Nano Octacosanol and Policosanol patent

5/13/2018 Nano Octacosanol and Policosanol patent

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(12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT)(19)World Intellectual Property Organization

International Bureau(43) International Publication Date

1 March 2012 (01.03.2012) peT(51) International Patent Classification:

A61K 9/14 (2006.01)(21) International Application Number:

PCT/US2011l047951(22) International Filing Date:

(25) Filing Language:(26) Publication Language:(30) Priority Data:

611376,194611376,645

16 August 2011 (16.08.2011)English (84)English

23 August 2010 (23.08.2010)24 August 2010 (24.08.2010) USUS

(71) Applicant (for all designated States except US):NANORX, INC. [US/uS]; 6 Devoe Place, Chappaqua,NY 10514 (US).

(72)(75)

Inventor; andInventor/Applicant (forPalayakotai, R. [US/uS];NY 10514 (US).

US only): RAGHAVAN,6 Devoe Place, Chappaqua,

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 11 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 11 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 11 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1(10) International Publication NumberWO 2012/027159 Al

HN, HR, HU, ill, lL, IN, IS, JP, KE, KG, KM, KN, KP,KR, KZ, LA, LC, LK, LR, LS, LT, LU, LY, MA, MD,ME, MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI,NO, NZ, OM, PE, PG, PH, PL, PT, QA, RO, RS, RU,SC, SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, ri,TM,TN,TR,TT,TZ, UA, UG,US, UZ, VC, VN,ZA,ZM,ZW.Designated States (unless otherwise indicated, for everykind of regional protection available): ARIPO (BW, GH,GM, KE, LR, LS, MW, MZ, NA, SD, SL, SZ, TZ, UG,ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, MD, RU, ri,TM), European (AL, AT, BE, BG, CH, CY, CZ, DE, DK,EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, LU,LV, MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK,SM, TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, GN, GQ,GW, ML, MR, NE, SN, TD, TG).

Declarations under Rule 4.17:as to applicant's entitlement to apply for and be granteda patent (Rule 4.17 (iit)as to the applicant's entitlement to claim the priority ofthe earlier application (Rule 4.17 (iiij)-(74) Agent: WONG, Ada, 0.; Morgan, Lewis & BockiusLLP, One Market, Spear Street Tower, San Francisco, CA Published:94105 (US).-; ; ; ; ; ; ; ; ; ; ; ; ; ;; ; ; ; ; ; ; ; ; ; ; ; ; ; ;

; ; ; ; ; ; ; ; ; ; ; ; ; ; ;---(81) Designated States (unless otherwise indicated, for everykind of national protection available): AE, AG, AL, AM,AO, AT, AU, AZ, BA, BB, BG, BH, BR, BW, BY, BZ,CA,CH,CL,CN,CO,CR,CU,CZ,DE,DK,DM,DO,DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT,--; ; ; ; ; ; ; ; ; ; ; ; ; ;----; ; ; ; ; ; ; ; ; ; ; ; ; ;---; ; ; ; ; ; ; ; ; ; ; ; ; ;-

with international search report (Art. 2 1 ( 3) )before the expiration of the time limit for amending theclaims and to be republished in the event of receipt ofamendments (Rule 48.2(h))

(54) Title: POLICOSANOL NANOPARTICLES(57) Abstract: The present invention provides nanoparticulate policosanol, and octacosanol formulations including these parti-cles, as a well as methods of using the particles and formulations for treatment and prophylaxis of various diseases and conditions.


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POLICOSANOL NANOPARTICLES

CROSS-REFERENCE TO RELATED APPLICATION

(0001) This application claims priority to United States Provisional Application No.611376,194, filed August 23,2010, and No. 611376,645, filed August 24,2010, the

5 disclosures of which are incorporated herein by reference in their entirety for all purposes.FIELD OF THE INVENTION

(0002) The present invention relates to nanoparticulate compositions comprising at leastone policosanol, novel nanoparticulate policosanol formulations, and uses thereof. In variousembodiments, the nanoparticulate policosanol particles have an effective average particle size

10 of less than about 100nm.BACKGROUND OF THE INVENTION

[0003) Policosanol is a complex mixture of concentrated long chain N-alkyl alcoholsderived from plant sources, such as sugar cane. Octacosanol is a major constituent ofnaturally derived policosanol, e.g. from sugarcane. Much of the published work on

15 policosanol has been directed to mixtures in which octacosanol was a major component.Early work in Cuba studying the effects of policosanol on serum lipid and lipoprotein levelsin healthy volunteers indicated that, at dosages of2-40 mg/d, policosanol administrationreduced serum lipid and lipoprotein levels (Hernandez et aI., Curro Ther. Res. Clin. Exp.1992; 51: 568), and reduced hypercholesterolemia (Pons et aI., Curro Ther. Res. Clin. Exp.;

20 1992; 52: 507). However, despite numerous subsequent studies, researchers outside Cubahave been unable to verify the claims made in conjunction with the original research. Theineffectiveness of policosanol on serum lipidlcholesterollevels has been comprehensivelydocumented in respected, peer-reviewed journals.[0004) For example, Francini-Pesenti and coworkers conducted double blinded,

25 randomized, placebo-controlled trials of policosanol in subjects with hypercholesterolemiaand concluded that doses of 10mg/d and 20 mg/d of policosano1showed no lipid loweringeffects (Complement Ther. Med.; 2008; 16(2): 61; andPhytother. Res.; 2008; 22(3): 318). Ina similar blind, placebo-controlled study, Berthold and coworkers showed that doses of 10,20,40 and 80 mg/d of policosanoI did not result in lower serum lipid levels than those seen in


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subjects to whom the placebo was administered (JAMA; 2006; 295(19): 2262). Dullens et al.found that neither individual policosanol components (C24, C26, C28, or C30) nor the naturalpolicosanol mixture (all components, 30 mg/IOOg diet) lowered serum cholesterolconcentrations in LDL receptor knockout mice (J. Lipid Res.; 2008; 49(4): 790). Kassis and

5 coworkers studied the efficacy of Cuban sugar cane policosanols for treatinghypercholesterolemia in humans at a dosage of 10mg/d and concluded that policosanol hadno beneficial effects on lipid indicators in hypercholesterolemia subjects (Am. J. Clin. Nutr;2006; 84(5): 1003). Lin and coworkers studied the effects of20 mg/d dosages of wheatgerm policosanol in subjects with normal to mildly elevated plasma cholesterol and detected

10 no lowering of plasma cholesterol (Metabolism; 2004; 53(10): 1309). Lukashevich et al.found that beeswax policosanol (l0 mg or 40 mg) administered daily in tablet or soft gelformulations had no effect on serum lipids in subjects with mild-to-moderatehypercholesterolemia. (Circulation; 2006; 114: 892). Murphy et al. found dietarysupplementation of rabbits with policosanol from sunflower oil did not have any cholesterol

15 lowering effect. (J. Am. College Nutr; 2008; 27(4): 476).[0005] Thus, despite the early apparently promising results of the Cuban research, theconclusion that must be reached from contemporary blinded, placebo-controlled studies isthat art-recognized policosanol formulations are not effective at modulating serum1ipid/chol esterol levels.

20 [0006] Research on the utility of policosanol formulations on other metabolic andphysiologic parameters has produced similarly negative results. For example, policosanolwas shown to have no effect on blood sugar levels, glycemic control (Crespo et al., Int. J.Clin. Pharm. Res.; 1999: 117)or diabetic status (Shinbori et al., Eur. J. Pharmacol; 2007;139-144).

25 [0007] A controversy existed for a time regarding whether the composition or formulationof the policosanol used in the Cuban studies was responsible for the inconsistent resultsbetween the Cuban research and that of other workers. This controversy has been put to restin seminal research. (Kassis, British Journal of Nutrition (2007), 97, 381-388; Kassis,LipidsHealth Dis.(2008); 7:17; Kassis Appl. Physiol. Nutr. Metab; (2008); 33(3): 540 and DuUensJ.

30 Lipid Res. (2008), 49: 790). They studied utilized different sugar cane derived policosanolformulations, including the formulation used in the Cuban research. Their studies concludedthat none of the tested policosanol formulations significantly improved lipid parameters in

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humans or animals relative to the control. Moreover, the in vivo assessment ofLDLoxidation showed no significant alteration in oxidized LDL concentration relative to thebaseline and control. Thus, as of mid-2008, the controversy regarding the serum lipidlowering effects of the Cuban as well as other sugar cane policosanol formulations is resolved

5 outside Cuba.[0008) There are very few reports in the literature on the therapeutic effects of pureoctacosanol. Kim et.al., (Journal of Medicinal Food. (December 2003) ,6(4): 345-351)evaluated the effects of octacosanol on running performance and related biochemicalparameters in exercise-trained rats run to exhaustion on a treadmill. Their results suggest that

10 the ergogenic properties of octacosanol include the sparing of muscle glycogen stores andincreasing the oxidative capacity in the muscle of exercise-trained rats. Ping-ping Zuoet.al.,(Acta Pharmacologica Sinica (July 2010) 31, 765-774) on their rat studies suggestedthat octacosanol may be a promising agent for treatment of Parkinson's disease. Fallat RJ,et.al., (Neurology 1986;36:1263-1264) found that administration of octacosanol in patients

15 with amyotrophic lateral sclerosis in a double-blind, placebo-controlled, crossover study didnot show any benefits to neurologic and pulmonary function. In Jahreis G. (Lipids 2008; 43(2): 109-15), octacosanol administration to humans decreases neutral sterol and bile acidconcentration in feces but serum cholesterol levels were not influenced. S. Kato.et.al.,(British Journal of Nutrition (1995), 73:433-441) fed a high-fat diet rats with octacosanol at

20 very high dose of 10 gmI kilo and found that lipid absorption was not affected by theinclusion ofoctacosanol. Thippeswamy (Eur J Pharmacol2008; 588 (2-3): 141-50) foundthat octacosanol inhibits angiogenesis. It is known that angiogenesis is involved in tumorgrowth and metastasis. Plat et.al (Journal of Lipid Research, Vol. 49, 790-796, April 2008)studied emulsified and fmely dispersed policosanol constituents individually and came to the

25 conclusion that the evaluated individual policosanols as well as the natural policosanolmixtures have no potential in reducing CHD risk through effects on serum lipoproteinconcentrations.

