MPE, while for Duloxetine and Reboxetine it is 24.41 and 9.32 mg/kg, respectively. These translate to a relative potency (relative to TTX) of 0.52 for Duloxetine and 1.36 for Reboxetine.
  • Table 4 [00211] Based on the relative potency, the TTX dose equivalence was calculated for each of the drug combinations, as shown in the Table 5 below. The table also shows the expected %MPE (assuming additive effect), the actual (observed) %MPE, the corresponding dose required to achieve the observed effect (for TTX alone), and the interaction index.
  • Oxaliplatin was dissolved in 5% dextrose in distilled water to 4 mg/ml_.
  • TTX for injection doses were prepared under aseptic conditions in the morning of administration. One or more vials, as required, were removed from the storage condition and allowed to come to ambient temperature (20°C to 25°C). Each vial of lyophilized product was reconstituted with sterile water for injection under aseptic conditions.
  • Duloxetine was dissolved in normal saline; crushed in a pestle, then normal saline will be added, with the material further ground to produce an even suspension.
  • Reboxetine for the study was crushed in a pestle, then normal saline added. The material was further ground to produce an even suspension.
  • Other solutions for example, 5% dextrose and normal saline, were prepared using standard aseptic methods.
  • Rat model for chemotherapy induced neuropathic pain Rats were anesthetized with 3% isoflurane mixed with medical oxygen (2 L/min). Oxaliplatin was intravenously injected through the tail vein. The rats were monitored for the development of neuropathic pain, characterized by significant mechanical allodynia. A series of graduated von Frey hairs were applied to the hind-paw to trigger a withdrawal response (Paw Withdrawal Threshold, PWT). Rats with significant mechanical allodynia (PWT ⁇ 4.0 g) were then selected for use in the drug testing experimental groups. The study groups are shown in Table 6 below.
  • TTX was administered to each rat by subcutaneous injection.
  • rats receiving either a SSRI or NRI the compound was administered orally.
  • TTX for injection and saline control were administered subcutaneously once daily for 14 days.
  • duloxetine or reboxetine were administered orally once daily for 14 days.
  • TTX was administered by subcutaneous injection first, then immediately followed by an oral dose of either duloxetine or reboxetine, depending on the particular animal.
  • Paw Withdrawal Threshold Rats were placed in individual perspex boxes on a raised metal mesh for at least 40 min before the PWT. The test was conducted by starting from the filament of lowest force (1 g), and applying each filament perpendicularly to the center of the ventral surface of the paw until the filament slightly bends, and held for 6 seconds. If the animal withdraws or lifts the paw upon stimulation, then the von Frey hair with the immediately next lower force than the one presently generating the withdrawal response is tested. If no response is observed, then test a filament with a force immediately higher is applied as described. The lowest amount of force required to induce reliable responses (positive in 2 out of 3 trials) was recorded as the PWT result.
  • Figures 17-18 show the effect of the combinations of TTX with Duloxetine or TTX with Roboxetine on paw withdrawal threshold compared to TTX, Duloxetine or Reboxetine alone. Statistical significance is illustrated by the asterisk.
  • the effect was significantly more evident than those of TTX at 3pg/kg or Reboxetine at 3 mg/kg alone, or their combinations.
  • the results suggest that the combination of low dose TTX with duloxetine or reboxetine leads to similar therapeutic effects with less adverse side-effects and less severity compared to the higher dose of each individual compound administered alone, as outlined in previous studies, and noted in the product monographs. Therefore, combinations of TTX with either duloxetine or reboxetine may be potentially produce a better result in relieving clinical chemotherapy-induced neuropathic pain.
  • TTX tetrodotoxin
  • duloxetine and reboxetine administered alone, and combinations comprised of TTX with either duloxetine or reboxetine, at low or high doses, respectively were tested by paw withdrawal threshold (PWT), assessed using a series of graduated von Frey hairs, over a 5- or 14-day dosing course observed over a 1 -2-week period post-treatment in rats with oxaliplatin-induced neuropathy.
  • Paw withdrawal threshold was asssed daily, and the data analyzed statistically as discussed above. Multidimensional dose plots were prepared according to the general methods of Tallarida as discussed above in the Synergisim section.
  • Figure 19A, 19B, and 19C show the dose-effect relations at Day 5 Hour 1 , as estimated using a hyperbolic model, for each of the three drugs (alone).
  • the lines represent the dose response curves and the dots represent the mean %MPE (over the post-baseline period) for each rat.
  • Figure 20 is the response surface plots showing the expected responses under additive assumption and the actual (observed) responses from the drug combinations at Day 5 Hour 1.
  • a point above the surface represent the existence of synergism in the drug combination.
  • the strong performance of TTX 5 pg/kg + Duloxetine 15 mg/kg is clearly noticeable in Figure 20B.
  • Figure 21A, 21 B, 21 C show the dose-effect relations at Day 14 Hour 1 , as estimated using a hyperbolic model, for each of the three drugs (alone).
  • the lines represent the dose response curves and the dots represent the mean % PE (over the post-baseline period) for each rat.
  • Figure 22 is the response surface plots showing the expected responses under additive assumption and the actual (observed) responses from the drug combinations at Day 14 Hour 1. A point above the surface represent the existence of synergism in the drug combination.
  • TTX 5 pg/kg + Duloxetine 15 mg/kg appears to be synergistic, with an interaction index of 0.579; while based on data from Hour 1 Day 14, drug combinations TTX 5 pg/kg + Reboxetine 5 mg/kg, TTX 3 pg/kg + Duloxetine 10 mg/kg, and TTX 5 pg/kg + Duloxetine 15 mg/kg are synergistic.

Abstract

The present invention relates to compositions and methods of treating pain comprising an amount of tetrodotoxin and an amount of a neurotransmitter reuptake inhibitor selected from the group consisting of a serotonin-norepinephrine reuptake inhibitor (SNRI), a norepinephrine reuptake inhibitor (NRI), and a selective serotonin reuptake inhibitor (SSRI). These compositions may also include additional analgesic compounds, and can be used in synergistic doses. The methods of using these compositions include the treatment of neuropathic pain.

Description

SYNERGISTIC TETRODOTOXIN FORMULATIONS AND METHODS OF TREATMENT
FOR NEUROPATHIC PAIN
STATEMENT OF RELATED APPLICATIONS
[001] This application claims priority to U S. Provisional Application No. 62/675,462, filed on May 23, 2018, which is incorporated by reference herein in its entirety.
FIELD OF THE INVENTION
[002] The invention described herein relates generally to the use of a sodium channel blocker, for example tetrodotoxin or its analogs and/or derivatives as well as their pharmaceutically acceptable salts, as a medicament for the treatment of neuropathic pain in combination with other compounds. More particularly, but not exclusively, to use such a medicament in synergistic doses to ameliorate pain.
BACKGROUND OF THE INVENTION
[003] Tetrodotoxin is a marine organic toxin which is mainly found in testis, ovaries, eggs, liver, spleen, gastrointestinal tract, skin, muscle, eyes, and blood of puffer fish. Tetrodotoxin is also found in several diverse animal species, including goby fish, some newts and frogs, the blue ringed octopus, and even in some marine alga. See, e.g., Bane et al , Tetrodotoxin: Chemistry, Toxicity, Source, Distribution and Detection, Toxins 6:693-755 (2014).
Tetrodotoxin (hereinafter“TTX”) is an alkaloid found in puffer fish ( Tetradontidae ). The chemical name of TTX is [4R-(4a,4aa,5a,7a,9a, 10a, 10a ,1 1 S*, 12S*)]-Octahydro-12- (hydroxymethyl)-2-imino-5, 9:7, 10a-dimethano- 10aH-[1 ,3]dioxocino[6,5-d]pyrimidine- 4,7, 10, 1 1 , 12-pentol with a molecular formula CiiHi7N308 and a molecular weight of 319.27 Da. It is a potent non-protein, small molecule neurotoxin and an indispensable tool for the study of neurobiology and physiology.
[004] The treatment of pain conditions is of great importance in medicine. Currently, there is a world-wide need for additional pain therapies. The pressing requirement for a specific treatment of pain conditions or a treatment of specific pain conditions which is tailored to the patient, which is to be understood as the successful and satisfactory treatment of pain for the patients, is documented in the large number of scientific works which have recently and over the years appeared in the field of applied analgesics or on basic research on nociception. SUMMARY OF THE INVENTION
[005] The present invention is directed to compositions and methods useful to treat pain in a subject mammal. An aspect of the present invention are synergistic combinations of a sodium channel blocker, for example tetrodotoxin, and modulators of synaptic signaling, for example, antidepressants, selective serotonin reuptake inhibitors, mixed serotonin and norepinephrine reuptake inhibitors, or norepinephrine reuptake inhibitors. Other aspects of the present invention are methods of using such synergistic combinations to treat pain.
[006] For example, a composition for the treatment of pain includes: (a) an amount of tetrodotoxin; and (b) an amount of either a serotonin-norepinephrine reuptake inhibitor (SNRI), selective serotonin reuptake inhibitor (SSRI), or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat pain.
[007] Another embodiment is a composition for the treatment of neuropathic pain includes: (a) an amount of tetrodotoxin; and (b) an amount of any one of duloxetine and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat neuropathic pain.
[008] In another embodiment, the present invention comprises a method of treating neuropathic pain in a mammal in need thereof, includes: (a) administering an amount of tetrodotoxin; and (b) administering an amount of either a serotonin-norepinephrine reuptake inhibitor (SNRI), a selective serotonin reuptake inhibitor (SSRI) or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and reboxetine exhibit synergism to treat neuropathic pain. In another embodiment, the present invention comprises a method of treating neuropathic pain in a mammal in need thereof, includes: (a) administering an amount of tetrodotoxin; and (b) administering an amount of either a serotonin-norepinephrine reuptake inhibitor (SNRI), a selective serotonin reuptake inhibitor (SSRI) or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and duloxetine exhibit synergism to treat neuropathic pain.
[009] Another embodiment is a composition for the treatment of nociceptive pain includes:
(a) an amount of tetrodotoxin; and (b) an amount of any one of duloxetine and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat nociceptive pain.
[0010] In another embodiment, the present invention comprises a method of treating nociceptive pain in a mammal in need thereof, includes: (a) administering an amount of tetrodotoxin; and (b) administering an amount of either a serotonin-norepinephrine reuptake inhibitor (SNRI), a selective serotonin reuptake inhibitor (SSRI) or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and reboxetine exhibit synergism to treat nociceptive pain. In another embodiment, the present invention comprises a method of treating nociceptive pain in a mammal in need thereof, includes: (a) administering an amount of tetrodotoxin; and (b) administering an amount of either a serotonin-norepinephrine reuptake inhibitor (SNRI), a selective serotonin reuptake inhibitor (SSRI) or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and duloxetine exhibit synergism to treat nociceptive pain.
[0011] These and other features, aspects, and advantages of the present invention will become better understood with reference to the following description, appended claims, and accompanying drawings.
BRIEF DESCRIPTION OF THE DRAWINGS
[0012] The foregoing summary, as well as the following detailed description of the invention, will be better understood when read in conjunction with the appended drawings.
[0013] Figure 1 shows the effects of TTX at 1 , 2 and 3 pg/kg on baseline Paw Withdrawal Test (PWT) (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0, D4-0, D5-0, D7, D10, D14: baseline value for the 2nd, 3rd, 4th, 5th, 7th, 10th, and 14th days after the start of dosing course; 1 h, 2 h: 1 hour and 2 hours after each dosing. This figure accompanies Example 1 below.
[0014] Figure 2 shows the effects of duloxetine at 5, 10 and 15 mg/kg on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin- induced neuropathy. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1 -0: pre-dosing control; D2-0, D3-0, D4-0, D5-0, D7, D10, D14: baseline value for the 2nd, 3rd, 4th, 5th, 7th, 10th, and 14th days after the start of dosing course; 1 h, 2 h: post-dosing value for 1 hour and 2 hours after each dosing.
[0015] Figure 3 shows the effects of Reboxetine at 1 , 3 and 5 mg/kg on baseline PWT (A) and baseline PWT, PWT at 1 and 2 hours after first dosing (B) in rats with oxaliplatin- induced neuropathy. The dosing schedule lasted for 5 days. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0, D4-0, D5-0, D7, D10, D14: baseline value for the 2nd, 3rd, 4th, 5th, 7th, 10th, and 14th days after the start of dosing course; 1 h, 2 h: post-dosing value for 1 hour and 2 hours after each dosing. This figure accompanies Example 1 below.
[0016] Figure 4 shows the effects of the combination of TTX and Duloxetine on baseline PWT (A) and baseline PWT, PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin- induced neuropathy. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1 -0: pre-dosing control; D2-0, D3-0, D4-0, D5-0, D7, D10, D14: baseline value for the 2nd, 3rd, 4th, 5th, 7th, 10th, and 14th days after the start of dosing course; 1 h, 2 h: post-dosing value for 1 hour and 2 hours after each dosing. This figure accompanies Example 1 below.
[0017] Figure 5 shows the effects of the combination of TTX and Reboxetine on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0, D4-0, D5-0, D7, D10, D14: baseline value for the 2nd, 3rd, 4th, 5th, 7th, 10th, and 14th days after the start of dosing course; 1 h, 2 h: post-dosing value for 1 hour and 2 hours after each dosing. This figure accompanies Example 1 below.
[0018] Figure 6, shows the effect of 3 pg/kg TTX and 2 mg/kg Reboxetine, administered alone and in combination for 5 days, on baseline PWT (A), and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0, D4-0, D5-0, D7, D10, D14: baseline value for the 2nd, 3rd, 4th, 5th, 7th, 10th, and 14th days after the start of the course of dosing; 1 h, 2 h: 1 hour and 2 hours after each dosing. N=2 for each group.
