(Hiv) Protease Inhibitors Using Cellulosic Surface

Europäisches Patentamt *EP001002065B1* (19) European Patent Office

Office européen des brevets (11) EP 1 002 065 B1

(12) EUROPEAN PATENT SPECIFICATION

(45) Date of publication and mention (51) Int Cl.7: C12N 9/99, A61K 38/55, of the grant of the patent: A61K 9/18 21.09.2005 Bulletin 2005/38 (86) International application number: (21) Application number: 98931809.2 PCT/US1998/014097

(22) Date of filing: 09.07.1998 (87) International publication number: WO 1999/002665 (21.01.1999 Gazette 1999/03)

(54) NANOCRYSTALLINE FORMULATIONS OF HUMAN IMMUNODEFICIENCY VIRUS (HIV) PROTEASE INHIBITORS USING CELLULOSIC SURFACE STABILIZERS AND METHODS OF MAKING SUCH FORMULATIONS NANOKRISTALLINE ZUBEREITUNGEN VON MENSCHLICHEN IMMUNSCHWÄCHEVIRUS (HIV)-PROTEASE-IN-HIBITOREN UNTER VERWENDUNG VON CELLULOSE-OBERFLÄCHENSTABILISATOREN UND VERFAHREN ZU DEREN HERSTELLUNG FORMULATIONS NANOCRISTALLINES D’INHIBITEURS DE LA PROTEASE DU VIRUS DE L’IMMUNODEFICIENCE HUMAINE (VIH) FAISANT APPEL A DES STABILISANTS SUPERFICIELS CELLULOSIQUES ET PROCEDES DE PREPARATION DE CES FORMULATIONS

(84) Designated Contracting States: (74) Representative: Dörries, Hans Ulrich et al AT BE CH CY DE DK ES FI FR GB GR IE IT LI LU Dörries Frank-Molnia Pohlman MC NL PT SE Postfach 22 16 61 80506 München (DE) (30) Priority: 09.07.1997 US 890602 (56) References cited: (43) Date of publication of application: WO-A1-95/09614 US-A- 5 145 684 24.05.2000 Bulletin 2000/21 • THAISRIVONGS, S. et al. Structure-Based (73) Proprietor: Elan Pharma International Limited Design of Novel HIV Protease Inhi- bitors: Shannon, Co. Clare (IE) Sulfonamide-Con- taining 4-Hydroxycoumarins and 4-Hydroxy-2-pyrones as Potent (72) Inventors: Non-Peptidic In- hibitors. Journal of Medicinal • LIVERSIDGE, Gary, G. Chemistry, 1996, Vol. 39, No. 12, pages West Chester, PA 19380 (US) 2400-2410, XP002900253 • ENGERS, David, A. • BENDER A. ET AL.: "Efficiency of nanoparticles Mystic, CT 06355 (US) as a carrier system for antiviral agents in Human • ROBERTS, Mary, E. Immunedeficiency Virus-infected human Downingtown, PA 19335 (US) monocytes/macrophages in vitro" • RUDDY, Stephen, B. ANTIMICROBIAL AGENTS AND Schwenksville, PA 19473 (US) CHEMOTHERAPY, vol. 40, no. 6, June 1996 • WONG, Sui-Ming (1996-06), pages 1467-1471, US Collegeville, PA 19426 (US) • XU, Shuqian Phoenixville, PA 19460 (US)

Note: Within nine months from the publication of the mention of the grant of the European patent, any person may give notice to the European Patent Office of opposition to the European patent granted. Notice of opposition shall be filed in a written reasoned statement. It shall not be deemed to have been filed until the opposition fee has been paid. (Art. 99(1) European Patent Convention). EP 1 002 065 B1

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Description duce particle size, but that flocculation restricts the lower particle size limit to approximately 10 microns (10,000 BACKGROUND OF THE INVENTION nm). Commercial airjet milling techniques have provided particles ranging in average particle size from as low Field of the Invention 5 as about 1 to 50 µm. [0005] Other techniques for preparing pharmaceuti- [0001] The present invention relates to formulations cal compositions to aid in dissolution and increase bio- of nanoparticulate human immunodeficiency virus (HIV) availability include loading drugs into liposomes or pol- protease inhibitor drug substances comprising a cellu- ymers, such as during emulsion polymerization. losic surface stabilizer. The nanoparticulate formula- 10 [0006] For example, Bender et al., "Efficiency of Na- tions have an increased rate of dissolution in vitro,an noparticles as a Carrier System for Antiviral Agents in increased rate of absorption in vivo, a decreased fed/ Human Immunodeficiency Virus-Infected Human Mono- fasted ratio variability, and a decreased variability in ab- cytes/Macrophages In Vitro", Antimicrobial Agents and sorption. The present invention is also directed to meth- Chemotherapy, 40(6):1467-1471 (1996), is directed to ods of making the novel formulations. In particular, na- 15 polyhexylcyanoacrylate nanoparticles loaded with the noparticulate formulations of HIV type 1 (HIV-1) and HIV protease inhibitor saquinavir. The nanoparticles are type 2 (HIV-2) protease inhibitors employing cellulosic prepared by emulsion polymerization of the monomer in stabilizers are disclosed. an acidic medium containing sodium sulfate and poloxamer 188. After polymerization, the nanoparticu- Description of the Related Art 20 late preparation is neutralized, ultrasonicated, and diluted. Thus, Bender et al. is directed to solubilizing an HIV [0002] It is known that with an increase in surface area protease inhibitor, followed by polymerizing the solubi- of a particulate therapeutic agent, the rate of dissolution lized HIV protease inhibitor within a polymer to form na- of the agent increases. Such an increase in surface area noparticles of entrapped HIV protease inhibitor and pol- can be obtained by decreasing the particle size of the 25 ymer. agent. Increasing the rate of dissolution of a therapeutic [0007] However, such techniques as in Bender et al. agent is often desirable because it can result in an in- exhibit problems and limitations. For example, a lipid creased rate of absorption in vivo, increased bioavaila- soluble drug is often required in preparing suitable lipo- bility, and decreased variability in absorption of the somes. Further, unacceptably large amounts of the lipo- agent. 30 somes or polymer are often required to prepare unit drug [0003] Bioavailability is the degree to which a drug be- doses. Further still, techniques for preparing such phar- comes available to the target tissue after administration. maceutical compositions tend to be complex. Many factors can affect bioavailability, including the dos- [0008] One solution to the problem of preparing wa- age form and dissolution rate of the drug. Poor bioavail- ter-insoluble drugs previously suggested is to provide ability is a significant problem encountered in the devel- 35 stable dispersible drug particles in the submicron size opment of pharmaceutical compositions, particularly range, as described in U.S. Patent No. 5,145,684 ("the those containing an active ingredient that is poorly sol- '684 patent"). The submicron sized particles can be pre- uble in water. Poorly water soluble drugs tend to be elim- pared by wet milling in the presence of grinding media inated from the gastrointestinal tract before being ab- in conjunction with a surface stabilizer. Such particles sorbed into the circulation of the patient. Moreover, 40 can be readily prepared, do not appreciably flocculate poorly water soluble drugs tend to be unsafe for intra- or agglomerate due to interparticle attractive forces, and venous administration techniques, which are used pri- do not require the presence of a cross-linked matrix. In marily in conjunction with fully soluble drug substances. the '684 patent, the inventors hypothesized that the sur- [0004] As noted above, it is known that by increasing face stabilizer hinders the flocculation and/or agglomer- the surface area of a particulate drug, such as by de- 45 ation of the drug particles by functioning as a mechani- creasing the particle size of the drug, the rate of disso- cal or stearic barrier between the particles, thereby min- lution of the particulate drug is increased. Consequently, imizing the close, interparticle approach necessary for efforts have been made to control the size and size agglomeration and flocculation. However, as noted in range of drug particles in pharmaceutical compositions the '684 patent, not all stabilizers will function to produce and methods of making fine particulate drugs have been 50 a nanoparticulate composition of any drug. studied. For example, dry milling techniques have been [0009] Drug substances which have been difficult to used to reduce particle size and thereby influence drug formulate prior to the present invention because of their absorption. However, in conventional dry milling, as dis- low solubility include HIV protease inhibitors, which are cussed by Lachman et al., "The Theory and Practice of useful in the treatment of AIDS (acquired immune defi- Industrial Pharmacy," Milling, 45 (1986), the limit of fine- 55 ciency syndrome), caused by HIV infection. ness is reached at about 100 microns (100,000 nm) [0010] AIDS was first recognized in 1981 as a clinical when material cakes on the milling chamber. Lachman syndrome consisting of opportunistic infection and/or et al. also note that wet grinding is useful to further re- neoplasia associated with unexplained immunodefi-

2 3 EP 1 002 065 B1 4 ciency. Shaw et al., AIDS: Etiology, Diagnosis, Treat- tion Institute Antiviral Agents Bulletin, March, 1995. It ment, and Prevention, 2nd Edition, DeVita, Jr. et al., has also been reported that many HIV protease inhibi- eds., 11-31 (J.B. Lippincott Co., 1988). AIDS is a com- tors, such as saquinavir, are poorly soluble and there- plex disease that includes progressive destruction of the fore not very good at getting into the bloodstream. Step immune system and degeneration of the central and pe- 5 By Step, The Economist Newspaper, Nov. 26, 1994, at ripheral nervous system. The discovery of HIV as the 93. Moreover, early trials of an HIV protease inhibitor etiological agent of AIDS followed several years later. developed by Searle were stopped because the drug Id. at 12-13. seemed unable to enter the body's cells. Id. [0011] HIV-1 and HIV-2 are retroviruses, and like all [0015] With insolubility and low bioavailability, HIV retroviruses, they have a RNA-dependent DNA 10 protease inhibitors have to be given in huge doses to polymerase. In the life cycle of the HIV virus, the cell- effect results. Id. For example, it has been reported that free virion first binds to the target cell. Following virus saquinavir is administered at 1800 mg/day, and indinavir adsorption, reverse transcription catalyzed by the viral (MK-639) is administered at 2400 mg/day, while the av- RNA-dependent DNA polymerase generates a double- erage amount of drug used to treat most diseases is stranded DNA copy of the single-strand viral RNA ge- 15 10-30 mg/day. Id. Thus, the dosage of a typical HIV pro- nome. Subsequent expression of viral DNA is controlled tease inhibitor can be up to 24 times that of a typical by a combination of viral and host cellular proteins in- drug. The low oral bioavailability and rapid biliary excre- teracting with viral DNA regulatory elements. tion of many HIV protease inhibitors have limited their [0012] One method of treating HIV infection targets utility as potential therapeutic agents. Thaisrivongs et the reverse transcriptase (RT). Drugs which interfere 20 al., J. Med. Chem., 39:2400-10 (1996). Such large dose with the HIV RT are nucleoside analogs, which include requirements also translate to expensive drug protocols AZT (azidothymidine), ddI (didanosine), and ddT (dithi- for patients. othreitol). However, such drugs are only modestly effec- [0016] HIV protease inhibitors have also been found tive and are plagued by severe side effects because to have an extremely poor taste. The poor taste, in con- they interfere with cellular enzymes similar to the HIV 25 junction with the necessity of large doses of the drugs, viral enzyme they target. has made it extremely difficult, if not impossible, to de- [0013] An alternative method of treating HIV infection velop formulations of the drugs which are palatable to targets the HIV protease. The HIV protease, also known patients. Even large doses of sweeteners have been un- as proteinase, is essential for the final assembly of via- successful in masking the unpleasant taste of the drugs. ble viral particles in HIV replication. Basically, the HIV 30 [0017] There is a need in the art for compositions of protease cuts and trims the proteins that give the virus HIV protease inhibitors which provide for high bioavail- its structure. More specifically, the HIV protease cleaves ability and which are soluble in water. In addition, there a number of proteins from the viral Gag-Pol polyprotein is a need in the art for methods of making such compo- precursor, including the HIV protease, the reverse tran- sitions. The present invention satisfies these needs. scriptase and integrase encoded in the pol gene, and 35 the matrix, capsid, and nucleocapsid encoded in the gag SUMMARY OF THE INVENTION gene. If the Gag-Pol precursor molecule is not proc- essed in this way, noninfectious particles are formed. [0018] The present invention is directed to the surpris- More importantly, HIV protease is unlike other enzymes ing and unexpected discovery that the use of cellulosic in human cells and, therefore, inhibition of the HIV pro- 40 surface stabilizers in nanoparticulate formulations of tease should not interfere with normal human cellular HIV protease inhibitors can form superior, stable, and action. Inhibition of the HIV protease therefore repre- dispersible compositions. This discovery is an improve- sents an attractive treatment of HIV infection. ment upon the invention described in U.S. Patent No. [0014] However, the identification of HIV protease in- 5,145,684. hibitors with antiretroviral activity in vivo has been ham- 45 [0019] It is an advantageous feature that the cellulosic pered by the poor bioavailability of many molecules in surface stabilizers can be used to prepare nanoparticu- this class. For example, it has been reported that the late formulations of a wide range of HIV protease inhib- following HIV protease inhibitors, which are some of the itors. largest and most complex drugs in development or mar- [0020] In particular, the present invention is directed keted, all suffer from low oral bioavailability: Hoffmann 50 to a nanoparticulate composition comprising a crystal- La-Roche's saquinavir (INVIRASE®), Vertex's VX-478, line HIV protease inhibitor having a solubility in water of Merck's MK-639 (L-735,524) (also known as indinavir, less than 10 mg/ml, and having an average particle size and marketed under the trade name CRIXIVAN®), of less than 1000 nm and a cellulosic surface stabilizer Agouron Pharmaceutical's AG1343, Abbott Labs' ABT- which is adsorbed to the surface of the HIV protease 538, Upjohn Co.'s U-103,017, Dupont Merck's DMP- 55 inhibitor. 450, and National Cancer Institute's and Japan Energy's [0021] The invention also provides a stable dispersion KNI-272. Early HIV Protease Inhibitors Difficult to Pro- comprising a liquid dispersion medium with the above- duce and Supplies Restricted, Biotechnology Informa- described nanoparticulate composition. The terms "dis-

