Small molecule prolactin receptor inhibitors, pharmaceutical compositions and treatment methods using such inhibitors

Abstract

Small molecule inhibitors (SMIs) of the Prolactin receptor, pharmaceutical compositions of the SMIs, and methods for treating patients suffering from disorders characterized increased expression or excitation of the Prolactin receptor, including breast cancer, prostate cancer and nociceptive pain disorders such as migraine headache, by administering pharmaceutical compositions of SMIs are provided.

Claims

1. A pharmaceutical composition comprising: a small molecule prolactin receptor (PRLR) inhibitor according to Formula I or a pharmaceutically acceptable salt thereof; and at least one pharmaceutically acceptable carrier, wherein Formula I is structurally depicted as: ##STR00007## and wherein: R.sub.1 and R.sub.2 are independently selected from —CY.sub.3, linear or cyclic alkyl, halogen, cycloalkyl, heterocycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, and H; a, b, and c are independently 0, 1, 2, or 3; and each Y is independently H or halogen selected from Cl, Br, F, and I.

2. The pharmaceutical composition according to claim 1, wherein R.sub.1 is selected from —CY.sub.3, halogen and H; R.sub.2 is selected from CY.sub.3, halogen, and linear C2-C3 alkyl; a, b, and c are independently 0, 1, or 2; and each Y is independently F or H.

3. The pharmaceutical composition according to claim 1 wherein the small molecule PRLR inhibitor is a compound according to Formula I and structurally depicted as: ##STR00008##

4. The pharmaceutical composition according to claim 1, formulated as an injectable solution or as an oral dosing form.

5. The pharmaceutical composition according to claim 4, wherein the molar concentration of inhibitor is up to 10 μM, up to 5 μM, between 1 μM and 10 μM, between 5 μM and 10 μM, or between 1 μM and 5 μM.

6. A method of treating breast cancer or prostate cancer in a subject in need thereof, the method comprising: administering to the subject an effective amount of a pharmaceutical composition according to claim 1.

7. The method according to claim 6, wherein administration of the pharmaceutical composition is by a parenteral route or an oral route.

8. The method according to claim 6 wherein the pharmaceutical composition is administered in conjunction with one or more chemotherapeutic agents that affect cellular microtubules.

9. The method according to claim 8, wherein the one or more chemotherapeutic agents are selected from the group consisting of cisplatin, demecolcine, nocodazole, vinblastine, and paclitaxel.

10. A method of reducing or substantially eliminating acquired resistance to chemotherapeutic drugs in a patient undergoing or about to undergo treatment for breast cancer or prostate cancer comprising a regimen of one or more chemotherapeutic drugs, the method comprising administering to the patient an effective amount of a pharmaceutical composition according to claim 1.

11. The method according to claim 10, wherein administering the pharmaceutical composition is before, during, and/or after initiation of the regimen of one or more chemotherapeutic drugs.

12. The method according to claim 10, wherein the one or more chemotherapeutic drugs are selected from the group consisting of cisplatin, demecolcine, nocodazole, vinblastine, and paclitaxel.

Description

BRIEF DESCRIPTION OF THE FIGURES

(1) FIG. 1A. Bar graph showing nhibition of PRL-induced invasion of breast cancer cells by SMI-1 and SMI-6; FIG. 1B. Bar graph showing only minimal cytotoxicity at indicated concentrations.

(2) FIG. 2. Bar graph showing suppression of autocrine PRL-dependent BrdU uptake in Jurkat lymphocytes by 1 M of each SMI, as determined by flow cytometry; the top panel indicates percent of live cells.

(3) FIG. 3. Bar graph demonstrating the effects of SMI-6 on thermal nociception; female mice were operated on the right hind paw; one day post-surgery, heat hyperalgesia was applied and paw withdrawal latency (PWL) was measured; BL—baseline: POP—postoperative pain; SMI-6 was applied either peripherally (5 g per), or spinally 5 g spin or 25 g spinSMI-6; each value is a mean±SME of 6 mice/treatment.