[0019] Figure 7, shows the effect of 3 pg/kg TTX and 10 mg/kg duloxetine, administered alone and in combination, for 14 days, on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0. ... D14-0, ... , D27-0: Baseline value for the 2nd, 3rd, ... 14th, ... 27th days after the start of the course of dosing;
1 h, 2 h: 1 hour and 2 hours after each dosing. N=2 for each group.
[0020] Figure 8, shows the effect of 3 pg/kg TTX and 2 mg/kg reboxetine, administered alone and in combination, for 14 days, on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0. ... D14-0, ... , D27-0: Baseline value for the 2nd, 3rd, ... 14th, ... 27th days after the start of the course of dosing;
1 h, 2 h: 1 hour and 2 hours after each dosing. N=2 for each group.
[0021] Figure 9, shows the effect of 5 pg/kg TTX, administered alone and in combination with 5 mg/kg Reboxetine or 15 mg/kg Duloxetine for 5 days, on baseline PWT (A) and baseline PWT, PWT at 1 and 2 hours after first dosing (B) in rats with oxaliplatin-induced neuropathy. The dosing schedule lasted for 5 days. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0, D4-0, D5-0, D7, D10, D14: baseline value for the 2nd, 3rd, 4th, 5th, 7th, 10th, and 14th days after the start of the course of dosing; 1 h,
2 h: post-dosing value for 1 hour and 2 hours after each dosing. N=2 for each group.
[0022] Figure 10, shows the effect of 5 pg/kg TTX and 15 mg/kg duloxetine, administered alone and in combination, for 14 days, on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0. ... D14-0. D27-0:
Baseline value for the 2nd, 3rd, ... 14th, ... 27th days after the start of the course of dosing;
1 h, 2 h: 1 hour and 2 hours after each dosing. N=2 for each group.
[0023] Figure 1 1 , shows the effect of 5 pg/kg TTX and 5 mg/kg reboxetine, administered alone and in combination, for 14 days, on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0. ... D14-0, .... D27-0: Baseline value for the 2nd, 3rd, ... 14th, ... 27th days after the start of the course of dosing;
1 h, 2 h: 1 hour and 2 hours after each dosing. N=2 for each group.
[0024] Figure 12 depicts the dose-effect relationships, as estimated using a hyperbolic model, for each of the three drugs administered alone. Figure 12A shows the dose-effect relationship for TTX alone. Figure 12B shows the dose-effect relationship for reboxetine alone. Figure 12C shows the dose-effect relationship for duloxetine alone. The hyperbolic curved lines in each panel represent the dose response curves. The dots represent the mean % PE (over the post-baseline period) for each rat.
[0025] Figure 13 shows the performance of the reboxetine/TTX drug combination, compared with the TTX alone dose effect curve. For the high dose combination (5:5, equivalent to 1 1 .8 pg/kg TTX) the observed effect is much higher above the curve, signifying strong synergism. The interaction index of 0.469 indicates the combination took less than half of the dose (if using TTX alone) to achieve the same response. That is, using TTX alone, a dose of 25.1 pg/kg would be required to achieve the same MPE of 66.5%.
[0026] Figure 14 shows the performance of duloxetine/TTX drug combination, compared with the TTX alone dose effect curve.
[0027] Figure 15 show response surface plots produced with the methods of Tallarida.
These methods were used to represent the combination data as 3-D plots. This response surface plots shows the expected responses under additive assumption and the actual (observed) responses from the drug combinations.
[0028] Figure 16 shows the Tallarida response surface plots depicting the expected responses under additive assumption and the actual (observed) responses. This figure shows the performance of the TTX/duloxetine combination.
[0029] Figure 17 shows the effects of 3 pg/kg TTX and Duloxetine administered alone and in combination for 14 days on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy. Pre1 , Pre2, Pre3: 1 st, 2nd and 3rd days before first oxaliplatin injection; W1 , W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0 ... D14-0 ... D27: baseline value for the 2nd, 3rd... 14th, and 27th days after the start of dosing course; 1 h, 2 h: 1 hour and 2 hours after each dosing. N=7 for each group.
[0030] Figure 18 shows the effects of 3 pg/kg or 5 pg/kg TTX and varying amounts of Reboxetine administered alone and in combination for 14 days on baseline PWT (A) and baseline PWT and PWT at 1 and 2 hours after dosing (B) in rats with oxaliplatin-induced neuropathy. Pre1 , Pre2, Pre3: 1st, 2nd and 3rd days before first oxaliplatin injection; W1 ,
W2: 1 st and 2nd week after first oxaliplatin injection, respectively; D1-0: pre-dosing control; D2-0, D3-0 ... D14-0 ... D27: baseline value for the 2nd, 3rd... 14th, and 27th days after the start of dosing course; 1 h, 2 h: 1 hour and 2 hours after each dosing n = 8 for TTX 5 pg/kg + Reboxetine 5 mg/kg group, 7 for any other groups.
[0031] Figure 19 depicts the dose response curves, as estimated using a hyperbolic model, at Day 5 Hour 1 , for each of the three drugs administered alone. Figure 19A shows the dose-effect relationship for TTX alone. Figure 19B shows the dose-effect relationship for reboxetine alone. Figure 19C shows the dose-effect relationship for duloxetine alone. The hyperbolic curved lines in each panel represent the dose response curves. The dots represent the mean %MPE (over the post-baseline period) for each rat. [0032] Figure 20 shows response surface plots depicting the expected responses under additive assumption and the actual (observed) responses from the drug combinations, produced with the methods of Tallarida. Figure 20A shows the expected responses under additive assumption and the actual (observed) responses from the drug combinations of TTX administered in combination with Reboxetine. Figure 20B shows the the expected responses under additive assumption and the actual (observed) responses from the drug combinations of TTX administered in combination with Duloxetine. A point above the surface represent the existence of synergism in the drug combination. The strong performance of TTX 5 pg/kg + Duloxetine 15 mg/kg is clearly noticeable.
[0033] Figure 21 depicts the dose response curves, as estimated using a hyperbolic model, at Day 14 Hour 1 , for each of the three drugs administered alone. Figure 21 A shows the dose-effect relationship for TTX alone. Figure 21 B shows the dose-effect relationship for reboxetine alone. Figure 21 C shows the dose-effect relationship for duloxetine alone. The hyperbolic curved lines in each panel represent the dose response curves. The dots represent the mean %MPE (over the post-baseline period) for each rat.
[0034] Figure 22 shows the Tallarida response surface plots depicting the expected responses under additive assumption and the actual (observed) responses from the drug combinations. Figure 22A shows the expected responses under additive assumption and the actual (observed) responses from the drug combinations of TTX administered in combination with Reboxetine. Figure 22B shows the the expected responses under additive assumption and the actual (observed) responses from the drug combinations of TTX administered in combination with Duloxetine. The strong performance of TTX 5 pg/kg + Reboxetine 5 mg/kg combination, TTX 3 pg/kg + Duloxetine 10 mg/kg, and TTX 5 pg/kg + Duloxetine 15 mg/kg are clearly seen.
DETAILED DESCRIPTION OF THE INVENTION
[0035] In the Summary of the Invention above and in the Detailed Description of the Invention, and in the claims below, and in the accompanying drawings, reference is made particular compositions, features and method steps of the invention. It is to be understood that the disclosure of the invention in this specification includes all possible combinations of such compositions, features and method steps. For example, where a particular feature is disclosed in the context of a particular aspect or embodiment of the invention, or a particular claim, that feature can also be used, to the extent possible, in combination with and/or in the context of the other particular aspects and embodiments of the invention, and in the invention generally. [0036] Each value referenced herein is understood to be the value as referenced or a value about the value referenced. “About” as used herein is understood to encompass values that are within +/- 10% of the value referenced.
[0037] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of skill in the art to which this invention belongs. All patents and publications referred to herein are incorporated by reference in their entireties, with particular aspects highlighted.
Synergism
[0038] Drug synergy describes the total effect of two drugs in combination. When the combined effect of a drug combination is greater than predicted (additive effect) by their individual potencies, it is called super-additive; when the combination is said to be synergistic (enhanced effect more than mere additivity). Synergy or synergism is a well- known pharmacological phenomenon with specific quantitative techniques to identify, explore, and characterize relationships between two different drugs. R. J. Tallarida discusses various methods to identify and characterize synergy between two drugs in“Drug Synergism: Its Detection and Applications,” Perspectives in Pharmacology, 298: 865-872 (2001). The methodical approaches discussed therein, in particular, the production and analysis of response surfaces to analyze dose/effect of two drugs is incorporated by reference in its entirety. Further fundamental quantitative analysis, experimental design, and theoretical modeling methods are discussed in depth by Ting-Chao Chou in“Theoretical Basis, Experimental Design, and Computerized Simulation of Synergism and Antagonism in Drug Combination Studies," Pharmacological Reviews 58: 621 -681 (2006). Chou is incorporated by reference in its entirety, particularly for experimental design and analysis of data where drug combinations are in constant ratios, non-constant ratios, and methods for determining optimal combination ratios of two drugs. Additional analytical methods, experimental methods, and general approaches are found, for example, in the following: Tallarida,“Statistical Analysis of Drug Combinations for Synergism,” Pain 49:93-97 (1992); Tallarida,“Response Surface Analysis of Synergism Between Morphine and Clonidine,” Journal of Pharmacology and Experimental Therapeutics 289:8-13 (1999); Tallarida, “Quantitative Methods for Assessing Drug Synergism,” Genes and Cancer 2:1003-1008 (201 1). It is well known to the skilled artisan that a combination of two drugs may show synergism, additive effects, or antagonism. Synergism is also referred to as positive synergy by some artisans. Likewise, antagonism may be referred to as negative synergy by some artisans. Compositions for Treating Pain
[0039] In an embodiment, a composition for the treatment of pain comprising: (a) an amount of tetrodotoxin; and (b) an amount of either a selective serotonin reuptake inhibitor (SSRI), a serotonin-norepinephrine reuptake inhibitor (SNRI), or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat pain.
[0040] In another embodiment, a composition for the treatment of neuropathic pain comprises: (a) an amount of tetrodotoxin; and (b) an amount of any one of paroxetine, fluoxetine, duloxetine, and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat neuropathic pain.
[0041] In another embodiment, a composition for the treatment of nociceptive pain comprises: (a) an amount of tetrodotoxin; and (b) an amount of any one of paroxetine, fluoxetine, duloxetine, and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat nociceptive pain.
Tetrodotoxin (TTX)
[0042] TTX is a well-known compound described for example in WO02/22129 as systemically acting as analgesic. Among the publications describing TTX are, for example, Tu, Anthony (Ed.) Handbook of Natural Toxins, Vol. 3: Marine Toxins and Venoms, 1988, 185-210, Kao (1966), Pharmacol. Rev. 18:997-1049, and Bane et al., Tetrodotoxin:
Chemistry, Toxicity, Source, Distribution and Detection, Toxins 6:693-755 (2014).
[0043] Several processes for producing TTX are known. Usually TTX is extracted from marine organisms (e.g. JP 270719 Goto and Takahashi), but numerous other methods of preparation or synthesis are also described (and used for the preparation of TTX in connection to this invention) in U.S. Pat. No. 6,552, 191 , U.S. Pat. No. 6,478,966, U.S. Pat. No. 6,562,968 or 2002/0086997, all of which are included here by reference. Each of the preceding patents are incorporated by reference in their entirety and in particular for the methods of isolation, synthesis, and purification of TTX and TTX analogs.
[0044] It is surprising that administration of tetrodotoxin, a sodium channel blocker, is highly effective for the treatment for pain, including neuropathic pain/allodynia/hyperalgesia.
[0045] The present invention relates to the use of sodium channel blockers. In one embodiment the sodium channel blocker is tetrodotoxin, its analogs/derivatives, optionally in the form of its racemate, pure stereoisomers, especially enantiomers or diastereomers or in the form of mixtures of stereoisomers, in any suitable mixing ratio; in neutral form, in the form of an acid or base or in the form of a salt, in particular a physiologically acceptable salt, or in the form of a solvate, especially a hydrate for the production of a medicament for the treatment of pain.
[0046] In some embodiments, the sodium channel blocker is an analog of TTX. In some embodiments, an analog may be one disclosed in Lin, U.S. Pat. No. 8,486,901 , which is incorporated by reference in its entirety, particularly for the identity, composition of matter, purification, and characterization of TTX analogs. Further embodiments may include a TTX analog as disclosed in Buschmann et al., U.S. Pat. No. 9,018,222, which is incorporated by reference in its entirety, particularly for the identity and composition of matter of various TTX analogs.
Serotonin-Noepinephrine Reuptake Inhibitors (SNRIs)
[0047] Any serotonin-norepinephrine reuptake inhibitor (SNRI) can be used in the present compositions and methods. In an embodiment, the SNRI is Duloxetine.
[0048] Duloxetine, molecular formula C-isH-igNOS, is a commercially available central nervous system penetrant serotonin-norepinephrine reuptake inhibitor. Duloxetine also has some effect on the reuptake of norepinephrine. Duloxetine is (+)-N-methyl-3-(1- naphthalenyloxy)-3-(2-thienyl)propanamine hydrochloride. Duloxetine was first taught by U.S. Patent 4,956,388, which discloses its high potency serotonin reuptake inhibition as well as its inhibition of norepinephrine uptake. U.S. Patent 4,956,388 is incorporated by reference in its entirety, in particular for the method of synthesis and purification of duloxetine. Herein,“duloxetine” is used to refer to the free base of the molecule as well as any salt, solvate, crystalline or amorphous form of the compound as well as commercially formulated and commercially sold unit doses of the molecule. The U.S. Food and Drug Administration marketing label for Duloxetine Delayed-Release Capsules, marketed in the U.S. under the trade name Cymbalta®, is incorporated by reference in its entirety, and particularly for the formulation and dosing information therein.