SUMMARY OF THE INVENTION[0009) The present invention provides nanoparticles and nanoparticulate formulations of

30 policosanol as well as methods for making these nanoparticles and formulations. Quitesurprisingly, the policosanol formulations of the invention lower cholesterol and serum lipids.and reduce systolic and diastolic blood pressure. Moreover, the formulations of the invention

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exert antioxidant effects, mitigate insulin resistance and its consequences, and raise vitamin Clevels. Accordingly, the invention also provides methods of treating disease and regulatingmetabolism by administering to a subject a formulation of the invention. The invention alsoprovides methods of regulating metabolism and treating hypertension, hypercholesterolemia

5 as well as several other diseases. For example, the formulation of the invention is of use incontrolling in vivo protein oxidation, regulating blood glucose levels, and can be used to treatand prevent insulin resistance and its consequences, e.g., diabetes, and the deleteriousdownstream effects thereof. Moreover, the formulations of the invention are of use inregulating in vivo vitamin C levels in a subject.

10 2. Octacosanol[0010) The present invention provides nanoparticles and nanoparticulate formulations ofoctacosanol as well as methods for making these nanoparticles and formulations. Quitesurprisingly, the octacosanol formulations of the invention lower cholesterol and serum lipidsand blood glucose levels. Moreover, the formulations mitigate insulin resistance and its

15 consequences. Accordingly, the invention also provides methods of treating disease andregulating metabolism by administering to a subject a formulation of the invention. Theinvention also provides methods of regulating metabolism and treating hypertension,hypercholesterolemia as well as several other diseases. For example, the formulation of theinvention is of use in, regulating blood glucose levels, and can be used to treat and prevent

20 insulin resistance and its consequences, e.g., diabetes, and the deleterious downstream effectsthereof. Moreover, the formulations of the invention are of use in regulating in vivo thelevels ofVEGF (Vascular Endothelial Growth Factor) in diseases such as coronary arterydisease, stroke, and chronic wounds. It can be used to deliver angiogenic growth factors tothe heart, limbs, and wounds in a subject.

25 (0011) Thus, in various embodiments, the invention provides a nanoparticle ofpolicosanol. A representative nanoparticle of the invention includes a policosanol fractioncomprising about 60% to about 95%, e.g., from about 50% to about 69% octacosanol; and astabilizer fraction. In an exemplary embodiment, the stabilizer fraction includes a poly(ethylene glycol) ester. In various embodiments, the stabilizer fraction includes a tocopheryl

30 ester. Exemplary components of the stabilizer fraction include tocopheryl poly (ethyleneglycol) esters, e.g., tocopheryl polyethylene glycol (1000) succinate ("TPGS"). Exemplarynanoparticles of the invention have a diameter of less than about 100 nm.

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[0012) Thus, in various embodiments, the invention provides a nanoparticle ofoctacosanol. A representative nanoparticle of the invention includes a policosanol fractioncomprising about 95% to about 100%, e.g., from about 95% to about 100%octacosanol; anda stabilizer fraction. In an exemplary embodiment, the stabilizer fraction includes a poly

5 (ethylene glycol) ester. In various embodiments, the stabilizer fraction includes a tocopherylester. Exemplary components of the stabilizer fraction include tocopheryl poly (ethyleneglycol) esters, e.g., tocopheryl polyethylene glycol (1000) succinate ("TPGS"). Exemplarynanoparticles of the invention have a diameter of less than about 100 nm[0013) Other objects, advantages, and embodiments of the invention are set forth in the

10 detailed description that follows.BRIEF DESCRIPTION OF THE DRAWINGS

[0014] So that the matter in which the above-recited features, advantages and objects ofthe invention, as well as others which will become clear, are attained and can be understoodin detail, more particular descriptions of the invention briefly summarized above may be had

15 by reference to certain embodiments thereof which are illustrated in the appended Figures.These Figures form a part of the specification. It is to be noted, however, that the appendedFigures illustrate preferred embodiments of the invention and therefore are not to beconsidered limiting in their scope.[0015] FIG. 1 shows the size of the size of the particles of the invention relative to other

20 nanoparticles,[0016] FIG. 2 shows the size distribution of nanoparticles of the invention with a peak atabout 58 nm as measured by light scattering.[0017] FIG. 3 shows the size distribution of nanoparticles of the invention with a peak atabout 61 nm as measured by light scattering.

25 [0018] FIG. 4 shows levels of insulin resistance in untreated rats and rats treated withparticles of the invention at 2 mg/kg and 5 mg/kg.[0019) FIG. 5 shows fasting plasma levels of insulin in untreated rats and rats treatedwith particles of the invention.(0020) FIG. 6 shows levels of total cholesterollHDL ratio in untreated rats and rats

30 treated with particles of the invention at 2 mg/kg and 5 mg/kg.5


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[0021] FIG. 7 shows blood levels of C-reactive protein in untreated rats and rats treatedwith particles of the invention at 1 mg/kg and 2 mg/kg.[0022) FIG. 8 shows levels of glycosylated hemoglobin (HhAIc) levels in untreated ratsand rats treated with particles of the invention at 2 mg/kg and 5 mg/kg.

5 [0023] FIG. 9 shows HDL levels in untreated rats and rats treated with particles of theinvention at 2 mg/kg and 5 mg/kg.[0024] FIG. 10 shows the ICAM-I (inter cellular adhesion molecule) levels in untreatedrats and rats treated with particles of the invention at 2 mg/kg and 5 mg/kg.

[0025] FIG. 11 shows MCP-I (monocyte chemotactic protein-I) levels in untreated rats10 and rats treated with particles of the invention at 2 mg/kg and 5 mg/kg.

[0026] FIG. 12 shows PAl-I (plasminogen activation inhibitor) levels in untreated ratsand rats treated with particles of the invention at 2 mg/kg and 5 mg/kg.[0027J FIG. 13 shows nitric oxide levels in untreated rats and rats treated with particlesof the invention at 2 mg/kg and 5 mg/kg.

15 [0028) FIG. 14 shows protein oxidation levels in untreated rats and rats treated withparticles of the invention at 2 mg/kg and 5 mg/kg.[0029) FIG. 15 shows RBP-4 (retinal binding protein) levels in untreated rats and ratstreated with particles of the invention at 2 mg/kg and 5 mg/kg.[0030) FIG. 16 shows vitamin-C levels in untreated rats and rats treated with particles of

20 the invention at 2 mg/kg and 5 mg/kg.[0031J Figure 17 shows levels of absorbance of DPP-IV assay in untreated rats and ratstreated with particles of invention[0032) FIG. 18 shows the size of the particles of the invention relative to othernanoparticles.

25 [0033] FIG. 19 shows the size distribution ofnanoparticles of the invention with a peakat about 57 nm as measured by light scattering.[0034] FIG. 20 shows levels of insulin resistance in untreated rats and rats treated withparticles of the invention at 5 mg/kg.

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[0035] FIG. 21 shows cholesterol levels in untreated rats and rats treated with particles ofthe invention at 5 mg/kg.[0036] FIG. 22 shows levels of total cholesterol/HDL ratio in untreated rats and ratstreated with particles of the invention 5 mg/kg.

5 (0037] FIG. 23 shows random glucose levels in untreated rats and rats treated withparticles of the invention at 5 mg/kg[0038] FIG. 24 shows fasting insulin levels in untreated rats and rats treated withparticles of the invention at 5 mg/kg.[0039] FIG. 25 shows fasting glucose levels in untreated rats and rats treated with

10 particles of the invention at 5 mg/kg.[0040] FIG. 26 shows triglyceride levels in untreated rats and rats treated with particlesof the invention at 5 mg/kg.[0041] FIG. 27 shows adiponectin levels in untreated rats and rats treated with particlesof the invention at 5 mg/kg.

15 [0042] FIG. 28 shows MCP-I (monocyte chemotactic protein-I) levels in untreated ratsand rats treated with particles of the invention 5 mg/kg.[0043J FIG. 29 shows protein oxidation levels in untreated rats and rats treated withparticles of the invention at 5 mg/kg.[0044J FIG. 30 shows VEGF (vascular endothelial growth factor) levels in untreated rats

20 and rats treated with particles of the invention at 5 mg/kg.[0045] FIG. 31 shows Glycated Hemoglobin levels in untreated and treated rats withparticles of invention at 5 mg/kg

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DETAILED DESCRIPTION OF THE INVENTION AND THEPREFERRED EMBODIMENTS

Definitions[0046] Unless defined otherwise, all technical and scientific tenus used herein generally

5 have the same meaning as commonly understood by one of ordinary skill in the art to whichthis invention belongs.[0047) The term "policosanol" refers to a mixture of concentrated N-alkyl alcohols.Exemplary sources of policosanol are sugar cane and bees wax. The policosanols areextracted by known methods. The long chain alcohols in policosanol are primarily I-

10 Octacosanol, 1-Triacontanol, 1-Tetracosanol, and l-Hexacosanol. Typical commerciallyavailable commercial compositions are composed of 90% minimum fatty alcohols of (a) 1-Tetracosanol: 0-10%; (b) l-Hexacosanol: 2-15%; (c) l-Heptacosanol: 0-0.5%; (d) 1-Octacosanol: 50-70%; (e) l-Nonacosanol: 0-10%; (f) l-Triacontanol: 5-20%; (g) 1-Dotriacontanol: 0.1-10%; and (h) 1-Tetratriacontanol: 0.1-10%.