(4) FIG. 4. Bar graph demonstrating effects of SMI-6 on mechanical allodynia; female mice were operated on the right hind paw; one day postsurgery, an anesthesiometer was used to measure leg withdrawal threshold against increasing mechanical pressure on the animal right hind paw. BL—baseline: POP—postoperative pain; SMI-6 was applied either peripherally (5 g per), or spinally (5 g or 25 g); each value is a mean±SME of 6 mice/treatment.

DETAILED DESCRIPTION OF THE INVENTION

(5) Embodiments of the invention provide small molecule inhibitors of PRLR signaling, pharmaceutical compositions comprising one or more of the small molecule inhibitor compounds, and methods for treating cancer, methods for reducing resistance to chemotherapy, and methods for treating patients suffering from conditions characterized by nociceptive pain by administering one or more of the small molecule inhibitor compounds. Based on previous in silico molecular docking work, the present investigators identified two structural formulae possessing enhanced binding and inhibition potential vis a vis the PRLR. Specific, exemplary embodiments of compounds of Formula I as defined herein include SMI-6 and derivatives of SMI-6, and specific embodiments of compounds of Formula II as defined herein include SMI-1 and derivatives of SMI-1. Both specific exemplary compounds strongly interact with the ligand binding domain of human PRLR (hPRLR) at binding affinities (Kd) of 1.2 and 3.3 for SMI-1 and SMI-6, respectively, and both were confirmed pharmacologically to block PRLR signaling in multiple cellular assays.

(6) One embodiment is directed to a pharmaceutical composition comprising: a small molecule PRLR inhibitor according to Formula I or Formula II; and, optionally, at least one pharmaceutically acceptable carrier, and, optionally, at least one pharmaceutically acceptable salt, wherein Formula I is structurally depicted as:

(7) ##STR00003##
and
Formula II is structurally depicted as:

(8) ##STR00004##
and further wherein, R.sub.1, and R.sub.2 are independently selected from —CY.sub.3, linear or cyclic alkyl, halogen, cycloalkyl, heterocycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, and H; and X is selected from —NH.sub.2, halogen, —OH, —SH, —NO.sub.2, —COOH, —SO.sub.3H, and —SO.sub.2NH.sub.2; a, b, and c are independently 0, 1, 2, or 3, n is 0 or an integer between 1 and 10; and each Y is independently H or halogen selected from Cl, Br, F, and I. According to specific embodiments, R.sub.1 is selected from —CY.sub.3, halogen and H, R.sub.2 is selected from CY.sub.3, halogen, linear C2-C3 alkyl, a, b and c are independently 0, 1 or 2, and each Y is independently F or H; and for Formula II, R.sub.1 is para and selected from —CY.sub.3, halogen and H, and n is 0, 1 or 2.

(9) As utilized herein, “halogen” or “halogen atom” means fluorine, chlorine, bromine, or iodine. The term “alkyl” refers to a substituted or unsubstituted saturated, straight-chain or branched hydrocarbon group that contains from 1 to 20 carbon atoms, preferably from 1 to 10 carbon atoms, for example a n-octyl group, especially from 1 to 6 carbon atoms, i.e. 1, 2, 3, 4, 5, or 6. Specific examples of alkyl groups include methyl, ethyl, propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, tert-butyl, n-pentyl, iso-pentyl, n-hexyl and 2,2-dimethylbutyl. Specific examples of suitable substitutions include —NH.sub.2, halogen, —OH, —SH, —NO.sub.2, —COOH, —SO.sub.3H, and —SO.sub.2NH.sub.2. Very specific examples are selected from Cl, F, and —OH. Exemplary, non-limiting preferred substituted alkyl included mono-, di- and tri-halogenated methyl and ethyl wherein the halogen is selected from Fluorine and Chlorine.

(10) The term “alkenyl” refers to an at least partially unsaturated straight-chain or branched hydrocarbon group that contains from 2 to 20 carbon atoms, preferably from 2 to 10 carbon atoms, especially from 2 to 6, i.e. 2, 3, 4, 5 or 6, carbon atoms. Specific examples of alkenyl groups include ethenyl (vinyl), propenyl (allyl), iso-propenyl, butenyl, ethinyl, propinyl, butinyl, acetylenyl, propargyl, iso-prenyl and hex-2-enyl group. Preferably, alkenyl groups have one or two double bonds.