3 5 EP 1 002 065 B1 6 persion" or "suspension" are synonymous and used in- ity. terchangeably herein and refer to a formulation in which [0028] The nanoparticulate composition of the the ingredient nanoparticles float undissolved in a fluid present invention comprise a drug substance as a dis- such as water. crete, crystalline phase. The crystalline phase differs [0022] In a preferred embodiment, a pharmaceutical 5 from a non-crystalline or amorphous phase which re- composition is provided comprising the above-de- sults from precipitation techniques. scribed nanoparticulate composition, and a pharmaceu- [0029] Poorly water soluble HIV protease inhibitor tically acceptable carrier therefor. The pharmaceutical drug substances, many of which could not have been compositions of the invention, which exhibit unexpect- administered intravenously prior to the present inven- edly high bioavailability, may be formulated as a liquid, 10 tion, may be safely and effectively administered by in- gel, solid, or any other appropriate formulation. travenous methods in formulations according to the in- [0023] In another embodiment of the invention, there vention. Additionally, many HIV protease inhibitor drug is provided a method of preparing the above-described substances which prior to the present invention could nanoparticulate composition or dispersion. The method not have been administered orally due to poor bioavail- comprises dispersing an HIV protease inhibitor in a liq- 15 ability, may also be safely and effectively administered uid dispersion medium and applying mechanical means orally in formulations according to the present invention. in the presence of grinding media to reduce the average Pharmaceutical compositions according to the present particle size of the HIV protease inhibitor to an effective invention include the nanoparticulate composition or average particle size of less than about 1000 nm and dispersion described above and a pharmaceutically ac- isolating the resultant HIV protease inhibitor composi- 20 ceptable carrier therefor. Suitable pharmaceutically action or dispersion from the grinding media The drug par- ceptable carriers are well known to those skilled in the ticles can be reduced in size in the presence of a cellu- art and include, for example, non-toxic physiologically losic surface stabilizer, or the drug particles can be con- acceptable carriers, adjuvants, or vehicles for parenter- tacted with a cellulosic surface stabilizer after attrition. al injection, for oral administration in solid or liquid form, [0024] In yet another embodiment of the invention, a 25 for rectal administration, and the like. Pharmaceutical method for preparing a nanoparticulate pharmaceutical [0030] Compositions or dispersions according to the composition in a tablet form is provided. In such a meth- present invention may also comprise binding agents, fill- od, the nanoparticles are compressed into tablets. The ing agents, lubricating agents, disintegrating agents, tablets generally also comprise at least one pharmaceu- suspending agents, sweeteners, flavoring agents, pre- tically acceptable carrier. 30 servatives, buffers, wetting agents, and other excipi- [0025] The present invention also provides for a nan- ents. Examples of filling agents are lactose monohy- oparticulate composition or dispersion according to the drate, lactose hydrous, and various starches; examples invention for use in therapy. The nanoparticulate com- of binding agents are various celluloses, preferably low- position or dispersion of the invention may be used for substituted hydroxylpropyl cellulose, and cross-linked the preparation of a pharmaceutical composition for the 35 polyvinylpyrrolidone; an example of a disintegrating treatment of acquired immune deficiency syndrome agent is croscarmellose sodium; and examples of lubri- (AIDS) or AIDS related complex (ARC) in a mammal ei- cating agents are talc, magnesium stearate, stearic ac- ther alone or in combination with other treatments. id, and silica gel. Examples of suspending agents are [0026] It is to be understood that both the foregoing hydroxypropyl cellulose, methyl cellulose, hydroxyethyl general description and the following detailed descrip- 40 cellulose, carboxymethyl cellulose sodium, hydroxypro- tion are exemplary and explanatory and are intended to pyl methylcellulose, acacia, alginic acid, carrageenin, provide further explanation of the invention as claimed. and other hydrocolloides. Examples of sweeteners are Other objects, advantages, and novel features will be any natural or artificial sweetener, such as sucrose, xy- readily apparent to those skilled in the art from the fol- litol, sodium saccharin, cyclamate, aspartame, and ac- lowing detailed description of the invention. 45 sulfame. Examples of flavoring agents are Magnas- weet® (trademark of MAFCO), bubble gum flavor, and DESCRIPTION OF THE PREFERRED fruit flavors, such as orange, grape, cherry, berry, lem- EMBODIMENTS on-lime, and the like. Examples of preservatives, which control microbial contamination, are potassium sorbate, [0027] The invention is directed to the unexpected 50 methylparaben, propylparaben, benzoic acid and its discovery that nanoparticulate formulations of highly in- salts, other esters of parahydroxybenzoic acid such as soluble HIV protease inhibitors having an extremely butylparaben, alcohols such as ethyl or benzyl alcohol, small effective average particle size can be prepared us- phenolic compounds such as phenol, or quaternary ing a cellulosic surface stabilizer. The nanoparticles are compounds such as benzalkonium chloride. stable and do not appreciably flocculate or agglomerate 55 [0031] The pharmaceutical compositions according due to interparticle attractive forces. In addition, such to the present invention are particularly useful in oral and nanoparticles can be formulated into pharmaceutical parenteral, including intravenous, administration formu- compositions exhibiting unexpectedly high bioavailabil- lations. The pharmaceutical compositions may also be

4 7 EP 1 002 065 B1 8 administered enterally. Examples of parenteral routes commercially available and/or can be prepared by tech- of administration include, but are not limited to, subcu- niques known in the art. taneous, intramuscular, respiratory, and intravenous in- [0035] As described below in the Examples, it has jection, as well as nasopharyngeal, mucosal, and been discovered that stable and dispersible nanopartic- transdermal absorption. It is a particularly advanta- 5 ulate formulations of HIV protease inhibitors can only be geous feature that the pharmaceutical compositions of formed using cellulosic surface stabilizers, and that the the present invention exhibit unexpectedly high bioavail- use of other known surface stabilizers results in a com- ability, they provide more rapid onset of drug action, and position which is not stable and dispersible. This discov- they provide decreased gastrointestinal irritancy. ery is significant in that, as noted above, the use of HIV [0032] The selected dosage level of the drug sub- 10 protease inhibitors in the treatment of HIV infection has stance for therapy or for the preparation of a pharma- been limited prior to the present invention because of ceutical composition for the treatment of acquired im- the low solubility and corresponding low bioavailability mune deficiency syndrome (AIDS) or AIDS related com- of the drugs. Moreover, the formation of nanoparticulate plex (ARC) in a mammal is effective to obtain a desired HIV protease inhibitor compositions enables the prepa- therapeutic response for a particular composition and 15 ration of pharmaceutical compositions which are more method of administration. The selected dosage de- palatable to patients. This is because as noted above, pends, therefore, upon the particular drug substance, the use of a smaller volume, but the same dose, of drug the desired therapeutic effect, the route of administra- allows for greater camouflage of the unpleasant taste of tion, the desired duration of treatment, and other factors. the HIV protease inhibitor using larger doses of sweet- In addition, because the particle size of the drug has 20 eners. been dramatically reduced, the pharmaceutical compo- [0036] Pharmaceutically acceptable salts of the HIV sition of the present invention can comprise an identical protease inhibitor drug substance can also be used in dosage to a prior art large drug size dosage in a much the invention and include the conventional non-toxic smaller volume. Alternatively, the pharmaceutical com- salts or the quaternary ammonium salts which are position of the present invention can deliver a smaller 25 formed, for example, from inorganic or organic acids or dosage as a greater percentage of the drug will be ab- bases. Examples of such acid addition salts include, but sorbed into the bloodstream of the patient as compared are not limited to, acetate, adipate, alginate, aspartate, to prior art large drug size dosage. Both pharmaceutical benzoate, benzenesulfonate, bisulfate, butyrate, citrate, compositions allow for greater camouflage of the poor camphorate, camphorsulfonate, cyclopentanepropion- taste of HIV protease inhibitors. The pharmaceutical 30 ate, digluconate, dodecylsulfate, ethanesulfonate, fu- compositions comprising the nanoparticulate drug sub- marate, glucoheptanoate, glycerophosphate, hemisul- stance are also useful in treatment of mammals in com- fate, heptanoate, hexanoate, hydrochloride, hydrobro- bination with other antivirals, immunomodulators, anti- mide, hydroiodide, 2-hydroxy-ethanesulfonate, lactate, biotics, vaccines, or the like. maleate, methanesulfonate, 2-naphthalenesulfonate, 35 nicotinate, oxalate, palmoate, pectinate, persulfate, Description of Drug Substance 3-phenylpropionate, picrate, pivalate, propionate, suc- cinate, tartrate, thiocyanate, tosylate, and undecanoate. [0033] The HIV protease inhibitor drug substance is Examples of base salts include, but are not limited to, preferably present in an essentially pure form. In addi- ammonium salts, alkali metal salts such as sodium and tion, the HIV protease inhibitor drug substance is poorly 40 potassium salts, alkaline earth metal salts such as cal- soluble, although it is dispersible in at least one liquid cium and magnesium salts, salts with organic bases medium. By "poorly soluble" it is meant that the drug such as dicyclohexylamine salts, N-methyl-D-glu- substance has a solubility in the liquid dispersion medi- camine, and salts with amino acids such as arginine, um of less than about 10 mg/ml, and preferably less than lysine, and the like. In addition, the basic nitrogen-con- about 1 mg/ml. A preferred liquid dispersion medium is 45 taining groups may be quaternized with such agents as water. However, the invention can be practiced with oth- lower alkyl halides, such as methyl, ethyl, propyl, and er liquid media in which the HIV protease inhibitor drug butyl chloride, bromides, and iodides; dialkyl sulfates substance is poorly soluble and dispersible including, such as dimethyl, diethyl, dibutyl, and diamyl sulfates; for example, aqueous salt solutions, safflower oil, and long chain halides such as decyl, lauryl, myristyl, and solvents such as ethanol, t-butanol, hexane, and glycol. 50 stearyl chlorides, bromides, and iodides; aralkyl halides The pH of the aqueous dispersion media can be adjust- such as benzyl and phenethyl bromides and the like. ed by techniques known in the art. Other pharmaceutically acceptable salts include the sul- [0034] Suitable HIV protease inhibitor drug substanc- fate salt ethanolate and sulfate salts. es can be selected from a variety of known drugs including but not limited to for example, saquinavir, retinovir, 55 Surface Stabilizer VX-478, (amprenavir) MK-639 (i.e, indinavir), AG1343 (nelfinavir) ABT-538 (ritonavir) U-103,017, DMP-450 [0037] The surface stabilizer of the present invention (mozenavir), and KNI-272. The drug substances are is a cellulosic stabilizer. Cellulosic surface stabilizers