15 [0048J The tenus "effective average particle size," "particle size" and "size" are usedinterchangeably. The tenus refer to the particle size essentially corresponding to the apex ofa peak produced in an assessment of particle size using light scattering. Useful methods fordetermining the size of the particles of the invention are not limited to light scattering.[0049J The methods and formulations may be used for prophylactic or therapeutic

20 purposes. In some embodiments, the tenus "treating" or "treatment" of any disease ordisorder refers to ameliorating the disease or disorder (i.e., arresting or reducing thedevelopment of the disease or at least one of the clinical symptoms thereof). In otherembodiments, "treating" or "treatment" refers to ameliorating at least one physical parameter,which may not be discernible by the subject. In yet other embodiments, "treating" or25 "treatment" refers to inhibiting the disease or disorder, either physically, (e.g., stabilization oreradication of a discernible symptom), physiologically, (e.g., stabilization or eradication of aphysical parameter) or both. In still other embodiments, "treating" or "treatment" refers todelaying the onset of the disease or disorder.[0050J "Therapeutically effective amount" is used interchangeably herein with "an amount

30 effective to," when referring to a method of the invention. When used in reference to apolicosanol dosage, these tenus refer to a dosage that provides the specific pharmacological

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response for which the policosanol is administered in a significant number of subjects in needof such treatment. It is emphasized that "therapeutically effective amount," administered to aparticular subject in a particular instance may not be effective for 100% of patients treated fora specific disease, and will not always be effective in treating the diseases described herein,

5 even though such dosage is deemed a "therapeutically effective amount" by those skilled inthe art. It is to be further understood that policosanol dosages are, in particular instances,measured as oral dosages, or with reference to drug levels as measured in blood.[0051] As used herein, the terms "individual," "subject," and "patient," are usedinterchangeably to refer to an animal, e.g. a mammal, e.g., a human.

lOA. The Compositions[0052J In various embodiments, the invention provides a nanopartic1e of policosanol. Arepresentative nanoparticle of the invention includes a policosanol fraction comprising about50% to 69% octacosanol and a stabilizer fraction. In an exemplary embodiment, thestabilizer fraction includes a poly (ethylene glycol) ester. In various embodiments, the

15 stabilizer fraction includes a tocopheryl ester. Exemplary components of the stabilizerfraction include tocopheryl poly (ethylene glycol) esters, e.g., tocopheryl polyethylene glycol(1000) succinate ("TPGS"). Exemplary nanopartic1es of the invention have a diameter oflessthan about 100 nm. Also provided are formulations incorporating a plurality of thenanoparticles of the invention, including pharmaceutical formulations.

20 [0053] In various embodiments, the invention provides a nanoparticle of octacosanol. Arepresentative nanoparticle of the invention includes a policosanol fraction comprising about95% to 100% octacosanol and a stabilizer fraction. In an exemplary embodiment, thestabilizer fraction includes a poly (ethylene glycol) ester. In various embodiments, thestabilizer fraction includes a tocopheryl ester. Exemplary components of the stabilizer

25 fraction include tocopheryl poly (ethylene glycol) esters, e.g., tocopheryl polyethylene glycol(1000) succinate ("TPGS"). Exemplary nanoparticles of the invention have a diameter of lessthan about 100 nm. Also provided are formulations incorporating a plurality of thenanoparticles of the invention, including pharmaceutical formulations.[0054] In various embodiments, the nanoparticles include a policosanol fraction that

30 includes at least about 50% octacosanol, at least about 51% octacosanol, at least about 52%octacosanol, at least about 53% octacosanol, at least about 54% octacosanol, at least about55% octacosanol, at least about 56% octacosanol, at least about 57% octacosanol, at least

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about 58% octacosanol, at least about 59% octacosanol, at least about 60% octacosanol, atleast about 61% octacosanol, at least about 62% octacosanol, at least about 63% octacosanol,at least about 64% octacosanol, at least about 65% octacosanol or at least about 66%octacosanol, at least about 67% octacosanol, at least about 68% octacosanol, at least about

5 69% octacosanol,[0055] In various embodiments, the nanoparticles include a policosanol fraction thatincludes not more than about 69% octacosanol, not more than about 68% octacosanol, notmore than about 67% octacosanol, not more than about 66% octacosanol, not more thanabout 65% octacosanol, not more than about 64% octacosanol, not more than about 63%

10 octacosanol, not more than about 62% octacosanol, not more than about 61% octacosanol,not more than about 60% octacosanol, not more than about 59% octacosanol, not more thanabout 58% octacosanol, not more than 57% octacosanol, not more than 56% octacosanol, notmore than 55% octacosanol, not more than 54% octacosanol, not more than 53%octacosanol, not more than 52% octacosanol, not more than 51% octacosanol, or not more

15 than 50% octacosanol.[0056J In an exemplary embodiment, the nanoparticles include a policosanol fractionhaving octacosanol in the range from about 50% to about 69%, from about 51% to about68%, from about 52% to about 67%, from about 53% to about 66%, from about 54% to about65%, from about 55% to about 64%, from about 56% to about 63 %, from about 57% to

20 about 62%, from about 58% to about 61%, from about 59% to about 60%, or about 60%.[0057J In various embodiments, the policosanol fraction includes both octacosanol andtriacontanol. In an exemplary embodiment, the policosanol used has an octacosanol-triacontanol ratio from about 2: 1 to about 7:1 , from about 3: 1 to about 6: 1, from about 4: 1 toabout 5: 1, from about 6:1 to about 7:1,

25 [0058J In various embodiments, the policosanol fraction includes both octacosanol andhexacosanol. In an exemplary embodiment, the policosanol used has an octacosanol:hexacosanol ratio ranging from about 4: 1 to about 9: 1; from about 5: 1 to about 8:1 ; fromabout 6: 1 to about 7: 1; from about 7: 1 to about 8:1 ; from about 8:1 to about 9: 1;(0059) In various embodiments, the policosanol fraction includes both triacontanol and

30 hexacosanol. In an exemplary embodiment, the policosanol used has a triacontanol:hexacosanol ratio of at most about 4:1, at most about 3:1; at most about 2:1; at most about1:1; at most about 0.8:1; at most about 0.6:1; or at most about 4:1.

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[0060] In an exemplary embodiment, the nanoparticles include a policosanol fraction thatincludes from 50% to about 69% octacosanol in admixture with triacontanol at a ratio ofabout 2: 1 to about 7: I and a stabilizer fraction that is essentially completely formed fromTPGS (e.g., at least about 90%,91 %,92%,93%,94%,95%,96%,97% 98% or 99% TPGS).

5 In various embodiments, the policosanol fraction and TPGS are in a ratio of about 1:2.8.[0061] In various embodiments, the invention provides a nanoparticle of octacosanoL Arepresentative nanoparticle of the invention includes a policosanol fraction comprising about95% to 100% octacosanol and a stabilizer fraction. In an exemplary embodiment, thestabilizer fraction includes a poly (ethylene glycol) ester. In various embodiments, the

10 stabilizer fraction includes a tocopheryl ester. Exemplary components of the stabilizerfraction include tocopheryl poly (ethylene glycol) esters, e.g., tocopheryl polyethylene glycol(1000) succinate ("TPGS"). Exemplary nanoparticles of the invention have a diameter of lessthan about 100 nm. Also provided are formulations incorporating a plurality of thenanoparticles of the invention, including pharmaceutical formulations.

15 [0062] In various embodiments, the nanoparticles include a policosanol fraction thatincludes at least about 95% octacosanol, at least about 96% octacosanol, at least about 97%octacosanol, at least about 98% octacosanol, at least about 99% octacosanol, at least about100% octacosanol.[0063] In various embodiments, the nanoparticles include a policosanol fraction that

20 includes not more than about 100% octacosanol, not more than about 99% octacosanol, notmore than about 98% octacosanol, not more than about 97% octacosanol, not more thanabout 96% octacosanol, not more than about 95% octacosanoL[0064] In an exemplary embodiment, the nanoparticles include a policosanol fractionhaving octacosanol in the range from about 95% to about 100%, from about 95% to about

25 96%, from about 96% to about 97%, from about 98% to about 99%, or about 100%.[0065] In various embodiments, the policosanol fraction includes both octacosanol andtriacontanol. In an exemplary embodiment, the policosanol used has an octacosanol-triacontanol ratio from about 45: 1 to about 99: 1[0066] In various embodiments, the policosanol fraction includes both octacosanol and

30 hexacosanol. In an exemplary embodiment, the policosanol used has an octacosanol:hexacosanol ratio ranging from about 40: I to about 99:0.5.

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(0067) In various embodiments, the policosanol fraction includes both triacontanol andhexacosanol. In an exemplary embodiment, the policosanol used has a triacontanol:hexacosanol ratio of at most about 2:1, at most about I:1;at most about 0.8:1; at most about0.6: 1; or at most about 2:1.

5 [0068) In an exemplary embodiment, the nanoparticles include a policosanol fraction thatincludes from 95% to about 100%octacosanol in admixture with triacontanol at a ratio ofabout 45: 1 to about 99:0.5 and a stabilizer fraction that is essentially completely formed fromTPGS (e.g., at least about 95%,96%,97%,98%,99%,99% TPGS). In variousembodiments, the octacosanol fraction and TPGS are in a ratio of about 1:2.8.

10 [0069] The present invention makes use of policosanol or a component of policosanolacquired or isolated from any appropriate source. For example, U.S. Pat. Nos. 5,663,156;5,856,316; 6,197,832; 6,225,354; and 6,596,776, all of which are incorporated herein byreference, disclose policosanol compositions that are specific to the starting material andextraction processes used. In various embodiments, the policosanol of use inmaking a

15 nanoparticle of the invention includes at least about 50% octacosanol, at least about 51%octacosanol, at least about 52% octacosanol, at least about 53% octacosanol, at least about54% octacosanol, at least about 55% octacosanol, at least about 56% octacosanol, at leastabout 57% octacosanol, at least about 58% octacosanol, at least about 59% octacosanol or atleast about 60% octacosanol. In various embodiments, the policosanol used includes between

20 about 60% and about 69% octacosanol. In an exemplary embodiment, the amount ofoctacosanol is from about 57% to about 58%.[0070) The present invention makes use of policosanol or a component of policosanolacquired or isolated from any appropriate source. For example, U.S. Pat. Nos. 5,663,156;5,856,316; 6,197,832; 6,225,354; and 6,596,776, all of which are incorporated herein by

25 reference, disclose policosanol compositions that are specific to the starting material andextraction processes used. In various embodiments, the policosanol of use in making ananoparticle of the invention includes at least about 95% octacosanol, at least about 96%octacosanol, at least about 97% octacosanol, at least about 99% octacosanol, at least about99% octacosanol, at least about 100% octacosanol. In various embodiments, the policosanol

30 used includes between about 91% and about 100% octacosanol. In an exemplaryembodiment, the amount of octacosanol is from about 98% to about 99%.