[0049] Duloxetine, C-ieHigNOS
Figure imgf000012_0001
Norepinephrine Reuptake Inhibitors (NRIs)
[0050] Any norepinephrine reuptake inhibitor (NRI) can be used in the present compositions and methods. In an embodiment the NRI is reboxetine.
[0051] Reboxetine is a commercially available central nervous system penetrant selective norepinephrine reuptake inhibitor (NRI). Reboxetine has the molecular formula C19H23NO3. Reboxetine is also known as 2-[(2-ethoxyphenoxy)phenylmethyl]-, (R*,R*)-Morpholine and (S*,S*) form, and mixtures thereof. Herein,“reboxetine” is used to refer to the free base of the molecule as well as any salt, solvate, crystalline or amorphous form of the compound, whether sterochemically pure or a mixture, and commercially formulated and commercially sold unit doses of the molecules. The European Medicines Agency Summary of Product Characteristics for reboxetine, marketed as Edronax® in Europe, is incorporated by reference in its entirety, in particular for its formulation and dosing information.
[0052] Reboxetine, C19H23NO3
Figure imgf000013_0001
Compositions
[0053] In an embodiment, the composition for the treatment of pain includes: (a) an amount of tetrodotoxin; and (b) an amount of either a selective serotonin reuptake inhibitor (SSRI), serotonin-norepinephrine reuptake inhibitor (SNRI), or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat pain. In an embodiment, the composition includes a singly therapeutically effective amount of tetrodotoxin or a singly subtherapeutic amount of tetrodotoxin. In an embodiment, the amount of component (b) is a singly therapeutically effective amount of component (b) or a singly subtherapeutic amount of component (b). In an embodiment, component (b) is an SSRI. In an embodiment, component (b) is a mixed SSRI/NRI. In an embodiment, component (b) is an NRI. In an embodiment, component (b) is one or more of a SSRI, mixed SSRI/NRI, and NRI. In an embodiment, component (b) is an SNRI. In an embodiment, component (b) is a mixed SNRI/NRI.
[0054] In an embodiment, component (a) and component (b) are formulated in the same pharmaceutically acceptable carrier. In an embodiment, component (a) and component (b) are formulated in the same pharmaceutically acceptable carrier and administered simultaneously via the same route of administration. In an embodiment, component (a) and component (b) are formulated in the same pharmaceutically acceptable carrier and form a unit dose.
[0055] In an embodiment, component (a) and component (b) are formulated in different pharmaceutically acceptable carriers. In an embodiment, component (a) and component (b) are formulated in different pharmaceutically acceptable carriers and administered via different routes of administration. In an embodiment, component (a) and component (b) are formulated in different pharmaceutically acceptable carriers and component (a) is administered subcutaneously and component (b) is administered orally. In an embodiment, component (a) and component (b) are formulated in different pharmaceutically acceptable carriers and component (a) is administered subcutaneously, intramuscularly, or
intravenously; and component (b) is administered orally. In an embodiment, component (a) and component (b) are formulated in different pharmaceutically acceptable carriers and component (a) is administered subcutaneously, intramuscularly, or intravenously; and component (b) is separately administered subcutaneously, intramuscularly, or intravenously.
[0056] In some embodiments, the formulation may include a second active pharmaceutical ingredient. The second active pharmaceutical ingredient may be an analgesic present in a therapeutically effective amount to ameliorate pain in conjunction with an embodiment of the present compositions or used in an embodiment of the present methods. Optionally, the embodied compositions may be co-administered with an analgesic; the analgesic being present at a therapeutically effective amount.
Dosages and Dosing Regimens
[0057] In one embodiment, a composition for the treatment of pain includes: (a) an amount of tetrodotoxin; and (b) an amount of either a selective serotonin reuptake inhibitor (SSRI) , serotonin-norepinephrine reuptake inhibitor (SNRI), or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat pain. TTX Doses
[0058] The amounts of TTX administered will be dependent on the human or mammal being treated, the severity of the disorder or condition, the rate of administration, the disposition of the compounds and the discretion of the prescribing physician. However, an effective dosage of each is in the range of 3.0 pg to about 150 pg of TTX per day, in single or divided doses. In some instances, dosage levels below the lower limit of the aforesaid range may be more than adequate, while in other cases still larger doses may be employed without causing any harmful side effect, e.g. , by dividing such larger doses into several small doses for administration throughout the day.
[0059] In another embodiment, TTX is administered in multiple doses. Dosing may be at least once, twice, three times, four times, five times, six times, or more than six times per day. Dosing may be every day for one, two, three, four, or more consecutive days with this regimen repeating.
[0060] In some embodiments, a daily dosage of TTX or an analog thereof is in the range of 3.0 pg to about 150 pg, about 144 pg, in the range of about 3.0 pg to about 50 pg, about 44 pg, in the range of about 3.0 pg to about 30 pg, in the range of about 3.0 pg to about 15 pg, in the range of about 3.0 pg to 7.5 pg, or about 3.0 pg.
[0061] In further embodiments, a daily dosage of TTX or an analog thereof is a mammal equivalent of a rat dosage, such that a daily dosage of TTX or an analog thereof is 3.0 pg/kg or 5.0 pg/kg. In some embodiments, the daily dosage in an adult human patient may be between about 0.5 to about 1 .5 of the mammal equivalent of a rat dosage, between about 0.75 to about 1.25 of the mammal equivalent of a rat dosage, or between about 0.9 to about 1 .1 of the mammal equivalent of a rat dosage. In other embodiments, the daily dosage in an adult human patient may be about 0.5, about 0.55, about 0.6, about 0.65, about 0.7, about 0.75, about 0.8, about 0.85, about 0.9, about 0.95, about 1 .0, about 1 .05, about 1.1 , about 1 .15, about 1.2, about 1 .25, about 1.3, about 1.35, about 1 .4, about 1 .45, or about 1.5 of the mammal equivalent of a rat dosage.
[0062] Extrapolating from the mammal equivalent of a rat dosage and assuming an adult human patient having a weight of 55 kg, in some embodiments a daily dosage of TTX or an analog thereof is in the range of 148.5 pg to about 181.5 pg, 123.75 pg to about 206.25 pg, 82.5 pg to about 247.5 pg, about 82.5 pg, about 90.75 pg, about 99.0 pg, about 107.25 pg, about 1 15.5 pg, about 123.75 pg, about 132.0 pg, about 140.25 pg, about 148.5 pg, about 156.75 pg, about 165 pg, about 173.25 pg, about 181.5 pg, about 189.75 pg, about 198.0 pg, about 206.25 pg, about 214.5 pg, about 222.75 pg, about 231.0 pg, about 239.25 pg, or about 247.5 pg. In other embodiments, a daily dosage of TTX or an analog thereof is in the range of 150 pg to about 180 pg, 125 pg to about 205 mg, 85 mg to about 250 mg, about 85 mg, about 90 mg, about 100 mg, about 105 mg, about 1 15 mg, about 125 mg, about 130 mg, about 140 mg, about 150 mg, about 155 mg, about 175 mg, about 180 mg, about 190 mg, about 200 mg, about 205 mg, about 215 mg, about 225 mg, about 230 mg, about 240 mg, or about 250 mg. In preferred embodiments, the daily dosage of TTX or an analog thereof is about 165 pg.
[0063] Extrapolating from the mammal equivalent of a rat dosage and assuming an adult human patient having a weight of 55 kg, in other embodiments a daily dosage of TTX or an analog thereof is in the range of 137.5 pg to about 412.5 pg, 206.25 pg to about 343.75 pg, 247.5 pg to about 302.5 pg, about 137.5 pg, about 151.25 pg, about 165.0 pg, about 178.75 pg, about 192.5 pg, about 206.25 pg, about 220.0 pg, about 233.75 pg, about 247.5 pg, about 261.25 pg, about 275.0 pg, about 288.75 pg, about 302.5 pg, about 316.25 pg, about 330.0 pg, about 343.75 pg, about 357.5 pg, about 371 .25 pg, about 385.0 pg, about 398.75 pg, or about 412.5 pg. In further embodiments, a daily dosage of TTX or an analog thereof is in the range of 140 pg to about 415 pg, 205 pg to about 345 pg, 250 pg to about 305 pg, about 140 pg, about 150 pg, about 180 pg, about 195 pg, about 205 pg, about 235 pg, about 250 pg, about 260 pg, about 290 pg, about 305 pg, about 315 pg, about 345 pg, about 360 pg, about 370 pg, about 400 pg, or about 415 pg. In preferred embodiments, the daily dosage of TTX or an analog thereof is about 275 pg.
[0064] In an embodiment, the amount of TTX is less than an individually therapeutic amount. In an embodiment, the amount of TTX is less than an individually effective amount. In an embodiment, the amount of TTX is a subtherapeutic amount. The doses above are understood to be therapeutically effective amounts of TTX.
[0065] In an embodiment, the total daily dose of TTX is a therapeutically effective amount. In a further embodiment, the total daily dose of TTX is a therapeutically effective amount administered subcutaneously, orally, intramuscularly, intravenously, intranasally, or transdermally.
[0066] In an embodiment, the total daily dose of TTX is a subtherapeutic amount. In an embodiment, the total daily dose is 95% of a therapeutically effective amount, 90% of a therapeutically effective amount, 85% of a therapeutically effective amount, 80% of a therapeutically effective amount, 75% of a therapeutically effective amount, 70% of a therapeutically effective amount, 65% of a therapeutically effective amount, 60% of a therapeutically effective amount, 55% of a therapeutically effective amount, 50% of a therapeutically effective amount, 45% of a therapeutically effective amount, 40% of a therapeutically effective amount, 35% of a therapeutically effective amount, 30% of a therapeutically effective amount, 25% of a therapeutically effective amount, 20% of a therapeutically effective amount, 18% of a therapeutically effective amount, 15% of a therapeutically effective amount, 10% of a therapeutically effective amount, or 5% of a therapeutically effective amount.
[0067] In some instances, dosage levels below the lower limit of the aforesaid ranges may be more than adequate, while in other cases still larger doses may be employed without causing any harmful side effect - for example, by dividing such larger doses into several small doses for administration throughout the day.
SNRI Doses
[0068] In an embodiment, the SNRI is duloxetine. In some embodiments, duloxetine is administered orally. In some embodiments, duloxetine is administered parenterally. In some embodiments, duloxetine is administered transdermally. In some embodiments, duloxetine is administered subcutaneously. In some embodiments, duloxetine is administered subcutaneously in the same pharmaceutically acceptable carrier as TTX.
[0069] In an embodiment, duloxetine is administered once per day. In an embodiment the once daily dose is 20 mg. In other embodiments the once daily dose is 30 mg. In other embodiments the once daily dose is 60 mg. In yet further embodiments, duloxetine is administered in divided doses throughout a single day. In an embodiment, the total daily dose is 20 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg, 80 mg, 90 mg, 100 mg, 1 10 mg, or 120 mg. In an embodiment, the mammal is given the mammal equivalent of a rat dosage of about 1 mg/kg to 50 mg/kg.
[0070] In other embodiments, a daily dosage of duloxetine is a mammal equivalent of a rat dosage, such that a daily dosage of duloxetine is 10 mg/kg or 15 mg/kg. In some embodiments, the daily dosage in an adult human patient may be between about 0.5 to about 1.5 of the mammal equivalent of a rat dosage, between about 0.75 to about 1 .25 of the mammal equivalent of a rat dosage, or between about 0.9 to about 1 .1 of the mammal equivalent of a rat dosage. In other embodiments, the daily dosage in an adult human patient may be about 0.5, about 0.55, about 0.6, about 0.65, about 0.7, about 0.75, about 0.8, about 0.85, about 0.9, about 0.95, about 1.0, about 1.05, about 1 .1 , about 1.15, about 1 .2, about 1.25, about 1 .3, about 1 .35, about 1.4, about 1.45, or about 1.5 of the mammal equivalent of a rat dosage.
[0071] Extrapolating from the mammal equivalent of a rat dosage and assuming an adult human patient having a weight of 55 kg, in some embodiments a daily dosage of duloxetine is in the range of 275 g to about 825 mg, 412.5 mg to about 687.5 mg, 495 mg to about 605 mg, about 275 mg, about 302.5 mg, about 330 mg, about 357.5 mg, about 385 mg, about 412.5 mg, about 440 mg, about 467.5 mg, about 495 mg, about 522.5 mg, about 550 mg, about 577.5 mg, about 605 mg, about 632.5 mg, about 660 mg, about 687.5 mg, about 715 mg, about 742.5 mg, about 770 mg, about 797.5 mg, or about 825 mg. In other embodiments, a daily dosage of duloxetine is in the range of 415 mg to about 690 mg, about 305 mg, about 360 mg, about 415 mg, about 470 mg, about 525 mg, about 580 mg, about 635 mg, about 690 mg, about 745 mg, or about 800 mg. In preferred embodiments, the daily dosage of duloxetine is about 550 mg.
[0072] Extrapolating from the mammal equivalent of a rat dosage and assuming an adult human patient having a weight of 55 kg, in other embodiments a daily dosage of duloxetine is in the range of 412.5 mg to about 1 .2375 g, 618.75 mg to about 1.03125 g, 742.5 mg to about 907.5 mg, about 412.5 mg, about 453.75 mg, about 495 mg, about 536.25 mg, about 577.5 mg, about 618.75 mg, about 660 mg, about 701.25 mg, about 742.5 mg, about 783.75 mg, about 825 mg, about 866.25 mg, about 907.5 mg, about 948.75 mg, about 990 mg, about 1.03125 g, about 1.0725 g, about 1.1 1375 g, about 1.155 g, about 1 .19625 g, or about 1 .2375 g. In still further embodiments, a daily dosage of duloxetine is in the range of 415 mg to about 1.240 g, 620 mg to about 1.030 g, 745 mg to about 910 mg, about 415 mg, about
455 mg, about 535 mg, about 580 mg, about 620 mg, about 700 mg, about 745 mg, about
785 mg, about 865 mg, about 910 mg, about 950 mg, about 1 .030 g, about 1.075 g, about
1 .1 15 g, about 1.195 g, or about 1.240 g. In preferred embodiments, the daily dosage of duloxetine is about 825 mg.