5 9 EP 1 002 065 B1 10 are non-ionic and water soluble. Examples of cellulosic stance with the cellulosic surface stabilizer adsorbed on stabilizers include, but are not limited to, hydroxypropyl the surface thereof. It has been discovered that the cel- cellulose (HPC), which is an ether of cellulose, HPC su- lulosic surface stabilizer physically adheres to the drug per low viscosity (HPC-SL), HPC-low viscosity (HPC-L), substance, but it does not chemically bond to or chem- and hydroxypropyl methyl cellulose (HPMC). 5 ically react with the drug. Such chemical bonding or in- [0038] The surface stabilizer is present at an amount teraction would be undesirable as it could result in alter- of about 1 to about 100 mg/ml, preferably, about 10 mg/ ing the function of the drug. The surface stabilizer is ad- ml to about 20 mg/ml, and most preferably at about 10 sorbed on the surface of the drug substance in an mg/ml. amount sufficient to maintain an effective average par- [0039] The cellulosic surface stabilizer can also be 10 ticle size of less than about 1000 nm, and more prefer- used in conjunction with one or more other surface sta- ably, less than about 400 nm. Furthermore, the individ- bilizers. Suitable additional surface stabilizers can pref- ually adsorbed molecules of the surface stabilizer are erably be selected from known organic and inorganic essentially free of intermolecular cross-linkages. pharmaceutical excipients. Such excipients include various polymers, low molecular weight oligomers, natural 15 Ouantities of Drug Substance and Surface Stabilizer products and surfactants. Preferred additional surface stabilizers include nonionic and anionic surfactants. [0042] The relative amount of the HIV protease inhib- Representative examples of excipients include gelatin, itor drug substance and cellulosic surface stabilizer can casein, lecithin (phosphatides), gum acacia, cholester- vary widely. The optimal amount of the cellulosic surface ol, tragacanth, stearic acid, benzalkonium chloride, cal- 20 stabilizer, and any other additional surface stabilizer, cium stearate, glyceryl monostearate, cetostearyl alco- can depend upon, for example, the particular HIV pro- hol, cetomacrogol emulsifying wax, sorbitan esters, tease inhibitor employed, the hydrophilic-lipophilic bal- polyoxyethylene alkyl ethers, e.g., macrogol ethers ance (HLB) of the stabilizer, the melting point and water such as cetomacrogol 1000, polyoxyethylene castor oil solubility of the stabilizer, the surface tension of water derivatives, polyoxyethylene sorbitan fatty acid esters, 25 solutions of the stabilizer, and the like. e.g., the commercially available Tweens, polyethylene [0043] The cellulosic surface stabilizer is preferably glycols, polyoxyethylene stearates, colloidol silicon di- present in an amount of about 0.1 to about 10 mg per oxide, phosphates, sodium dodecylsulfate, car- square meter of surface area of the drug substance, or boxymethylcellulose calcium, carboxymethylcellulose in an amount of about 0.1 to about 90%, and more pref- sodium, methylcellulose, hydroxyethylcellulose, hy- 30 erably about 20 to about 60% by weight based upon the droxypropyl methycellulose phthalate, noncrystalline total weight of the dry drug particle. Additional stabilizer, cellulose, magnesium aluminum silicate, trieth- such as Aerosol OT®, in an amount of 0.01 % to 1% by anolamine, polyvinyl alcohol, and polyvinylpyrrolidone weight based upon the total weight of the dry particle (PVP). Most of these excipients are described in detail are also preferable. in the Handbook of Pharmaceutical Excipients, pub- 35 lished jointly by the American Pharmaceutical Associa- Reducing the Particle Size of the HIV Protease tion and The Pharmaceutical Society of Great Britain Inhibitor Drug Substance to Nanoparticles (The Pharmaceutical Press, 1986). The surface stabilizers are commercially available and/or can be pre- [0044] The nanoparticulate composition of the pared by techniques known in the art. 40 present invention can be prepared by first dispersing an [0040] Particularly preferred surface stabilizers which HIV protease inhibitor in a liquid dispersion medium fol- can be used in conjunction with the cellulosic surface lowed by applying mechanical means in the presence stabilizer, include polyvinyl pyrrolidone, Pluronic F68® of grinding media to reduce the particle size of the drug and F108®, which are block copolymers of ethylene ox- substance to an effective average particle size of less ide and propylene oxide, Tetronic 908®, which is a 45 than about 1000 nm, and more preferably, less than tetrafunctional block copolymer derived from sequential about 400 nm. The HIV protease inhibitor particles can addition of ethylene oxide and propylene oxide to ethyl- be reduced in size in the presence of the cellulosic sur- enediamine, dextran, lecithin, Aerosol OT®, which is a face stabilizer or the drug particles can be contacted dioctyl ester of sodium sulfosuccinic acid, available from with the cellulosic surface stabilizer following attrition. American Cyanamid, Duponol P®, which is a sodium 50 [0045] A general procedure for preparing the HIV pro- lauryl sulfate, available from DuPont, Triton X-200®, tease inhibitor nanoparticulate composition of the inven- which is an alkyl aryl polyether sulfonate, available from tion is set forth below. The HIV protease inhibitor is ei- Rohm and Haas, Tween 80®, which is a polyoxyethyl- ther obtained commercially or prepared by techniques ene sorbitan fatty acid ester, available from ICI Specialty known in the art in a conventional coarse form. It is pre- Chemicals, and Carbowax 3350® and 934®, which are 55 ferred, but not essential, that the particle size of the se- polyethylene glycols available from Union Carbide. lected drug be less than about 100 µm as determined [0041] The nanoparticles of the invention contain a by sieve analysis. If the coarse particle size of the drug discrete phase of an HIV protease inhibitor drug sub- is greater than about 100 µm, then it is preferred that

6 11 EP 1 002 065 B1 12 the drug particles be reduced in size to less than about cooled with conventional cooling equipment. Generally, 100 µm using a conventional milling method, such as the methods of the invention can be conveniently carried airjet or fragmentation milling, prior to reducing the par- out under conditions of ambient temperature and at ticulate drug to submicron particle size. processing pressures which are safe and effective for [0046] The coarse HIV protease inhibitor particles can 5 the milling process. For example, ambient processing then be added to a liquid medium in which the drug is pressures are typical of ball mills, attritor mills, and vi- essentially insoluble to form a premix. The concentra- bratory mills. Control of the temperature, for example, tion of the drug in the liquid medium can vary from about by jacketing or immersion of the milling chamber in ice 0.1 to about 60%, but is preferably from about 5 to about water, is encompassed by the invention. 30% (w/w). It is preferred, but not essential, that the cel- 10 [0051] Processing pressures from about 1 psi (0.07 lulosic surface stabilizer is present in the premix. The kg/cm2) up to about 50 psi (3.5 kg/cm2) are encom- concentration of the cellulosic surface stabilizer can passed by the invention. Processing pressures typically vary from about 0.1 to about 90%, but it is preferably range from about 10 psi to about 20 psi. from about 1 to about 75%, and more preferably from [0052] The surface stabilizer, if not present in the about 20 to about 60%, by weight based upon the total 15 premix, must be added to the dispersion after attrition combined weight of the HIV protease inhibitor and sur- in an amount as described for the premix above. There- face stabilizer. The apparent viscosity of the premix sus- after, the dispersion can be mixed by, for example, shak- pension is preferably less than about 1000 centipoise. ing vigorously. Optionally, the dispersion can be subject- [0047] The premix can be used directly by subjecting ed to a sonication step using, for example, an ultrasonic it to mechanical means to reduce the average particle 20 power supply. In such a method, the ultrasonic power size in the dispersion to less than about 1000 nm, and supply can, for example, release ultrasonic energy hav- more preferably, less than about 400 nm. It is preferred ing a frequency of about 20 to about 80 kHz for a time that the premix be used directly when a ball mill is used of about 1 to about 120 seconds. for attrition. Alternatively, the HIV protease inhibitor, and [0053] After attrition is completed, the grinding media optionally the cellulosic surface stabilizer, can be dis- 25 is separated from the milled particulate product using persed in the liquid medium using suitable agitation, conventional separation techniques, such as by filtra- such as a roller mill or a Cowles-type mixer, until a ho- tion, sieving through a mesh screen, and the like. The mogeneous dispersion is observed. In a homogeneous surface stabilizer may be added to the milled particulate dispersion, no large agglomerates are visible to the na- product either before or after the milled product is sep- ked eye. It is preferred that the premix be subjected to 30 arated from the grinding media. such a premilling dispersion step when a recirculating [0054] In a preferred grinding process, the particles media mill is used for attrition. are made continuously. In such a continuous method, [0048] The mechanical means applied to reduce the the slurry of HIV protease inhibitor/cellulosic surface particle size of the HIV protease inhibitor can be a dis- stabilizer and optionally an additional surface stabilizer persion mill. Suitable dispersion mills include, but are 35 is continuously introduced into a milling chamber, the not limited to, a ball mill, an attritor mill, a vibratory mill, HIV protease inhibitor is continuously contacted with and media mills such as a sand mill or a bead mill. A grinding media while in the chamber to reduce the par- media mill is preferred due to the relatively shorter mill- ticle size of the HIV protease inhibitor, and the HIV pro- ing time required to provide the desired reduction in par- tease inhibitor is continuously removed from the milling ticle size. For media milling, the apparent viscosity of 40 chamber. The cellulosic surface stabilizer, either alone the premix is preferably from about 100 to about 1000 or in conjunction with one or more additional surface sta- centipoise. For ball milling, the apparent viscosity of the bilizers, can also be continuously added to the media premix is preferably from about 1 to about 100 chamber along with the HIV protease inhibitor, or it can centipoise. Such ranges tend to afford an optimal bal- be added to the HIV protease inhibitor which is removed ance between efficient particle fragmentation and media 45 from the chamber following grinding. erosion. [0055] The resulting dispersion of the present inven- [0049] The attrition time can vary widely and depends tion is stable and comprises the liquid dispersion medi- primarily upon the particular mechanical means and um described above. The dispersion of surface stabiliz- processing conditions selected. For ball mills, process- er and nanoparticulate HIV protease inhibitor can be ing times of up to five days or longer may be required. 50 spray dried, spray coated onto sugar spheres or onto a Using a high shear media mill, processing times of less pharmaceutical excipient using techniques well known than 1 day (residence times of from one minute up to in the art. several hours) have provided the desired results. [0050] The HIV protease inhibitor particles must be Grinding Media reduced in size at a temperature which does not signif- 55 icantly degrade the drug substance. Processing temper- [0056] The grinding media for the particle size reduc- atures of less than about 30-40°C are ordinarily pre- tion step can be selected from rigid media which is pref- ferred. If desired, the processing equipment can be erably spherical or particulate in form and which has an