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[0071] Various exemplary surfactants of use as a stabilizer fraction in the nanoparticle ofthe invention and its formulations include vitamin E TPGS (tocopherol propylene glycolsuccinate, a water-soluble form of vitamin E), sorbitan monolaurate (Span 20), sorbitanmonopalmitate (Span 40), poloxamer, sorbitan monostearate (Span 60), sorbitan monooleate

5 (Span 80), polyoxyethylene (20) sorbitan monolaurate (Tween 20, polysorbate 20),polyoxyethylene (20) monopalmitate (Tween 40, polysorbate 40), polyoxyethylene (20)monostearate (Tween 60, polysorbate 60), polyoxyethylene (20) tri-stearate (Tween 65,polysorbate 65), polyoxyethylene (20) monooleate (Tween 80, polysorbate 80), sucrosemonomyristate, sucrose palmitate/stearate, sucrose stearate, dioctylsulfosuccinate sodium

10 salt, monoglyceride monooleate, monoglyceride monolaurate, monoglyceride monopalmitate,lecithin, diglyceride mixtures, citric acid esters of monoglycerides, acetic acid esters ofmonoglycerides, lactic acid esters of monoglycerides, diacetyl tartaric esters ofmonoglycerides, polyglycerol esters of fatty acids, cyclodextrins, propylene glycol esters offatty acids, stearoyl lactylates, C S - 1 s free fatty acids, PTS (US Patent No 6.045.826) or

15 combinations thereof. In various embodiments, the stabilizer fraction does not include acyclodextrin. In other embodiments, the stabilizer fraction does not include apolyoxyethylene sorbitan fatty acid ester.

[0072J The nanoparticles of the invention can include any useful ratio of policosanolfraction to stabilizer fraction that provides a nanoparticle having a diameter of less than or20 equal to about 100nm. In an exemplary embodiment, the ratio of policosanol fraction:

stabilizer fraction is from about I:1 to about 1:4, for example, from about 1:2 to about 1:3.5.Similarly, in various embodiments, the ratio of octacosanol: stabilizer ranges from about1:1.6 to about I:3.5, for example, from about 1:2 to about 1:2.5. In an exemplaryembodiment, the ratio is about 1:2.8. The ratio of triacontanoI: stabilizer in exemplary

25 nanoparticles of the invention ranges from about 1:5 to about I:30, for example, from about1:6: to about I:30. In an exemplary embodiment, the stabilizer is an ester of vitamin E, suchas TPGS (d-alpha-tocopheryl polyethylene glycol 1000 succinate).[0073) Nanoparticles of the invention can include any useful ratio of policosanol fraction tostabilizer fraction that provides a nanoparticle having a diameter of less than or equal to about

30 100 nm. In an exemplary embodiment, the ratio of policosanol fraction: stabilizer fraction isfrom about 1:I to about I:4, for example, from about 1:2 to about I:3.5. Similarly, in variousembodiments, the ratio of Policosanol: stabilizer ranges from about I:1.6 to about I:3.5, forexample, from about 1:2 to about 1:2.5. In an exemplary embodiment, the ratio is about

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1:2.S. The ratio of triacontanol: stabilizer in exemplary nanoparticles of the invention rangesfrom about 1:225 to about 1:3000, for example, from about I :200: to about 1:3000. In anexemplary embodiment, the stabilizer is an ester of vitamin E, such as TPGS (d-alpha-tocopheryl polyethylene glycol lOOOsuccinate).

5 [0074] The mixture from which the nanoparticles are produced can also include asurfactant in addition to the stabilizer fraction. Exemplary surfactants are set forth above andare generally known to those of skill in the art, e.g., TWEEN 20, TWEEN SO,esters (e.g.,sucrose esters of palmitate and stearic acid monoesters), pectin, agar and the like.10075] The resultant nanoparticulate policosanol formulation can be utilized in solid or

10 liquid dosage formulations, such as liquid dispersions, gels, aerosols, ointments, creams,controlled release formulations, fast melt formulations, lyophilized formulations, tablets,capsules, delayed release formulations, extended release formulations, pulsatile releaseformulations, mixed immediate release and controlled release formulations, etc.(0076] Nanoparticles and pharmaceutical formulations according to the invention may also

15 comprise one or more binding agents, filling agents, lubricating agents, suspending agents,sweeteners, flavoring agents, preservatives, buffers, wetting agents, disintegrants,effervescent agents, and other excipients. Such excipients are known in the art.[0077] Exemplary excipient(s) that can be used in a nanoparticle or a formulation of thenanoparticles include, but are not limited to, soybean lecithin, soybean lecithin derivatives,

20 caprylocaproyl macrogol-S glycerides, medium chain triglycerides, refined olive oil, liquidflavors, polyoxyethylene sorbitan fatty acid esters, sugar esters, polyoxyethylene alkyl ethers,propylene glycol, dexpanthenol, almond oil, rice oil, sunflower oil, soybean oil, sesame oil,glycerin, glyceryl palmitostearate, sweet almond oil, oleic acid, polyglyceryl oleate,saccharose, poloxamer, macrogol-l S hydroxystearate, sorbitan fatty acid ester, ascorbyl

25 palmitate, polethylene glycol, ceralution F, ceralution H, ceralution C, lauroyl macrogol-32glycerides, glycerides, C12-C18 mono-, di- and triglycerides, glyceryl stearate, propyleneglycol laureate, propylene glycol caprylate, propylene glycol dipergonate. In someembodiments, the formulation includes one or more excipients selected from citric acidanhydrous, potassium sorbate, sodium benzoate, and sucrose laureate.

30 [0078] Examples of filling agents are lactose monohydrate, lactose anhydrous, and variousstarches; examples of binding agents are various celluloses and cross-linkedpolyvinylpyrrolidone, microcrystalline cellulose, such as Avicel, PH 10 I and AviceJTM

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PH I02, microcrystalline cellulose, and silicified microcrystalline cellulose (ProSolvSMCCTM).[0079) Suitable lubricants, including agents that act on the flowability of the powder to becompressed, are colloidal silicon dioxide, such as Aerosil200, talc, stearic acid, magnesium

5 stearate, calcium stearate, and silica gel.(0080] Examples of sweeteners are any natural or artificial sweetener, such as sucrose,xylitol, sodium saccharin, cyclamate, aspartame, and acsulfame. Examples of flavoringagents are Magnasweet (trademark of MAFCO) , bubble gum flavor, and fruit flavors, andthe like.

10 (00811 Examples of preservatives are potassium sorbate, methylparaben, propylparaben,benzoic acid and its salts, other esters of para hydroxybenzoic acid such as butylparaben,alcohols such as ethyl or benzyl alcohol, phenolic compounds such as phenol, or quaternarycompounds such as benzalkonium chloride. Exemplary preservatives include, but are notlimited to, potassium nitrite, sodium nitrite, benzoic acid, sodium benzoate, potassium

15 benzoate and calcium benzoate(0082J Suitable diluents include pharmaceutically acceptable inert fillers, such asmicrocrystalline cellulose, lactose, dibasic calcium phosphate, saccharides, and/or mixturesof any of the foregoing. Examples of diluents include microcrystalline cellulose, such asAvicelt'" PHIOI and Avicel PHI02; lactose such as lactose monohydrate, lactose

20 anhydrous, and Pharmatose DCL2I; dibasic calcium phosphate such as Emcompress;mannitol; starch; sorbitol; sucrose; and glucose.(0083] Suitable disintegrants include lightly cross linked polyvinyl pyrrolidone, comstarch, potato starch, maize starch, and modified starches, croscarmellose sodium, cross-povidone, sodium starch glycolate, and mixtures thereof.

25 [0084] Examples of effervescent agents are effervescent couples such as an organic acidand a carbonate or bicarbonate. Suitable organic acids include, for example, citric, tartaric,malic, fumaric, adipic, succinic, and alginic acids and anhydrides and acid salts. Suitablecarbonates and bicarbonates include, for example, sodium carbonate, sodium bicarbonate,potassium carbonate, potassium bicarbonate, magnesium carbonate, sodium glycine

30 carbonate, L-Iysine carbonate, and arginine carbonate. Alternatively, only the sodiumbicarbonate component of the effervescent couple may be present.

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[0085] In various embodiments, the compositions of the invention contain nanoparticulatepolicosanol nanoparticles, which have an average particle size of less than about 100 nm, lessthan about 90 nm, less than about 80 nm, less than about 70 nm, less than about 60 nm, orless than about 50 nm, as measured by light-scattering methods, microscopy, or other

5 appropriate methods.[0086] In one embodiment of the invention, there is provided a formulation in which atleast 99% of the policosanol nanoparticles have a particle size less than about 100 nm, lessthan about 90 nm, less than about 80 nm, less than about 70 nrn, less than about 60 nm, orless than about 50 nm, when measured by the above-mentioned methods.