[0073] In some embodiments, duloxetine is given once per day. In some embodiments, reboxetine is given twice per day. In some embodiments the dose is oral. In other embodiments the dose may be administered via the routes of administration as described below. The doses above are understood to be therapeutically effective amounts of duloxetine.
[0074] In an embodiment, the total daily dose of duloxetine is from about 5 mg to 120 mg. In an embodiment, the total daily doses of duloxetine are selected from the group consisting of about 5 mg, about 10 mg, about 20 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 1 10 mg, and about 120 mg. In an embodiment, the total daily dose of duloxetine is in the range of about 1 mg to 120 mg. The doses above are understood to be therapeutically effective amounts of duloxetine.
[0075] An amount of TTX and an amount of a serotonin-norepinephrine reuptake inhibitor may be given by the same route of administration or a different route of administration. An amount of TTX and an amount of a serotonin-norepinephrine reuptake inhibitor may be given by the same route of administration or a different route of administration. In an embodiment, the amount of TTX administered is not a therapeutically effective amount of TTX when administered alone. In an embodiment, the amount of TTX administered is a therapeutically effective amount of TTX when administered alone.
[0076] In an embodiment, the amount of an SNRI administered is not a therapeutically effective amount of the SNRI when administered alone. In an embodiment, the amount of an SNRI administered is a therapeutically effective amount of the SNRI when administered alone.
[0077] In an embodiment, the total daily dose is subtherapeutic for duloxetine alone. In an embodiment, the total daily dose is 95% of a therapeutically effective amount, 90% of a therapeutically effective amount, 85% of a therapeutically effective amount, 80% of a therapeutically effective amount, 75% of a therapeutically effective amount, 70% of a therapeutically effective amount, 65% of a therapeutically effective amount, 60% of a therapeutically effective amount, 55% of a therapeutically effective amount, 50% of a therapeutically effective amount, 45% of a therapeutically effective amount, 40% of a therapeutically effective amount, 35% of a therapeutically effective amount, 30% of a therapeutically effective amount, 25% of a therapeutically effective amount, 20% of a therapeutically effective amount, 18% of a therapeutically effective amount, 15% of a therapeutically effective amount, 10% of a therapeutically effective amount, or 5% of a therapeutically effective amount.
[0078] In an embodiment the total daily dose of duloxetine is a subtherapeutic amount in the range of about 5% of a therapeutically effective amount to about 98% of a therapeutically effective amount.
NRI Doses
[0079] In an embodiment, the NRI is reboxetine. In some embodiments, reboxetine is administered orally. In some embodiments, reboxetine is administered parenterally. In some embodiments, reboxetine is administered transdermally. In some embodiments, reboxetine is administered subcutaneously. In some embodiments, reboxetine is administered subcutaneously in the same pharmaceutically acceptable carrier as TTX.
[0080] In an embodiment, the method further comprises the administration of an analgesic. In an embodiment, the composition further comprises an analgesic.
[0081] In an embodiment, reboxetine is administered once or twice per day (b.i.d.); for example, each dose 4 mg, for a total daily dose of 8 mg. In other embodiments, reboxetine is administered twice per day in evenly divided doses for a total daily dose of 10 mg. In other embodiments, reboxetine is administered twice per day in evenly divided doses for a total daily dose of 12 mg. In some embodiments, reboxetine is administered in total daily doses of 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 1 1 mg, or 12 mg. In an embodiment, the mammal is given the mammal equivalent of a rat dosage of about 1 mg/kg to 6 mg/kg.
[0082] In an embodiment, the daily dose of reboxetine is the mammal equivalent of a rat dosage, such that the daily dosage of reboxetine is 2 mg/kg or 5 mg/kg. In some embodiments, the daily dosage in an adult human patient may be between about 0.5 to about 1.5 of the mammal equivalent of a rat dosage, between about 0.75 to about 1 .25 of the mammal equivalent of a rat dosage, or between about 0.9 to about 1 .1 of the mammal equivalent of a rat dosage. In other embodiments, the daily dosage in an adult human patient may be about 0.5, about 0.55, about 0.6, about 0.65, about 0.7, about 0.75, about 0.8, about 0.85, about 0.9, about 0.95, about 1.0, about 1.05, about 1 .1 , about 1.15, about 1 .2, about 1.25, about 1 .3, about 1 .35, about 1.4, about 1.45, or about 1.5 of the mammal equivalent of a rat dosage.
[0083] Extrapolating from the mammal equivalent of a rat dosage and assuming an adult human patient having a weight of 55 kg, in some embodiments a daily dosage of reboxetine is in the range of 55 mg to about 165 mg, 82.5 mg to about 137.5 mg, 99 mg to about 121 mg, about 55 mg, about 60.5 mg, about 66 mg, about 71.5 mg, about 77 mg, about 82.5 mg, about 88 mg, about 93.5 mg, about 99 mg, about 104.5 mg, about 1 10 mg, about 1 15.5 mg, about 121 mg, about 126.5 mg, about 132 mg, about 137.5 mg, about 143 mg, about 148.5 mg, about 154 mg, about 159.5 mg, or about 165 mg. In other embodiments, a daily dosage of reboxetine is in the range of 80 mg to about 140 mg, 100 mg to about 120 mg, about 60 mg, about 65 mg, about 70 mg, about 75 mg, about 80 mg, about 85 mg, about 90 mg, about 95 mg, about 100 mg, about 105 mg, about 1 15 mg, about 120 mg, about 125 mg, about 130 mg, about 135 mg, about 140 mg, about 145 mg, about 150 mg, about 155 mg, or about 160 mg. In preferred embodiments, the daily dosage of reboxetine is about 1 10 mg.
[0084] Extrapolating from the mammal equivalent of a rat dosage and assuming an adult human patient having a weight of 55 kg, in other embodiments a daily dosage of reboxetine is in the range of 137.5 mg to about 412.5 mg, 206.25 mg to about 343.75 mg, 247.5 mg to about 302.5 mg, about 137.5 mg, about 151.25 mg, about 165.0 mg, about 178.75 mg, about 192.5 mg, about 206.25 mg, about 220.0 mg, about 233.75 mg, about 247.5 mg, about 261.25 mg, about 275.0 mg, about 288.75 mg, about 302.5 mg, about 316.25 mg, about 330.0 mg, about 343.75 mg, about 357.5 mg, about 371.25 mg, about 385.0 mg, about 398.75 mg, or about 412.5 mg. In further embodiments, a daily dosage of reboxetine is in the range of 140 mg to about 415 mg, 205 mg to about 345 mg, 250 mg to about 300 mg, about 140 mg, about 150 mg, about 180 mg, about 195 mg, about 205 mg, about 235 mg, about 250 mg, about 260 mg, about 290 mg, about 300 mg, about 315 mg, about 345 mg, about 360 mg, about 370 mg, about 400 mg, or about 415 mg. In preferred embodiments, the daily dosage of reboxetine is about 275 mg.
[0085] In some embodiments, reboxetine is given once per day. In some embodiments, reboxetine is given twice per day. In some embodiments reboxetine is administered orally.
In some embodiments, reboxetine is administered parenterally. The doses above are understood to be therapeutically effective amounts of reboxetine.
[0086] In an embodiment, reboxetine is administered at a total daily dose selected from about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 6 mg, about 7 mg, about 8 mg, about 9 mg, about 10 mg, about 1 1 mg, about 12 mg, about 13 mg, about 14 mg, about 15 mg. The doses above are understood to be therapeutically effective amounts of reboxetine.
[0087] In an embodiment, reboxetine is administered orally and the total daily dose is subtherapeutic for reboxetine alone. In an embodiment, the total daily dose is 95% of a therapeutically effective amount, 90% of a therapeutically effective amount, 85% of a therapeutically effective amount, 80% of a therapeutically effective amount, 75% of a therapeutically effective amount, 70% of a therapeutically effective amount, 65% of a therapeutically effective amount, 60% of a therapeutically effective amount, 55% of a therapeutically effective amount, 50% of a therapeutically effective amount, 45% of a therapeutically effective amount, 40% of a therapeutically effective amount, 35% of a therapeutically effective amount, 30% of a therapeutically effective amount, 25% of a therapeutically effective amount, 20% of a therapeutically effective amount, 18% of a therapeutically effective amount, 15% of a therapeutically effective amount, 10% of a therapeutically effective amount, or 5% of a therapeutically effective amount.
[0088] In an embodiment the total daily dose of reboxetine is a subtherapeutic amount in the range of about 5% of a therapeutically effective amount to about 98% of a therapeutically effective amount.
[0089] In an embodiment, the amount of a NRI administered is not a therapeutically effective amount of the NRI for analgesia when administered alone. In an embodiment, the amount of a NRI administered is a therapeutically effective amount for analgesia of the NRI when administered alone. Methods for Treating Pain
[0090] In certain aspects, the invention relates generally to methods for treating pain by administering one or more doses of a sodium channel blocker in combination with one or more selective serotonin reuptake inhibitors. In certain aspects, the invention relates generally to methods for treating pain by administering a sodium channel blocker in combination with one or more mixed serotonin and/or norepinephrine reuptake inhibitors. In certain aspects, the invention relates generally to methods for treating pain by administering a sodium channel blocker in combination with one or more serotonin-norepinephrine reuptake inhibitors. In certain aspects, the invention relates generally to methods for treating pain by administering one or more amounts of a sodium channel blocker in combination with one or more norepinephrine reuptake inhibitors. In preferred embodiments, the sodium channel blocker is TTX.
[0091] In some embodiments, the method is used to treat a subject undergoing treatment for chemotherapy induced neuropathic pain.
[0092] In an embodiment, a method for the treatment of neuropathic pain comprises: (a) an amount of tetrodotoxin; and (b) an amount of any one of paroxetine, fluoxetine, duloxetine, and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat neuropathic pain.
[0093] In an embodiment, the method for the treatment of neuropathic pain, wherein the neuropathic pain is selected from the group consisting of pain arising from diabetic peripheral neuropathy, fibromyalgia, and chemotherapy induced neuropathic pain.
[0094] In an embodiment, the method for the treatment of neuropathic pain, wherein the mammal is a human patient undergoing treatment for chemotherapy induced neuropathic pain.
[0095] In a particular embodiment, the composition or method is used to treat a subject undergoing treatment for cancer pain and wherein said subject also receives at least one low dose of an analgesic or an opioid. In some embodiments, the subject is further treated with about 500 mg morphine equivalent dose of an opioid.
[0096] In some embodiments, the method is used to treat a subject undergoing treatment for nociceptive pain.
[0097] In an embodiment, a method for the treatment of nociceptive pain comprises: (a) an amount of tetrodotoxin; and (b) an amount of any one of paroxetine, fluoxetine, duloxetine, and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat nociceptive pain.
[0098] In an embodiment, the method for the treatment of nociceptive pain, wherein the nociceptive pain is selected from the group consisting of radicular pain, somatic pain, and visceral pain.
[0099] In an embodiment, the method for the treatment of nociceptive pain, wherein the mammal is a human patient undergoing treatment for nociceptive pain.
Routes of Administration
[00100] As noted above, compositions disclosed herein may be administered
subcutaneously, orally, intramuscularly, intravenously, transdermally, topically, or via any route know to the art. Likewise, the methods of the present invention may include administration subcutaneously, orally, or any other route know to the art.
[00101] In some embodiments, the composition for the treatment of neuropathic pain comprising: (a) an amount of tetrodotoxin; and (b) an amount of any one of duloxetine and reboxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat neuropathic pain.
[00102] In some embodiments, compositions disclosed herein include a combination of (a) a dosage amount of tetrodotoxin and (b) a dosage amount of duloxetine or reboxetine, wherein component (a) and component (b) exhibit synergism to treat neuropathic and/or nociceptive pain. In one embodiment, a composition includes a combination of (a) 5 pg per kg TTX and (b) 5 mg per kg Reboxetine, wherein the 5 pg/kg TTX and 5 mg/kg Reboxetine exhibit synergism to treat neuropathic and/or nociceptive pain. In another embodiment, a composition includes a combination of (a) 3 pg per kg TTX and (b) 2 mg per kg Reboxetine, wherein the 3 pg/kg TTX and 2 mg/kg Reboxetine exhibit synergism to treat neuropathic and/or nociceptive pain. In a further embodiment, a composition includes a combination of (a) 3 pg per kg TTX and (b) 10 mg per kg Duloxetine, wherein the 3 pg/kg TTX and 10 mg/kg Duloxetine exhibit synergism to treat neuropathic and/or nociceptive pain. In yet another embodiment, a composition includes a combination of (a) 5 pg per kg TTX and (b)
15 mg per kg Duloxetine, wherein the 5 pg/kg TTX and 15 mg/kg Duloxetine exhibit synergism to treat neuropathic and/or nociceptive pain.
[00103] In some embodiments, the composition is a single formulation with both an amount of TTX and an amount of any one of paroxetine, fluoxetine, duloxetine, and reboxetine. In some embodiments such a single formulation is administered subcutaneously. In some embodiments such a single formulation is administered intramuscularly. In some embodiments such a single formulation is administered intravenously. In some
embodiments such a single formulation is administered transdermally. In some
embodiments such a single formulation is administered orally.
[00104] In one embodiment, the TTX is administered via the same route of administration as the SSRI. In one embodiment, the TTX is administered via the same route of administration as the SNRI. In one embodiment, the TTX is administered via the same route of administration as the NRI.
[00105] In one embodiment, the TTX is administered via a different route of administration as the SSRI. In one embodiment, the TTX is administered via a different route of administration as the SNRI. In one embodiment, the TTX is administered via a different route of administration as the NRI.