(11) The term “alkynyl” refers to an at least partially unsaturated substituted or unsubstituted, straight-chain or branched hydrocarbon group that contains from 2 to 20 carbon atoms, preferably from 2 to 10 carbon atoms, especially from 2 to 6 carbon atoms, i.e. 2, 3, 4, 5 or 6. Specific examples of alkynyl groups are ethynyl, propynyl, butynyl, acetylenyl and propargyl groups. More specifically, alkynyl groups have one or two triple bonds, and even more specifically alkynyl groups have one triple bond.

(12) The term “cycloalkyl” refers to a saturated or partially unsaturated (for example, a cycloalkenyl group) substituted or unsubstituted cyclic group that contains one (or more, in the case of polycyclic) rings (preferably 1 or 2), and contains from 3 to 14 ring carbon atoms, preferably from 3 to 10 (especially 3, 4, 5, 6 or 7) ring carbon atoms. Specific examples of cycloalkyl groups are cyclopropyl, cyclobutyl, cyclopentyl, spiro[4,5]decanyl, norbornyl, cyclohexyl, cyclopentenyl, cyclohexadienyl, decalinyl, bicyclo[4.3.0]nonyl, tetraline, adamantane (i.e. tricycle[3.3.1.1.sup.3,7]decane), cyclopentylcyclohexyl and cyclohex-2-enyl. Specific examples of suitable substitutions include —NH.sub.2, halogen, —OH, —SH, —NO.sub.2, —COOH, —SO.sub.3H, and —SO.sub.2NH.sub.2. Very specific examples are selected from Cl, F, and —OH.

(13) The term “heterocycloalkyl” refers to a substituted or unsubstituted cycloalkyl group as defined above in which one or more (preferably 1, 2 or 3) ring carbon atoms, each independently, have been replaced by an oxygen, nitrogen, silicon, selenium, phosphorus or sulfur atom (preferably by an oxygen, sulfur or nitrogen atom). A heterocycloalkyl group has preferably 1 or 2 rings containing from 3 to 10 (especially 3, 4, 5, 6 or 7) ring atoms (preferably selected from C, O, N and S). Specific examples include piperidyl, prolinyl, imidazolidinyl, piperazinyl, morpholinyl, urotropinyl, pyrrolidinyl, tetra-hydrothiophenyl, tetrahydropyranyl, tetrahydrofuryl and 2-pyrazolinyl group and also lactames, lactones, cyclic imides and cyclic anhydrides. Specific examples of suitable substitutions include —NH.sub.2, halogen, —OH, —SH, —NO.sub.2, —COOH, —SO.sub.3H, and —SO.sub.2NH.sub.2. Very specific examples are selected from Cl, F, and —OH.

(14) The term “aryl” refers to a substituted or unsubstituted aromatic group that contains one (or more, in the case of polycyclic aryl) rings containing from 6 to 14 ring carbon atoms, preferably from 6 to 10 (especially 6) ring carbon atoms. Examples are phenyl, naphthyl and biphenyl groups. Specific examples of suitable substitutions include —NH.sub.2, halogen, —OH, —SH, —NO.sub.2, —COOH, —SO.sub.3H, and —SO.sub.2NH.sub.2. Very specific examples are selected from Cl, F, and —OH.