7 13 EP 1 002 065 B1 14 average size of less than about 3 mm and, more pref- [0060] The grinding media is separated from the erably, less than about 1 mm. Such media can provide milled particulate HIV protease inhibitor using conven- the desired drug particles of the invention with shorter tional separation techniques in a secondary process, processing times and impart less wear to the milling such as by filtration, sieving through a mesh filter or equipment. The selection of material for the grinding 5 screen, and the like. Other separation techniques such media is not believed to be critical. Zirconium oxide, as centrifugation may also be employed. such as 95% ZrO stabilized with yttrium and 95% ZrO stabilized with magnesia, zirconium silicate, and glass Particle Size grinding media have been found to provide particles having acceptable minimal levels of contamination for 10 [0061] As used herein, particle size is determined on the preparation of pharmaceutical compositions. Other the basis of the average particle size as measured by media, such as stainless steel, titania, and alumina can conventional techniques well known to those skilled in also be used. Preferred grinding media have a density the art, such as sedimentation field flow fractionation, greater than about 3 g/cm3. photon correlation spectroscopy, or disk centrifugation. 15 When photon correlation spectroscopy (PCS) is used Polymeric Grinding Media as the method of particle sizing, the average particle diameter is the Z-average particle diameter known to [0057] The grinding media can comprise particles, those skilled in the art. By "an average effective particle preferably spherical in shape, such as beads, consisting size of less than about 1000 nm," it is meant that at least of essentially polymeric resin. Alternatively, the grinding 20 90% of the particles, by weight, have a particle size of media can comprise particles having a core with a coat- less than about 1000 nm when measured by the above- ing of the polymeric resin adhered thereto. The media noted techniques. In a preferred embodiment of the in- can range in size from about 0.1 to about 3 mm. For fine vention, the effective average particle size is less than grinding, the particles preferably are from about 0.2 to about 400 nm, more preferred is less than about 250 about 2 mm, and more preferably, from about 0.25 to 25 nm, and in an even more preferred embodiment, the ef- about 1 mm in size. fective average particle size is less than about 100 nm. [0058] The polymeric resin can have a density from It is preferred that at least 95% and, more preferably, at about 0.8 to about 3.0 g/cm3. Higher density resins are least 99% of the particles have a particle size less than preferred as such resins can provide more efficient par- the effective average, e.g., 1000 nm. In a particularly ticle size reduction. 30 preferred embodiment, essentially all of the particles [0059] In general, polymeric resins suitable for use in have a size of less than about 400 nm, in a more pre- the present invention are chemically and physically in- ferred embodiment, essentially all of the particles have ert, substantially free of metals, solvent, and monomers, a size of less than about 250 nm, and in a most preferred and of sufficient hardness and friability to enable them embodiment, essentially all of the particles have a size to avoid being chipped or crushed during grinding. Suit- 35 of less than about 100 nm. able polymeric resins include, but are not limited to, cross-linked polystyrenes, such as polystyrene cross- Preparing Tablet Formulations linked with divinylbenzene, styrene copolymers, polycarbonates, polyacetals such as Delrin®, vinyl chloride [0062] An exemplary process for preparing nanopar- polymers and copolymers, polyurethanes, polyamides, 40 ticulate compositions of the invention in a tablet formu- poly(tetrafluoroethylenes), such as Teflon® and other lation comprises: (1) using the method described below fluoropolymers, high density polyethylenes, polypropyl- to obtain spray-dried nanoparticles of the drug sub- enes, cellulose ethers and esters such as cellulose ac- stance; (2) sieve-screening the spray-dried nanoparti- etate, polyhydroxymethacrylate, polyhydroxyethyl acr- cles to obtain uniform particles of less than about 20 ylate, silicone containing polymers such as polysi- 45 mesh; (3) blending the nanoparticulate drug substance loxanes, and the like. The polymer can also be biode- with tableting excipients; (4) compressing the uniform gradable. Exemplary biodegradabe polymers include, particles into tablets using a tableting apparatus; and (5) but are not limited to, poly(lactides), poly(glycolides), film coating the tablets. copolymers of lactides and glycolides, polyanhydrides, [0063] The spray drying process is used to obtain an poly(hydroxyethyl methacylate), poly(imino carbon- 50 "intermediate" nanoparticulate powder subsequent to ates), poly(N-acylhydroxyproline)esters, poly(N-palmi- the milling process used to transform the HIV protease toyl hydroxyproline) esters, ethylene-vinyl acetate co- inhibitor into nanoparticles. In an exemplary spray dry- polymers, poly(orthoesters), poly(caprolactones), and ing process, the high-solids drug substance nanosus- poly(phosphazenes). For biodegradable polymers, con- pension and the cellulosic surface stabilizer are fed to tamination of the resultant composition from the media 55 an atomizer using a peristatic pump and atomized into itself can advantageously metabolize in vivo into biolog- a fine spray of droplets. The spray is contacted with hot ically acceptable products that can be eliminated from air in the drying chamber resulting in the evaporation of the body. moisture from the droplets. The resulting spray is

8 15 EP 1 002 065 B1 16 passed into a cyclone where the powder is separated and collected. [0064] At the completion of the spray drying process, the collected spray-dried intermediate comprises the nanoparticles of the HIV protease inhibitor suspended 5 in a solid polymer matrix of the cellulosic surface stabilizer. The moisture content of the intermediate is controlled by the operating conditions of the spray drying process. The characteristics of the nanoparticulate powder are critical to the development of a free flowing pow- 10 der that can be blended with other excipients suitable for a directly compressible tablet formulation. [0065] Tablets can be made using a direct compression tablet process or using a roller compaction process. In an exemplary direct compression process, the spray- 15 dried intermediate and stabilizer are sieved through a screen and the screened material is blended. The desired excipients are sieved and added to the blender. At the completion of blending, the contents of the blender can be discharged into a tared collection container and 20 compression of tablet cores can be completed on a tablet press. The blended material can be fed into a feed hopper and force-fed into the die cavities using an au- tomatic feeder. The tablet press operating conditions can be set to meet thickness, hardness, and weight 25 specifications. Upon completion of the compression operation, a film-coating can be applied to the tablet cores using, for example, a Vector-Freund Hi-Coater machine. [0066] In an exemplary roller compaction process following the media milling process, the nanoparticulate 30 HIV protease inhibitor suspension can be spray dried to form an intermediate. The spray dryer can be assem- bled in a co-current configuration using a rotary atomi- zation nozzle and the nanosuspension can be fed to the rotary atomizer using a peristaltic pump. Acceptable 35 spray-dried product has a moisture content which does not exceed 1.0% (w/w). [0067] A dry granulation operation can be used to manufacture tablets comprising the HIV protease inhibitor drug substance. Required amounts of the HIV pro- 40 tease inhibitor drug substance/cellulosic surface stabilizer spray-dried intermediate and appropriate excipients can be screened and blended. The blended material is then compacted using, for example, a roller com- pactor. The compacted material can then be granulated. 45 Following granulation, additional excipients can be screened and blended with the granulation. The blended materials can then be compressed into tablets using a tablet press followed by coating. [0068] Process Flow Diagram A show a process of 50 manufacturing tablets by direct compression, and Proc- ess Flow Diagram B shows a process of manufacturing tablets by roller compaction.

9 17 EP 1 002 065 B1 18

in the art, and it is commercially available from Merck & Co., Inc., Rahway, NJ. [0072] A surface stabilizer solution comprising 90 ml 2.5% (w/w) of HPC-SL in purified water was added into 5 a 500 ml Pyrex® roller mill bottle. 10% (w/w, 10.0 g) of the indinavir was dispensed incrementally into the HPC-SL stabilizer solution until the entire amount was added. 250 ml of 0.5 mm YTZP media (Yttria doped Zir- conia beads commercially available from Toray, Japan) 10 was charged into the 500 ml roller mill bottle. The bottle was placed on a roller mill (U.S. Stoneware, Akron, OH) with the roller speed set at 160 rpm and milled for 4 days. [0073] A sample of the indinavir/HPC-SL slurry was removed and analyzed for particle size. The particle size 15 of the sample was measured with a Microtrack UPA siz- er according to the following procedures. First, the chamber of the instrument was rinsed three times with DI water. Next, the chamber was filled with 50 ppm DOSS (dioctyl sodium sulfosuccinate) and one drop of 20 204 nm standard was placed into the chamber. Using a disposable transfer pipette, the contents of the chamber were gently agitated for ten seconds. The signal level was checked to be within 0.2-0.3 (unitless). If the level was higher, the sample was diluted by removing some 25 of the contents in the chamber and adding DOSS solution. Once the sample was in the signal level range, it was run by selecting "run w/o save" for 180 seconds. The chamber of the instrument was then rinsed three times and filled with 50 ppm DOSS solution. A sample 30 of the nanoformulation to be tested was added carefully into the DOSS solution in the chamber to gradually increase the signal level to within 0.2-0.3 (unitless). Once the sample was in the signal level range, it was run for 180 seconds to determine the size of the particle. The 35 chamber of the instrument was then rinsed three times and filled with 50 ppm DOSS solution in preparation for the next test. [0074] It was determined that the mean size of the indinavir/HPC-SL nanoparticles was 186 nm. [0069] The following examples are given to illustrate 40 the present invention. It should be understood, however, Example 2 that the invention is not to be limited to the specific conditions or details described in these examples. [0075] The purpose of this example was to demon- [0070] Examples 1-7 describe the formation of nano- strate the formation of a nanoparticulate composition of particulate compositions of indinavir. 45 indinavir. [0076] A surface stabilizer solution comprising 90 ml Example 1 3.30% (w/w) of HPC-SL in purified water was added into each of four 500 ml Pyrex® roller mill bottles. 10% (w/ [0071] The purpose of this example was to demon- w, 10.0 g) of the indinavir was dispensed incrementally strate the formation of a nanoparticulate composition of 50 into the HPC-SL stabilizer solution until the entire indinavir, the sulphate salt of which is marketed as amount was added and the solution appeared mixed. CRIXIVAN® (trademark of Merck & Co., Inc.), i.e., indi- 250 ml of 0.5 mm YTZP media (Yttria doped Zirconia navir sulfate. Indinavir, or N-(2-(R)-hydroxy-1(S)-inda- beads commercially available from Toray, Japan) was nyl)-2(R)-phenylmethyl-4-(S)-hydroxy-5-(1-(4-(3-pyridyl- charged into each of the four 500 ml Pyrex roller mill methyl)-2(S)-N'-(t-butylcarboxamido)-piperazinyl))- 55 bottles. The bottles were placed on a roller mill (U.S. pentaneamide, is a HIV protease inhibitor useful in treat- Stoneware, Akron, OH) with the roller speed set at 160 ing AIDS and ARC (AIDS Related Complex). Indinavir rpm and milled for 5 days. can be chemically synthesized using techniques known [0077] Following particle size analysis, it was deter-

