WO2008091704A2

WO2008091704A2 - Treatment of cushing's syndrome and autism

Info

Publication number: WO2008091704A2
Application number: PCT/US2008/001030
Authority: WO
Inventors: Vassilios Papadopoulos; Janet Greeson
Original assignee: Georgetown University; Samaritan Pharmaceuticals, Inc.
Priority date: 2007-01-25
Filing date: 2008-01-25
Publication date: 2008-07-31
Also published as: WO2008091704A3

Abstract

A method for the treatment of Cushing's syndrome in a patient in need thereof, the method comprising administering to the patient an effective amount of a combination of a pharmaceutically acceptable salt of procaine and zinc sulfate heptahydrate, at a frequency and for a duration of time sufficient to provide a beneficial effect to the patient.

Description

TREATMENT OF CUSHING'S SYNDROME AND AUTISM

Cross-reference to Related Application and Claim of Priority

This application claims the priority of U.S. Ser. No. 60/886,642, filed Jan. 25, 2007, which is incorporated herein by reference in its entirety.

Background Cushing's syndrome is a hormonal disorder caused by prolonged exposure of the body's tissues to high levels of the hormone Cortisol (corticosteroids). Sometimes called "hypercortisolism" or "hyperadrenocorticism," it is relatively rare and most commonly affects adults aged 20 to 50. An estimated 10 to 15 of every million people are affected each year.

Overexposure to Cortisol due to the body's inability to regulate corticosteroid production has been associated with numerous symptoms including upper body obesity, rounded face, increased fat around the neck, thinning arms and legs, fragile and thin skin, weakened bones, fatigue, high blood pressure, high blood sugar, excessive hair growth, irritability, anxiety, and depression.

Autism is a developmental disorder of the human brain that first gives signs during infancy or childhood and follows a steady course without remission or relapse. Impairments result from maturation-related changes in various systems of the brain. Autism is characterized by widespread abnormalities of social interactions and communication, and severely restricted interests and highly repetitive behavior. The manifestations of autism cover a wide spectrum, ranging from individuals with severe impairments — who may be silent, mentally disabled, and locked into hand flapping and rocking — to less impaired individuals who may have active but distinctly odd social approaches, narrowly focused interests, and verbose, pedantic communication. Despite extensive investigation, how autism occurs is not well understood. Its mechanism can be divided into two areas: the pathophysiology of brain structures and processes associated with autism, and the neuropsychological linkages between brain structures and behaviors.

Neuroanatomical studies and the associations with teratogens strongly suggest that autism's mechanism includes alteration of brain development soon after conception. This localized anomaly appears to start a cascade of pathological events in the brain that are significantly influenced by environmental factors. Many major structures of the human brain have been implicated. Consistent abnormalities have been found in the development of the cerebral cortex; and in the cerebellum and related inferior olive, which have a significant decrease in the number of Purkinje cells. Brain weight and volume and head circumference tend to be greater in autistic children. There is no cure. Medications are often used to treat problems associated with ASD. More than half of U.S. children diagnosed with ASD are prescribed psychoactive drugs or anticonvulsants, with the most common drug classes being antidepressants, stimulants, and antipsychotics. In the United States, the antipsychotic risperidone is approved for treating symptomatic irritability in autistic children aged 5-16 years. Other drugs are prescribed off-label, which means they have not been approved for treating ASD. For example, some selective serotonin reuptake inhibitors and dopamine blockers can reduce some maladaptive behaviors associated with ASD. However, there is scant reliable research about the effectiveness or safety of drug treatments for adolescents and adults with ASD. A person with ASD may respond atypically to medications, the medications can have adverse side effects, and no known medication relieves autism's core symptoms of social and communication impairments.

Brief Description of the Figures

Figure 1 shows an assessment and comparison of the toxicity of SPOl, AZT and ddl on human T lymphocytes.

Figure 2 shows an assessment and comparison of the toxicity of SPOl, AZT and ddl on human macrophages.

Figure 3 is a bar graph depicting the proportion of subjects with abnormally low Cortisol secretion attaining normal values at week 8.

Figure 4 shows the in vitro toxicity of 3TC, ddl, AZT, procaine hydrochloride and SPOlA on HeLa cells. Figure 5 shows potential procaine metabolite pathways. Figure 6 is a bar graph depicting the proportion of subjects with abnormally low Cortisol secretion attaining normal values at week 8. Figure 7 depicts a study schema for clinical trials of SPOlA.

Description of the Invention

The present invention is directed to methods of treatment of Cushing's syndrome and autism, using a pharmaceutical composition comprising a combination of a pharmaceutically acceptable salt of procaine and zinc sulfate heptahydrate, and to those compositions adapted for treatment of those conditions.

An embodiment of the invention provides a method for the treatment of Cushing's syndrome in a patient in need thereof, the method comprising administering to the patient an effective amount of a combination of a pharmaceutically acceptable salt of procaine and zinc sulfate heptahydrate, at a frequency and for a duration of time sufficient to provide a beneficial effect to the patient.

An embodiment of the invention provides a method of treatment a malcondition in a patient wherein a reduction of Cortisol production is medically indicated, comprising administering to the patient an effective amount of a combination of a pharmaceutically acceptable salt of procaine and zinc sulfate heptahydrate, at a frequency and for a duration of time sufficient to provide a beneficial effect to the patient.