(15) The term “heteroaryl” refers to a substituted or unsubstituted aromatic group that contains one (or more, in the case of polycyclic heteroaryl) rings containing from 5 to 14 ring atoms, preferably from 5 to 10 (especially 5 or 6) ring atoms, and contains one or more (preferably 1, 2, 3 or 4) oxygen, nitrogen, phosphorus or sulfur ring atoms (preferably O, S or N). Examples are pyridyl (for example, 4-pyridyl), imidazolyl (for example, 2-imidazolyl), phenylpyrrolyl (for example, 3-phenylpyrrolyl), thiazolyl, iso-thiazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, oxadiazolyl, thiadiazolyl, indolyl, indazolyl, tetrazolyl, pyrazinyl, pyrimidinyl, pyridazinyl, oxazolyl, isoxazolyl, triazolyl, tetrazolyl, isoxazolyl, indazolyl, indolyl, benzimidazolyl, benzoxazolyl, benzisoxazolyl, benzthiazolyl, pyridazinyl, quinolinyl, isoquinolinyl, pyrrolyl, purinyl, carbazolyl, acridinyl, pyrimidyl, 2,3′-bifuryl, pyrazolyl (for example, 3-pyrazolyl) and iso-quinolinyl groups. Specific examples of suitable substitutions include —NH.sub.2, halogen, —OH, —SH, —NO.sub.2, —COOH, —SO.sub.3H, and —SO.sub.2NH.sub.2. Very specific examples are selected from Cl, F, and —OH.

(16) Small molecule inhibitors may be present in a pharmaceutical composition as a pharmaceutically acceptable salt. By pharmaceutically acceptable salt it is meant those salts, within the scope of medical science, which are suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, suitable pharmaceutically acceptable salts of compounds according to the present invention may be prepared by mixing the compounds of the invention with a pharmaceutically acceptable acid (including inorganic and organic acids). Suitable pharmaceutically acceptable salts of the compounds of the present invention therefore include acid addition salts. Suitable pharmaceutically acceptable organic acids for making this salts include, but are not limited to the following: acetic acid, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, citric acid, ethenesulfonic acid, fiimaric acid, gluconic acid, glutamic acid, hydrobromic acid, hydrochloric acid, isethionic acid, lactic acid, maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonic acid, mucic acid, nitric acid, pamoic acid, pantothenic acid, phosphoric acid, oxalic acid, succinic acid, sulfuric acid, tartaric acid acid, p-toluenesulfonic acid, and the like. Inorganic acid addition salts are hydrochloric, hydrobromic, phosphoric, and sulfuric acid salts.

(17) The salts can be prepared in situ during the final isolation and purification of the compounds of the invention, or separately by reacting the free base function with a suitable organic acid. Representative acid addition salts include acetate, adipate, alginate, ascorbate, asparate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptonate, hexanoate, hydrobromide, hydrochloride, hydroiodide, 2-hydroxyethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, toluenesulfonate, undecanoate, valerate salts, and the like.

(18) Embodiments of the invention contemplate that the small molecule inhibitors may be formulated into a pharmaceutical composition comprising a pharmaceutically acceptable carrier. Preferably the SMI is provided in pharmaceutical grade purity.

(19) The term “pharmaceutically acceptable carrier” is intended to include solvents, dispersion media, coatings, antibacterial and anti-fungal agents, isotonic and absorption delaying agents, and the like. The use of such media and agents for pharmaceutically active substances is well known in the art. Except in so far as any conventional media or agent is incompatible with the compound, use thereof in the therapeutic compositions and methods of treatment is contemplated. Supplementary active compounds may also be incorporated into the compositions according to embodiments of the present invention. It is especially advantageous to formulate parenteral compositions in dosage unit form for ease of administration and uniformity of dosage.

(20) The term “dosage unit form” as used herein refers to physically discrete units suited as unitary dosages for the individual to be treated; each unit containing a predetermined quantity of compound(s) calculated to produce the desired therapeutic effect in association with the required pharmaceutical earner. The compound(s) may be formulated for convenient and effective administration in effective amounts with a suitable pharmaceutically acceptable carrier in an acceptable dosage unit. In the case of compositions containing supplementary active ingredients, the dosages are determined by reference to the usual dose and manner of administration of the said ingredients.

