Classifications

• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07K—PEPTIDES
  • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
  • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
  • C07K14/575—Hormones
  • C07K14/655—Somatostatins
  • C07K14/6555—Somatostatins at least 1 amino acid in D-form
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
  • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
  • A61K51/04—Organic compounds
  • A61K51/08—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
  • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
  • A61K51/04—Organic compounds
  • A61K51/08—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins
  • A61K51/083—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins the peptide being octreotide or a somatostatin-receptor-binding peptide
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
  • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
  • A61K51/04—Organic compounds
  • A61K51/08—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins
  • A61K51/088—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins conjugates with carriers being peptides, polyamino acids or proteins
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P35/00—Antineoplastic agents
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K2123/00—Preparations for testing in vivo
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K38/00—Medicinal preparations containing peptides

Definitions

• the invention relates to a method of intraoperativt; / detecting and locating tumoura tissues in the body of a warm-blooded living being, to a method of radioguide surgery of said being, and to a radiopharmaceutical composition to be used for th latter method.
• the invention further relates to a labelled peptide compound to b used in said composition and to a kit for preparing said composition.
• tumours in particular malignant tumours
• tumours or thei me astases are extremely small «1 cm), and because of this small size they are no readily detectable and distinguishable using conventional imaging techniques.
• advanced imaging techniques such as SPECT acquisition techniques
• tumour-selective imaging agents is frequently unable to show all lesions, because of disturbing background activity that makes accurate image interpretation difficult.
• abdominal area it is often difficult t distinguish benign for malignant tissues using conventional imaging methods.
• An example of such a subgroup of tumours are gastro-enteropancreatic tumours that produce hormones, which result in sometimes life-threatening symptoms, e.g. massive diarrhea.
• a relatively new technique provides surgical aid: a gamma detecting probe, e.g. Neoprobe ⁇ that can be used to detect sources of gamma radiation that are ver small. After parenteral administration of a radiolabelled substance, the surgeon can intraoperatively, use this probe to find the lesions in which uptake of this substanc has taken place.
• E.W. Martin and coworkers have investigated this new technique e.g. Amer. J. Surgery 1 ⁇ 6, 1988, 386-392; Antibody Immunocon. Radiopharm. 4 1991 , 339-358. These investigators have observed that antibodies or antibod fragments, labelled with iodine-125, a low-energy gamma photon emittor, ar promising substances to be used in this technique.
• this techniqu may successfully target 80% of colorectal cancer and detect occult tumours in th abdomen in 20% of the surgical cases involving cancer of the colon.
• this improvement in diagnosing enables the surgeon t better resect tumour deposits, in particular those tumours and metastases which cannot be seen or palpated, and so contributes to the chance of curing cancer patients, the results of this technique are not yet satisfactory.
• the known radiolabelled substances generally show an insufficiently selective tumour uptake and, in particular, a not sufficiently fast blood clearance, so that the tumour to background ratio is often inadequate for accurate detection.
• a method as mentioned above comprising (a) parenterally administering to said being a pharmaceutical composition comprising, in a quantity sufficient for detection by a gamma detecting probe, a peptide compound labelled with a low-energy gamma photon emitting radionuclide, said peptide compound being derived from a peptide selected from the following groups: (i) peptides having a selective neurokinin 1 receptor affinity and having the general formula
• R is a hydrogen atom or a C ⁇ C* alkylcarbonyl group
• R 2 is a carbamoyl group, a carboxy group, a C,-C 4 alkoxycarbonyl group, a hydroxymethyl group or a C ⁇ C* alkoxymethyl group
• A is Arg, Gly or 5-oxo-Pro (pGlu);
• a 2 is Pro or ⁇ -Ala
• a 3 is Lys or Asp; A « is Gin, Asn or 5-oxo-Pro;
• a 5 is Gin, Lys, Arg, N-ac ⁇ lated Arg or 5-oxo-Pro; or wherein A 5 together with A 3 forms a cystine moiety;
• a ⁇ is Phe or Tyr;
• a 7 is Gly, Sar or Pro;
• a 8 is Leu or Pro; and
• R ⁇ is a straight or branched C 2 -C 4 alkyl group, which group may be inter rupted by thio, sulphinyl or sulphonyl; and their Tyr 0 derivatives;
• B, and B 2 are each independently Phe, MePhe, EtPhe, Tyr, Trp and Nal,
• B 3 is Lys or MeLys
• B 4 is Thr or Val
• R 7 is a 1 -hydroxyethyl group or an indol-3-ylmethyl group; and their Tyr 0 derivatives;
• peptides selected from cytokines, growth factors and hormones, as well as thei derivatives and analogues;
• Nal means a naphthylalan group
• Sar means a sarcosyl group
• Suitable examples of substituent R ⁇ are (CH 2 ) 2 S(0),CH 3 , wherein s is 0,1 or 2, an CH 2 CH(CH 3 ) 2 .