An embodiment of the invention provides a method for the treatment of autism in a patient in need thereof, the method comprising administering to the patient an effective amount of a combination of a pharmaceutically acceptable salt of procaine and zinc sulfate heptahydrate, at a frequency and for a duration of time sufficient to provide a beneficial effect to the patient.

An embodiment of the invention provides a combination of a pharmaceutically acceptable salt of procaine, such as procaine hydrochloride, and zinc sulfate heptahydrate, adapted for treatment of Cushing's syndrome or any malcondition wherein inhibition of Cortisol production or secretion is medically indicated. In an embodiment the combination can be comprised by an oral dosage form adapted for oral ingestion by patients suffering from Cushing's syndrome. Another embodiment of the invention further provides a pharmaceutically acceptable salt of procaine, such as procaine hydrochloride, and zinc sulfate heptahydrate, adapted for treatment of autism. In an embodiment the combination can be comprised by an oral dosage form adapted for oral ingestion by patients suffering from autism.

The inventive methods make use of a formulation adapted for use as an oral dosage form, wherein the pharmaceutically acceptable salt of procaine is stabilized for oral administration. An embodiment of the formulation is a synergistic combination of procaine HCl and zinc sulfate heptahydrate. SPOlA is an orally administered therapeutic comprising procaine HCl and zinc sulfate heptahydrate that, as disclosed herein, has Cortisol secretion modulation properties. Procaine hydrochloride is a monohydrochloride salt of 2-diethylaminoethyl 4-aminobenzoate. The molecular weight of procaine hydrochloride is 272.77 and the empirical formula is Ci₃H₂₀N₂O₂, HCl with the following structural formula:

Procaine hydrochloride is an odorless, small white crystal or crystalline powder, with a melting point range of 153°- 158° and a solubility of lg/mL in water.

SPOlA also can have a beneficial effect in the treatment of autism. As is well known in the art, the terms "Tl" and "T2" as used herein refer to specific responses ("type 1" and "type 2" respectively) of the immune system that each have distinct characteristics. It is known that the two types of responses are interactive; down-regulation of one type of response can lead to inappropriate overactivation of the other type (MR Gyetko, et al., Infection and Immunity, January 2004, p. 461-467, Vol. 72, No. 1). The Tl response has been described as cell-mediated, and the T2 response as humoral (W.C. Davis, et al., Annals of the New York Academy of Sciences 969:119-125 (2002)). It is believed by the inventors herein that compounds of the present invention such as SPOlA can act to prevent the overexpression of the T2 response with a concomitant reduction of the Tl response. It is also believed by the inventors herein that the condition of autism is induced, mediated, or influenced by a pathological overexpression of the T2 immune response and consequent reduction of the Tl response, and that administration of an effective dose of a compound of the present invention will result in a remission, amelioration, or stabilization of the symptoms of autism. A therapeutically effective amount of the compounds of the present invention is contemplated to be any amount which would serve to regulate immune response, including Tl -mediated and T2-mediated immune responses. Thus, the inventors herein believe that a pharmaceutically acceptable salt of procaine, such as an HCl salt, in its orally available complexed form with zinc sulfate heptahydrate, can act as an immunomodulatory compound that has the ability to modulate pathological Tl and T2 immune responses and is therefore useful in the treatment of autism when administered to a patient in need thereof. The combination of the procaine salt and zinc sulfate heptahydrate can be administered by oral ingestion.

Oral administration of the compounds disclosed herein is particularly desirable. By oral administration, there is contemplated preparation of the compounds disclosed herein in any dosage form capable of oral administration. Such dosage forms include tablets, capsules, caplets, solutions, sublingual dosage forms, suppositories, nasal sprays and the like.

The oral dosage forms of the present invention may contain pharmaceutically acceptable inert ingredients. As such inert ingredients there are contemplated pharmaceutical carriers, excipients, fillers, etc. which do not interfere with the activity of the compound. Also, fillers such as clays or siliceous earth may be utilized if desired to adjust the size of dosage form.

Further ingredients such as excipients and carriers may be necessary to impart the desired physical properties of the dosage form. Such physical properties are, for example, release rate, texture and size of the dosage form. Examples of excipients and carriers useful in oral dosages forms are waxes such as beeswax, castor wax, glycowax and carnauba wax, cellulose compounds such as methylcellulose, ethylcellulose, carboxymethylcellulose, cellulose acetate, hydroxypropylcellulose and hydroxypropylmethylcellulose, polyvinyl chloride, polyvinyl pyrrolidone, stearyl alcohol, glycerin monostearate, methacrylate compounds such as polymethacrylate, methyl methacrylate and ethylene glycol dimethacrylate, polyethylene glycol and hydrophilic gums. Also in accordance with the present invention, there is provided a liquid- based dosage form suitable for the administration of the composition to a patient. The liquid base for this dosage form may be any liquid capable of transporting the composition into the body of a patient without disrupting the activity of the compound or harm the patient. Exemplary of such a liquid is an isotonic solution. The isotonic solution may also contain conventional additives therein such as sugars. These solutions can be used in the preparation of oral compositions.