[00106] In some embodiments, at least one analgesic is administered via the same route of administration as the TTX. In some embodiments, at least one analgesic is administered via a different route of administration than the TTX. In some embodiments, at least one analgesic is administered via the same route of administration as the SSRI. In some embodiments, at least one analgesic is administered via the same route of administration as the SNRI. In some embodiments, at least one analgesic is administered via the same route of administration as the NRI. In some embodiments, the combination, an embodiment of the present compositions and an analgesic, is administered in a single dose. Such
administration may be by injection, e.g., intravenous injection, in order to introduce pain control quickly. However, other routes, including the preferred oral route, may be used as appropriate. A single dose of the combination of an embodiment of the present compositions and an analgesic may also be used for treatment of acute pain.
[00107] The amount of TTX and the amount of either a SSRI, SNRI or NRI may be given simultaneously or at differing times.
[00108] An amount of TTX in a first pharmaceutically acceptable formulation, the second pharmaceutically acceptable formulation including a neurotransmitter reuptake inhibitor, or a combination of TTX and a neurotransmitter reuptake inhibitor, may be administered in either single or multiple doses by any of the accepted modes of administration of agents having similar utilities, including rectal, buccal, intranasal and transdermal routes, by intra-arterial injection, intravenously, intraperitoneally, parenterally, intramuscularly, subcutaneously, orally, topically, or as an inhalant. Kits
[00109] An embodiment may be a kit comprising a unit dose of an amount of TTX and an amount of an SNRI. An embodiment may be a kit comprising a daily dose of an amount of TTX, which may be more than one unit dose, and a daily dose of an amount of an SNRI, which may be more than one unit dose.
[00110] In one embodiment the kit comprises one or more unit doses of TTX and one or more oral doses of the SNRI duloxetine. The one or more unit doses of TTX may be for subcutaneous injection, or for administration orally, intramuscularly, intravenously, intranasally, or transdermally.
[00111] An embodiment may be a kit comprising a unit dose of an amount of TTX and an amount of an NRI. An embodiment may be a kit comprising a daily dose of an amount of TTX, which may be more than one unit dose, and a daily dose of an NRI, which may be more than one unit dose.
[00112] In one embodiment, the kit comprises one or more unit doses of TTX and one or more oral doses of the NRI reboxetine. The one or more unit doses of TTX may be for subcutaneous injection, or for administration orally, intramuscularly, intravenously, intranasally, or transdermally.
[00113] An embodiment may be a kit comprising a unit dose of an amount of TTX and an amount of an SSRI. An embodiment may be a kit comprising a daily dose of an amount of TTX, which may be more than one unit dose, and a daily dose of an amount of an SSRI, which may be more than one unit dose.
[00114] In one embodiment the kit comprises one or more unit doses of TTX and one or more oral doses of an SSRI (e.g., paroxetine or fluoxetine). The one or more unit doses of TTX may be for subcutaneous injection, or for administration orally, intramuscularly, intravenously, intranasally, or transdermally.
[00115] In one embodiment, the kit comprises one or more unit doses of TTX and one or more SNRIs. In one embodiment, the kit comprises one or more unit doses of TTX, one or more SNRIs, and one or more analgesics. In one embodiment, the kit comprises one or more unit doses of TTX, an amount of at least one SNRI, and an amount of at least one NRI. In one embodiment, the kit comprises one or more unit doses of TTX, at least one SNRI, at least one NRI, and at least one analgesic. In one embodiment, the kit comprises one or more unit doses of TTX, an amount of at least one SNRI, an amount of at least one NRI , and an amount of one or more analgesics. In one embodiment, the kit comprises one or more unit doses of TTX and one or more NRIs. In one embodiment, the kit comprises one or more unit doses of TTX, one or more NRIs, and one or more analgesics. In any of the preceding embodiments, the one or more unit doses of TTX may be for subcutaneous injection, or for administration orally, intramuscularly, intravenously, intranasally, or transdermally. In preferred embodiments, the one or more unit doses of TTX is for subcutaneous injection.
Definitions
[00116] As used in this application,“tetrodotoxin” refers collectively to tetrodotoxin, its analogs, derivatives, metabolites, degradation products, hydrates thereof, salts thereof, and combinations thereof.
[00117] Synergism or drug synergy as discussed above in paragraph [0024], means that the combined effect of two drugs in combination is greater than predicted (additive effect) by their individual potencies, it is called super-additive, and the combination is said to be synergistic (i.e. enhanced effect).
[00118] The term“analogs” as used in this application is defined here as meaning a chemical compound that is a derivative of a compound which has similar biochemical activity to that compound.
[00119] The term“derivatives” as used in this application is defined here as meaning a chemical compound having undergone a chemical derivatization such as substitution or addition of a further chemical group to change (for pharmaceutical use) any of its physicochemical properties, such as solubility or bioavailability. Derivatives include so-called prodrugs, e.g. ester and ether derivatives of an active compound that yield the active compound per se after administration to a subject.
[00120] The term“analgesic” refers to a substance that acts to relieve pain. Analgesics, without limitation, may be drawn from any of the following: Cyclo-oxygenase-2 inhibitors (e.g. valdecoxib, rofecoxib, or celecoxib); antimigraine agents (e.g. sumatriptan, methylsergide maleate, frovatriptan, naratriptan, almotriptan, ergotamine, rizatriptan, zolmitriptan, or dihydroergotamine); non-steroidal anti-inflammitory agents (e.g. ibuprofen, naproxen, sulindac, ketoprofen, tolmetin, etodolac, diclofenac, flurbiprofen, ketorolac, piroxicam, indomethacin, or nabumetone); salicylates (e.g. aspirin or magnesium salicylate); or narcotic agents (e.g. morphine, oxycodone, fentanyl, oxymorphone, hydromorphone, meperidine, buprenorphine, methadone, tramadol, butorphanol, tapentadol, propoxyphene, alfentanil, liposomal morphene, sufentanil, remifentanil, or pentazocine).
[00121] The term“effective amount” or“therapeutically effective amount” refers to that amount of a compound or combination of compounds as described herein that is sufficient to effect the intended application including, but not limited to, disease treatment. A
therapeutically effective amount may vary depending upon the intended application (in vitro or in vivo), or the subject and disease condition being treated (e.g., the weight, age and gender of the subject), the severity of the disease condition, the manner of administration, etc. which can readily be determined by one of ordinary skill in the art. The term also applies to a dose that will induce a particular response in target cells (e.g., the reduction of platelet adhesion and/or cell migration). The specific dose will vary depending on the particular compounds chosen, the dosing regimen to be followed, whether the compound is administered in combination with other compounds, timing of administration, the tissue to which it is administered, and the physical delivery system in which the compound is carried.
[00122] “Subtherapeutic” doses are defined as doses that are less than doses that would be an individually effective amount or an individually therapeutically effective amount of the compound, composition, or combinations of compounds.
[00123] A“therapeutic effect” as that term is used herein, encompasses a therapeutic benefit and/or a prophylactic benefit. A prophylactic effect includes delaying or eliminating the appearance of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing, halting, or reversing the progression of a disease or condition, or any combination thereof.
[00124] The terms“QD,”“qd,” or“q.d.” mean quaque die, once a day, or once daily. The terms“BID,”“bid,” or“b.i.d.” mean bis in die, twice a day, or twice daily. The terms“TID,”
“tid,” or“t.i.d.” mean ter in die, three times a day, or three times daily. The terms“QID,”“qid,” or“q.i.d.” mean quater in die, four times a day, or four times daily.
[00125] The term“pharmaceutically acceptable formulations” refer to compositions that contain pharmaceutically acceptable salts of TTX and optionally pharmaceutically acceptable carriers and/or pharmaceutically acceptable excipients, and/or non-active ingredients known generally to the art to serve non-therapeutic purposes in medicaments.
[00126] The term“mammal equivalent dose” is used to indicate the conversion of a dose determined in one mammal species for another subject mammal species. For example, and without limitation, converting a dose determined in a rat to the equivalent human dose. The methods for the determination of such equivalent doses are known to those skilled in the art and are summarized in the U.S. Food and Drug Administration Guidance to Industry, available at https://www.fda.gov/downloads/drugs/guidances/ucm078932.pdf, entitled “Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers.” The document is incorporated by reference in its entirety, with particular attention to the method summarized in Table I.
[00127] Pharmaceutically acceptable formulations may include formulations wherein at least one component is a composition disclosed by Zhang et al. in U.S. Pat. No. 8, 124,608, which is incorporated by reference in its entirety. Pharmaceutically acceptable formulations may include formulations wherein at least one component is a composition disclosed by Lin et al. in U.S. Pat. No. 8,222,258, which is incorporated by reference in its entirety.
Pharmaceutically acceptable formulations may include formulations wherein at least one component is a composition disclosed by Lin et al. in U.S. Pat. No. 8,530,481 , which is incorporated by reference in its entirety.
[00128] The term‘‘pharmaceutically acceptable salt” refers to salts derived from a variety of organic and inorganic counterions known in the art. Pharmaceutically acceptable acid addition salts can be formed with inorganic acids and organic acids. Preferred inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid and phosphoric acid. Preferred organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p- toluenesulfonic acid and salicylic acid. Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases. Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese and aluminum. Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins. Specific examples include isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. In some embodiments, the
pharmaceutically acceptable base addition salt is chosen from ammonium, potassium, sodium, calcium, and magnesium salts. The term“cocrystal” refers to a molecular complex derived from a number of cocrystal formers known in the art. Unlike a salt, a cocrystal typically does not involve hydrogen transfer between the cocrystal and the drug, and instead involves intermolecular interactions, such as hydrogen bonding, aromatic ring stacking, or dispersive forces, between the cocrystal former and the drug in the crystal structure.
[00129] ‘‘Pharmaceutically acceptable carrier" or“pharmaceutically acceptable excipient" is intended to include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and inert ingredients. The use of such pharmaceutically acceptable carriers or pharmaceutically acceptable excipients for active pharmaceutical ingredients is well known in the art. Except insofar as any
conventional pharmaceutically acceptable carrier or pharmaceutically acceptable excipient is incompatible with the active pharmaceutical ingredient, its use in the therapeutic compositions of the invention is contemplated. Additional active pharmaceutical ingredients, such as other drugs, can also be incorporated into the described compositions and methods.
[00130] It is understood that some embodiments according to the method include the specified ranges of TTX or other active pharmaceutical ingredient amounts per dosage unit.
It is further understood that use of a medicament according to the method includes the manufacture of a medicament wherein the medicament contains from 3.0 pg to about 150 pg of TTX or a pharmaceutically acceptable analog thereof per dosage unit.
[00131] While preferred embodiments of the invention are shown and described herein, such embodiments are provided by way of example only and are not intended to otherwise limit the scope of the invention. Various alternatives to the described embodiments of the invention may be employed in practicing the invention. Therefore, the spirit and scope of the appended claims should not be limited to the description of the preferred versions contained herein.
[00132] The reader’s attention is directed to all papers and documents which are filed concurrently with this specification and which are open to public inspection with this specification, and the contents of all such papers and documents incorporated herein by reference. All the features disclosed in the specification (including any accompanying claims, abstract, and drawings) may be replaced by alternative features serving the same, equivalent or similar purpose, unless expressly stated otherwise. Thus, unless expressly stated otherwise, each feature disclosed is one example only of a generic series of equivalent or similar features.
EXAMPLES
[00133] The embodiments encompassed herein are now described with reference to the following examples. These examples are provided for the purpose of illustration only and the disclosure encompassed herein should in no way be construed as being limited to these examples, but rather should be construed to encompass any and all variations which become evident as a result of the teachings provided herein.
[00134] In these studies, the effects of TTX, duloxetine and reboxetine, administered alone, and combinations of TTX with either duloxetine or reboxetine, at low doses, were investigated on rats with oxaliplatin-induced neuropathy. Paw withdrawal threshold (PWT) was monitored and assessed using a series of graduated Von Frey hairs, during a 5 day or 14 day dosing course and 2 weeks post-treatment observation in these models.
Example 1
[00135] In this study, a behavioral investigation was made of the synergetic effects of combinations of tetrodotoxin (TTX) and duloxetine or reboxetine as compared to TTX, duloxetine, or reboxetine alone on mechanical allodynia in rats with oxaliplatin-induced neuropathy. The purpose of this study was to conduct behavioral tests using von Frey hairs to evaluate the effects of tetrodotoxin (TTX) in combination with either duloxetine or reboxetine, in comparison to any of TTX, duloxetine, or reboxetine alone, on mechanical allodynia in rats with oxaliplatin-induced neuropathy.
[00136] Materials and methods— animals: male Sprague-Dawley Rattus norvegicus, 7-8 weeks old, were used, two animals per experimental group. Where needed, additional rats were added to the protocol in appropriate numbers to compensate for rats not developing oxaliplatin-induced peripheral neuropathy. All rats were housed in groups of 4 in an air- conditioned room on a 12-hour light/dark cycle. Food and water was available ad libitum.
[00137] Oxaliplatin was dissolved in 5% dextrose in distilled water to 4 mg/ml_; then further diluted the stock solution to 2 mg/ml_ with normal saline before use to induce neuropathy.
[00138] TTX for injection: doses were prepared under aseptic conditions in the morning of administration. One or more vials, as required, was removed from the storage condition and allowed to come to ambient temperature (20°C to 25°C). Each vial of lyophilized product was reconstituted with sterile water for injection under aseptic conditions.
[00139] Duloxetine was dissolved in normal saline; capsules were crushed in a pestle, then normal saline was added. The material was further ground to produce an even suspension.
[00140] Reboxetine tablets were crushed in a pestle, then normal saline was added. The material was further ground to produce an even suspension.
[00141] Other solutions, for example, 5% dextrose and normal saline, were prepared using standard aseptic research methods.