• cytokines are tumour necrosis factor (TNF), in particular TN o , interleukines (IL), in particular IL-1 , IL-2, IL-4, IL-5 and IL-6, and interferons.
• growth factors are epidermal growth factor (EGF), insulin-like growth-factor (IGF), in particular IGF-I (somatedin C) and IGF-II, bombesin, transforming growth factor (TGF), in particular TGF-oCand TGF- i, platelet-derived growth factor, fibroblast growth factor and nerve growth factor.
• hormones are luteinizing hormone-releasing hormone (LHRH), gastrin, gastrin-releasing peptide, angiotensin, thyroid-stimulating hormone, vasoactive intestinal polypeptide, prolactin, thyrotropin-releasing hormone, insulin, adrenocorticotropic hormone (ACTH), in particular 0 -MSH (melanocyte-stimulating hormone) and i -(meth ⁇ lsulfonyl)-L- oi-aminobutyryl-L- ⁇ -glutamyl-L-histidyl-L- phenylalanyl-D-lysyl-L-phenylalanine, cholecystokinin, corticotropin-releasing hormone (CRH), growth hormone-releasing hormone (GRH), arginine and lysine vasopressin, oxytocin, glucagon, secretin, parathyroid hormone (PTH) and PTH related peptide.
• tumours By using the above method of the invention, virtually all malignant tumours can be detected and located and can be distinguished from benign tissues, because these tumours contain substantially large numbers of receptors for binding the above peptide compound.
• the above-defined peptides are composed of amino acids, of which at least one may have the D-configuration.
• the peptides may also comprise so-called pseudo peptide bonds, viz. -CH 2 -NH- bonds, in addition to the natural amide bonds, viz. -CO-NH- bonds.
• Suitable examples of peptides sub (i) above are: (1 ) H-Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met-NH 2 (Substance P), (2) H-Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-Sar-Leu-Met(0 2 )-NH 2 ,
• Suitable examples of peptides sub(ii) above, which can be used as indicated above, are:
• Suitable examples of peptides sub (iii) above, which can be used as indicated above, are: EGF, TGF-c , gastrin, bombesin and derivatives of these peptides.
• Suitable low-energy gamma photon emitting radionuclides which can be used as labels for the peptide compounds to be used in the method of the present invention should have a gamma energy of approx. 80 keV at most.
• Such radionuclides are well tuned to the gamma detecting microprobe to be manipulated by the surgeon and intended to register the emitted gamma radiation.
• a hand-held microprobe e.g. Neoprobe 1000®
• Neoprobe 1000® is at present equipped with a miniature cadmium telluride crystal detector.
• Such a detector requires for optimum detection properties gamma energies in the range of approx. 30-80 KeV. Higher energies may cause excessive scattering so that the accuracy of the detection is considerably decreased.
• the surgeon can use his gamma detecting probe during operation to be sure that he/she does not overlook small-sized tumours.
• the well-tuned label enables the surgeon to accurately detect and locate such small tumours with the aid of the hand-held microprobe in order to guide the surgery treatment.
• suitable radionuclides for labelling the above peptide compounds are 1-125, As-73, Sb- 1 19, Cs-131 , Dy-159, W-181 and Hg-197.
• the desired radioisotope should be firmly attached to the peptide molecule to reduce the chance of detaching this label after administration to the living being.