Thus, the compositions of the present invention may be admixed according to known procedures using known excipients. A therapeutically effective amount of the compounds of the present invention, there is contemplated any amount which would serve to regulate Cortisol levels or treat Cortisol related diseases, including Cushing's syndrome, or to provide a beneficial effect to a patient in treatment of autism. hi one embodiment of the invention, the composition is a mixture of procaine HCl and zinc sulfate heptahydrate. The weight ratio of procaine HCl to zinc sulfate heptahydrate can range from about 40:1 to about 160:1. In one embodiment, the total adult dose of the mixture is between 100 mg and 2.0 gm per day, more preferably 500 mg and 1.0 gm per day with 750 mg to 1.0 gm being most preferred. The infant/child dose can typically range from 50-500 mg per day with 50-200 mg per day being preferred. Typically daily dosages for a 60 kg human can range from 200-3,000 mg.

The compositions of the present invention may include other materials such as protein, fats, carbohydrates, vitamins, minerals, sweeteners, flavoring agents and the like. For example, the composition of the present invention, anti- HIV drug plus Cortisol blockers, may be combined with known food ingredients or dispersed in a liquid such as orange juice, and consumed orally. The composition of the present invention may also be in the form of a powder, liquid, tablet, capsule, pill, candy, sublingual dosage form, suppositories, confection, food additive or gel cap.

Procaine hydrochloride, 2-diethylaminoethyl p-aminobenzoate hydrochloride, is also known as Novocain^®, Neocaine, Planocaine, and Ethocaine^®. Procaine, (β-diethylaminoethyl p-aminobenzoate) is one of the oldest and most used of the synthetic local anesthetics, having been developed in 1906. The free ester is an oil, but is isolated and used as a salt, typically the hydrochloride salt. It occurs as an odorless, white crystalline powder that is stable in air, soluble in water and alcohol, but much less soluble in organic solvents. Procaine is most stable at pH 3.6 and becomes less stable as the pH is increased or decreased from this value. The procaine molecule is also subject to oxidative decomposition.

Procaine is an anti-cortisol compound that has the ability to decrease the level of Cortisol previously elevated in the blood. Other anti-cortisol compounds will also have use in the methods of the invention. Additional compounds having anti-cortisol effects include lidocaine HCl, zinc, zinc salts, zinc sulfate heptahydrate, ascorbic acid, dilantin (also referred to as phenytoin), clonidine, phosphatidylserine, DHEA, RU-486, HMB, ketaconazole, pregnenalone and Ipriflavone. Additional Cortisol blockers include pantothenic acid, acetylsalicylic acid (aspirin), dimethyl sulphoxide (DMSO), retinol (vitamin A), co-enzyme QlO, acetyl-L-carnitine and ginko beloba.

Examples

The following examples are provided in order to demonstrate and further illustrate certain embodiments and aspects of the present invention and are not to be construed as limiting the scope thereof.

Example 1

In one embodiment, the composition of a gelatin capsule is provided below:

Example 2

Mechanism of Action for Cortisol Modulation In the publication by the inventors herein, Xu J, Lecanu L, Han Z, Yao Z,

Greeson J, and Papadopoulos V. Inhibition of Adrenal Cortical Steroid Formation by Procaine Is Mediated by Reduction of the cAMP-Induced 3- Hydroxy-3-methylglutaryl-coenzyme A Reductase Messenger Ribonucleic Acid Levels. JPET 307:1148-1157, 2003, evidence is provided about the proposed mechanism of action of SPO 1 A (a formulation of procaine hydrochloride and a complexing agent, zinc sulfate heptahydrate, capable of forming a protected complex with procaine).

Procaine hydrochloride was found to act, in a dose-dependent manner, by reducing the hormone/stress-induced increase of the rate-limiting enzyme HMG- CoA reductase mRNA expression, leading to decreased intracellular cholesterol and corticosteroid biosynthesis. Procaine hydrochloride did not affect basal corticosteroid formation, suggesting that only pathological states of abnormal glucocorticoid formation would be affected.

This unique action was observed in an 8-week, Phase II clinical investigation characterized by daily administration of SPOlA. Basal metabolic levels were re-established in individuals with abnormally high and low Cortisol secretion levels at baseline. Conversely, individuals with normal Cortisol secretion levels at baseline continued to sustain normal secretion levels. The in-vitro, in-vivo data and clinical studies discussed below demonstrate that SPOlA directly effects glucocorticoid production and restores Cortisol secretion levels to basal metabolism levels.

Thus, SPOlA represents a new and novel treatment for Cushing's Syndrome. Demonstration of Cortisol Modulation Activity In a Phase II investigation of SPOlA, patients with sufficient adrenal function as demonstrated by an increase in basal Cortisol levels after administration of ACTH were monitored for changes from baseline in 24-hour urinary Cortisol excretion and changes from baseline in Cortisol levels over an 8- week period. A total of 29 subjects were evaluated for 24-hour urinary Cortisol secretion. Fourteen (9 in the low dose (200 or 400 mg daily) group and 5 in the high dose (600 or 800 mg daily) group had abnormally low Cortisol secretion levels (below 20 μg/24hr) at baseline. Two additional subjects, both from the high dose group, had secretion levels in excess of normal values levels (above 90 μg/24hr). The remaining 6 subjects (2 and 4 subjects, respectively) demonstrated normal secretion levels at baseline.