10 [0087] In one embodiment of the invention, there is provided a formulation in which atleast 95% of the policosanol nanoparticles have a particle size less than about 100 nm, lessthan about 90 nm, less than about 80 nm, less than about 70 nm, less than about 60 nm, orless than about 50 nm, when measured by the above-mentioned methods.[0088] In one embodiment of the invention, there is provided a formulation in which at

15 least about 90% of the policosanol nanoparticles have a particle size less than about 100 nm,less than about 90 nm, less than about 80 nm, less than about 70 nm, less than about 60 nm,or less than about 50 nm, when measured by the above-mentioned methods.[0089) In one embodiment of the invention, there is provided a formulation in which atleast about 85% (e.g., from about 85% to about 99%,95%, or 90%) of the policosanol

20 nanoparticles have a particle size less than about 100 nm, less than about 90 nm, less thanabout 80 nm, less than about 70 nm, less than about 60 nm, or less than about 50 nm, whenmeasured by the above-mentioned methods.[0090J In one embodiment of the invention, there is provided a formulation in which atleast about 80% (e.g., from about 80% to about 99%,95%,90%, or 85%) of the policosanol

25 nanoparticles have a particle size less than about 100 nm, less than about 90 nm, less thanabout 80 nm, less than about 70 nm, less than about 60 nm, or less than about 50 nm, whenmeasured by the above-mentioned methods.[0091] In one embodiment of the invention, there is provided a formulation in which atleast about 75% (e.g., from about 75% to about 99%,95%,90%,85%, or 80%) of the

30 policosanol nanoparticles have a particle size less than about 100 nm, less than about 90 nm,

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less than about 80 run, less than about 70 run, less than about 60 nm, or less than about 50run, when measured by the above-mentioned methods.[0092] In one embodiment of the invention, there is provided a formulation in which atleast about 70% (e.g., from about 70% to about 99%,95%,90%,85%,80%, or 70%) of the

5 policosanol nanoparticles have a particle size less than about 100 run, less than about 90 run,less than about 80 run, less than about 70 run, less than about 60 run, or less than about 50run, when measured by the above-mentioned methods.[0093] In one embodiment of the invention, there is provided a formulation in which atleast about 65% (e.g., from about 65% to about 99%,95%,90%,85%,80%, 75%, or 70%) of

10 the policosanol nanoparticles have a particle size less than about 100 run, less than about 90run, less than about 80 run, less than about 70 run, less than about 60 run, or less than about50 run, when measured by the above-mentioned methods.[0094] In one embodiment of the invention, there is provided a formulation in which atleast about 60% (e.g., from about 60% to about 99%,95%,90%,85%,80%, 75%, or 65%) of

15 the policosanol nanoparticles have a particle size less than about 100 run, less than about 90run, less than about 80 run, less than about 70 run, less than about 60 run, or less than about50 run, when measured by the above-mentioned methods.(0095] In one embodiment of the invention, there is provided a formulation in which atleast 55% (e.g., from about 55% to about 99%,95%,90%,85%,80%, 75%, 70%,65%, or

20 60%) of the policosanol nanoparticles have a particle size less than about 100 run, less thanabout 90 run, less than about 80 run, less than about 70 run, less than about 60 run, or lessthan about 50 nm, when measured by the above-mentioned methods.[0096] Methods to determine the size of the nanoparticle are well known in the art. Forexample, optical diffraction (i.e. optical scatterometry) techniques can be used. These

25 techniques include broadband scatterometry (U.S. Pat. Nos. 5,607,800; 5,867,276 and5,963,329), spectral ellipsometry (U.S. Pat. No.5, 739,909) as well as spectral and single-wavelength beam profile reflectance and beam profile ellipsometry (U.S. Pat. No. 6,429,943).In addition it may be possible to employ single-wavelength laser BPR or BPE to obtain CDmeasurements on isolated lines or isolated vias and mesas (See U.S. patent application Ser.

30 No. 10/243,245, filed Sep. 13,2002).

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[0097) In an exemplary embodiment, the invention provides a unit dosage formulation ofpolicosanol nanoparticles of the invention containing a therapeutically effective amount ofpolicosanoL In an exemplary embodiment, the unit dosage formulation is a formulation ofnanoparticles containing a policosanol fraction and a stabilizer fraction and the unit dosage

5 formulation includes from about 10 mg to about 100 mg, for example from about 10 mg toabout 50 mg. In various embodiments, the unit dosage is a daily dosage. One of ordinaryskill will appreciate that therapeutically effective amounts of policosanol can be determinedempirically and can be employed in pure form or, where such forms exist, inpharmaceutically acceptable salt, ester, or pro-drug form. Actual dosage levels of

10 policosanol in the nanoparticulate compositions of the invention may be varied to obtain anamount of the policosanol that is effective to obtain a desired therapeutic response for aparticular composition and method of administration. The selected dosage level thereforedepends upon the desired therapeutic effect, the route of administration, the potency of theadministered policosanol, the desired duration of treatment, and other factors.

15 [0098) Dosage unit compositions may contain such amounts of such submultiples thereofas may be used to make up the daily dose. Itwill be understood, however, that the specificdose level for any particular patient will depend upon a variety of factors: the type and degreeof the cellular or physiological response to be achieved; activity of the specific agent orcomposition employed; the specific agents or composition employed; the age, body weight,

20 general health, sex, and diet of the patient; the time of administration, route of administration,and rate of excretion of the agent; the duration of the treatment; drugs used in combination orcoincidental with the specific agent; and like factors well known in the medical arts.[0099) In an exemplary method for making a nanoparticle of the invention, a pre-selectedquantity of a stabilizer and policosanol are melted together with stirring to ensure

25 homogeneity. Water or an aqueous solution of an additive, preservative, excipient, etc. at anelevated temperature is added to the melt, and maintained at a desired temperature, generallyfrom about 60C to about 90 C while stirring. In an exemplary method, the mixture ofwater to solids includes more water than solids, e.g., is approximately 10: I by weight. Themixture is stirred while cooling to room temperature. In various embodiments, the method

30 produces a suspension of the particles

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A. The .Methods(0100] In addition to nanoparticles of policosanol and octacosanol and formulationsincluding these nanoparticles, the present invention provides methods of using thesenanopartic1es and formulations to treat and prevent disease and to regulate metabolism. In

5 various embodiments, the nanopartic1es of the invention are of use to regulate hypertension,cholesterol metabolism and treat hyperlipidemia, hypercholesterolemia, inflammation, etc. Invarious embodiments, the formulations are use to regulate or reduce protein oxidation. In anexemplary embodiment, the formulation are of use to manage glycemic levels by reducing,insulin resistance, diabetes, and other conditions related to blood sugar levels. In still further

10 embodiments, the formulations are of use to regulate, e.g., increase, plasma vitamin C levelsand to reduce systolic and diastolic blood pressure. In other embodiments, the formulationsare of use to inhibit PAI-I (Plasminogen activation inhibitor). In other embodiments theformulations are of use in increasing DDP-IV that could be useful in cancer treatment.(0101] In addition to nanoparticles of octacosanol and formulations including these

15 nanoparticles, the present invention provides methods of using these nanoparticles andformulations to treat and prevent disease and to regulate metabolism. In variousembodiments, the nanoparticles of the invention are of use to regulate hypertension,cholesterol metabolism and treat hyperlipidemia, hypercholesterolemia, inflammation, etc. Invarious embodiments, the formulations are use to regulate or reduce protein oxidation. In an

20 exemplary embodiment, the formulation are of use to manage glycemic levels by reducing,insulin resistance, diabetes, and other conditions related to blood sugar levels. In still furtherembodiments, the formulations are of use to regulate, e.g., increase, plasma adiponectin andregulate body weight, and regulate VEGF(vascular endothelial growth factor) levels.[0102] In an exemplary embodiment, the formulations are administered in a therapeutically

25 effective amount to a subject to treat a particular disease or disorder and wherein the subjectis not otherwise in need of treatment with a policosanoL In various embodiments, thenanoparticles are administered to treat a single disease or regulate a single metabolic factor.Thus, in an exemplary embodiment, the invention provides a method to treat insulinresistance in a subject not in need of treatment for hyperlipidemia, hypercholesterolemia,

30 hypertension, etc. In an exemplary embodiment, the invention provides a method ofregulating blood sugar in a subject not in need of treatment for hyperlipidemia,hypercholesterolemia, etc. In various embodiments, the invention provides a method oftreating diabetes (e.g., Type II diabetes) in a subject not in need of treatment for

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hyperlipidemia, hypercholesterolemia, etc. In various embodiments, the invention provides amethod to decrease or prevent protein oxidation in a subject who is not in need of treatmentfor treatment for hypertension, hyperlipidemia, hypercholesterolemia, etc. In an exemplaryembodiment, the invention provides a method of increasing serum vitamin C levels in a

5 subject not in need of treatment for hyperlipidemia, hypercholesterolemia, etc(0103) Non-limiting examples of methods of the invention are set forth below:Protein Oxidation(0104) The invention provides a method of decreasing protein oxidation in a subject and,therefore, reducing the deleterious consequences of this oxidation. The method includes

10 administering to a subject a therapeutically effective amount of a policosanol formulation ofthe invention to decrease protein oxidation in a subject.[0105J Oxidative stress has been implicated in the pathogenesis of acute and chronicdiseases and injury in a variety of pathophysiological conditions such as hepatotoxinexposures, intrahepatic cholestasis, alcoholic liver injury, liver ischemia/reperfusion injury

15 and viral hepatitis (Stehbens, Exp. Mol. Pathol; 2003; 75(3): 265; Jaeschke et aI., Toxieol.Lett; 2003; 144(3): 279-88; McDonough, Toxieology.2003; 189(1-2): 89; Jaeschke et aI.,JClin. Invest; 1988; 81(4): 1240). Overproduction of reactive oxygen species (ROS) andnitrogen species (RNS), along with significant decrease of antioxidant defense in thesepathological conditions, impairs various cellular functions through the processes of lipid per

20 oxidation, protein oxidation and nucleic base oxidation. Lipid peroxidation, for example,causes changes in the physical and chemical properties of cellular membranes, thus alteringtheir fluidity and permeability, leading to impairment in membrane signal transduction andion exchange, resulting in swelling, cytolysis and finally cell death. The oxidation of proteinsand DNA also relates directly to cellular dysfunction and death (Fang Y Z et a1.Nutrition.

25 2002; 18(10): 872-9).[0106J Diseases related to protein oxidation include, but are not limited to, rheumatoidarthritis (IgG, u-l-proteinase inhibitor), ischemia reperfusion injury, emphysema (u-I-proteinase inhibitor, elastase), neurodegenerative diseases (e.g., Alzheimer, Parkinson's),muscular dystrophy, disorders associated with aging (glutamine synthetase, carbonic

30 anhydrase Ill, aconitase), acute pancreatitis, cataractogenesis (n-crystallins), cancer, chronicethanol ingestion, adult respiratory distress syndrome. The formulations of the invention can

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also be used to treat or ameliorate the effects of Kwashiorkor (Manory, J Pediatr; 2000; 137:421).[0107] An exemplary therapeutically relevant dose is one that leads to a reduction in astandard clinical marker of protein oxidation, e.g., protein carboxylation. Protein carbonyl

5 content (PCC) is the most widely used marker of oxidative modification of proteins. Thereare several methodologies for the quantitation of PCC; in various conventional methods, 2,4-dinitrophenyl hydrazine is allowed to react with the protein carbonyls to form thecorresponding hydrazone, which can be analyzed optically by radioactive counting orimmunohistochemically. See, e.g., Yan et al., Arch.Biochem. Biophys.327: 330-334, 1996.