10 19 EP 1 002 065 B1 20 mined that the mean size of the indinavir/HPC-SL nan- [0086] Following particle size analysis, it was deter- oparticles was 142 nm - 152 nm. mined that the mean size of the indinavir/HPC-SL nanoparticles was 172 nm. Example 3 5 Example 6 [0078] The purpose of this example was to demon- strate the formation of a nanoparticulate composition of [0087] The purpose of this example was to demon- indinavir. strate the formation of a nanoparticulate composition of [0079] A surface stabilizer solution comprising 13.5 g indinavir. 3.73% (w/w) of HPC-SL in purified water was added into 10 [0088] A surface stabilizer solution comprising 3.75 a 60 ml roller mill bottle. 10% (w/w, 1.5 g) of the indinavir ml 1.0% (w/w) of HPMC in purified water was added into was dispensed incrementally into the HPC-SL surface a 15 ml roller mill bottle. 2% (w/w) of the indinavir was stabilizer solution until the entire amount was added. 30 dispensed incrementally into the HPMC surface stabi- ml of 0.5 mm YTZP media (Yttria doped Zirconia beads lizer solution until the entire amount was added. 7.5 ml commercially available from Toray, Japan) was charged 15 of 0.5 mm YTZP media (Yttria doped Zirconia beads into the 60 ml roller mill bottle. The bottle was placed on commercially available from Toray, Japan) was charged a roller mill (U.S. Stoneware, Akron, OH) with the roller into the 15 ml roller mill bottle. The bottle was placed on speed set at 160 rpm and milled for 9 days. a roller mill (U.S. Stoneware, Akron, OH) with the roller [0080] Following particle size analysis, it was deter- speed set at 160 rpm and milled for 12 days. mined that the mean size of the indinavir/HPC-SL nan- 20 [0089] Following particle size analysis, it was deter- oparticles was 267 nm. mined that the mean size of the indinavir/HPMC nanoparticles was 240 nm. Example 4 Example 7 [0081] The purpose of this example was to demon- 25 strate the formation of a nanoparticulate composition of [0090] The purpose of this example was to demon- indinavir. strate the use of high energy milling to provide a nano- [0082] A surface stabilizer solution comprising 2.75 particulate composition of indinavir at higher than 30% ml 1.0% (w/w) of HPC-SL in purified water was added w/w. into a 15 ml roller mill bottle. 2% (w/w, 75 mg) of the 30 [0091] A Dyno Mill (Type KDL, cat. # 6094-AEA) was indinavir was dispensed incrementally into the HPC-SL used in the size reduction process. 510 ml of 500 µm surface stabilizer solution until the entire amount was SDy20 polymeric media were added into a 600 ml mill- added. 7.5 ml of 0.5 mm YTZP media (Yttria doped Zir- ing chamber. An agitator composed of four 64 mm di- conia beads commercially available from Toray, Japan) ameter polyblades and a shaft rotation of 2500 rpm was was charged into the 15 ml roller mill bottle. The bottle 35 used in the milling process. DI water was used to cool was placed on a roller mill (U.S. Stoneware, Akron, OH) the seal and the milling chamber. DI water at 4°C was with the roller speed set at 160 rpm and milled for 12 circulated through the jacketed milling chamber and the days. seals of the mill during the milling operation. [0083] Following particle size analysis, it was deter- [0092] The size reduction process was performed in mined that the mean size of the indinavir/HPC-SL nan- 40 a recirculation mode. Suspension or slurry was pumped oparticles was 127 nm. at a flow rate of 150 ml/min from a 1.5 L jacketed holding tank into the milling chamber and back out into the hold- Example 5 ing tank. [0093] Added to the holding tank were 323.19 gm of [0084] The purpose of this example was to demon- 45 indinavir, 31.42 gm of HPC-SL, and 1277 gm of diluent, strate the formation of a nanoparticulate composition of which comprised of xylitol at 151 mg/ml, Magnasweet® indinavir. at 3.05 mg/ml, Tween 80® at 0.57 mg/ml, sodium citrate [0085] A surface stabilizer solution comprising 118.25 at 1.67 mg/ml, orange flavor at 10.1 mg/ml, potassium g 1.07% (w/w) of HPC-SL in purified water was added sorbate at 3.26 mg/ml, methylparaben at 0.76 mg/ml, into a 500 ml Pyrex® roller mill bottle. 5% (w/w, 6.25 g) 50 propylparaben at 0.076 mg/ml, and water at 880 mg/ml, of the indinavir was dispensed incrementally into the with the pH adjusted to 7.0 using 0.1 N NaOH. The in- HPC-SL surface stabilizer solution until the entire gredients inside the holding tank were cooled by circu- amount was added. 250 ml of 0.5 mm YTZP media (Yt- lating DI water at 4°C through the jacket. A Servo Dyne tria doped Zirconia beads commercially available from mixer was inserted into the holding tank for 1 hour to Toray, Japan) was charged into the 500 ml roller mill bot- 55 dissolve the HPC-SL and to wet the drug powder. The tle. The bottle was placed on a roller mill (U.S. Stone- slurry was then pumped into the Dyno mill chamber with ware, Akron, OH) with the roller speed set at 160 rpm a peristaltic pump at a flow rate of 150 ml/min. After the and milled for 5 days. milling chamber was filled, the Dyno mill was turned on

11 21 EP 1 002 065 B1 22 and the suspension was milled in the recirculation mode between 5:3 and 14:10. for 2 υ hours to provide an indinavir nanocrystalle sus- [0101] The thoroughly mixed suspension was distrib- pension at a pH of 7.68. The particle size of the harvest- uted into Teflon® containers covering 1/8 of an inch in ed indinavir formulation was 50% at 100 nm and 90% height. The suspension was then allowed to dry under at 147 nm when measured with a Horiba LA910 instru- 5 clean air to afford the dried film. The dried film was re- ment, and it was 50% at 157 nm and 90% at 231 nm suspended in WFI or SGF to provide a nanocrystalle when measured with a Microtrack UPA instrument. suspension with a 50% particle size between 180 nm to [0094] Examples 8-12 are directed to the formation of 200 nm and a 90% size between 250 nm to 300 nm nanoparticulate compositions of VX478, a HIV protease when measured with a Horiba LA910 instrument. inhibitor. 10 Example 10 Example 8 [0102] The purpose of this example was to demon- [0095] The purpose of this example was to demon- strate the formation of a nanoparticulate composition of strate the formation of a nanoparticulate composition of 15 VX478. VX-478, which is a HIV protease inhibitor, using Dyno [0103] A stock surface stabilizer solution consisting of milling. VX-478 is commercially available from Vertex 2.5% (w/v) HPC-L (Nisso DI-0031) was prepared in dis- Pharmaceuticals, Inc., Cambridge, MA, and methods of tilled water. A 15 mL polycarbonate mini-DC milling tube making VX-478 are described in U.S. Patent No. was charged with 2.5 mL (1.625 grams) of 0.5 mm Sdy- 5,585,397. 20 20 polymeric milling media. Approximately 2.5 mL of a [0096] A Dyno Mill (Type KDL cat. # 6094-AEA) was 1.0% (w/v) VX478/1.0% (w/v) HPC-L/water slurry was used in the process. A 300 ml milling chamber contain- prepared directly in the DC milling tube by combining ing 240 ml of 500 µm SDy20 polymeric media and an the following ingredients: agitator composed of two 64 mm diameter polyblades having a shaft rotation of 2500 rpm were used in the 25 0.025 grams VX478 milling process. DI water at 4°C was circulated through 1.0 mL stock HPC-L stabilizer solution (@ 2.5% w/ the jacketed milling chamber and the seals of the mill v) during the size reduction process. 1.5 mL water [0097] The size reduction process was done in a recirculation mode. Suspension or slurry was pumped at 30 The mini-DC milling tube was water jacketed and cooled a flow rate of 150 ml/min from a jacketed holding tank with tap water. The slurry and media were milled for 5 (1.5 liter) into the milling chamber and back out into the hours using a mini-DC mill stainless steel shaft at a mill- holding tank. ing speed of 2000 rpm. [0098] 120 gm of VX478, 18 g of HPC-L, 60 mg of [0104] The dispersion settled upon standing over- DOSS, and 500 ml of WFI were added to the holding 35 night but was easily resuspended. The particle size was tank. The ingredients inside the holding tank were measured using a Horiba LA 900 Particle Size Analyzer, cooled by circulating DI water at 4°C through the jacket. with a limited volume fraction cell, 0.2 µm filtered 0.01% A Servo Dyne mixer was inserted into the holding tank DOSS as the sizing diluent, and a sonication time of 30 for 1 hour to dissolve the HPC-L and DOSS and to wet seconds. The mean particle size was 582 nm with 90% the drug powder. The slurry was then pumped into the 40 of the particles having a particle size of less than 880 Dyno mill chamber with a peristaltic pump at a flow rate nm. The nanosuspension was stable in SGF and SIF. of 157 ml/min. After the milling chamber was filled, the After 13 days storage at ambient room temperature the Dyno mill was turned on and the suspension was milled nanosuspension had a mean particle size of 571 nm in the recirculation mode for 4 hours to provide a VX478 with 90% of the particles having a particle size of less nanocrystalline suspension at a pH of 6. The particle 45 than 830 nm. size of the harvested VX478 formulation was 50% at 169 nm and 90% at 221 nm when measured with a Horiba Example 11 LA910 instrument. [0105] The purpose of this example was to demon- Example 9 50 strate the formation of a nanoparticulate composition of VX478. [0099] The purpose of this example was to demon- [0106] A stock surface stabilizer solution consisting of strate the formation of a dry film nanoparticulate com- 2.5% (w/w) HPC-L (Nisso DI-0031) was prepared in dis- position of VX-478. tilled water. A stock stabilizer solution of 1.0% (w/w) [0100] A VX-478 nanoparticulate formulation as pre- 55 SDS was prepared in distilled water. A 100 mL polycar- pared in Example 84 was mixed with sugar (preferably bonate DC milling tube was charged with 10.0 mL (6.5 sucrose or mannose), HPC-L, DOSS, and WFI to pro- grams) of 0.5 mm Sdy20 polymeric milling media. Ap- vide a suspension with the drug:sugar ratio preferably proximately 10 grams of a 2.0% (w/w) VX478/1.0% (w/