All subjects with abnormal baseline secretion levels in the high dose group, regardless of whether they were above or below normal values, normalized by Week 8. Conversely, none of the subjects in the low dose group with abnormally low secretion levels at baseline normalized by Week 8 remaining effectively at the same low level they had at baseline. Subjects who had normal levels at baseline remained normal at Week 8.

These results demonstrate that SPOlA (procaine hydrochloride / zinc sulfate heptahydrate) has a modulatory effect on Cortisol, restoring secretion levels to basal metabolism levels. Urinary Cortisol

In an 8-week investigation designed to study the effect of SP-OlA treatment on urinary Cortisol, urinary Cortisol secretion was determined by 24- hour Cortisol secretion observations at Week 1 (baseline) and Week 8 (end-of- treatment). The change in urinary Cortisol levels was measured comparing Week 1 to Week 8 values.

This evaluable population included all subjects with 24-hour Cortisol secretion observations at Weeks 1 and Week 8. This population was comprised of 22 subjects (11 in the low dose group from Cohorts A and B and 11 in the high dose group from Cohorts C and D). Each dose group had a multi-racial profile and all subjects were male. The average age of the low and high dose groups was 43 years and 41 years, respectively.

Normal 24-hour urinary Cortisol secretion levels range between 20.0 and 90.0 μg/24hr. A comparison of the proportion of subjects with abnormally low values at baseline that attained normal values by Week 8 is presented in Table 1 and Figure 3. Table 1 : Proportion of Subjects with Abnormally low Cortisol Secretion Attaining Normal Values at Week 8

^*Statistical testing was done using Fisher's Exact test.

Of the 22 subjects who were evaluated for 24-hour urinary Cortisol secretion, 14 subjects (9 in the low dose group and 5 in the high dose group) had abnormally low Cortisol secretion levels (below 20 μg/24hr) at baseline. Two additional subjects, both from the high dose group, had secretion levels in excess of normal values levels (above 90 μg/24hr). The remaining 6 subjects (2 and 4 subjects, respectively) demonstrated normal secretion levels at baseline.

All of the subjects with abnormal baseline secretion levels in the high dose group, regardless of whether they were above or below normal levels, normalized by Week 8. This finding was statistically significant (p=0.0005). Conversely, none of the subjects in the low dose group with abnormally low secretion levels at baseline normalized by Week 8, remaining effectively at the same low level they had at baseline. Of the 6 subjects who had normal levels at baseline in the high dose group at baseline, all remained within normal levels at Week 8. Of the 2 subjects who had normal levels at baseline in the low dose group, 1 subject remained within normal levels at Week 8 and 1 subject dropped below normal levels.

There was no difference in total urine Cortisol measurements between Cohort E (controls) and subjects from Cohorts A, B, C, and D. Table 2: Comparison of Baseline Urine Cortisol* between Case and Control⁴*

♦ Summed over 7 measurements taken within 24-hours

♦ ♦ Case includes Cohort A, B, C, and D; Control is Cohort E

These results support findings that procaine hydrochloride (SPOlA) has a modulatory effect on Cortisol, restoring secretion levels to basal metabolism levels. Serum Cortisol

Serum Cortisol measurements were also examined comparing the low and high dose groups from Week 1 to Week 8 and Week 8 to Week 10. Mean Cortisol levels decreased for both the low and high dose groups from Week 1 to Week 8 (12.7 to 8.8 μg/dL and 15.9 to 9.1 μg/dL, respectively). Mean Cortisol levels increased for both the low and high dose groups from Week 8 to Week 10 (8.8 to 12.6 μg/dL and 9.1 to 11.6 μg/dL, respectively). In both cases, the values remained within normal range (5 to 25 μg/dL). In Vitro Cytotoxicity

An assessment of the toxicity of SPOlA on monocytes and macrophages and the methods used to assess the toxicity are described in Samaritan Preclinical Report Samaritan Preclinical Report, Effect of SPOl on Monocytes and Lymphocytes, Report Number: SPOl A-l-102-04, November 2004, Data on

File.

Human T-Lymphocytes

SPOl induced a decrease of T lymphocytes viability when present at 6.2 to 500 μM, reaching a maximal inhibition of 10%. Under the same conditions, AZT and ddl displayed a cytotoxic effect even at the very low concentrations of 0.228 and 0.68 μM reaching maximal inhibitions of 14% and 11%, respectively. Figure 4-6 shows that AZT was significantly more toxic than SPOl at almost every concentration tested, whereas, 0.68 μM, represented the concentration at which the highest toxicity of ddl was displayed. Human Macrophages

SPOl displayed a cytotoxic effect only at the high concentrations of 166.6 and 500 μM reaching maximal inhibition of 7.5%. Under the same conditions, AZT decreased human macrophage viability even at the lowest concentration of 0.228 μM, reaching a maximal inhibition of 9%, and ddl at 0.68 μM, reaching a maximal inhibition of 5%.