10 [0108] In various embodiments, the nanoparticles of the invention are administered orally.In an exemplary embodiment, the nanoparticles are administered at a dosage of from about10 mg to about 100 mg per day, for example, from about 10 mg to about 50 mg per day.Increase in Serum Vitamin C10109] The invention also provides a method of increasing serum vitamin C in a subject.

15 The method includes administering to a subject a therapeutically effective amount of apolicosanol formulation of the invention to increase or regulate serum vitamin C levels in asubject.[0110J Because insulin resistance and diabetes are accompanied by decreased serumvitamin C levels as compared to the levels observed in healthy subjects, orally administered

20 supplemental vitamin C has been suggested as a treatment for the consequences of insulinresistance and diabetes, including endothelial dysfunction. However, supplemental vitamin Cwas not found to be effective when used alone (Kaneto et al., Diabetes; 1999; 48(12): 2398),quite possibly because orally administered vitamin C supplements does not improveendothelial dysfunction or insulin resistance (Chen et al., Am. J Physiol. Heart Cire.

25 Physiol.; 2006; 290(1): H137).10111 J The present invention provides a method of regulating endogenous vitamin C and,therefore, a method of treating insulin resistance, diabetes and the consequences of thesesyndromes.[0112J An exemplary therapeutically relevant dose is one that leads to an increase in a

30 standard clinical marker of serum Vitamin C concentration, e.g., a sandwich ELISA method

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using commercially available kits from Fisher Thermo Scientific Co, Rockford, Ill. See, e.g.,Washko et al., Anal. Biochem.1992; 204:1-14.[0113] In various embodiments, the nanoparticles of the invention are administered orally.In an exemplary embodiment, the nanoparticles are administered at a dosage of from about

5 10 mg to about 100 mg per day, for example, from about 10 mg to about 50 mg per day.10114] The invention provides a method of increasing VEGF levels in a subject and,therefore, reducing the deleterious consequences in diseases such as coronary artery disease,stroke, and chronic wounds. The method includes administering to a subject atherapeutically effective amount of an octacosanol formulation of the invention to increase

10 VEGF levels in a subject.[0115] Vascular endothelial growth factor (VEGF) is a sub-family of growth factors,specifically the platelet-derived growth factor family of cystine-knot growth factors. Theyare important signaling proteins involved in both vasculogenesis (the "de novo" formation ofthe embryonic circulatory system) and angiogenesis (the growth of blood vessels from pre-

IS existing vasculature). Vascular endothelial growth factor (VEGF) is a chemical signalproduced by cells that stimulates the growth of new blood vessels. It is part of the systemthat restores the oxygen supply to tissues when blood circulation is inadequate. (Folkman J.Harrisiorr's Texbook of Internal Medicine, 15th McGraw-Hill, New York, NY, 2000 pp.132-

20 [0116] VEGF's normal function is to create new blood vessels during embryonicdevelopment, new blood vessels after injury, and new vessels (collateral circulation) tobypass blocked vessels. The normal, healthy body maintains a perfect balance ofangiogenesis modulators. In general, angiogenesis is "turned oft" by the production of moreinhibitors than stimulators. (Li V. et. all., The Role of Therapeutic Angiogenesis in Tissue

25 Repair and Regeneration Adv Skin Wound Care 2005;18:491-500).

[0117] When VEGF is overexpressed, it can contribute to disease. Excessiveangiogenesis occurs in diseases such as cancer, diabetic blindness, age-related maculardegeneration, rheumatoid arthritis, psoriasis, and more than 70 other conditions produceabnormal amounts of angiogenic growth factors, overwhelming the effects of natural

30 angiogenesis inhibitors. Antiangiogenic therapies, aimed at halting new blood vessel growth,

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are used to treat these conditions.

[0118] Insufficient angiogenesis occurs in diseases such as coronary artery disease,stroke, and chronic wounds. In these conditions, blood vessel growth is inadequate, andcirculation is not properly restored, leading to the risk of tissue death. Insufficient

5 occurs when tissues cannot produce adequate amounts of angiogenic growthfactors. Therapeutic angiogenesis, aimed at stimulating new blood vessel growth withgrowth factors, is being developed to treat these conditions. Angiogenic gene therapy is alsobeing developed as a method to deliver angiogenic growth factors to the heart, limbs, andwounds. There are at least 30 known natural angiogenesis inhibitors found in the body

10 Folkman, et.al (Journal of Experimental Medicine 1975;141) demonstrated the firstangiogenesis inhibitor molecule in a study of cartilage.

[0119] Angiogenesis-dependent disease was treated by using interferon alfa2a, anangiogenesis inhibitor, was used to regress the abnormal blood vessels growing in the lungsof a boy with a benign disease called pulmonary hemangiornatosis.I White et.al.,(N Engl J

15 Med 1992;326:1456-1463)

[0120] Pro-angiogenic Therapy fur the Ischemic heart disease has been demonstrated(Stegmann T, et.al., ( Circulation 1998;97(7):645-650)

[0121] VEGF can stimulate endothelial cells to synthesize osteogenic factors and thusindirectly promote bone formation ( Sato K. et.al. Anabolic Endocrinology 1997; 138:2953-

20 2962: Longaker MT. et.al., Plast Reconstr Surg 2002; 109:2384-2397).

[0122] Low levels of (VEGF) ihas been implicated in diabetes (Gurtner, et.al., PNAS,2009, vol. 106, 13505-13510). Both fibroblasts isolated from type 2 diabetic patients, andnormal fibroblasts exposed chronically to high glucose, were defective in their capacity to up-regulate VEGF in response to hypoxia.

25 10123] Exemplary markers of a "therapeutically effective amount" as used herein refers tothe amount of octacosanol in a formulation of the invention which, upon administration (e.g.,oral administration) raises VEGF levels in the subject,

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Insulin Resistance10124] The present invention also provides a method of treating insulin resistance and theconsequences arising out of insulin resistance including, but not limited to, diabetes. Themethod includes administering to a subject an amount of a particulate formulation of the

5 invention sufficient to treat insulin resistance.[0125] Insulin resistance is defined as an inadequate response by insulin target tissues, suchas skeletal muscle, liver, and adipose tissue, to the physiologic effects of circulating insulin.The hallmarks of impaired insulin sensitivity in these three tissues are decreased insulin-stimulated glucose uptake into skeletal muscle, impaired insulin-mediated inhibition of

10 hepatic glucose production in liver, and a reduced ability of insulin to inhibit lipolysis inadipose tissue. In fact, insulin resistance is a major predictor for the development of Type IIdiabetes.10126] In Type II diabetes, it has been widely established that insulin resistance precedesthe development of overt hyperglycemia. The causes of insulin resistance can be genetic

15 and/or acquired. Type II diabetes also predisposes patients to elevated cholesterol andcardiovascular disease. In Western cultures, the most common acquired factors causinginsulin resistance are obesity, sedentary lifestyle, and aging, all of which are interrelated. Inthe presence of a robust compensatory insulin secretory response to insulin resistance,glucose levels can remain relatively normal. However, when insulin-producing pancreatic p

20 cells can no longer compensate for the decreased tissue insulin sensitivity, glucosehomeostasis deteriorates and impaired glucose tolerance and eventually Type II diabetesdevelop.(0127] Common diseases or disorders associated with insulin resistance include acanthosisnigricans, acne vulgaris, allergies, asthma, Alzheimer's Disease, atherosclerosis, bipolar

25 disorder, breast cancer, cardiovascular disease, cataracts, cervical cancer, depression, diabetesmellitus, dyslipidemia, fatty liver disease, childhood Type-2 diabetes, chronic fatigue, colonand rectal cancer, dandruff, Graves' disease, heart disease, high LDL cholesterol, hightriglycerides, hirsutism, hypoglycemia, hypothyroidism, inflammation, kidney disease, lowHDL cholesterol, lupus, neuropathy, neuritis, osteoporosis, pancreatic cancer, Parkinson's

30 disease, polycystic ovary syndrome, prostate cancer, rheumatoid arthritis, scleroderma,seborrhea, strokes, and varicose veins.

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(0128) In addition to the diseases and disorders set forth above, some of the mainconsequences of insulin resistance include, but are not limited to, Type 2 diabetes mellitus,hypertension, arteriosclerosis, polycystic ovarian syndrome, non-alcoholic fatty liver disease,disturbances in the function of the vascular endothelium, elevation of triglycerides and

5 cholesterol, disturbances of clotting, disturbances in kidney function, disturbances in heartrhythm, and elevated uric acid levels.[0129] An exemplary therapeutically relevant dose is one that leads to a decrease in astandard clinical marker of serum insulin resistance, e.g., insulin and insulin levels in theplasma can be determined by the sandwich ELISA method using commercially available kits

10 from Fisher Thermo Scientific Co, Rockford, Ill).10130) In various embodiments, the nanoparticles of the invention are administered orally.In an exemplary embodiment, the nanoparticles are administered at a dosage of from about10 mg to about 100 mg per day, for example, from about 10 mg to about 50 mg per day.Cholesterol-Related Diseases

15 [0131) In various embodiments, the present invention provides a method of treatinghypercholesterolemia and/or regulating lipid metabolism in a subject. The method includesadministering to a subject an amount of a nanoparticulate formulation of the inventionsufficient to treat hypercholesterolemia.10132] Hypercholesterolemia, hyperlipidemia and cardiovascular disease are increasingly

20 prevalent in Western industrial societies. The reasons for this are not completely understood,but may relate partly to a genetic predisposition to these conditions and partly to a diet highin saturated fats, together with an increasingly sedentary lifestyle as manual labor becomesincreasingly less necessary. Hypercholesterolemia and hyperlipidemia are very significant,because they predispose individuals to cardiovascular disease, including atherosclerosis,

25 myocardial infarction (heart attack), and stroke.10133] Specific forms of hyperlipidemia include, for example, hypercholesterolemia,familial dysbetalipoproteinemia, diabetic dyslipidemia, nephrotic dyslipidemia and familialcombined hyperlipidemia. Hypercholesterolemia is characterized by an elevation in serumlow-density lipoprotein-cholesterol and serum total cholesterol. Low-density lipoprotein

30 (LDL cholesterol) transports cholesterol in the blood. Familial dysbetalipoproteinemia, alsoknown as Type III hyperlipidemia, is characterized by an accumulation of very low-density

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lipoprotein-cholesterol (VLDL-cholesterol) particles called beta VLDLs in the serum. Alsoassociated with this condition is a replacement of normal apolipoprotein E3 with abnormalisoform Apo lipoprotein E2. Diabetic dyslipidemia is characterized by multiple lipoproteinabnormalities, such as an overproduction ofVLDL-cholesterol, abnormal VLDL triglyceride

5 lipolysis, reduced LDL-cholesterol receptor activity and, on occasion, Type IIIhyperlipidemia. Nephrotic dyslipidemia, associated with malfunction of the kidneys, isdifficult to treat and frequently includes hypercholesterolemia and hypertriglyceridemia.Familial combined hyperlipidemia is characterized by multiple phenotypes of hyperlipidemia,i.e., Type IIa, lIb, IV, V or hyperapobetalipoproteinemia.