12 23 EP 1 002 065 B1 24 w) HPC-L/water slurry was prepared directly in the DC 0.5 mm Sdy20 polymeric milling media. Approximately milling tube by combining the following ingredients: 2.5 mL of a 1.0% (w/v) VX478/1.0% (w/v) stabilizer/water slurry was prepared directly in each of the DC milling 0.20 grams VX478 tubes by combining the following ingredients: 4.0 grams stock HPC-L stabilizer solution (@ 2.5% 5 w/w) 0.025 grams VX478 5.8 grams water 1.0 mL stock stabilizer solution (@ 2.5% w/v) 1.5 mL water The DC milling tube was water jacketed and cooled with tap water. The slurry and media were milled for 4 hours 10 The mini-DC milling tubes were water jacketed and using a standard size, tefzel-coated DC milling shaft at cooled with tap water. The slurry and media were milled a milling speed of 2300 rpm. for various time intervals using mini-DC mill stainless [0107] Following the completion of the milling proc- steel shafts at a milling speeds of 2000 rpm. ess, the dispersion was separated from the media by filtering through a stainless steel mesh syringe filter. The 15 a) 1.0% VX478 in 1.0% PVP K-29/32®: Following mass of the collected dispersion was calculated and an 8 hours of DC milling, the resultant mixture con- appropriate amount of 1.0% (w/w) SDS was added such tained unmilled drug and agglomerates. No size re- that the concentration of SLS in the final dispersion was duction was observed. 3.75 mL of an identical slurry 0.01% (w/w). (The SDS was added because it was preparation was also roller milled in a 15 mL bottle found that upon scaling up to a 2:1 concentration in the 20 with 7.5 mL of 0.5 mm YTZ media. After 3 days, standard DC mill, the dispersion formed loose agglom- unmilled drug with no size reduction was observed. erates that were visualized microscopically, but which The particle size was measured using a Horiba LA were not represented on the particle size distributions 900 Particle Size Analyzer, with a limited volume due to the sonication step during the size measurement fraction cell, 0.2 µm filtered 0.01% DOSS as the siz- process. Addition of SDS to a final concentration of 0.0 25 ing diluent, and a sonication time of 30 seconds. 1 % prevented the formation of the loose agglomerates.) The mean particle size was measured to be greater The VX478/HPC-L/SDS composition was mixed well than 7 µm. and the particle size, fluid stability, and shelf stability were determined. b) 1.0% VX478 in 1.0% Pluronic F68®: Following 4 [0108] The particle size was measured using a Horiba 30 hours of DC milling, the resultant mixture contained LA 900 Particle Size Analyzer, with a limited volume unmilled drug and no size reduction was observed. fraction cell, 0.2 µm filtered 0.01% DOSS as the sizing The particle size was measured as described in diluent, and a sonication time of 30 seconds. The mean subsection (1) above. The mean particle size was particle size was 230 nm with 90% of the particles hav- measured to be greater than 20 µm. ing a particle size of less than 315 nm. The nanosus- 35 pension was stable when mixed 1:1 with SGF and SIF. c) 1.0% VX478 in 1.0% Pharmacoat 603®: Follow- After 14 days storage at ambient room temperature the ing 4 hours of DC milling, the resultant mixture con- nanosuspension had a mean particle size of 268 nm tained large, tight agglomerates. The particle size with 90% of the particles having a particle size of less was measured as described in subsection (1) than 384 nm. 40 above. The mean particle size was measured to be [0109] In a repetition of this experiment, the nanosus- approximately 400 nm (agglomerates broken dur- pension had a mean particle size of 157 nm with 90% ing sonication). The size distribution had a long tail of the particles having a particle size of less than 300 extending beyond 10 µm. nm. 45 d) 1.0% VX478 in 1.0% Methocel K100LV®: Follow- Example 12 ing milling for 11 hours, the resultant mixture had a mean particle size of 302 nm with 90% of the parti- [0110] The purpose of this example is to summarize cles having a particle size of less than 408 nm. The surface stabilizers and size reduction techniques which particle size was measured as described in subsec- are inadequate or insufficient to produce a nanopartic- 50 tion (1) above. When this dispersion was scaled up ulate composition of VX478. to a 2:1 concentration ratio in a standard size DC [0111] Procedures: Stock surface stabilizer solutions mill, excessive foaming was encountered, although consisting of 2.5% (w/v) PVP K-29/32, 2.5% (w/v) an acceptable particle size was obtained (Mean Pluronic F68®, 2.5% (w/v) Pharmacoat 603®, 2.5% (w/ size of 297 nm, 90 % less than 400 nm). After 3 days v) Methocel K100LV®, and 2.5% (w/v) Pluronic F108® 55 on the shelf, a peak possibly representing large ag- were prepared in distilled water. glomerates appeared during the size measurement [0112] Five 15 mL polycarbonate mini-DC milling indicating potential shelf instability of the nanosus- tubes were charged with 2.5 mL (1.625 grams) each of pension.

13 25 EP 1 002 065 B1 26

e) 1.0% VX478 in 1.0% Pluronic F108®: Following of claims 2 to 6, wherein the surface stabilizer is se- milling for 8 hours, the resultant suspension had a lected from the group consisting of hydroxypropyl mean particle size of 400 nm, with 90% of the par- cellulose, hydroxypropyl methyl cellulose, hydroxy- ticles having a size of less than 530 nm. After 3 propyl cellulose super low viscosity, and hydroxy- days, the nanosuspension had a mean particle size 5 propyl cellulose low viscosity. of 375 nm, with 90% of the particles having a size of less than 490 nm. The particle sizes were meas- 8. The composition according to any one of claims 1 ured as described in subsection (1) above. The na- and 4 to 7, or the dispersion according to any one nosuspension was stable in the presence of SGF of claims 2 to 7, wherein the composition or disper- and SIF. When this dispersion was scaled to a 2:1 10 sion comprises at least one additional surface sta- concentration in a standard DC mill, excessive bilizer for the HIV protease inhibitor. foaming was encountered, the drug substance was difficult to wet into the stabilizer phase, and the dis- 9. The composition or the dispersion of claim 8, persion quickly settled. wherein the additional surface stabilizer is selected 15 from the group consisting of gelatin, casein, lecithin, gum acacia, cholesterol, tragacanth, stearic acid, Claims benzalkonium chloride, calcium stearate, glyceryl monostearate, cetostearyl alcohol, cetomacrogol 1. A nanoparticulate composition comprising: emulsifying wax, sorbitan esters, polyoxyethylene 20 alkyl ethers, polyoxyethylene castor oil derivatives, (a) a crystalline HIV protease inhibitor having a polyoxyethylene sorbitan fatty acid esters, polyeth- solubility in water of less than 10 mg/ml, and ylene glycols, polyoxyethylene stearates, colloidol having an effective average particle size of less silicon dioxide, phosphates, sodium dodecylsulfate, than 1000 nm; and carboxymethylcellulose calcium, carboxymethyl- (b) a cellulosic surface stabilizer which is ad- 25 cellulose sodium, methylcellulose, hydroxyethylcel- sorbed to the surface of the HIV protease inhib- lulose, hydroxypropyl methycellulose phthalate, itor. noncrystalline cellulose, magnesium aluminum silicate, triethanolamine, polyvinyl alcohol, and poly- 2. A stable dispersion comprising: vinylpyrrolidone. 30 (a) a liquid dispersion medium; and 10. The composition or the dispersion of claim 8, (b) a nanoparticulate composition of claim 1. wherein the additional surface stabilizer is selected from the group consisting of polyvinyl pyrrolidone, 3. The dispersion of claim 2, wherein the dispersion Pluronic F68®, Pluronic F108®, Tetronic 908®, medium is selected from the group consisting of wa- 35 dextran, lecithin, Aerosol® OT, Duponol® P, Triton ter, safflower oil, ethanol, t-butanol, hexane, and X-200®, Tween 80®, and Carbowax 3350® and glycol. Carbowax 934®.

4. The composition of claim 1, or the dispersion of 11. The composition or the dispersion according to any claims 2 or 3, wherein the HIV protease inhibitor is 40 one of claims 8 to 10, wherein the additional stabi- VX478 (amprenavir). lizer is present in an amount of 0.01% to 1%, by weight, based upon the total weight of the dry par- 5. The composition of claim 1, or the dispersion of ticle. claims 2 or 3, wherein the HIV protease inhibitor is selected from the group consisting of saquinavir, 45 12. The composition or the dispersion according to any retinovir, AG1343 (nelfinavir), ABT-538 (ritonavir), one of claims 1 and 4 to 11, or the dispersion ac- U-103,017, DMP-450 (mozenavir), MK-639 (indina- cording to any one of claims 2 to 11, wherein the vir), and KNI-272. HIV protease inhibitor has an effective average particle size of less than 400 nm, less than 250 nm or 6. The composition according to any one of claims 1, 50 less than 100 nm. 4, or 5, or the dispersion according to any one of claims 2 to 5, wherein the cellulosic surface stabi- 13. The composition or the dispersion according to any lizer is present in an amount of 0.1 to 90% or in an one of claims 1 and 4 to 12, or the dispersion ac- amount of 20 to 60%, by weight, based upon the cording to any one of claims 2 to 12, wherein at least total weight of the dry drug particle. 55 90%, at least 95%, or at least 99% of the HIV protease inhibitor particles, by weight, have a particle 7. The composition according to any one of claims 1 size less than the effective average particle size. and 4 to 6, or the dispersion according to any one

14 27 EP 1 002 065 B1 28

14. The composition according to any one of claims 1 20. The method of claim 19, wherein the cellulosic sur- and 4 to 13, or the dispersion according to any one face stabilizer is not present in step (a), but rather of claims 2 to 13, further comprising a pharmaceu- is added to the HIV protease inhibitor following par- tically acceptable carrier. ticle size reduction. 5 15. The composition according to any one of claims 1 21. The method according to any one of claims 17 to and 4 to 14, or the dispersion according to any one 20, wherein the mechanical means comprises wet of claims 2 to 14 for use in therapy. grinding.

16. Use of the composition according to any one of 10 22. The method according to any one of claims 17 to claims 1 and 4 to 14, or the dispersion according to 21, wherein the mechanical means is a dispersion any one of claims 2 to 14, for the preparation of a mill. pharmaceutical composition for the treatment of acquired immune deficiency syndrome (AIDS) or 23. The method of claim 22, wherein the dispersion mill AIDS related complex (ARC) in a mammal. 15 is selected from the group consisting of a ball mill, an attritor mill, a vibratory mill, and media mills. 17. A method of preparing the nanoparticulate composition according to any one of claims 1 and 4 to 14, 24. The method according to any one of claims 17 to or the dispersion according to any one of claims 2 23, wherein the grinding media has an average par- to 14 comprising: 20 ticle size of 3 mm or less or of 1 mm or less.

(a) dispersing a HIV protease inhibitor in a liq- 25. The method according to any one of claims 17 to uid dispersion medium; 24, wherein the grinding media is selected from the group consisting of zirconium oxide, 95% ZrO sta- (b) applying mechanical means in the presence 25 bilized with yttrium, 95% ZrO stabilized with mag- of rigid grinding media and a cellulosic surface nesia, zirconium silicate, glass, stainless steel, tita- stabilizer to reduce the particle size of the HIV nia, and alumina. protease inhibitor to an effective average particle size of less than 1000 nm; and 26. The method according to any one of claims 17 to 30 24, wherein the grinding media comprises a poly- (c) isolating the resultant nanoparticulate HIV meric resin. protease inhibitor composition or dispersion from the grinding media. 27. The method of claim 26, wherein the polymeric resin is selected from the group consisting of cross- 18. The method of claim 17, wherein the cellulosic sur- 35 linked polystyrenes, styrene copolymers, polycar- face stabilizer is not present in step (b), but rather bonates, polyacetals, vinyl chloride polymers, vinyl is added to the HIV protease inhibitor composition chloride copolymers, polyurethanes, polyamides, or dispersion either before or after the composition poly(tetrafluoroethylenes), fluoropolymers, high or dispersion is separated from the grinding media. density polyethylenes, polypropylenes, cellulose 40 ethers, cellulose esters, polyhydroxymethacrylate, 19. A method of preparing the nanoparticles according polyhydroxyethyl acrylate, and silicone containing to any one of claims 1 and 4 to 14, or the dispersion polymers. according to any one of claims 2 to 14 comprising: 28. The method of claim 27, wherein the polymeric res- (a) continuously introducing into a milling 45 in is selected from the group consisting of polysty- chamber a slurry of an HIV protease inhibitor/ rene cross-linked with divinylbenzene, cellulose ac- a cellulosic surface stabilizer; etate, and polysiloxanes.