Animal study design
[00142] Rat model for chemotherapy induced neuropathic pain. Rats were anesthetized with 3% isoflurane mixed with medical oxygen (2 L/min). Oxaliplatin was intravenously injected through the tail vein. The rats were monitored for the development of neuropathic pain, characterized by significant mechanical allodynia. A series of graduated von Frey hairs were applied to the hind-paw to trigger a withdrawal response (Paw Withdrawal Threshold, PWT). Rats with significant mechanical allodynia (PWT < 4.0 g) were then selected for use in the drug testing experimental groups. The study groups are shown in Table 1 below.
Table 1.
Figure imgf000031_0001
[00143] TTX was administered to each rat by subcutaneous injection. For rats receiving a SSRI or NRI , either duloxetine or reboxetine was administered orally.
[00144] TTX for injection and the saline control were administered subcutaneously once daily for 14 days. Depending on the particular animal, duloxetine or reboxetine was administered orally once daily for 14 days. For the combination groups (no. 10-1 1), TTX was administered by subcutaneous injection first, then immediately followed by an oral dose of either duloxetine or reboxetine, depending on the particular animal.
[00145] Paw Withdrawal Threshold (PWT)— Rats were placed in individual perspex boxes on a raised metal mesh for at least 40 min before the PWT. The test was conducted by starting from the filament of lowest force (1 g), and applying each filament perpendicularly to the center of the ventral surface of the paw until the filament slightly bends, and held for 6 seconds. If the animal withdraws or lifts the paw upon stimulation, then the von Frey hair with the immediately next lower force than the one presently generating the withdrawal response is tested. If no response is observed, then test a filament with a force immediately higher is applied as described. The lowest amount of force required to induce reliable responses (positive in 2 out of 3 trials) was recorded as the PWT result.
[00146] The baseline paw-withdrawal threshold (PWT) of both hind-limbs was monitored for
3 consecutive days before the first injection of oxaliplatin. PWT was re-evaluated every 3 to
4 days before repeat of the oxaliplatin injection until successful establishment of a neuropathic pain state (PWT < 4.0g).
[00147] On the days of drug dosing, PWT was assessed pre-dose, 60 and 120 min following TTX or placebo administration. Animals were returned to their home cages for a break (about 40 min) between two neighboring testing time points. Baseline PWT was reassessed on the 7th, 10th, and 14th days from the start of the dosing course.
[00148] Data Analysis— PWT measured from both hind paws was averaged for each rat, and the mean of the 2 rats in each group was averaged for determining a trend between groups rather than a statistical comparison.
[00149] The effect of TTX at 1 , 2, and 3 pg/kg on PWT in rats with oxaliplatin-induced neuropathy on analgesia— TTX at 1 and 2 pg/kg did not significantly change the baseline PWT over the 5 days of dosing undertaken in this pilot study, whilst TTX at 3 pg/kg slightly increased the baseline PWT on Day 5 of the dosing course, compared to the pre-dosing value. After the cessation of dosing, the baseline PWT declined to the pre-dosing level on Day 7. For post-dosing effect on each dosing day, over the dose range of 1 , 2 and 3 pg/kg, TTX increased PWT at 1-hour post-dosing. The PWT declined after 2 hours post-dosing.
The effect of TTX in reversing the mechanical allodynia appeared to be dose-related. Figure 1 shows this dose relationship. There were no significant adverse effects observed during the dosing/observation period.
[00150] Effects of duloxetine at 5, 10 and 15 mg/kg on PWT in rats with oxaliplatin-induced neuropathy on analgesia— Duloxetine, at 5 mg/kg, appeared to marginally increased the baseline PWT on Day 4 to Day 5 of the dosing course. At higher doses of 10 and 15 mg/kg, duloxetine, produced a clear increase of the baseline PWT on days 4 to 7. Over the doses of 5, 10 and 15 mg/kg, duloxetine increased PWT from day 2 to day 5 post-dosing, in a dose-dependent manner. The effect reached a peak at 2 hours post-dosing. Figure 2 shows this dose-dependent relationship. No significant adverse effects observed during the dosing/observation period.
[00151] Effects of reboxetine at 1 , 3 and 5 mg/kg on the PWT in rats with oxaliplatin-induced neuropathy on analgesia— Reboxetine at 1 mg/kg did not change the baseline PWT compared to the pre-dosing level, whilst at doses of 3 and 5 mg/kg, reboxetine clearly increased the baseline PWT from day 3 to day 7. Reboxetine at 1 mg/kg slightly increased the PWT, post-dosing from day 3 onward. At the doses of 3 and 5 mg/kg, reboxetine increased the PWT from day 1 to day 5, in a dose-dependent manner. The effect peaked at about 2 hours post-dosing. Figure 3 clearly shows the peak at 2 hours. No significant adverse effects observed during the dosing or observation period.
[00152] Effects of the combination of TTX with duloxetine on the PWT in rats with oxaliplatin-induced neuropathy on analgesia— A combination of TTX at 3 pg/kg with duloxetine at 10 mg/kg increased the baseline PWT from day 2 to day 7, with a trend towards showing an effect greater than that induced by either TTX or duloxetine, at the same doses, administered alone.
[00153] The combination of TTX at 3 pg/kg in combination with duloxetine at 10 mg/kg induced an increase of PWT from day 1 to day 5, post-dosing, with a trend towards showing an effect greater than that induced by TTX or duloxetine administered alone. The data derived from this treatment group is plotted in Figure 4. No significant adverse effects observed during the dosing/observation period.
[00154] Effects of the combination of TTX with reboxetine on the PWT in rats with oxaliplatin-induced neuropathy on analgesia— A combination of TTX at 3 pg/kg with reboxetine at 3 mg/kg increased the baseline PWT from day 3 to day 7, with a trend towards showing an effect greater than that induced by TTX or duloxetine administered alone.
[00155] Regarding post-dosing effects, the combination of TTX at 3 pg/kg and reboxetine at 3 g/kg induced an increase of PWT from day 1 to day 5, with a trend towards showing an effect greater than that induced by TTX or reboxetine administered alone. Figure 5 depicts this data. No significant adverse effects observed during the dosing/observation period.
[00156] Summary— TTX, administered at 1 , 2 and 3 pg/kg, increased the PWT 1 hour after dosing in rats with oxaliplatin-induced neuropathy, in a dose-dependent manner.
[00157] Duloxetine, administered at 5, 10 and 15 mg/kg, increased the baseline PWT and PWT post-dosing in rats with oxaliplatin-induced neuropathy in a dose-dependent manner.
[00158] Reboxetine, administered at 1 , 3 and 5 mg/kg, increased the baseline PWT and PWT post-dosing in rats with oxaliplatin-induced neuropathy, in a dose-dependent manner.
[00159] The combination of TTX at 3 pg/kg with duloxetine at 10 mg/kg produced an increase in baseline PWT and PWT post-dosing, with a trend towards showing an effect greater than that induced by TTX or duloxetine administered alone. [00160] The combination of TTX at 3 pg/kg and reboxetine at 3 mg/kg increased the baseline PWT and PWT post-dosing, with a trend towards showing an effect greater than that induced by TTX or reboxetine administered alone.
[00161] The results of the pilot study suggest that combinations of TTX with either duloxetine or reboxetine produced effects greater than those induced by these compounds administered alone.
[00162] The results suggest that the combination of low dose TTX with duloxetine or reboxetine leads to less adverse effects and less severity compared to the higher dose of each individual compound administered alone in previous studies and noted in the product monographs.
[00163] The results of the pilot study suggest that combinations of TTX with either duloxetine or reboxetine produced effects greater than those induced by these compounds administered alone.
Example 2
[00164] This study is a further pilot study to determine the most effective dose ranges for the combinations of tetrodotoxin (TTX) with duloxetine or reboxetine on mechanical allodynia in rats with oxaliplatin-induced neuropathy. The study purpose was to determine the most effective dose ranges for the combinations of TTX with duloxetine (Dul) or reboxetine (Reb) on mechanical allodynia in rats with oxaliplatin-induced neuropathy.
[00165] Materials and methods: Oxaliplatin; Description: White powder; Vehicle: 5% dextrose, sterile; Preparation: Oxaliplatin was dissolved in 5% dextrose in distilled water, then diluted to 2 mg/ml_ with normal saline before use.
[00166] TTX for injection; Description: Lyophilized white powder; Preparation: The dose was prepared under aseptic conditions in the morning of the administration. One or more vials, as required, was removed from storage condition and allowed to equilibrate to ambient temperature (20°C to 25°C). Each vial of lyophilized product was reconstituted with sterile water for injection under aseptic conditions.
[00167] Duloxetine; Description: Capsules; Preparation: Duloxetine was dissolved in normal saline. Capsules were opened into a mortar, with normal saline added, and the content was ground to form an even suspension.
[00168] Reboxetine; Description: Tablets; Preparation: Reboxetine was dissolved in normal saline. Other solutions, for example, 5% dextrose and normal saline, were prepared using standard aseptic research methods. [00169] Animal study design— Model system for neuropathic pain— Rats were anaesthetized with 3% isoflurane mixed with oxygen (2 L per min). Oxaliplatin was administered intravenously through the tail vein. The development of neuropathic pain, characterized by significant mechanical allodynia, was monitored using a series of graduated von Frey hairs applied to the hind-paw to trigger a withdrawal response (Paw Withdrawal Threshold, PWT). Only those rats with significant mechanical allodynia (PWT < 4 g) were selected for further drug testing.
[00170] Table 2 below lists the 16 study groups; each with two rats.
Table 2.
Figure imgf000035_0001
[00171] The baseline PWT was assessed every day during the dosing course and on day 7, 10 and 14 after the first dosing; and the PWT change post dosing was assessed at 1 and 2 hours on day 1 , 2, 3, 4 and 5.
[00172] Paw Withdrawal Threshold (PWT)— Rats were placed in individual perspex boxes on a raised metal mesh for at least 40 min before the PWT. The test was conducted by starting from the filament of lowest force (1 g), and applying each filament perpendicularly to the center of the ventral surface of the paw until the filament slightly bends, and held for 6 seconds. If the animal withdraws or lifts the paw upon stimulation, then the von Frey hair with the immediately next lower force than the one presently generating the withdrawal response is tested. If no response is observed, then test a filament with a force immediately higher is applied as described. The lowest amount of force required to induce reliable responses (positive in 2 out of 3 trials) was recorded as the PWT result.
[00173] The baseline paw-withdrawal threshold (PWT) of both hind-limbs was monitored for
3 consecutive days before the first injection of oxaliplatin. PWT was re-evaluated every 3 to
4 days before repeat of the oxaliplatin injection until successful establishment of a neuropathic pain state (PWT < 4.0g).
[00174] On the days of drug dosing, PWT was assessed pre-dose, 60 and 120 min following TTX or placebo administration. At this point, animals were returned to their home cages for a break (about 40 min) between two neighboring testing time points. Baseline PWT was reassessed on the 7th, 10th, and 14th days from the start of the dosing course.
[00175] Data Analysis— PWT measured from both hind paws was averaged as the value for each rat, and the mean of the 2 rats in each group was averaged.
[00176] Effect of 3 pg/kg TTX and 2 mg/kg reboxetine, administered alone and in combination for 5 days, on PWT in a rat model of oxaliplatin-induced neuropathy on analgesia— Baseline PWT in the animals receiving 2 mg/kg reboxetine alone did not show any clear changes from day 2 to day 4. However, on day 5 there was a slight increase in PWT, and the increase became clear on day 7. On days 10 and 14, PWT declined to the pre-dosing value.
[00177] In the 3 pg/kg TTX alone treatment group, baseline PWT increased marginally from day 2 to day 4, and notably on day 5, reaching a peak on day 7. On day 10 and 14, PWT declined but remained higher than the pre-dosing level.
[00178] In the group receiving a combination treatment of 3 pg/kg TTX and 2 mg/kg reboxetine, PWT appeared to remain at a similar level as that of the 3 pg/kg TTX alone group at all time points, except where it was slightly higher than that of the TTX alone group on day 7, but slightly lower on day 14.
[00179] The post-dosing PWT in all groups increased slightly at 1 and 2 hours after dosing. The effect became most evident on day 4 in the group administered reboxetine alone and in the combination groups. On day 5, PWT in all groups increased sharply after dosing, reached a peak at 2 hours post-dosing, and remained slightly higher in the combination and TTX administered alone groups compared to that of the reboxetine only-treated group.
[00180] The data, plotted in Figure 6, shows the day 5 peak. No significant adverse effects observed during the dosing/observation period. [00181] 1 : Effects of 3 pg/kg TTX and 10 mg/kg duloxetine, administered alone and in combination for 14 days, on PWT in a rat model of oxaliplatin-induced neuropathy on analgesia— In the 3 pg/kg TTX-alone treated group, baseline PWT marginally increased on day 2 and further increased on day 3, and remained at a level around 4 - 6 g from day 3 to day 18, except on day 13 when a sudden increase in PWT was noted. On days 20 and 27, 1 and 2 weeks after cessation of dosing, the PWT was also increased.
[00182] In the 10 mg/kg duloxetine-alone treated group, baseline PWT increased on day 3 and remained at the same level on day 4, further increased on day 5 and remained at this level on day 6. However, PWT further increased on day 7 and remained at this new level of 810 g, towards day 27.
[00183] In the group treated with a combination of 10 mg/kg duloxetine and 3 pg/kg TTX, baseline PWT appeared to follow a similar trend as that observed in the duloxetine-alone and TTX-alone treated groups, although on day 4, PWT increased more markedly than that observed in the duloxetine-alone and TTX-alone treated groups. PWT then steadily increased between 6-8 g up to day 14, when a marked increase in PWT to 10 g was observed, before declining on day 16 and remaining at a level of 7-8 g between days 18-27. Baseline PWT in this group appeared to be higher than that of the TTX-alone treated group from day 4 to day 20, but lower than that observed in the duloxetine-alone treated group.
