USOO6331289B1 (12) United States Patent (10) Patent No.: US 6,331,289 B1 Klaveness et al. (45) Date of Patent: Dec. 18, 2001 (54) TARGETED DIAGNOSTIC/THERAPEUTIC 5,716,594 2/1998 Elmaleh et al. .................... 424/1.41 AGENTS HAVING MORE THAN ONE 5,733,572 3/1998 Unger et al. ......................... 424/450 DIFFERENT VECTORS 5,780,010 7/1998 Lanza et al. ... 424/9.32 5,846,517 12/1998 Unger ........... ... 424/9.52 (75) Inventors: Jo Klaveness; Pál Rongved; Anders 5,849,727 12/1998 Porter et al. ......................... 514/156 Høgset; Helge Tolleshaug, Alan 5,910,300 6/1999 Tournier et al. .................... 424/9.34 Cuthbertson; Lars Hoff, Klaus Bryn; 6.245,318 6/2001 Klibanov et al. ................... 424/9.52 Halldis Hellebust; Magne Solbakken, FOREIGN PATENT DOCUMENTS all of Olso (NO) 2 145 SOS 4/1994 (CA). 19 626 530 7/1996 (DE). (73) Assignee: Nycomed Imaging AS, Oslo (NO) O 727 225 8/1996 (EP). WO 93/20802 10/1993 (WO). * Y NotOtice: Subjubject to anyy disclaimer,disclai theh term off thisthi WO 94/07539 4/1994 (WO). patent is extended or adjusted under 35 WO 94/28873 12/1994 (WO). U.S.C. 154(b) by 0 days. WO 94/28874 12/1994 (WO). WO95/03356 2/1995 (WO). WO95/03357 2/1995 (WO). (21) Appl. No.: 08/959,206 WO95/07072 3/1995 (WO). (22) Filed: Oct. 28, 1997 WO95/15118 6/1995 (WO). WO 96/39149 12/1996 (WO). Related U.S. Application Data WO 96/40277 12/1996 (WO). (60) Provisional application No. 60/049.263, filed on Jun. 7, WO 96/40285 12/1996 (WO). 1997, provisional application No. 60/049.264, filed on Jun. WO 96/41647 12/1996 (WO). 6, 1997, and provisional application No. 60/049.266, filed WO 97/23855 7/1997 (WO). on Jun. 7, 1997. WO 97/33474 9/1997 (WO). WO 97/41898 11/1997 (WO). (30) Foreign Application Priority Data WO 98/OO172 1/1998 (WO). Oct. 28, 1996 (GB) .................................................. 9622366 WO 98/04293 2/1998 (WO). Oct. 28, 1996 (GB) ... ... 9622369 WO 98/19705 5/1998 (WO). Feb. 4, 1997 (GB) ... ... 97.02195 WO 98/20856 5/1998 (WO). Apr. 24, 1997 (GB) ... ... 9708265 WO 98/42384 10/1998 (WO). Jun. 6, 1997 (GB) ... ... 9711837 OTHER PUBLICATIONS Jun. 6, 1997 (GB) .................................................. 9711839 Worthington Enzyme Manual, Worthington Biochemical (51) Int. Cl. ............................. A61B 8/00; A61K 51/00; Corporation, 1972. A61K 9/127 U.S. application No. 08/640,464, Unger, filed May 1, 1996. (52) U.S. Cl. ......................... 424/9.52; 424/1.21; 424/9.4; U.S. application No. 08/660,032, Unger, filed Jun. 6, 1996. 424/9.6; 424/450 U.S. application No. 08/712,173, Unger, filed Sep. 11, 1996. (58) Field of Search .................................. 424/9.52, 9.51, Medline database extract No. XP-002063692, 1997. 424/9.5, 450, 489, 498, 499, 502, 1.21, Biosis database extract No. XP-002063693, Nov. 1997. 9.4, 9.6, 9.1; 530/300, 324, 326, 327: 514/14 Medline database extract No. KP-002064110, 1997. Muzykantov et al., J. Nuclear Medicine, 35(8): 1358–1365 (56) References Cited (1994). U.S. PATENT DOCUMENTS Klibanov et al., Acta Radiologica, 38(Supp. 412): 113-120 (1997). 4,927,916 5/1990 Matsueda ............................. 530/387 5,013,556 5/1991 Woodle et al. ...................... 424/450 * cited by examiner 5,154,924 10/1992 Friden ........ 424/85.91 5,198.424 3/1993 McEver .................................. 514/13 Primary Examiner Michael G. Hartley 5,356,633 10/1994 Woodle et al. ... 424/450 (74) Attorney, Agent, or Firm-Bacon & Thomas 5,362.478 11/1994 Desai et al. .............................. 424/9 (57) ABSTRACT 5,498,421 3/1996 Grinstaff et al. ..................... 424/450 5,505,932 4/1996 Grinstaff et al. ... ... 424/9.3 Targetable diagnostic and/or therapeutically active agents, 5,534.241 7/1996 Torchillin et al. ... 424/9.321 e.g. ultrasound contrast agents, comprising a Suspension in 5,612,057 3/1997 Lanza et al. ......................... 424/450 an aqueous carrier liquid of a reporter comprising gas 5,632,986 5/1997 Tait et al. ....... ... 424/94.64 5,643,553 7/1997 Schneider et al. ... 424/9.52 containing or gas-generating material, Said agent being 5,650,156 7/1997 Grinstaff et al. ... ... 424/400 capable of forming at least two types of binding pairs with 5,656.211 8/1997 Unger et al. ....... ... 264/4.1 a target. 5,665,383 9/1997 Grinstaff et al. ... 424/450 5,690,907 11/1997 Lanza et al. .......................... 