10 10134] It is well known that the likelihood of cardiovascular disease can be decreased if theserum lipids, and in particular HDL-cholesterol, can be increased. It is also well known thatthe progression of atherosclerosis can be retarded or the regression of atherosclerosis can beinduced if total cholesterol to HDL ratio can be lowered. In such cases, individualsdiagnosed with hyperlipidemia or hypercholesterolemia should consider lipid-lowering

15 therapy to retard the progression or induce the regression of atherosclerosis for purposes ofreducing their risk of cardiovascular disease, and in particular coronary artery disease. Suchtherapy will reduce the risk of stroke and myocardial infarction, among other consequences.In addition, certain individuals with what are considered normal serum lipid levels candevelop cardiovascular disease. In these individuals other factors like lipid per oxidation and

20 high levels of Lp(a) or lipoprotein A can lead to atherogenesis despite relatively normalcholesterol and lipid levels.10135] Exemplary markers of a "therapeutically effective amount" as used herein refers tothe amount of policosano I in a formulation of the invention which, upon administration (e.g.,oral administration) to the subject, maintains healthy serum lipid profiles, illustratively by

25 raising HDL cholesterol, lowering the total cholesterol! HDL ratio, and/or loweringtriglycerides, or aids in maintaining a healthy body weight.[0136] Another exemplary therapeutically relevant dose is one that leads to a decrease ofC-reactive protein, a standard clinical marker of lipid metabolism and cardiovascular riskfactor.

30 Plasminogen Activation Inhibitor10137] In various embodiments, the present invention provides a method of regulating, e.g.lowering, the PAI-l protein level in a subject. The method includes administering to a

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subject an amount of a nanoparticulate formulation of the invention sufficient to reduce thePAI-1 level in the subject. PAl -1 is a serine protease inhibitor that inhibits fibrinolysis byinactivating urokinase-type and tissue-type plasminogen activator. Plasma PAI-I activity ishighest between 12 midnight and 6 am. (Kluft C, et al., Thromb Haemost. 1988 Apr 8; 59(2):

5 329-32).[0138] PAI-l is present in increased levels in various disease states (such as a number offorms of cancer), as well as in obesity and the metabolic syndrome. Ithas been linked to theincreased occurrence of thrombosis in patients with these conditions. (Mimuro J(1991)"[Type 1 plasminogen activator inhibitor: its role in biological reactions]". Rinsho

10 Ketsueki 32 (5): 487-9. PMID 1870265.): Binder BR, et al. (2002). "Plasminogen activatorinhibitor 1: physiological and pathophysiological roles". News Physiol, Sci. 17: 56-61. PMID11909993: Hoekstra T, et aI., (2004). "Plasminogen activator inhibitor-type 1: its plasmadeterminants and relation with cardiovascular risk". Thromb. Haemost. 91 (5): 861-72:Lijnen HR (2005). "Pleiotropic functions of plasminogen activator inhibitor-I" . J. Thromb.

15 Haemost. 3 (1): 35-45; De Taeye B, et al., (2005). "Plasminogen activator inhibitor-I: acommon denominator in obesity, diabetes and cardiovascular disease". Current opinion inpharmacology 5 (2): 149-54: Dellas C, LoskutoffDJ (2005). "Historical analysis ofPAI-lfrom its discovery to its potential role in cell motility and disease". Thromb. Haemost. 93 (4):631-40.

20 Retinol Binding Protein[0139J In various embodiments, the present invention provides a method of regulating, e.g.lowering, the RBP-4levels in a subject. The method includes administering to a subject anamount of a nanoparticulate formulation of the invention sufficient to reduce the RBP-4levels in a subject and thereby treat diseases associated with elevated RBP-4 levels.Retinol

25 binding protein 4 (RBP4) has recently been described as an adipokine that contributes toinsulin resistance in the AG4KO mouse model. See Yang Q, et aI., Serum retinol bindingprotein 4 contributes to insulin resistance in obesity and type 2 diabetes", Nature 436 (7049):356-62. (2005). RBP4 is elevated in the serum before the development of diabetes andappears to identify insulin resistance and associated cardiovascular risk factors in subjects

30 with varied clinical presentations. These findings provide a rationale for antidiabetic therapiesaimed at lowering serum RBP41evels. See Graham TE, et.al N Engl J Med. 2006 Jun 15;354(24):2552-63.

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Nitric Oxide[0140] In various embodiments, the present invention provides a method of increasing nitricoxide levels in a subject. The method includes administering to a subject an amount of ananoparticulate formulation of the invention sufficient to increase nitric oxide levels in a

5 subject.

[01411 Endothelial NOS (eNOS), also known as nitric oxide synthase 3 (NOS3), generatesNO in blood vessels and is involved with regulating vascular function and in reducing bloodpressure. See Alderton WK,et aI., Biochem J. 2001 Aug 1; 357(Pt 3):593-615. NO is knownto have various vascular effects, includingdirect vasodilatation (flow dependent and receptor

10 mediated); indirect vasodilatation by inhibiting vasoconstrictor influences (e.g., inhibitsangiotensin II and sympathetic vasoconstriction), anti-thrombotic effect (inhibits plateletadhesion to the vascular endothelium), anti-inflammatory effect (inhibits leukocyte adhesionto vascular endothelium), the ability to scavenge for superoxide anion; and anti-proliferativeeffect (e.g., inhibits smooth muscle hyperplasia).

15 [01421 Because of the above-mentioned actions of NO, its impaired production or reducedbioavailability can result in vasoconstriction (e.g., coronary vasospasm, elevated systemicvascular resistance, hypertension); thrombosis due to platelet aggregation and adhesion tovascular endothelium; inflammation due to up-regulation of leukocyte and endothelialadhesion molecules; vascular hypertrophy and stenosis. See Nitric Oxide: Biology and

20 Pathobiology; By: Louis J. Ignarro (Editor) ISBN-I0: 0123738660; ISBN-13:9780123738660 Publisher: Academic Press 2009. Diseases or conditions associated withabnormal NO production and bioavailability include, without limitation, hypertension;obesity; dyslipidemias (particularly hypercholesterolemia and hypertriglyceridemia); diabetes(types I and II); heart failure; atherosclerosis; and conditions associated with aging. See

25 Dessy, C, et aI., (September 2004). "Pathophysiological Roles of Nitric Oxide: In the Heartand the Coronary Vasculature" and Current Medical Chemistry Anti-Inflammatory &Anti-Allergy Agents in Medicinal Chemistry (Bentham Science Publishers Ltd.) 3 (3): 207-216.MCP-l (Monocyte Chemotactic Protein-I)101431 In various embodiments, the present invention provides a method of regulating, e.g.

30 lowering, the MCP-I levels in a subject. The method includes administering to a subject anamount of a nanoparticulate formulation of the invention sufficient to reduce the MCP-l

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levels in a subject and thereby treat diseases associated with elevated MCP-1 levels.MCP-1 isa chemokine that recruits monocytes to sites of inflammation. MCP-1 is expressed andsecreted by adipocytes and stromal vascular cells in white adipose tissue. Obese rodents havehigher circulating MCP-1 levels with increased adipose tissue expression of MCP-I. See J

5 Biol Chern. 2003 Nov 21,278(47): 46654-60. Furthermore, MCP-1 can directly contribute toinsulin resistance by decreasing insulin stimulated glucose uptake and insulin induced insulinreceptor tyrosine phosphorylation and can decrease the expression of adipogenic genes so asto inhibit adipocyte growth and differentiation.[0144] Administration ofMCP-1 to mice peripherally increases circulating monocytes, and

10 increases monocyte accumulation in arteries with neo-intimal formation suggesting a role forMCP-1 in atherogenesis. See Cardiovasc Res. 2003 Jan; 57(1): 178-85.ICAM-l (Inter cellular adhesion molecule-I)10145] In various embodiments, the present invention provides a method of regulating, e.g.reducing, elevated ICAM-1 levels in a subject. The method includes administering to a

15 subject an amount of a nanoparticulate formulation of the invention sufficient to reduceICAM-1 levels in a subject and thereby treat diseases associated with elevated ICAM-1levels.ICAM-l is the same receptor molecule used by the vast majority of viruses that causethe common cold. Rhinoviruses are the frequent cause of the common cold. Adhesionmolecules playa major role in many fields of medicine including embryology, immunology,

20 and malignancy.10146] Many physiological processes require that cells come into close contact with andadhere to other cells or the extracellular matrix. Cell-cell and cell-matrix interactions aremediated through several families of intercellular adhesion molecules or "ICAMs." See NewCell adhesion research, Patrick Nott and other contributors, ISBN-10: 1606923781; ISBN-13:

25 9781606923788 Publisher: Nova Biomedical Books - 2009-04. ICAM-1 therefore plays anessential role in both normal and pathophysiological processes (Springer et aI., 1987, Ann.Rev. Immuno!. 5: 223-252). Strategies have therefore been developed to mediate celladhesion by blocking ICAM-1 function or expression. Such strategies typically employ anti-ICAM-1 antibodies, ligands which competitively block ICAM-1 binding, or antisense nucleic

30 acid molecules directed against ICAM-1 mRNA. However, the agents used in such therapiesproduce only a stoichiometric reduction in ICAM-1, and are typically overwhelmed by theabnormally high production of ICAM-1 by the diseased or activated cells.