[00184] Post-dosing PWT slightly increased in the duloxetine-alone treated and the combination groups, but not in the TTX-alone treated group on day 1. From day 5 to day 14 a marked increase in post-dosing PWT at 1 and 2 hours was observed in the combination group, which was more notable than that observed in the TTX-alone treated group, where a marked increase in post-dosing PWT was only apparent on days 5, 8, 10, and 14. A marked increase in post-dosing PWT was also apparent in the duloxetine-alone treated groups on days 7, 8 and 9, although this effect was not as abrupt in occurrence as those noted in the TTX alone or the combination-treated groups. The data for this group is plotted in Figure 7. No significant adverse effects observed during the dosing/observation period.
[00185] Effects on analgesia of 3 pg/kg TTX and 2 mg/kg reboxetine, administered alone and in combination for 14 days, on PWT in a rat model of oxaliplatin-induced neuropathy— The baseline PWT in the 2 mg/kg reboxetine-alone treated group remained at a similar level as that was observed in the 3 pg/kg TTX-alone treated group (stated above) between days 2 to 16, except on day 13 where a sudden marked increase in PWT was observed in the TTX- alone treated group. The PWT in this group subsequently declined to a pre-dosing level from day 18 to 27. [00186] In the group administered a combination of 2 mg/kg reboxetine and 3 pg/kg TTX, the baseline PWT steadily increased from day 3 to day 6, and remained at a level of 8-10 g from day 7 to day 27 with no obvious decline in PWT.
[00187] The PWT post-dosing in the group treated with reboxetine-alone showed a slight increase from day 3 and was most apparent on days 10 and 14. A sharp increase in postdosing PWT could be seen on days of 3, 4, 5, 6 and 10 in the group administered a combination treatment. The data for this treatment group is plotting in Figure 8. No significant adverse effects observed during the dosing/observation period.
[00188] Effect of 5 pg/kg TTX, administered alone and in combination with 5 mg/kg reboxetine or 15 mg/kg duloxetine for 5 days, on PWT in a rat model of oxaliplatin-induced neuropathy on Analgesia— In the 5 pg/kg TTX-alone treated group, baseline PWT steadily increased between days 3 to 7 and then declined on day 10 and 14 although still remaining at a level higher than that observed during pre-dosing time. In the combination group of 5 pg/kg TTX and 15 mg/kg duloxetine, baseline PWT increased sharply compared to that observed in the TTX-alone treated group from day 2 to day 5, before declining sharply on day 7 to similar levels of PWT to that observed in the TTX-alone treated group. PWT remained around this new level until day 14. In the combination group of 5 pg/kg TTX and 5 mg/kg reboxetine, baseline PWT increased even more sharply from day 2 than that observed in the group administered a combination of TTX and duloxetine group. This effect reached a peak value on day 4, remained high on day 5, and then gradually declined towards day 14 to PWT levels similar to those observed in the TTX and duloxetine group.
[00189] PWT post-dosing increased sharply on each dosing day in the 5 pg/kg TTX-alone treated group, the combination group of 5 pg/kg TTX and duloxetine, and in the combination group of 5 pg/kg TTX and reboxetine. The data for this group is plotted in Figure 9. No significant adverse effects observed during the dosing/observation period.
[00190] Effect of 5 pg/kg TTX and 15 mg/kg duloxetine, administered alone and in combination for 14 days, on PWT in a rat model of oxaliplatin-induced neuropathy on Analgesia— Baseline PWT in the 5 pg/kg TTX-alone treated group increased steadily from day 3 and reached a plateau on day 8 of around 10-12 g, remaining at this level up to day 14. PWT then subsequently declined gradually to day 27 to a level slightly higher than that observed in the pre-dosing period. In both of the 15 mg/kg duloxetine-alone treated group and the combination group of 5 pg/kg TTX and 15 mg/kg duloxetine, baseline PWT increased steadily from day 2 to day 5 in a similar manner. PWT in the duloxetine-alone treated group then increased to a slightly higher level than the combination group from day 5 to day 13, but on day 14 it declined to a level lower than that observed in the combination group. From day 14, PWT in both groups declined gradually, but remained at a higher level than those observed in the 5 pg/kg TTX alone treated group.
[00191] PWT post-dosing at 1 and 2 hours increased sharply from day 4 to day 14 in the 5 pg/kg TTX-alone treated group. A sharp increase in PWT was observed and more notable in the combination group of TTX and duloxetine than that observed in the TTX-alone treated group, from day 2 to day 6. In the duloxetine-alone treated group, PWT post-dosing also increased from day 1 on each dosing day, but it was less marked than the PWT observed in the combination treated group from day 2 to day 1 1 . The data for this group is plotted in Figure 10. No significant adverse effects observed during the dosing/observation period.
[00192] Effect of 5 pg/kg TTX and 5 mg/kg reboxetine, administered alone and in combination for 14 days, on PWT in a rat model of oxaliplatin-induced neuropathy on Analgesia— Baseline PWT in the 5 mg/kg reboxetine-alone treated group increased steadily from day 2, a trend similar to that observed in the 5 pg/kg TTX-alone treated group (see Part D1 above). PWT subsequently declined from days 18 to 27, but at a rate slower than that observed in the TTX-alone treated group. The baseline PWT in the 5 mg/kg reboxetine- alone treated group also remained higher after cessation of dosing than that observed during the pre-dosing period. Baseline PWT in the combination group of TTX and reboxetine appeared to be similar, between days 2 to 12, to those recorded in the TTX-alone and reboxetine-alone treated groups, except on days 13, 14, 16, 18 and 27 where it appeared to be higher than the PWT of the TTX- and reboxetine-alone treated groups.
[00193] PWT post-dosing in the reboxetine-alone treated group increased from day 1 and on each dosing day. The post-dosing PWT in the combination group increased from day 1 and was notable from day 2 to day 14, the extent of the increase in PWT and peak value being clearly more marked than those of the TTX- and reboxetine-alone treated groups over the same period. The data for this group is plotted in Figure 1 1.
[00194] There were no significant adverse effects observed over the entire
dosing/observation period or with the doses used here.
[00195] Summary - The combination treatment comprising of 3pg/kg TTX and 2 mg/kg reboxetine, increased the baseline PWT and PWT post-dosing more prominently than either 3pg/kg TTX or 2 mg/kg reboxetine administered alone. This effect was observed over the 14-day dosing course and 2 weeks post-treatment in rats with oxaliplatin-induced neuropathy.
[00196] TTX at 5 pg/kg increased the baseline PWT and post-dosing PWT in rats with oxaliplatin-induced neuropathy. When this dose of TTX was combined with 15 mg/kg duloxetine or 5 mg/kg reboxetine, the baseline PWT and post-dosing PWT increased to a level higher than that observed when TTX at 5 pg/kg was administered alone. Furthermore, this effect was more prominent when TTX at 5 pg/kg was administered in combination with reboxetine at 5 mg/kg than when it was combined with duloxetine at 15 mg/kg; observed over the 5-day dosing course and 1 -week post-treatment observation period in rats with oxaliplatin-induced neuropathy.
[00197] The combination treatment comprising 5 pg/kg TTX and 15 mg/kg duloxetine increased the post-dosing PWT, an effect more prominent than that observed with 5 pg/kg TTX or 15 mg/kg duloxetine administered alone from day 1 to day 6, in rats with oxaliplatin- induced neuropathy.
[00198] The combination treatment comprising of 5 pg/kg TTX and 5 mg/kg reboxetine increased the baseline PWT on days 13, 14,16, 18 and 27, and PWT post-dosing more prominently than dosing with 5 pg/kg TTX or 5 mg/kg reboxetine-alone, over the 14 day dosing course and 2 week post-treatment observation period in rats with oxaliplatin-induced neuropathy.
Example 3 - Synergism analysis of data from pilot studies [00199] The data from Examples 1 and 2 described above were analyzed for synergism. In both of these studies there were two rats were selected for each drug dose, and for each drug combination.
[00200] Drug synergism analysis— Statistical analyses were performed using SAS/STAT® software version 9.4. Drug synergism analysis were conducted following the methods proposed by Tallarida et al. (1999) and Tallarida (1992, 2001). The analysis was performed separately for TTX with Duloxetine, and TTX with Reboxetine. Specifically, following the approach by Tallarida et al. (1999), the post baseline response measure (PWT) was converted to percent of maximum percent effect (MPE), using the following formula:
PWT - Baseline PWT
%MPE = 100 X
15— Baseline PWT
[00201] Baseline is defined as the time point of measurement immediately prior to the treatment, that is, Day 1 Hour 0. The cut-off value of 15 is the maximum PWT that can be achieved. %MPE measures the improvement on PWT, with 0 indicating no improvement and 100 indicating complete recovery.
[00202] PWT data were collected at various time points for each rat. For synergism analysis, the first step is to assess the dose effect relationship, i.e. the response from a rat under a given dose level. Hence it is necessary to estimate the mean response (%MPE) for each rat. Initially, an attempt was made to estimate the mean %MPE for each rat using a repeated measure mixed model, taking account the correlation of the repeated measures. The response was estimated for each rat by averaging the %MPE over all measurements, resulting in a single data point representing the response for each rat under each drug and dose (individual drug alone), and for the combinations. A hyperbolic model was then constructed for the dose-effect relations:
Dose
% MPE = MPEmax
{Dose + C)
[00203] Here MPEmax, the maximum effect, is set to equal to 100. C is the constant that represents the dose corresponding to the 50% of the MPE (or D50). The dose-effect data for each drug alone (TTX, duloxetine, and reboxetine) were fitted to the above model to estimate C, using the SAS non-linear procedure PROC NLIN. The relative potency of Drug B to Drug A, R, was then calculated based on the estimated C: R = Ca/Cb. The relative potency measures the relative effect (strength) of two drugs. For example, if Ca=20 and Cb=40, then the relative potency of Drug B is 0.5 (or 50%) of Drug A, meaning that either Drug B needs to double the dose to achieve the same effect as Drug A, or Drug B is a diluted dose (to 50%) of Drug A.
[00204] Based on the relative potency, any combination of two drugs (A and B) can be converted to an equivalent dose of one drug:
Dose Equivalance Drug A = Dose of Drug A + R X {Dose of Drug B )
[00205] The equivalent dose of a combination was then calculated in the hyperbolic model to obtain the expected effect (%MPE) - the effect that is expected if the two drugs are additive. The expected effect was compared with the actual (observed) effect to determine if the combination drug is super-additive (synergistic, which can be an observed effect that is greater than the expected), additive (observed effect is the same as the expected), or subadditive (observed effect is lower than the expected).
[00206] The expected effect of the two drug combinations (TTX and Duloxetine or TTX and Reboxetine) was calculated for the dose combinations of two drugs to construct a response surface. This response surface, presented as a 3-D plot, allows for a visual comparison of the observed effect of the drug combination against the expected effect under the assumption of additivity from the two drugs.
[00207] An interaction index, defined as the ratio of the equivalent dose of the drug combination to the dose of the single drug that results in the same effect, was calculated. An index of less than 1 indicates that the effect of a combination is super-additive (exhibiting synergism), 1 indicates additive, and greater than 1 indicates sub-additive (Tallarida, 2001).
[00208] The descriptive statistics for %MPE, calculated from the PWT data collected from the experiments, are shown in the table below for each treatment group.
Table 3.
Figure imgf000042_0001
Figure imgf000042_0002
[00209] Figure 12 shows the dose-effect relations, as estimated using a hyperbolic model, for each of the three drugs administered alone. Figure 12A shows the dose-effect relationship for TTX alone. Figure 12B shows the dose-effect relationship for reboxetine alone. Figure 12C shows the dose-effect relationship for duloxetine alone. The hyperbolic curved lines in each panel represent the dose response curves. The dots represent the mean % PE (over the post-baseline period) for each rat. [00210] The parameter C (or D50) in the hyperbolic function is estimated from the non-linear model; the results are shown in T able 4 below. For TTX, it was estimated that a dose of 12.65 pg/kg is required to achieve 50% MPE, while for Duloxetine and Reboxetine it is 24.41 and 9.32 mg/kg, respectively. These translate to a relative potency (relative to TTX) of 0.52 for Duloxetine and 1.36 for Reboxetine.
Table 4.
Figure imgf000043_0001
[00211] Based on the relative potency, the TTX dose equivalence was calculated for each of the drug combinations, as shown in the Table 5 below. The table also shows the expected %MPE (assuming additive effect), the actual (observed) %MPE, the corresponding dose required to achieve the observed effect (for TTX alone), and the interaction index.
Table 5.
Figure imgf000043_0002
[00212] The performance of drug combinations can also be seen in Figures 13 and 14, compared with the TTX dose effect curve. For the high dose combination (5:5, equivalent to 1 1.8 pg/kg TTX), the observed effect is much higher above the curve, signifying strong synergism. The interaction index of 0.469 indicates the combination took less than half of the dose (if using TTX alone) to achieve the same response. That is, using TTX alone, a dose of 25.1 pg/kg would be required to achieve the same MPE of 66.5% (plotted in Figure 13). For the two TTX Duloxetine combinations, the observed %MPE, plotted in Figure 14.
[00213] The same information can also be visualized in the 3-D response surfaces. The methods of Tallarida may be used to represent the data in 3-D plots. The same data represented in this 3-D format is shown in Figure 15 for Reboxetine and Figure 16 for Duloxetine. Example 4
[00214] In this study, a behavioral investigation was made of the synergetic effects of combinations of tetrodotoxin (TTX) and duloxetine or reboxetine on mechanical allodynia in rats with oxaliplatin-induced neuropathy. The purpose of this study is to conduct behavioral tests using von Frey hairs to evaluate the effects of tetrodotoxin (TTX) in combination with duloxetine or reboxetine, on mechanical allodynia in rats with oxaliplatin-induced neuropathy. This study expands on the pilot study to determine the most effective dose ranges for the combinations of tetrodotoxin (TTX) with duloxetine or reboxetine. The following experimental groups were used:
[00215] 1 . Placebo.
[00216] 2. Tetrodotoxin (TTX).