(b) continuously contacting the slurry with 29. The method of claims 26 or 27, wherein the poly- grinding media while in the chamber to reduce 50 meric resin is biodegradable. the particle size of the HIV protease inhibitor; and 30. The method of claim 29, wherein the polymeric resin is selected from the group consisting of poly(lac- (c) continuously removing the HIV protease in- tides), poly(glycolides), copolymers of lactides and hibitor from the milling chamber, which has 55 glycolides, polyanhydrides, poly(hydroxyethyl been reduced to an effective average particle methacylate), poly(imino carbonates), poly(N-acyl- size of less than 1000 nm. hydroxyproline)esters, poly(N-palmitoyl hydroxyproline) esters, ethylene-vinyl acetate copolymers,

15 29 EP 1 002 065 B1 30

poly(orthoesters), poly(caprolactones), and poly lulose mit geringer Viskosität. (phosphazenes). 8. Die Zusammensetzung gemäß einem der Ansprü- 31. The method according to any one of claims 17 to che 1 und 4 bis 7, oder die Dispersion gemäß einem 30, further comprising forming the composition or 5 der Ansprüche 2 bis 7, wobei die Zusammenset- dispersion into a tablet. zung oder Dispersion wenigstens ein zusätzliches Oberflächenstabilisierungsmittel für den HIV-Pro- teaseinhibitor umfasst. Patentansprüche 10 9. Die Zusammensetzung oder die Dispersion gemäß 1. Eine nanopartikuläre Zusammensetzung, umfas- Anspruch 8, wobei das zusätzliche Oberflächensta- send: bilisierungsmittel ausgewählt ist aus der Gruppe bestehend aus Gelatine, Casein, Lecithin, Gummi- a) einen kristallinen HN-Proteaseinhibitor, der arabicum, Cholesterin, Tragant, Stearinsäure, Ben- eine Löslichkeit in Wasser von weniger als 10 15 zalkoniumchlorid, Calciumstearat, Glycerylmono- mg/ml und eine effektive durchschnittliche Par- stearat, Cetostearylalkohol, Cetomakrogol-Emul- tikelgröße von weniger als 1000 nm besitzt; und gierwachs, Sorbitanester, Polyoxyethylen-Alkyle- b) ein Zellulose-Oberflächenstabilisierungsmit- ther, Polyoxyethylen-Rizinusölderivate, Polyoxye- tel, das an die Oberfläche des HIV-Proteasein- thylensorbitan-Fettsäureester, Polyethylenglykole, hibitors adsorbiert ist. 20 Polyoxyethylenstearate, kolloidales Siliziumdioxid, Phosphate, Natriumdodecylsulfat, Carboxymethyl- 2. Eine stabile Dispersion, umfassend: zellulose-Calcium, Carboxymethylzellulose-Natri- um, Methylzellulose, Hydroxyethylzellulose, Hydro- a) ein flüssiges Dispersionsmedium; und xypropylmethylzellulose-Phthalat, nichtkristalline b) eine nanopartikuläre Zusammensetzung ge- 25 Zellulose, Magnesiumaluminiumsilikat, Triethano- mäß Anspruch 1. lamin, Polyvinylalkohol und Polyvinylpyrrolidon.

3. Die Dispersion gemäß Anspruch 2, wobei das Di- 10. Die Zusammensetzung oder die Dispersion gemäß spersionsmedium ausgewählt ist aus der Gruppe Anspruch 8, wobei das zusätzliche Oberflächensta- bestehend aus Wasser, Distelöl, Ethanol, t-Butanol, 30 bilisierungsmittel ausgewählt ist aus der Gruppe Hexan und Glykol. bestehend aus Polyvinylpyrrolidon, Pluronic F68®, Tetronic 908®, Dextran, Lecithin, Aerosol® OT, Du- 4. Die Zusammensetzung gemäß Anspruch 1 oder die ponol® P, Triton X-200®, Tween 80®, und Carbo- Dispersion gemäß Ansprüchen 2 oder 3, wobei der wax 3350® und Carbowax 934®. HIV-Proteaseinhibitor VX478 (Amprenavir) ist. 35 11. Die Zusammensetzung oder die Dispersion gemäß 5. Die Zusammensetzung gemäß Anspruch 1 oder die einem der Ansprüche 8 bis 10, wobei das zusätzli- Dispersion gemäß Ansprüchen 2 oder 3, wobei der che Oberflächenstabilisierungsmittel in einem Mas- HIV-Proteaseinhibitor ausgewählt ist aus der Grup- senanteil von 0,01% bis 1%, basierend auf dem Ge- pe bestehend aus Saquinavir, Retinovir, AG1343 40 samtgewicht des trockenen Partikels, vorhanden (Nelfinavir), ABT-538 (Ritonavir), U-103.017, DMP- ist. 450 (Mozenavir), MK-639 (Indinavir) und KNI-272. 12. Die Zusammensetzung oder die Dispersion gemäß 6. Die Zusammensetzung gemäß einem der Ansprü- einem der Ansprüche 1 und 4 bis 11, oder die Di- che 1, 4 oder 5, oder die Dispersion gemäß einem 45 spersion gemäß einem der Ansprüche 2 bis 11, wo- der Ansprüche 2 bis 5, wobei das Zellulose-Ober- bei der HIV-Proteaseinhibitor eine effektive durch- flächenstabilisierungsmittel in einem Massenanteil schnittliche Partikelgröße von weniger als 400 nm, von 0,1 bis 90% oder in einem Massenanteil von 20 weniger als 250 nm oder weniger als 100 nm hat. bis 60%, basierend auf dem Gesamtgewicht des trockenen Wirkstoffpartikels, vorhanden ist. 50 13. Die Zusammensetzung oder die Dispersion gemäß einem der Ansprüche 1 und 4 bis 12, oder die Di- 7. Die Zusammensetzung gemäß einem der Ansprü- spersion gemäß einem der Ansprüche 2 bis 12, wo- che 1 und 4 bis 6, oder die Dispersion gemäß einem bei ein Massenanteil von wenigstens 90%, wenig- der Ansprüche 2 bis 6, wobei das Oberflächensta- stens 95%, oder wenigstens 99% der HIV-Proteas- bilisierungsmittel ausgewählt ist aus der Gruppe 55 einhibitor-Partikel, eine Partikelgröße, die kleiner ist bestehend aus Hydroxypropylzellulose, Hydroxy- als die effektive durchschnittliche Partikelgröße, propylmethylzellulose, Hydroxypropylzellulose mit besitzt. äußerst geringer Viskosität und Hydroxypropylzel-

16 31 EP 1 002 065 B1 32

14. Die Zusammensetzung gemäß einem der Ansprü- (c)fortlaufendesEntfernendesHIV-Proteasein- che 1 und 4 bis 13, oder die Dispersion gemäß ei- hibitors, der auf einen effektive durchschnittli- nem der Ansprüche 2 bis 13, ferner umfassend ei- che Partikelgröße von weniger als 1000 nm re- nen pharmazeutisch akzeptablen Träger. duziert worden ist, aus der Mahlkammer. 5 15. Die Zusammensetzung gemäß einem der Ansprü- 20. Das Verfahren gemäß Anspruch 19, wobei das Zel- che 1 und 4 bis 14, oder die Dispersion gemäß ei- lulose-Oberflächenstabilisierungsmittel in Schritt nem der Ansprüche 2 bis 14 zur Verwendung im (a) nicht vorhanden ist, sondern vielmehr zum Heilverfahren. HIV-Proteaseinhibitor nach der Reduktion der Par- 10 tikelgröße zugegeben wird. 16. Die Verwendung der Zusammensetzung gemäß einem der Ansprüche 1 und 4 bis 14, oder der Disper- 21. Das Verfahren gemäß einem der Ansprüche 17 bis sion gemäß einem der Ansprüche 2 bis 14 zur Her- 20, wobei die mechanischen Mittel Naßvermahlung stellung einer pharmazeutischen Zusammenset- umfassen. zung zur Behandlung von Erworbene-Immun- 15 schwäche-Syndrom (AIDS) oder von AIDS-related- 22. Das Verfahren gemäß einem der Ansprüche 17 bis Komplex (ARC) in einem Säugetier. 21, wobei das mechanische Mittel eine Dispersi- onsmühle ist. 17. Ein Verfahren zur Herstellung der nanopartikulären Zusammensetzung gemäß einem der Ansprüche 1 20 23. Das Verfahren gemäß Anspruch 22, wobei die Di- und 4 bis 14, oder die Dispersion gemäß einem der spersionsmühle ausgewählt ist aus der Gruppe be- Ansprüche 2 bis 14, umfassend: stehend aus einer Kugelmühle, einem Attritor, einer Vibrationsmühle und Medienmühlen. (a) Dispergieren eines HIV-Proteaseinhibitors in einem flüssigen Dispersionsmedium; 25 24. Das Verfahren gemäß einem der Ansprüche 17 bis (b) Anwendung mechanischer Mittel in Gegen- 23, wobei das Mahlmedium eine durchschnittliche wart von starren Mahlmedien und eines Zellu- Partikelgröße von 3 mm oder weniger oder von 1 lose-Oberflächenstabilisierungsmittels zur Re- mm oder weniger besitzt. duktion der Partikelgröße des HIV-Proteasein- hibitors auf eine effektive durchschnittliche Par- 30 25. Das Verfahren gemäß einem der Ansprüche 17 bis tikelgröße von weniger als 1000 nm; und 24, wobei das Mahlmedium ausgewählt ist aus der (c) Isolierung der sich daraus ergebenden na- Gruppe bestehend aus Zirkoniumoxid, 95% ZrO nopartikulären HIV-Proteaseinhibitor-Zusam- stabilisiert mit Yttrium, 95% ZrO stabilisiert mit Ma- mensetzung oder -Dispersion aus dem Mahl- gnesium, Zirkonium-Silikat, Glas, rostfreiem Stahl, medium. 35 Titandioxid und Aluminiumoxid.

18. Das Verfahren gemäß Anspruch 17, wobei das Zel- 26. Das Verfahren gemäß einem der Ansprüche 17 bis lulose-Oberflächenstabilisierungsmittel in Schritt 24, wobei das Mahlmedium ein polymeres Harz um- (b) nicht vorhanden ist, sondern vielmehr zu der fasst. HIV-Proteaseinhibitor-Zusammensetzung oder -Di- 40 spersion, entweder bevor oder nachdem die Zu- 27. Das Verfahren gemäß Anspruch 26, wobei das po- sammensetzung oder Dispersion vom Mahlmedi- lymere Harz ausgewählt ist aus der Gruppe beste- um getrennt wird, zugegeben wird. hend aus quervernetzten Polystyrolen, Styrol-Kop- olymeren, Polycarbonaten, Polyacetalen, Vinyl- 19. Ein Verfahren zur Herstellung der Nanopartikel ge- 45 chlorid-Polymeren, Vinylchlorid-Kopolymeren, Po- mäß einem der Ansprüche 1 und 4 bis 14, oder die lyurethanen, Polyamiden, Poly(tetrafluorethyle- Dispersion gemäß einem der Ansprüche 2 bis 14, nen), Fluorpolymeren, Polyethylenen mit hoher umfassend: Dichte, Polypropylenen, Zelluloseethern, Zellulo- seestem, Polyhydroxymethacrylat, Polyhydroxye- (a) fortlaufendes Einleiten einer Aufschläm- 50 thylacrylat und Silikon, das Polymere enthält. mung eines HIV-Proteaseinhibitors / eines Zel- lulose-Oberflächenstabilisierungsmittels in ei- 28. Das Verfahren gemäß Anspruch 27, wobei das po- ne Mahlkammer; lymere Harz ausgewählt ist aus der Gruppe beste- (b) fortlaufendes Kontaktieren der Aufschläm- hend aus Polystyrol quervernetzt mit Divinylbenzol, mung mit einem Mahlmedium während des 55 Zelluloseacetat und Polysiloxanen. Aufenthalts in der Kammer, um die Partikelgrö- ße des HIV-Proteaseinhibitors zu reduzieren; 29. Das Verfahren gemäß Ansprüchen 26 oder 27, wo- und bei das polymere Harz bioabbaubar ist.