Comparison of SPOl A cytotoxicity to the anti-viral compounds AZT, 3TC and ddl used as reference compounds on human cervix carcinoma HeLa cells

SPOlA did not induce a significant decrease of HeLa cell viability when present at 1 nM to 100 μM. Under the same conditions, AZT, ddl and 3TC were significantly more toxic than SPOl and SPOlA at 10 μM and above. 3TC was toxic even at concentration as low as 0.1 μM. The IC_5O of the cytotoxic effect of AZT, 3TC and ddl were respectively 81.7, 75.7 and 189.7 μM whereas ICs₀S for SPOl and SPOlA could not be calculated. These results suggest that the use of SPOl as a therapeutic will not be hampered by significant cytoxicity .

Example 3 Pharmacokinetics

Procaine is an ester that is known to be hydrolysed when added to human plasma. The enzymes involved were formerly called procaine esterases. These are identified as non-specific acetylcholine esterases, the butylcholinesterases. Because these esterases are non-specific, they hydrolyse various esters including procaine, acetylcholine derivatives, and succinylcholine. They are secreted by hepatocytes and excreted in plasma, their main location in the body, but they can also be found in the liver and the gut. Although their structures are closely related, these enzymes may have different spectra of activity, and thus they act differently on the same substrate. Other enzymes such as oxonases and aliesterases are involved in the hydrolysis of ester functions as well. Generally, the hydrolysis process is an inactivation process, which means that procaine hydrolysis ends its pharmacological activity. It has been determined that the hydrolysis of procaine in the plasma of humans is quick and total in physiological conditions. It is also believed that procaine metabolites are in turn metabolized (Figure 5). Procaine is extensively metabolized in- vivo. Only about 2% of unchanged procaine is found in urine. Most of the metabolites formed in- vitro after incubation of procaine with isolated hepatocytes are no longer found in urine. Moreover, the concentrations of most of them decrease during the incubation time, suggesting that they are in turn metabolized. Thus, the occurrence of tertiary metabolites is likely. In rat hepatocytes the overall half- lives (Ti_/2) of transformations are 2.26 h and 6.77 h at the concentrations of 25 and 250 μM, respectively and were concentration dependent. In human hepatocytes, Tj_/2 are 3.24 h and 5.34 h at the concentrations of 25 and 250 μM, respectively and were also concentration dependent. When administered by IV injection (10 mg/kg in rats), the data indicates

2 compartments; distribution (T _1/2 of 12 min) and elimination (Ti_/2 of 55 min). The corresponding V_D (elimination) was 43 L/kg, suggesting that procaine is distributed in tissues, especially in the most irrigated organ in the CNS and the adrenals, as previously reported in published literature. Following IV administration, two plasmatic peaks were observed, which is in accordance with previous findings and suggests the existence of 2 different sites of absorption in the gut.

When administered by oral route, procaine is partly absorbed by the gut without being transformed and then distributed into the systemic circulation. As a consequence, procaine partly escapes both intestinal and liver first-pass effects and can reach various organs such as the CNS and the adrenals. Using a comparison of the respective plasmatic AUC, a relative bioavailability of 21% for procaine hydrochloride in SPOlA seems to be a reasonable estimation (Table 4). Table 4: Relative Bioavailability of Procaine and SP01 A in Rats

Lymphoid tissues are found underneath the gut epithelium inside the gut wall. Data shows that procaine at least partly crosses the gut wall. Therefore, procaine is able to reach the local lymphoid tissues which can be a target for an inhibitory effect on HIV replication. This also suggests that procaine may develop its effects at least partly before reaching the systemic circulation.

In summary, pharmacokinetic studies in rats show that a part of the initial oral dose, about 21%, is distributed into the body before transformations, escaping intestinal and liver transformations. Published studies have shown that the plasma binding percentage of Procaine is low, 19%, which cannot limit tissue distribution. V_D is high, about 43 L/kg which indicates a large tissue distribution. We may expect that a similar behavior of procaine occurs in humans. It also appears that high doses are needed to get significant clinical effects. A likely explanation for this is that such doses saturate the various transformation processes in the gut and the liver, which allows for at least a part of the remaining drug to reach its targets. This saturation process may, in all probability, be increased by daily repeated administrations of SPOl A throughout a chronic treatment.

Example 4

DESCRIPTION OF CLINICAL STUDIES

8-Week Open Label Clinical Study with SPOlA

The safety and dose response of orally administered SPOlA in HIV- infected patients was assessed in a Phase I/II study. The study was an 8-week non-randomized, open-label study conducted at a single investigational site in patients infected with HIV-I who were being treated with concomitant triple combination antiretroviral therapy for at least 8 weeks prior to study initiation. The original protocol was primarily designed to study the effect of SPOl A treatment on urinary Cortisol, assess its effect on Quality of Life, assess the safety of SPOlA at escalating doses, and evaluate pharmacokinetic data.

The evaluable population for urinary Cortisol secretion included all subjects with 24-hour Cortisol secretion observations at Weeks 1 and Week 8. This population was comprised of 22 subjects (11 in the low dose group from Cohorts A and B and 11 in the high dose group from Cohorts C and D). Each dose group had a multi-racial profile and all subjects were male. The average age of the low and high dose groups was 43 years and 41 years, respectively. Patients received the following doses of SPOlA and were divided into low dose and high dose groups for further analyses (see Table 5).