(21) Sterile injectable solutions can be prepared by incorporating the SMI compound in the required amount in an appropriate solvent with one or a combination of the ingredients discussed herein, as required, followed by filtered sterilisation. Generally, dispersions are prepared by incorporating the SMI compound into a sterile vehicle which contains a basic dispersion medium and the required other ingredients. Tablets, troches, pills, capsules and the like may also comprise the following: a binder such as gum tragacanth, acacia, corn starch or gelatin; excipients such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid and the like; a lubricant such as magnesium stearate; and a sweetening agent such as sucrose, lactose or saccharin or a flavouring agent such as peppermint, oil of wintergreen, or cherry flavouring. When the dosage unit form is a capsule, it may further comprise a liquid carrier. Various other materials can be present as coatings or to otherwise modify the physical form of the dosage unit. For instance, tablets, pills, or capsules can be coated with shellac, sugar or both. A syrup or elixir may comprise the SMI inhibitor, sucrose as a sweetening agent, methyl and propylparabens as preservatives, a dye and flavouring such as cherry or orange flavour. Of course, any material used in preparing any dosage unit form should be pharmaceutically pure and substantially non-toxic in the amounts employed. In addition, the SMI compound may be incorporated into sustained/extended-release, controlled release, delayed release preparations and formulations.

(22) In very specific embodiments, the pharmaceutical composition comprises a small molecule PRLR inhibitor compound according to Formula I:

(23) ##STR00005##

(24) In other very specific embodiments, the pharmaceutical composition comprises a small molecule PRLR inhibitor compound according to Formula II:

(25) ##STR00006##

(26) According to some embodiments, the pharmaceutical composition may be formulated as an injectable solution or as an oral dosing form. SMI compound may be included in the pharmaceutical composition in any therapeutically relevant amount, which may vary based on specifics of the patient, such as size, gender, health status and the like. Specific compositions are formulated with a molar concentration of SMI compound up to 10 μM, up to 5 μM, between 1 μM and 10 μM, between 5 μM and 10 μM or between 1 μM and 5 μM. As will be apparent to an ordinary clinician, amounts may be adjusted higher if lesser amounts are well-tolerated by the patient. Concentrations of total SMI compound up to 25 μM, including continuously all values in-between, are contemplated.

(27) Other embodiments of the invention are directed to methods of treating patients suffering from a disorder characterized by increased expression of Prolactin receptor (PRLR). According to specific embodiments, the disorder is cancer. According to more specific embodiments, the disorder comprises breast or prostate cancer. Method generally comprises administering a pharmaceutical composition comprising one or more SMI compounds according to the invention. According to very specific embodiments, The SMI compound comprises SMI-1 or SMI-6.

(28) Convenient modes of administration of SMI compounds of the invention include parenteral (for example, subcutaneous, intravenous, intramuscular, intradermal, intraperitoneal, intrathecal, intraocular, intranasal, intraventricular injection or infusion techniques), oral, pulmonary (e.g. inhalation), transdermal application, topical (e.g. creams or gels or powders), or rectal administration. Depending on the route of administration, the formulation and/or compound may be coated with a material to protect the compound from the action of enzymes, acids and other natural conditions which may inactivate the therapeutic activity of the compound. In other formulations the dosage form may be coated for extended, controlled, or delayed release by methods well-known in the art.

(29) Dispersions of the SMI compounds according to the invention may also be prepared in glycerol, liquid polyethylene glycols, oils, and mixtures thereof. Under ordinary conditions of storage and use, pharmaceutical preparations may contain a preservative to prevent the growth of microorganisms. Pharmaceutical compositions suitable for injection include: sterile aqueous solutions (where water soluble), or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. Ideally, the composition is stable under the conditions of manufacture and storage and may include a preservative to stabilise the composition against the contaminating action of microorganisms such as bacteria and fungi.

(30) SMI compounds of the invention may be administered orally, for example, with an inert diluent or an assimilable edible carrier. The compound(s) and other ingredients may also be enclosed in a hard or soft shell gelatin capsule, compressed into tablets, or incorporated directly into an individual's diet. For oral therapeutic administration, the SMI compound(s) may be incorporated with excipients and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like. Suitable dosages may be obtained by single or multiple administrations.

(31) According to some embodiments of the invention, the pharmaceutical composition is administered in conjunction with one or more chemotherapeutic agents that affect cellular microtubules. Chemotherapeutic agents may be selected from cisplatin, demecolcine, nocodazole, vinblastine, paclitaxel (taxol), and combinations thereof.

