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 Printed by Jouve, 75001 PARIS (FR) 1 EP 1 002 065 B1 2 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 provid- ed 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 dilut- ed. 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.