Table 5 : Dose Groups and Cohorts

Inclusion criteria for the study included CD4⁺ count >200 and stable triple therapy antiretroviral regimen for the preceding 8 weeks.

Treatment changes for end of study treatment to baseline (Week 8 - Week 1) were compared to assess whether there was a dose response between the high and low dose groups. Changes for post-treatment (Week 10 - Week 8) were compared to assess whether there was a "rebound effect" on observed changes after the drug was removed. Efficacy Evaluation Urinary Cortisol

Normal 24-hour urinary Cortisol secretion levels range between 20.0 and 90.0 μg/24hr. A comparison of the proportion of subjects with abnormally low values at baseline that attained normal values by Week 8 is presented in Table 6 and Figure 6. Table 6: Proportion of Subjects with Abnormally low Cortisol Secretion Attaining Normal Values at Week 8

'Statistical testing was done using Fisher's Exact test.

Of the 22 subjects who were evaluated for 24-hour urinary Cortisol secretion, 14 subjects (9 in the low dose group and 5 in the high dose group) had abnormally low Cortisol secretion levels (below 20 μg/24hr) at baseline. Two additional subjects, both from the high dose group, had secretion levels in excess of normal values levels (above 90 μg/24hr). The remaining 6 subjects (2 and 4 subjects, respectively) demonstrated normal secretion levels at baseline. All of the subjects with abnormal baseline secretion levels in the high dose group, regardless of whether they were above or below normal levels, normalized by Week 8. This finding was statistically significant (p=0.0005). Conversely, none of the subject in the low dose group with abnormally low secretion levels at baseline normalized by Week 8 remaining effectively at the same low level they had at baseline. Of the 6 subjects who had normal levels at baseline in the high dose group at baseline, all remained within normal levels at Week 8. Of the 2 subjects who had normal levels at baseline in the low dose group, 1 subject remained within normal levels at Week 8 and 1 subject dropped below normal levels. There was no difference in total urine Cortisol measurements between controls (HIV-) and HIV+ subjects. Serum Cortisol

Serum Cortisol measurements were also examined comparing the low and high dose groups from Week 1 to Week 8 and Week 8 to Week 10. Mean Cortisol levels decreased for both the low and high dose groups from Week 1 to Week 8 (12.7 to 8.8 μg/dL and 15.9 to 9.1 μg/dL, respectively). Mean Cortisol levels increased for both the low and high dose groups from Week 8 to Week 10 (8.8 to 12.6 μg/dL and 9.1 to 11.6 μg/dL, respectively). In both cases, the values remained within normal range (5 to 25 μg/dL). Whalen Symptom Index

Study Treatment (Week 1 to Week 8)

Favorable changes (decreases) were seen in Whalen Symptom Index Scores for both the low and high dose groups. The high dose group had a clinically meaningful decrease in adverse health symptoms, meaning an improvement in Quality of Life for both the evaluable and subgroup analysis. The change for the high dose group was from 4.4 points at Week 1 to 0.0 points at Week 8 (an improvement of 4.4 points) and 6.3 points at Week 1 to 0.0 points at Week 8 (an improvement of 6.3 points), respectively. The change for the low dose group was from 7.0 points at Week 1 to 5.6 points at Week 8 (an improvement of 1.4 points) and 7.9 points at Week 1 to 6.7 points at Week 8 (an improvement of 1.2 points), respectively. These data demonstrate that patients, especially in the high dose group, felt their Quality of Life improved during the 8 weeks of treatment with SPOlA (see Table). Post-Treatment (Week 8 to Week 10) hi spite of continued treatment with the patients' original anti-retro viral therapy, adverse health symptoms (Quality of Life) of patients in the high dose group rebounded when SPOlA was withdrawn, approaching baseline levels. Mean scores rose for both the evaluable and subgroup analysis from 0 points at Week 8 to 4.8 points at Week 10 (a 4.8 point increase) and 0 points at Week 8 to 6.7 points at Week 10 (a 6.7 point increase) in the high dose group and from 5.6 points at Week 8 to 6.2 points at Week 10 (a 0.6 point increase) and 6.7 points at Week 8 to 7.0 points at Week 10 (a 0.3 point increase) for the low dose group, respectively (see Table 7).

Table 7: Mean (SD) Whalen Symptom Index Scores (Weeks 1 to 8 and Weeks 8 to 10)

*Mean (sd)

Safety Evaluation

Of the 29 patients included in the safety analysis, 23 patients (12/17 patients in low dose group and 11/12 patients in high dose group) experienced at least one adverse event during the study. The breakdown of adverse events by dose group and severity are shown in Table 8. There was no relationship found between the dose groups and the severity of adverse events.

Table 8: Adverse Events by Severity

^* Patients could report more than one adverse event.

A total of 22 adverse events (20%) had severity ratings by the investigators of moderate (16 events) or severe (6 events). Adverse events rated as moderate in severity included headache, nausea, bronchitis, hypophosphothema, rectal pain, groin fungal rash, left TM erythema, increased diarrhea, parasites, shingles, avascular necrosis bilateral hips and shoulders, and increased bilirubin. Adverse events rated as severe included depression, nausea, headache, lightheadedness, and vomiting. There were no adverse events rated as very severe. A total of 16 adverse events (12%) were considered by the investigators to be possibly (13 events) or probably related (3 events) to the study medication. Adverse events considered possibly related to study medication included headache, nausea, fatigue, insomnia, lightheadedness, intermittent nervousness, and skin rash. Adverse events considered probably related to study treatment included headache and nausea. There were no adverse events reported as definitely related to study treatment.