• the peptide can be labelled with the desired isotope directly or indirectly, i.e. via a so-called linker. Direct labelling may be carried out, for example, by introducing a halogen atom or radioactive halogen atom, i.e. iodine-125, into an activated aromatic group (e.g. tyrosyl or imidazolyl) .present in the peptide, into the peptide molecule in a manner known per se, if desired followed by exchange with 1-125.
• an activated aromatic group e.g. tyrosyl or imidazolyl
• Tyrosine and histidine are suitable amino acids which, if present in the peptide molecule, allow an easy substitution with (radioactive) halogen.
• the labelling procedure is performed via a suitable linker, being capable of reacting with an amino group, preferably a terminal amino group, of said peptide, and having a functional group for binding said radioisotope.
• a suitable linker By using a suitable linker, the desired isotope can generally better be introduced into the peptide molecule.
• a suitable linker for labelling the peptide with 1-125 is derived from tyrosine or from Bolton-Hunter reagent, i.e. N-succinimidyl-3-(4-hydroxy-3- halo*phenyl)propionate, wherein halo* means iodine-125.
• the peptide is first reacted with tyrosine or with a "halo-deprived" Bolton-Hunter reagent, viz.
• N-succinimidyl-3-(4-hydroxyphenyl)propionate after which the derivatised peptide, thus obtained, is substituted by the desired halogen radioisotope by an appropriate reaction.
• the peptide can be labelled with the desired radioactive haiogen isotope without affecting its biological properties.
• the above radioiodinating reaction is preferably performed by reacting the peptide in question with a solution of an alkali metal radionuclide from 1-125 iodide, under the influence of a halide-oxidizing agent, such as chloramine T or iodogen.
• a halide-oxidizing agent such as chloramine T or iodogen.
• the above substitution reaction can be carried out with a non-radioactive halogenide, after which halo-exchange with radioactive halogen is performed, e.g. as described in European patent 165630.
• tumours and metastases which can be detected and located by using the method of the present invention, are small soft tissue tumours which are firmly attached to the surrounding benign tissues. This makes the surgical removal of such tumours after their detection often difficult. It is another object of the present invention to facilitate the therapeutic treatment of such tumoural tissues and consequently to improve the radioguided surgery.
• the present invention also and in particular relates to a method of radioguided surgery of a warm-blooded living being, which method, in addition to the method as discussed hereinbefore and intended to detect and locate tumoural tissues, comprises (i) parenterally administering to said being a pharmaceutical composition comprising, in a quantity sufficient for at least partial necrosis of tumoural tissues, a peptide compound derived from a peptide as defined hereinbefore and labelled with an isotope with sufficiently high specific activity and emitting corpuscular radiation, preferably selected from the group consisting of radionuclides as reviewed by Schubiger et al. (in 6th Int. Symp. Radiopharm.
• Suitable examples of the last-mentioned lanthanide radionuclides are Pr-142, Pr-143, Pm-149, Pm-151, Sm-153, Gd-159, Tb-161 , Dy-165, Ho-166, Er- 169, Tm-172, Yb-169, Yb-175 and Lu-177.
• an injected dose has a cell killing effect due to the uptake in the tumoural tissues in question, leading to an at least partial necrosis of the tumour cells. This enables the surgeon to more easily remove these tumours by excision during surgery.
• the overall result can be a treatment schedule, wherein an optimum use is made of this combination of detection and therapy.
• the gamma emissions permit the accurate detection of these tissues with a microprobe.
• lanthanide radionuclides are very suitable to perform both functions at the same time, viz. have a suitable gamma energy for detection and have a corpuscular radiation emitting effectiveness sufficient for tumour cell necrosis.
• terbium-1 61 Tb-161
• This lanthanide radionuclide combines an optimum gamma energy for microprobe detection, viz. in the range of 25-74 keV, with a beta emission suitable for therapeutic treatment of tumour cells, viz.
• terbium-161 in the range of 250-590 keV.
• the half-life of terbium-161 is extremely appropriate for the intended purpose, viz. 6.91 days. This means that sufficiently long after injection the blood clearance of radioactivity is sufficiently complete to permit accurate detection of the tumoural tissues, while at the same time necrosis of the malignant tissue cells has advanced sufficiently to allow easy excision of the detected tumours.