17 33 EP 1 002 065 B1 34

30. Das Verfahren gemäß Anspruch 29, wobei das po- 7. Composition suivant l'une quelconque des revendi- lymere Harz ausgewählt ist aus der Gruppe beste- cations 1 et 4 à 6, ou dispersion suivant l'une quel- hend aus Poly(lactiden), Poly(glykoliden), Kopoly- conque des revendications 2 à 6, dans laquelle le meren von Lactiden und Glykoliden, Polyanhydri- stabilisant de surface est choisi dans le groupe con- den, Poly(hydroxyethylmethacrylat), Poly(imino- 5 sistant en hydroxypropylcellulose, hydroxypropyl- carbonaten), Poly(N-acylhydroxyprolin)estern, Po- méthylcellulose, hydroxypropylcellulose d'ultra- ly(N-palmitoylhydroxyprolin)estern, Ethylen- basse viscosité et hydroxypropylcellulose de faible Vinylacetat-Kopolymeren, Poly(orthoestern), Poly viscosité. (caprolactonen) und Poly(phosphazenen). 10 8. Composition suivant l'une quelconque des revendi- 31. Das Verfahren gemäß einem der Ansprüche 17 bis cations 1 et 4 à 7, ou dispersion suivant l'une quel- 30, ferner umfassend das Umformen der Zusam- conque des revendications 2 à 7, la composition ou mensetzung oder Dispersion in eine Tablette. dispersion comprenant au moins un stabilisant de surface supplémentaire pour l'inhibiteur de protéa- 15 se du VIH. Revendications 9. Composition ou dispersion suivant la revendication 1. Composition en nanoparticules, comprenant: 8, dans laquelle le stabilisant de surface supplé- mentaire est choisi dans le groupe consistant en gé- (a) un inhibiteur cristallin de protéase du VIH 20 latine, caséine, lécithine, gomme arabique, choles- ayant une solubilité dans l'eau inférieure à 10 térol, gomme adragante, acide stéarique, chlorure mg/ml et ayant un diamètre moyen efficace de de benzalkonium, stéarate de calcium, monostéa- particules inférieur à 1000 nm; et rate de glycéryle, alcool cétostéarylique, cire émul- (b) un stabilisant cellulosique de surface qui est sionnable cétomacrogol, esters de sorbitanne, adsorbé à la surface de l'inhibiteur de protéase 25 éthers alkyliques de polyoxyéthylène, dérivés de du VIH. polyoxyéthylène d'huile de ricin, esters d'acides gras de sorbitanne-polyoxyéthylène, polyéthylène- 2. Dispersion stable comprenant: glycols, stéarates de polyoxyéthylène, dioxyde de silicium colloïdal, phosphates, dodécylsulfate de (a) un milieu liquide de dispersion; et 30 sodium, carboxyméthylcellulose calcique, carboxy- (b) une composition en nanoparticules de la re- méthylcellulose sodique, méthylcellulose, hydroxy- vendication 1. éthylcellulose, phtalate d'hydroxypropylméthylcel- lulose, cellulose non cristalline, silicate d'aluminium 3. Dispersion suivant la revendication 2, dans laquelle et de magnésium, triéthanolamine, alcool polyviny- le milieu de dispersion est choisi dans le groupe 35 lique et polyvinylpyrrolidone. consistant en l'eau, l'huile de carthame, l'éthanol, le tertiobutanol, l'hexane et glycol. 10. Composition ou dispersion suivant la revendication 8, dans laquelle le stabilisant de surface supplé- 4. Composition suivant la revendication 1, ou disper- mentaire est choisi dans le groupe consistant en po- sion suivant la revendication 2 ou 3, dans laquelle 40 lyvinylpyrrolidone, Pluronic F68®, Pluronic F108®, l'inhibiteur de protéase du VIH est le VX478 (am- Tetronic 908®, dextrane, lécithine, Aerosol® OT, prénavir). Duponol®, Triton X-200®, Tween 80® et Carbowax 3350® et Carbowax 934®. 5. Composition suivant la revendication 1, ou dispersion suivant la revendication 2 ou 3, dans laquelle 45 11. Composition ou dispersion suivant l'une quelcon- l'inhibiteur de protéase du VIH est choisi dans le que des revendications 8 à 10, dans laquelle le sta- groupe consistant en saquinavir, rétinovir, AG1343 bilisant supplémentaire est présent en une quantité (nelfinavir), ABT-538 (ritonavir), U-103 017, DMP- de 0,01 % à 1 %, en poids, sur la base du poids total 450 (mozénavir), MK-639 (indinavir) et KNI-272. des particules sèches. 50 6. Composition suivant l'une quelconque des revendi- 12. Composition ou dispersion suivant l'une quelcon- cations 1, 4 et 5, ou dispersion suivant l'une quel- que des revendications 1 et 4 à 11, ou dispersion conque des revendications 2 à 5, dans laquelle le suivant l'une quelconque des revendications 2 à 11, stabilisant cellulosique de surface est présent en dans laquelle l'inhibiteur de protéase du VIH a un une quantité de 0,1 à 90 % ou en une quantité de 55 diamètre moyen efficace de particules inférieur à 20 à 60 %, en poids, sur la base du poids total de 400 nm, inférieur à 250 nm ou inférieur à 100 nm. la particule sèche de médicament. 13. Composition ou dispersion suivant l'une quelcon-

18 35 EP 1 002 065 B1 36

que des revendications 1 et 4 à 12, ou dispersion chambre de broyage une suspension d'un inhi- suivant l'une quelconque des revendications 2 à 12, biteur de protéase du VIH/stabilisant cellulosi- dans laquelle une proportion d'au moins 90 %, d'au que de surface; moins 95 % ou d'au moins 99 % des particules d'in- (b) mettre en contact de manière continue la hibiteur de protéase du VIH, en poids, a un diamètre 5 suspension avec un milieu de broyage tandis de particules inférieur au diamètre moyen efficace qu'elle est présente dans la chambre pour ré- de particules. duire le diamètre de particules de l'inhibiteur de protéase du VIH; et 14. Composition suivant l'une quelconque des revendi- (c) évacuer de manière continue de la chambre cations 1 et 4 à 13, ou dispersion suivant l'une quel- 10 de broyage d'inhibiteur de protéase du VIH qui conque des revendications 2 à 13, comprenant en a été réduit à un diamètre moyen efficace de outre un support pharmaceutiquement acceptable. particules inférieur à 1000 nm.

15. Composition suivant l'une quelconque des revendi- 20. Procédé suivant la revendication 19, dans lequel le cations 1 et 4 à 14, ou dispersion suivant l'une quel- 15 stabilisant cellulosique de surface n'est pas présent conque des revendications 2 à 14, destinée à être dans l'étape (a) mais, en variante, est ajouté à l'in- utilisée en thérapie. hibiteur de protéase du VIH après réduction des diamètres de particules. 16. Utilisation de la composition suivant l'une quelconque des revendications 1 et 4 à 14, ou de la disper- 20 21. Procédé suivant l'une quelconque des revendica- sion suivant l'une quelconque des revendications 2 tions 17 à 20, dans lequel le moyen mécanique à 14, pour la préparation d'une composition phar- comprend un broyage à l'état humide. maceutique destinée au traitement du syndrome d'immunodéficience acquise (SIDA) ou du comple- 22. Procédé suivant l'une quelconque des revendica- xe associé au SIDA (ARC) chez un mammifère. 25 tions 17 à 21, dans lequel le moyen mécanique est un broyeur de dispersion. 17. Procédé pour la préparation de la composition en nanoparticules suivant l'une quelconque des reven- 23. Procédé suivant la revendication 22, dans lequel le dications 1 et 4 à 14, ou de la dispersion suivant broyeur de dispersion est choisi dans le groupe l'une quelconque des revendications 2 à 14, com- 30 consistant en un broyeur à bille, un broyeur à usure prenant les étapes consistant à: par frottement, un broyeur vibrant et des broyeurs renfermant un milieu de broyage. (a) disperser un inhibiteur de protéase du VIH dans un milieu liquide de dispersion; 24. Procédé suivant l'une quelconque des revendica- (b) appliquer un moyen mécanique en présen- 35 tions 17 à 23, dans lequel le milieu de broyage a un ce d'un milieu rigide de broyage et d'un stabili- diamètre moyen de particules égal ou inférieur à 3 sant cellulosique de surface pour réduire le dia- mm ou égal ou inférieur à 1 mm. mètre de particules de l'inhibiteur de protéase du VIH à un diamètre moyen efficace de parti- 25. Procédé suivant l'une quelconque des revendica- cules inférieur à 1000 nm; et 40 tions 17 à 24, dans lequel le milieu de broyage est (c) isoler du milieu de broyage la composition choisi dans le groupe consistant en oxyde de zirco- en nanoparticules ou dispersion résultante d'in- nium, ZrO à 95 % stabilisé avec de l'yttrium, ZrO à hibiteur de protéase du VIH. 95 % stabilisé avec de la magnésie, silicate de zirconium, verre, acier inoxydable, oxyde de titane et 18. Procédé suivant la revendication 17, dans lequel le 45 alumine. stabilisant cellulosique de surface n'est pas présent dans l'étape (b) mais, en variante, est ajouté à la 26. Procédé suivant l'une quelconque des revendica- composition ou dispersion d'inhibiteur de protéase tions 17 à 24, dans lequel le milieu de broyage com- du VIH avant ou après la séparation de la compo- prend une résine polymère. sition ou dispersion du milieu de broyage. 50 27. Procédé suivant la revendication 26, dans lequel la 19. Procédé pour la préparation de nanoparticules sui- résine polymère est choisie dans le groupe consis- vant l'une quelconque des revendications 1 et 4 à tant en des polystyrènes réticulés, des copolymè- 14, ou de la dispersion suivant l'une quelconque res de styrène, des polycarbonates, des polyacé- des revendications 2 à 14, comprenant les étapes 55 tals, des polymères de chlorure de vinyle, des co- consistant à: polymères de chlorure de vinyle, des polyuréthan- nes, des polyamides, des poly-(tétrafluoréthylè- (a) introduire de manière continue dans une nes), des fluoropolymères, des polyéthylènes hau-

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te densité, des polypropylènes, des éthers de cellulose, des esters de cellulose, un polyhydroxymé- thacrylate, un poly(acrylate d'hydroxyéthyle) et des polymères contenant une silicone. 5 28. Procédé suivant la revendication 27, dans lequel la résine polymère est choisie dans le groupe consistant en un polystyrène réticulé avec du divinylben- zène, l'acétate de cellulose et des polysiloxanes. 10 29. Procédé suivant la revendication 26 ou 27, dans lequel la résine polymère est biodégradable.

30. Procédé suivant la revendication 29, dans lequel la résine polymère est choisie dans le groupe consis- 15 tant en des poly(lactides), de poly(glycolides), des copolymères de lactides et de glycolides, des polyanhydrides, un poly(méthacrylate d'hydroxyéthyle), des poly(iminocarbonates), des polyesters de N- acylhydroxyproline, des polyesters de N-palmitoyl- 20 hydroxyproline, des copolymères éthylène-acétate de vinyle, des poly(orthoesters), des poly(caprolactones) et des poly(phosphazènes).

31. Procédé suivant l'une quelconque des revendica- 25 tions 17 à 30, comprenant en outre la transforma- tion de la composition ou dispersion en un compri- mé.