There was one serious adverse event that was considered unrelated to the study medication. No deaths or other significant adverse events occurred during study treatment. Laboratory abnormalities reported during the study were considered mild and not considered clinically significant for this study population. Mean vital sign measurements (temperature, systolic and diastolic blood pressure, and pulse rate), height, and weight at baseline were within normal limits. There were no significant changes in vital signs, physical findings, or other observations during the study treatment or post-treatment period. 10-Day Monotherapy Study with SPOlA

A 10-day multi-center, double-blind, randomized, placebo-controlled monotherapy study in HIV-infected subjects with evidence of resistance to currently available antiretroviral therapy was conducted to assess the safety of orally administered SPOlA. Safety was assessed at each visit by laboratory evaluations, physical examination and questioning for adverse events (AEs). In the event of toxicity, dosing of study drug was to be stopped according to the provisions outlined in the protocol.

SPOlA was well tolerated in this clinical study. AU adverse events reported were rated mild or moderate in severity. Adverse events reported by subjects in the active arms were similar in scope, severity, frequency, and likelihood of occurrence to those reported by subjects in the placebo arm. Of the 41 adverse events reported; 24 (59%) were rated by the investigators as mild and 17 (41%) with a severity of moderate. No events were rated severe or very severe. Only one event (a mild improvement in the subject's pre-existing condition) was assessed as possibly related to study drug. All other adverse events were categorized as "unlikely" to be related to study drug. The most common adverse events reported by subject were myalgia, arthralgia, diarrhea, and nausea. No serious adverse events or deaths were reported during this study. Any laboratory abnormalities reported during the study were not considered clinically significant and no significant changes in mean vital sign measurements or physical examination findings were reported during the study period. Extent of Exposure

The study conducted was a Phase II randomized, placebo controlled, monotherapy clinical study of 10 days duration using SPOlA in HIV positive patients. Three doses (200 mg, 400 mg, and 800 mg daily) of SPOlA and placebo were studied in a total of 35 study subjects. All 35 subjects were included in the safety adverse events analysis.

All subjects in the SPOlA treatment arms and all subjects in the placebo group, with the exception of one subject (S6624) who withdrew from the study, reported that they had taken the medication as planned. The total daily dose of SPOlA in each of the treatment groups is shown in Table 9.

Table 9: Treatment Groups and Exposure

Adverse Events

SPOlA was well tolerated in this clinical study. At the doses studied in this HIV positive patient population, all of the adverse events reported were rated mild or moderate in severity. The most common adverse events reported by subject were myalgia, arthralgia, diarrhea, and nausea. A total of 24 adverse events (59%) were rated by the investigators with a severity of mild and 17 adverse events (41%) were rated as moderate. No severe or very severe events were reported. Laboratory abnormalities seen were mostly mild in severity and unrelated to dose.

Vital Signs and General Chemistry Parameters

There were no significant changes in mean vital sign measurements (systolic and diastolic blood pressure, pulse, respiration, and temperature) and no significant physical examination findings during the study period. The values for change from baseline to Day-11 for hematology, biochemistry and urinalysis laboratory parameters, were similar across treatment groups for the majority of laboratory parameters and no trends were detectable. Any laboratory abnormalities reported during the study were considered characteristic of the study population and not clinically significant. 28-Day Monotherapy Study with SPOlA

A 28-day multi-center, double-blind, randomized, placebo-controlled monotherapy study in HIV-infected subjects with evidence of resistance to currently available antiretroviral therapy was conducted to assess the safety of orally administered SPOlA. Safety was assessed at each visit by laboratory evaluations, physical examination and questioning for adverse events (AEs). hi the event of toxicity, dosing of study drug was to be stopped according to the provisions outlined in the protocol.

Example 5

The safety and tolerability of orally administered SPOlA has already been demonstrated in an 8-week Phase Ib/IIa investigation of 29 subjects, a 10- day monotherapy investigation of 34 subjects, and a 28-day monotherapy investigation of 46 subjects. Moreover, SPOl, the putative active ingredient in SPOlA, has been used clinically for more than 40 years (primarily as an injectable local anesthetic) and has been evaluated in numerous pre-clinical and clinical investigations as safe and well tolerated.

Accordingly, the investigation of SPOlA as a beneficial therapeutic in the treatment of Cushing's Syndrome will initiate with a single-center, double-blind, randomized, placebo-controlled, dose escalation study to investigate the dose response, maximum tolerated dose, maximum effective dose, and safety of SPOlA in individuals inflicted with Cushing's Disease. Additionally, a cohort of healthy volunteers (Cohort E) will be recruited as a control for Cortisol secretion. Cohort E subjects will not receive study medication.