[00217] 3. Duloxetine dose 1.
[00218] 4. Reboxetine dose 1.
[00219] 5. TTX + Duloxetine dose 1 .
[00220] 6. TTX + Duloxetine dose 2.
[00221] 7. TTX + Reboxetine dose 1 .
[00222] 8. TTX + Reboxetine dose 2.
[00223] Materials and methods— Animals: male Rattus norvegicus, Sprague-Dawley, 7-8 weeks old seven animals were used per experimental group. Where needed, additional rats were added to the protocol in appropriate numbers to compensate for rats not developing oxaliplatin-induced peripheral neuropathy. All rats were housed in groups of 4 in an air- conditioned room on a 12-hour light/dark cycle. Food and water will be available ad libitum.
[00224] Oxaliplatin was dissolved in 5% dextrose in distilled water to 4 mg/ml_.
[00225] TTX for injection: doses were prepared under aseptic conditions in the morning of administration. One or more vials, as required, were removed from the storage condition and allowed to come to ambient temperature (20°C to 25°C). Each vial of lyophilized product was reconstituted with sterile water for injection under aseptic conditions.
[00226] Duloxetine was dissolved in normal saline; crushed in a pestle, then normal saline will be added, with the material further ground to produce an even suspension.
[00227] Reboxetine for the study was crushed in a pestle, then normal saline added. The material was further ground to produce an even suspension. [00228] Other solutions, for example, 5% dextrose and normal saline, were prepared using standard aseptic methods.
[00229] Animal study design— Rat model for chemotherapy induced neuropathic pain— Rats were anesthetized with 3% isoflurane mixed with medical oxygen (2 L/min). Oxaliplatin was intravenously injected through the tail vein. The rats were monitored for the development of neuropathic pain, characterized by significant mechanical allodynia. A series of graduated von Frey hairs were applied to the hind-paw to trigger a withdrawal response (Paw Withdrawal Threshold, PWT). Rats with significant mechanical allodynia (PWT < 4.0 g) were then selected for use in the drug testing experimental groups. The study groups are shown in Table 6 below.
Table 6.
Figure imgf000045_0001
[00230] TTX was administered to each rat by subcutaneous injection. For rats receiving either a SSRI or NRI, the compound was administered orally.
[00231] TTX for injection and saline control were administered subcutaneously once daily for 14 days. Depending on the particular animal, duloxetine or reboxetine were administered orally once daily for 14 days. For the combination groups (5-8), TTX was administered by subcutaneous injection first, then immediately followed by an oral dose of either duloxetine or reboxetine, depending on the particular animal.
[00232] Animal monitoring and evaluation: All animals were weighed each week during the study to monitor body weight as a proxy for general health.
[00233] Paw Withdrawal Threshold (PWT)— Rats were placed in individual perspex boxes on a raised metal mesh for at least 40 min before the PWT. The test was conducted by starting from the filament of lowest force (1 g), and applying each filament perpendicularly to the center of the ventral surface of the paw until the filament slightly bends, and held for 6 seconds. If the animal withdraws or lifts the paw upon stimulation, then the von Frey hair with the immediately next lower force than the one presently generating the withdrawal response is tested. If no response is observed, then test a filament with a force immediately higher is applied as described. The lowest amount of force required to induce reliable responses (positive in 2 out of 3 trials) was recorded as the PWT result.
[00234] The baseline paw-withdrawal threshold (PWT) of both hind-limbs was monitored for
3 consecutive days before the first injection of oxaliplatin. PWT was re-evaluated every 3 to
4 days before repeat of the oxaliplatin injection until successful establishment of a neuropathic pain state (PWT < 4.0g).
[00235] On the days of drug dosing, PWT was assessed pre-dose, 60 and 120 min following TTX or placebo administration. Baseline PWT will be reassessed on the 16th,
18th, 20th and 27th days from the start of the dosing course.
[00236] Data Analysis— Data obtained from behavioral experiments were grouped based on treatment and expressed as a mean ± standard error measurement (SEM). The different groups were compared at the same time points using one-way analysis of variance (ANOVA, PASW statistics SPSS, Version 18). Different time points in the same group were compared using paired Student’s t-test (Microsoft Excel 2007). For all tests, a P value lower than 0.05 (P < 0.05) were considered statistically significant. The data were further analyzed for drug synergism by applying similar methods as exemplified in Example 3.
[00237] Figures 17-18 show the effect of the combinations of TTX with Duloxetine or TTX with Roboxetine on paw withdrawal threshold compared to TTX, Duloxetine or Reboxetine alone. Statistical significance is illustrated by the asterisk.
[00238] Summary - The combination of 3 pg/kg TTX and 6 mg/kg duloxetine treatment, increased the baseline PWT and PWT post-dosing more noticeably than those of 3 pg/kg TTX-alone and 6 mg/kg duloxetine-alone in the 14-day dosing course and 2 weeks posttreatment observation. The combination of 3 pg/kg TTX and 3 mg/kg reboxetine, increased the baseline PWT and PWT post-dosing more clearly than those treated with 3 pg/kg TTX- alone and 3 mg/kg reboxetine-alone in the 14-day dosing course and 2 weeks posttreatment observation, in these models. A combination of TTX at 5pg/kg and Reboxetine at 5mg/kg, dosed chronically over 14 days, significantly reversed the reduction in PWT associated with model rats with Oxaliplatin-induced neuropathy. The effect was significantly more evident than those of TTX at 3pg/kg or Reboxetine at 3 mg/kg alone, or their combinations. The results suggest that the combination of low dose TTX with duloxetine or reboxetine leads to similar therapeutic effects with less adverse side-effects and less severity compared to the higher dose of each individual compound administered alone, as outlined in previous studies, and noted in the product monographs. Therefore, combinations of TTX with either duloxetine or reboxetine may be potentially produce a better result in relieving clinical chemotherapy-induced neuropathic pain.
Example 5 - Analysis of pooled set of synergism data
[00239] Data from various pilot studies and main studies were pooled to power the synergism analysis.
[00240] In general, the effects of TTX (tetrodotoxin), duloxetine and reboxetine, administered alone, and combinations comprised of TTX with either duloxetine or reboxetine, at low or high doses, respectively were tested by paw withdrawal threshold (PWT), assessed using a series of graduated von Frey hairs, over a 5- or 14-day dosing course observed over a 1 -2-week period post-treatment in rats with oxaliplatin-induced neuropathy.
[00241] Paw withdrawal threshold was asssed daily, and the data analyzed statistically as discussed above. Multidimensional dose plots were prepared according to the general methods of Tallarida as discussed above in the Synergisim section.
[00242] Figure 19A, 19B, and 19C show the dose-effect relations at Day 5 Hour 1 , as estimated using a hyperbolic model, for each of the three drugs (alone). The lines represent the dose response curves and the dots represent the mean %MPE (over the post-baseline period) for each rat. Figure 20 is the response surface plots showing the expected responses under additive assumption and the actual (observed) responses from the drug combinations at Day 5 Hour 1. A point above the surface represent the existence of synergism in the drug combination. The strong performance of TTX 5 pg/kg + Duloxetine 15 mg/kg is clearly noticeable in Figure 20B. Figure 21A, 21 B, 21 C show the dose-effect relations at Day 14 Hour 1 , as estimated using a hyperbolic model, for each of the three drugs (alone). The lines represent the dose response curves and the dots represent the mean % PE (over the post-baseline period) for each rat. Figure 22 is the response surface plots showing the expected responses under additive assumption and the actual (observed) responses from the drug combinations at Day 14 Hour 1. A point above the surface represent the existence of synergism in the drug combination. The strong performance of TTX 5 pg/kg + Reboxetine 5 mg/kg combination (Figure 22A), TTX 3 pg/kg + Duloxetine 10 mg/kg, and TTX 5 pg/kg + Duloxetine 15 mg/kg (Figure 22B) are clearly seen.
[00243] Summary - The results from the drug synergism analyses, conducted based on the data collected from the combinations of tetrodotoxin (TTX) with duloxetine or reboxetine on mechanical allodynia in rats with oxaliplatin-induced neuropathy, show that some drug combinations exhibit synergism. Particularly, based on data from Hour 1 on Day 5, the drug combination of TTX 5 pg/kg + Duloxetine 15 mg/kg appears to be synergistic, with an interaction index of 0.579; while based on data from Hour 1 Day 14, drug combinations TTX 5 pg/kg + Reboxetine 5 mg/kg, TTX 3 pg/kg + Duloxetine 10 mg/kg, and TTX 5 pg/kg + Duloxetine 15 mg/kg are synergistic.

Claims

CLAIMS We claim:
1 . A composition for the treatment of pain comprising:
(a) an amount of tetrodotoxin; and
(b) an amount of either a selective serotonin reuptake inhibitor (SSRI), serotonin- norepinephrine reuptake inhibitor (SNRI) or a norepinephrine reuptake inhibitor (NRI); formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat pain.
2. The composition of claim 1 , wherein the amount of tetrodotoxin is a singly therapeutically effective or singly subtherapeutic amount of tetrodotoxin.
3. The composition of either claim 1 or 2, wherein the amount of component (b) is a singly therapeutically effective or singly subtherapeutic amount of component (b).
4. The composition of any one of claims 1 to 3, wherein the amount of tetrodotoxin is between 82.5 pg to about 247.5 pg.
5. The composition of any one of claims 1 to 3, wherein the amount of tetrodotoxin is between 137.5 pg to about 412.5 pg.
6. The composition of any one of claims 1 to 5, wherein the SNRI is duloxetine.
7. The composition of claim 6, wherein the amount of duloxetine is between 275 mg to about 825 mg.
8. The composition of claim 6, wherein the amount of duloxetine is between 412.5 mg to about 1.2375 g.
9. The composition of any one of claims 1 to 5, wherein the NRI is reboxetine.
10. The composition of claim 9, wherein the amount of reboxetine is between 55 mg to about 165 mg.
1 1. The composition of claim 9, wherein the amount of reboxetine is between 137.5 mg to about 412.5 mg.
12. The composition of any one of claims 1 to 5, wherein the SSRI includes one or both of paroxetine and fluoxetine.
13. The composition of any one of claims 1 to 1 1 , wherein the pain is neuropathic pain.
14. The composition of any one of claims 1 to 1 1 , wherein the pain is nociceptive pain.
15. A composition for the treatment of neuropathic pain and/or nociceptive pain comprising:
(a) an amount of tetrodotoxin; and
(b) an amount of any one of duloxetine and reboxetine;
formulated in a pharmaceutically acceptable carrier, wherein the amounts of component (a) and component (b) exhibit synergism to treat neuropathic pain and/or nociceptive pain.
16. The composition of claim 15, wherein the neuropathic pain is selected from the group consisting of pain arising from diabetic peripheral neuropathy, fibromyalgia, and
chemotherapy induced neuropathic pain.
17. The composition of claim 15, wherein the nociceptive pain is selected from the group consisting of radicular pain, somatic pain, and visceral pain.
18. A composition for the treatment of neuropathic pain arising from chemotherapy comprising:
(a) an amount of tetrodotoxin; and
(b) an amount of reboxetine;
formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and reboxetine exhibit synergism to treat neuropathic pain.
19. A composition for the treatment of neuropathic pain arising from chemotherapy comprising:
(a) an amount of tetrodotoxin; and
(b) an amount of duloxetine; formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and duloxetine exhibit synergism to treat neuropathic pain.
20. A method of treating neuropathic pain and/or nociceptive pain in a mammal in need thereof, comprising:
(a) administering an amount of tetrodotoxin; and
(b) administering an amount of either a selective serotonin reuptake inhibitor (SSRI), a serotonin-norepinephrine reuptake inhibitor (SNRI), or a norepinephrine reuptake inhibitor (NRI);
wherein the amount of tetrodotoxin and the amount of either a selective serotonin reuptake inhibitor (SSRI), a serotonin-norepinephrine reuptake inhibitor (SNRI), or a norepinephrine reuptake inhibitor (NRI) are each formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and SSRI, SNRI or NRI exhibit synergism to treat neuropathic pain and/or nociceptive pain.
21. The method of claim 20, wherein the SNRI is duloxetine and the NRI is reboxetine.
22. A method of treating neuropathic pain and/or nociceptive pain in a mammal in need thereof, comprising:
(a) administering an amount of tetrodotoxin; and
(b) administering an amount of the SNRI duloxetine, formulated in a pharmaceutically acceptable carrier, wherein the amounts of tetrodotoxin and duloxetine exhibit synergism to treat neuropathic pain and/or nociceptive pain.
23. The method of claim 21 , wherein a dosage of the SNRI duloxetine is a subject mammal equivalent of a rat dose of about 1 mg/kg to 50 mg/kg.
24. The method of claim 21 , wherein a dosage of the NRI reboxetine is a subject mammal equivalent of a rat dose of about 1 mg/kg to 6 mg/kg.
25. The method of claim 21 , wherein a dosage of the NRI reboxetine is a subject mammal equivalent of a rat dose of about 4 mg/kg to 6 mg/kg.
26. The method according to any one of claims 20-25, wherein the mammal is a human patient undergoing treatment for chemotherapy induced neuropathic pain.
27. The method according to any one of the claims 20-26, wherein the mammal is a human patient undergoing treatment for cancer pain and wherein said patient also receives at least one low dose of an opioid.
28. A kit comprising an amount of tetrodotoxin, and an amount of an SSRI.
29. A kit comprising an amount of tetrodotoxin, and an amount of an SNRI.
30. A kit comprising an amount of tetrodotoxin, and an amount of an NRI.
31. A kit comprising an amount of tetrodotoxin for subcutaneous injection, and an amount of at least one of an SSRI, SNRI and a NRI for oral administration.
PCT/CA2019/050702 2018-05-23 2019-05-23 Synergistic tetrodotoxin formulations and methods of treatment for neuropathic pain WO2019222850A1 (en)

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