424/9.5 22 Claims, 1 Drawing Sheet U.S. Patent Dec. 18, 2001 US 6,331,289 B1 256 Gm: 15.62 CW. 34.39 144-1023) 18430 (92.2/) FITClog F. G. 1. US 6,331,289 B1 1 2 TARGETED DIAGNOSTIC/THERAPEUTIC Surfactant material but rather that this is incorporated in AGENTS HAVING MORE THAN ONE liquid-filled liposomes which stabilise the microbubbles. It DIFFERENT VECTORS will be appreciated that lamellar or laminar Surfactant mate rial Such as phospholipids present in Such liposomes will This application claims benefit of the filing dates under 5 inevitably be present in the form of one or more lipid 35 USC 119(e) of U.S. provisional applications 60/049.263 bilayers with the lipophilic tails “back-to-back” and the and 049,264 filed on Jun. 6, 1997 and 60/049,266 filed on hydrophilic heads both inside and outside (see e.g. Jun. 7, 1997. Schneider, M. on "Liposomes as drug carriers: 10 years of It is well known that ultraSonic imaging comprises a research” in Drug targeting, Nyon, Switzerland, 3-5 Octo potentially valuable diagnostic tool, for example in Studies ber 1984, Buri, P. and Gumma, A. (Ed), Elsevier, Amsterdam of the vascular System, particularly in cardiography, and of 1984). tissue microvasculature. A variety of contrast agents has EP-A-0727225 describes targeted ultrasound contrast been proposed to enhance the acoustic images So obtained, agents in which the reporter comprises a chemical having a including Suspensions of Solid particles, emulsified liquid Sufficient vapour pressure Such that a proportion of it is a gas droplets, gas bubbles and encapsulated gases or liquids. It is 15 at the body temperature of the subject. This chemical is generally accepted that low density contrast agents which asSociated with a Surfactant or albumin carrier which are easily compressible are particularly efficient in terms of includes a protein-, peptide- or carbohydrate-based cell the acoustic backScatter they generate, and considerable adhesion molecule ligand as vector. The receptor moieties in interest has therefore been shown in the preparation of Such contrast agents correspond to the phase shift colloid gas-containing and gas-generating Systems. systems described in WO-A-9416739; it is now recognised Gas-containing contrast media are also known to be that administration of Such phase shift colloids may lead to effective in magnetic resonance (MR) imaging, e.g. as generation of microbubbles which grow uncontrollably, susceptibility contrast agents which will act to reduce MR possibly to the extent where they cause potentially danger Signal intensity. Oxygen-containing contrast media also rep ouS embolisation of, for example, the myocardial vascula resent potentially useful paramagnetic MR contrast agents. 25 ture and brain (see e.g. Schwarz, Advances in Echo-Contrast Furthermore, in the field of X-ray imaging it has been 1994(3)], pp. 48–49). observed that gases Such as carbon dioxide may be used as WO-A-9320802 proposes that tissue-specific ultrasonic negative oral contrast agents or intravascular contrast image enhancement may be achieved using acoustically agents. reflective oligolamellar liposomes conjugated to tissue The use of radioactive gases, e.g. radioactive isotopes of Specific ligands Such as antibodies, peptides, lectins etc. The inert gases Such as Xenon, has also been proposed in liposomes are deliberately chosen to be devoid of gas and So Scintigraphy, for example for blood pool imaging. will not have the advantageous echogenic properties of Targeted ultrasound contrast agents may be regarded as gas-based ultrasound contrast agents. Further references to comprising (i) a reporter moiety capable of interacting with this technology, e.g. in targeting to fibrin, thrombi and ultrasound irradiation to generate a detectable signal; (ii) one 35 atherosclerotic areas are found in publications by or more vectors having affinity for particular target Sites Alkanonyuksel, H. et al. in J. Pharm. Sci. (1996) 85(5), and/or structures within the body, e.g. for Specific cells or 486-490, J. Am. Coll. Cardiol. (1996)27(2) Suppl A, 298A; areas of pathology; and (iii) one or more linkers connecting and Circulation, 68 Sci. Sessions, Anaheim 13-16 Novem said reporter and vector(s), in the event that these are not ber 1995. directly joined. 40 There is also a number of publications concerning ultra The molecules and/or structure to which the contrast Sound contrast agents which refer in passing to possible use agent is intended to bind will hereinafter be referred to as the of monoclonal antibodies as vectors without giving Signifi target. In order to obtain specific imaging of a Selected cant practical detail and/or to reporters comprising materials region/structure in the body the target must be present and which may be taken up by the reticuloendothelial System available in this region/structure. Ideally it will be expressed 45 and thereby permit image enhancement of organs Such as the only in the region of interest, but usually will also be present liver-see, for example WO-A-9300933, WO-A-94.01140, at other locations in the body, creating possible background WO-A-9408.627, WO-A-9428874, U.S.