Following a 4-week washout period (if required), subjects will be randomized into an appropriate cohort in accordance with the study schema. Patients will be admitted to an in-patient facility for 72 hours. After an initial night to acclimate to the facility, a 24-hour measurement of Cortisol secretion in blood and urine will be conducted. Patients will then receive an initial single dose of SPOlA orally. Each patient will receive one of four doses as follows: placebo for cohort A, 400 mg for cohort B, 800 mg for cohort C, and 1,200 mg for cohort D. Blood and urine samples will be collected for 24 hours to evaluate the single dose pharmacokinetics of the study medication. At the end of 72 hours, patients will be discharged from the facility.

Six healthy volunteer subjects will also be enrolled as Cohort E. After an initial night to acclimate to the facility, samples (serum and urine) will be collected for baseline 24 hour blood and urine Cortisol secretion determinations and patients will then be discharged. No study medication will be given to patients in Cohort E.

Following a 4-day washout, patients in Cohorts A, B, C, and D will initiate an 8- week study using the four doses described above as follows: placebo bid for Cohort A, 200 mg bid for Cohort B, 400 mg bid for Cohort C, and 400 mg bid for Cohort D.

At the end of the 8-week drug administration period, patients will again be admitted to an in-patient facility for 72 hours. As before, patients will have an initial night to acclimate to the facility followed by a 24-hour measurement of Cortisol secretion in blood and urine. In the morning following and ending the 24 hour basal Cortisol secretion, patients in the four successive cohorts (A, B, C, and D), will receive their last dose of medication and samples (blood and urine) will be collected over the next 24 hours for pharmacokinetic sample collection (blood and urine). Two weeks after their last dose administration, patients in Cohorts A, B,

C, and D will return to the study center for post-treatment examinations and specimen collection as well as evaluation of reactions to study treatment. The total duration of study subject participation will be 11 weeks. In all, subjects will visit the study center at Screening, Initial Dosing, Week 1 (baseline), Weeks 2, 3, 4, 5, 6, and 7, Week 8 (end of treatment), and Week 10 (post-treatment). (See Study Schema, Figure 7).

All publications, patents and patent applications are incorporated herein by reference. While in the foregoing specification this invention has been described in relation to certain preferred embodiments thereof, and many details have been set forth for purposes of illustration, it will be apparent to those skilled in the art that the invention is susceptible to additional embodiments and that certain of the details described herein may be varied considerably without departing from the basic principles of the invention.

Claims

WHAT IS CLAIMED IS:

1. A method for the treatment of Cushing's syndrome in a patient in need thereof, the method comprising administering to the patient an effective amount of a combination of a pharmaceutically acceptable salt of procaine and zinc sulfate heptahydrate, at a frequency and for a duration of time sufficient to provide a beneficial effect to the patient.

2. A method of treatment a malcondition in a patient wherein a reduction of Cortisol production is medically indicated, comprising administering to the patient an effective amount of a combination of a pharmaceutically acceptable salt of procaine and zinc sulfate heptahydrate, at a frequency and for a duration of time sufficient to provide a beneficial effect to the patient.

3. A method for the treatment of autism in a patient in need thereof, the method comprising administering to the patient an effective amount of a combination of a pharmaceutically acceptable salt of procaine and zinc sulfate heptahydrate, at a frequency and for a duration of time sufficient to provide a beneficial effect to the patient.

4. The method of any one of claims 1-3 wherein the pharmaceutically acceptable salt of procaine is procaine HCl.

5. The method of any one of claims 1-4 wherein the effective amount of the combination of a pharmaceutically acceptable salt of procaine and zinc sulfate heptahydrate is administered by oral ingestion.

6. The method of any one of claim 1-5, further comprising administration to the patient of a second medicament, selected from the group consisting of lidocaine HCl, zinc salts other than zinc sulfate heptahydrate, ascorbic acid, phenytoin, clonidine, phosphatidylserine, DHEA, RU-486, HMB, ketaconazole, pregnenalone Ipriflavone, pantothenic acid, acetylsalicylic acid, dimethyl sulphoxide (DMSO), retinol (vitamin A), co-enzyme QlO, acetyl-L-carnitine and ginko beloba.

7. The method of any one of claims 1-6 wherein the weight ratio of procaine HCl to zinc sulfate heptahydrate is about 40:1 to about 160:1.

8. The method of any one of claims 1-7 wherein the patient is an adult and wherein the total dose of the mixture is about 100 mg per day to about 2.0 gm per day.

9. The method of claim 8 wherein the total dose is about 500 mg per day to about 1.0 gm per day.

10. The method of claim 9 wherein the total dose is about 750 mg per day to about 1.0 gm per day.

11. The method of any one of claims 1 -7 wherein the patient is an infant or a child and wherein the total dose of the mixture is about 50 mg per day to about 500 mg per day.

12. The method of claim 11 wherein the total dose of the mixture is about 50 mg per day to about 200 mg per day.

13. A composition comprising a pharmaceutically acceptable salt of procaine and zinc sulfate heptahydrate, adapted for oral ingestion by a patient in the treatment of Cushing's syndrome or autism.

14. The composition of claim 13 wherein the pharmaceutically acceptable salt of procaine is procaine HCl.

15. A pharmaceutical formulation comprising the composition of claim 13 or 14 and a pharmaceutically acceptable carrier.

16. An oral dosage form comprising the pharmaceutical composition of claim 13 or 14 or the pharmaceutical formulation of claim 15.