(32) Other embodiments of the invention are directed to methods for reducing or substantially eliminating acquired resistance to chemotherapeutic drugs in a patient undergoing or about to undergo treatment for cancer comprising a regimen of one or more chemotherapeutic drugs by administering one or more of the SMI compounds according to embodiments of the invention. According to specific embodiments, the cancer comprises breast or prostate cancer. Depending on whether the treatment is intended for prophylactic purposes or as a response to observation of resistance, the SMI compounds may be administered before, during, and/or after initiation of a regimen of one or more chemotherapeutic drugs. Non-limiting examples of chemotherapeutic drugs with a problematic resistance profile but which are effectively addressed by methods disclosed herein include cisplatin, demecolcine, nocodazole, vinblastine, and paclitaxel (taxol), and generally any chemotherapeutic agent that acts on microtubules.

(33) The nociceptive pain response may also be controlled via inhibition of PRLR. Nociceptive pain is that which results from triggering/stimulating/activating a pain-receptor and may be considered a sensory nerve response; whereas non-nociceptive pain does not implicate pain receptors. A specific example of a condition characterized by nociceptive pain comprise migraine headache. Although the term “migraine headache” encompasses various types of headaches, all implicate stimulation of pain receptors. Embodiments of the invention provide methods of treating a patient suffering from a condition characterized by nociceptive pain by administering one or more of the SMI compounds according to Formula I or II as defined herein, and more specifically by administering SMI-1 or SMI-6. According to specific embodiments, administering comprises a peripheral parenteral or oral route and the small molecule inhibitor is formulated as an injectable solution or an oral dosing form at a concentration of between 1 μM and 5 μM. In other specific embodiments administering comprises a central route and the small molecule inhibitor is formulated as an injectable solution at a concentration of between 1 μM and 25 μM. Although endpoints are provided, the scope of the embodiment includes all values between the endpoints. As will be readily understood by a clinician of ordinary skill in the art, doses may be divided as necessary depending on the health and tolerance level of an individual patient.

(34) The following Examples are set forth to illustrate specific embodiments and should not be construed as narrowing the scope of the invention as defined by the appended claims.

EXAMPLES

Example 1

(35) Blockade of PRL-induced invasion of breast cancer cells is confirmed.

(36) Both SMI-1 and SMI-6 at 1 μM concentrations abrogated PRL-induced invasion of MDA-MB-468 breast cancer cells, using Boyden chambers (FIG. 1A). The LDH cytotoxicity assay revealed that the SMIs had minimal toxicity, especially at 10-fold higher doses (FIG. 1B).

Example 2

(37) Blockade of autocrine PRL-induced cell proliferation is demonstrated.

(38) To examine whether the SMIs can suppress cell proliferation that is driven by autocrine PRL, Jurkat human lymphocytes were utilized, which produce large amounts of autocrine PRL. Autocrine PRL in lymphocytes is well known to promote cell proliferation. The cells were incubated for 24 hrs in 2% charcoal-stripped serum (which is devoid on exogenous lactogenic hormones) with 1 μM of SMI-1 or SMI-6, and then with 1 nM PRL for 24 hrs. BrdU uptake was analyzed by flow cytometry. As shown in FIG. 2, added PRL had no effect, likely because of receptor saturation by autocrine PRL. Notably, both SMI-6 and SMI-1 completely blocked cell proliferation, while reducing cell viability by only 10-20%.

Example 3

(39) Abrogation of peripheral pain by exemplary compound SMI-6 is demonstrated.

(40) Female mice were operated on the right hind paw. One day after surgery, 5 μg SMI-6 was applied either peripherally or centrally/spinally (5 μg or 25 μg) and mice were subjected to thermal nociception or mechanical allodynia (increased sensitization to tactile stimuli). As evident in FIG. 3, SMI-6 showed a dose-dependent alleviation of thermal pain, which was applied to the inured paw by means of a short radiant heat beam and measurement of leg withdrawal latency. SMI-6 at the 25 μg dose was effective in alleviating thermal pain by as much as 75%.

(41) For evaluating allodynia, an anesthesiometer was used, which applied a constant ramp of increasing mechanical pressure to the injured paw. Withdrawal latency was measured in grams. FIG. 4 shows that SMI-6 was equally as effective in abrogating mechanical pain.