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[0147] Diseases characterized by infiltration of neutrophils are often associated withchronic conditions wherein ICAM-l or VCAM-l expression predominates. See Adams DH,Shaw S 1994 Leukocyte endothelial interactions and regulation of leukocyte migration.Lancet 343:831-836. An increase in local expression as well as in serum-soluble adhesion

5 molecules has been reported in diverse pathologic conditions including arteriosclerosis,vasculitis, arthritis, renal and hepatic diseases, ischemia reperfusion conditions, organrejection, metastasis, and many more pathologic conditions. See Bevilacqua MP, et aI., 1994Endothelial leukocyte adhesion molecules in human disease. Annu Rev Med 45:361-378.[0148] Adhesion molecules may be important in specific forms of inflammation. See

10 Gorski A 1994 the role of cell adhesion molecules in immunopathology. Immunol Today15:251-255. What is needed, therefore, are agents in catalytic or sub-stoichiometric amountswhich selectively inhibit expression ofICAM-l, in order to effectively decrease or blockICAM-l-mediated cell adhesion.[0149] One of the many different ways of inhibiting viral infection is to stop the virus from

15 binding to cells. Most of the rhinovirus serotypes use a single cellular receptor, i.e. theIntercellular Adhesion Molecule-I (ICAM-l) for attachment to the cells. This could lead tothe development of blockers of this receptor in an effort to find a cure for the common cold.10150] Expression of ICAM-l has also been associated with a variety of inflammatory skindisorders such as allergic contact dermatitis, fixed drug eruption, lichen planus, and psoriasis.

20 See Ho et aI., 1990, J. Am. Acad. Dermatoi. 22: 64-68; Griffiths and Nickoloff, 1989, Am. J.Pathology 135: 1045-1053; Lisby et aI., 1989, Br. 1. Dermatoi. 120: 479-484; and Shiohara etaI., 1989, Arch. Dermatoi. 125: 1371-1376. In addition, ICAM-l expression has beendetected in patients with rheumatoid arthritis (Hale et aI., 1989, Arth. Rheum. 32: 22-30); inpancreatic B-cells of diabetics (Campbell et aI., 1989, P.N.A.S. USA 86: 4282-4286); in

25 thyroid follicular cells of patients with Graves' disease (Weetman et aI., 1989, J. Endocrinoi.122: 185-191); in renal and liver allograft rejection (Faull and Russ, 1989, Transplantation48: 226-230; Adams et aI., 1989, Lancet 1122-1125); and in inflammatory bowel disease(IBD) tissue (Springer T, 1990, Nature 346: 425-34).10151] Complications commonly observed in type I diabetes also involve expression of

30 ICAM-l. For example, ICAM-l-mediated adhesion of leukocytes to capillary endotheliumcan cause microvascular ischemia in certain tissues of diabetics, such as the retina, peripheralnerves, and kidney. This results in capillary non-perfusion of these tissues, which in tum

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leads to diabetic retinopathy, neuropathy or nephropathy, or angiogenesis induced byadhesion between polymorphonuclear leukocyte and endothelial cell via intercellularadhesion molecule-l , (ICAM-l). It is therefore believed that inhibition of ICAM-l-mediatedleukostasis can prevent retinal abnormalities associated with diabetes. See Miyamoto K et al.

5 (2000), Am. J. Pathol. 156: 1733-1739; Miyamoto K et al. (1999), P.N.AS USA 96: 10836-1084; Jude E B et al. (1998), Diabetologia 41 :330-6; Miyamoto et al. 1999, P.N.AS USA 96:10836-10841; and Yong Song Gho et.al Cancer Research 59, 5128 ~5132, October 15,1999.[0152] DPPIV (DPP4, CD26) is a member of the class of proteases known as prolylpeptidases, which cleave proteins after proline residues (lS.Rosenblum et al.,Curr. Opin.

10 Chern. BioI. 2003 7,496). DPPIV, a serine dipeptidyl peptidase, cleaves the N-terminal X-Ala or X-Pro from target polypeptides, such as chemokines (e.g. CXCLlI) and peptidehormones (e.g., glucagon-like peptide-I, GLP-l).[0153] Dipeptidyl peptidase IV (DPP-IV) is involved in diverse biological processes suchas cellular differentiation, T cell activation, and cell-matrix interaction. DPPIV inactivates

15 certain growth factors, chemokines, and neuropeptides by peptide cleavage. It is widelyexpressed in various normal tissues including normal melanocytes, lung, and prostateepithelial cells. Thus it is emerging as a target for cancer and autoimmune diseases.[0154] DPPIV possesses a transmembrane region and a very short cytoplasmic domain, butis often cleaved and released as a soluble, circulating form. It is found as a dimer with itself

20 or with FAP (fibroblast activation protein-a, seprase), another prolyl peptidase. It also hasnon-peptidase functions: through its interaction with adenosine deaminase (ADA) andextracellular matrix components, it influences T-cell activation and proliferation (I.Ben-Shooshan et al. Biochem. Biophys. Acta 2002 158721 ). Though its role in diabetes is wellknown and drugs like januviaa in the market for diabetes ( US patent 7,026,316 B2, 2006).

25 Interestingly DPPIV expression is markedly decreased or completely extinguished in cellsderived from melanomas, lung cancer, prostate cancer, and tumorigenic, in vitro transformedepithelial cells. Ithas became a target of interest in cancer and auto immune diseases(Wesley UV.et.al .,2008. Frontiers in Bioscience 13,2435-2443: B. Pro, et al., HistolHistopathol (2004) 19: 1345-1351; Havre PA, et.al., Front. Biosci. 13: 1634-45.). The

30 results support the view that down regulation of DPPIV is an important event in progressionof melanoma, lung, and prostate cancers. Re-expression of DPPIV using tetracyclineinducible system, in melanomas and in non-small cell lung cancer cells, resulted in profound

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phenotypic changes that are characteristics of normal cells. DPPIV re-expression led toremoval of block in differentiation, acquired dependence on exogenous growth factor for cellsurvival, and loss of tumorigenicity.[0155] In vitro studies confirm that DPP-4 inhibition increases the metastatic potential of

5 colon [Masur K, et aI., Reg Pep. 2006; 137: 147-155.5]. In addition, dipeptidyl peptidaseinhibits the maligant phenotype of prostate cancer cells by blocking basic fibroblast growthfactor signaling pathway. [Wesley UV, et aI., Cancer Res. 2005;65:1325-1334. 6]. Asignificant decrease in serum DPPIV activity has been reported in patients with oral cancersas compared to healthy subjects.(Uetmasu et aI., 1. Oral PathoI. Med. 27, 106-110). Patients

10 with colorectal cancer (Cordero et aI., 2000; Br. 1. Cancer 83, 1139-1146.). have lower levelsof serum CD26 than normal donors.(0156] These results suggest that DPPIV is a tumor suppressor gene and, given the multiplefunctions ofCD26 and its potential involvement in tumor biology, the development oftherapeutic modalities targeting it may prove to be a useful strategy in the treatment of

15 selected tumors.(0157] In various embodiments, the nanoparticles of the invention are administered orally.In an exemplary embodiment, the nanoparticles are administered at a dosage of from about10mg to about 100 mg per day, for example, from about 10mg to about 50 mg per day.(0158] In various embodiments of each of the methods of the invention, including the

20 exemplary methods set forth above, the subject treated with the formulations of the inventionis not in need of vitamin E supplementation. In various other embodiments, the metabolicparameter that is regulated or the disease that is treated by administration of the formulationis not a parameter or disease recognized as treatable or known to be ameliorated by vitamin Esupplementation of the subject.

25 (0159] Art-accepted assays for alteration in blood chemistry or metabolism are of use toconfirm the efficacy of therapeutically relevant (or other) dosages of the particles of theinvention. The following provide examples illustrating just some of the conventional assaysthat can be used to analyze the efficacy of the present invention in varying dosages. Standardassays for cytokines, insulin, lipid peroxidation and vitamin C, CRP, MCP-l, IL-6, TNF-a,

30 leptin, VEGF, DPP-IV, retinol binding protein and insulin levels in the plasma can take theform of the sandwich ELISA method using commercially available kits from Fisher ThermoScientific Co, Rockford, Ill. VEGF was measured using ELISA kit purchased from Rand 0

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system (Minneapolis, MN). DPPV-IV was measured using kit purchased from CaymanChemical (Ann Arbor Mlj.Dxidative stress can be determined by measuringmalondialdehyde (an end product oflipid peroxidation) by its reaction with thiobarbituricacid. See Jain, J. Bioi. Chern. 264:21340-21345, 1989; Jain et al., Diabetes 38: 1539-1543,

5 1989). Protein oxidation can be determined by the methods disclosed in Yan et al., Arch.Biochem Biophys. 327:330-334, 1996. Insulin resistance can for instance be determined bythe HOMA method (Yaturu et al., Cytokine 34:219-23, 2006). Vitamin C concentration inthe plasma can be determined by the method of Nino and Shaw. See Alan Wu (Ed). TeitzClinical Guide to Laboratory Tests (Fourth Edition), Philadelphia, WB Saunders Co. 2006.

10 Glycosylated hemoglobin can be determined using Glyco- Tek Affinity column kits andreagents (cat # 5351) purchased from Helena Laboratories (Beaumont, TX). Glucose levelscan be determined using glucose oxidase by Accu-check Advantage glucometer (BoehringerManheim Corporation, Indianapolis, IN).[0160] The effects on biological parameters of an exemplary formulation of the invention

15 are forth in Table 1.

95-100% CommercialOctacosanol nano Octacosanol nano Policosanolparticle+ TPGS

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