• this preferred lanthanide radionuclide is readily accessible and can be produced carrier-free by irradiation of highly enriched gadolinium Gd-160 in a nuclear reactor.
• the labelled peptide compound to be used in the method of the invention is provided, directly or through a spacing group, with a chelating group.
• This chelating group is attached by an amide bond to an amino group of said peptide and is derived from ethylene diamine tetra-acetic acid (EDTA), di ⁇ ethylene triamine penta-acetic acid (DTPA), ethyleneglycol-0,0'-bis(2- aminoethyl)-N,N,N',N'-tetra-acetic acid (EGTA), N,N-bis(hydroxybenzyl)- ethylenediamine-N,N'-diacetic acid (HBED), triethylene tetramine hexa-acetic acid (TTHA), 1 ,4,7,10-tetraazacyclododecane-N,N',N",N'"-tetra-acetic acid (DOTA), 1 ,4,8,1 1 -t
• EDTA ethylene diamine tetra
• TETA 1 ,2-diaminoc ⁇ clohexane tetra-acetic acid
• DCTA 1 ,2-diaminoc ⁇ clohexane tetra-acetic acid
• DTPA 1 ,2-diaminoc ⁇ clohexane tetra-acetic acid
• EDTA substituted EDTA
• R is a branched or non-branched, optionally substituted hydrocarbyl radical, which may be interrupted by one or more hetero-atoms selected from N, 0 and S and/or by one or more NH groups
• Y is a group which is capable of reacting with an amino group of the peptide and which is preferably selected from the group consisting ofcarbonyl,carbimidoyl,N-(C,-C ⁇ )alkylcarbimidoyl, N-hydroxycarbi- midoyl and N-(C,-C 6 )-aIkoxycarbimidoyl; and wherein said optionally present spacing group has the general formula
• R 5 is a C,-C 10 alkylene group, a C,-C 10 alkylidene group or a C 2 -C 10 alkenylene group
• X is a thiocarbonyl group or a methylcarbonylgroup.
• Suitable chelators of the general formula HI are unsubstituted or substituted 2-iminothioIanes and 2-iminothiacyclohexanes, in particular 2- ⁇ mino-4-mercaptomethylthiolane.
• the invention further relates to a radiopharmaceutical composition to be used for the method of radioguided surgery as defined above, which composition comprises in addition to a pharmaceutically acceptable carrier and, if desired, at least one pharmaceutically acceptable adjuvant, as the active substance a peptide compound labelled with a low-energy gamma photon emitting radionuclide and a corpuscular radiation emitting isotope as defined hereinbefore.
• the composition can be brought into a form more suitable for parenteral administration, e.g. by adding a pharmaceutically acceptable liquid carrier material.
• the solution should of course be in a sterile condition.
• the above composition preferably comprises as the active substance a peptide compound provided with a chelating group as defined above, said chelating group chelating a metal radionuclide.
• the preferred radionuclides of the lanthanide group have excellent characteristics for being chelated with the above aminoacetic acid based chelating agents, thus assuring a good and stable binding to the chelating group of the peptide compound.
• the radioisotope is presented to the chelating group comprising peptide compound in the form of a salt of a suitable acid, e.g. a mineral acid or acetic acid.
• the complex-forming reaction can generally be carried out in a simple manner and under conditions which are not detrimental to the peptide.
• the invention further relates to a labelled peptide compound to be used as an active ingredient in the above composition, said peptide compound having a selective affinity to endocrinically active tumours and being labelled with a low-energy gamma photon emitting radionuclide and a corpuscular radiation emitting isotope, as defined hereinbefore, preferably with a suitable lanthanide radionuclide having the above-defined radiation characteristics.
• the invention finally relates to a so-called cold kit for preparing a radiopharmaceutical composition, comprising (i) a peptide provided with a chelating group as defined hereinbefore, to which substance, if desired, an inert pharmaceutically acceptable carrier and/or formulating agent(s) and/or adjuvant(s) is/are added, (ii) a solution of a salt of a lanthanide radionuclide having the above-defined radiation characteristics, and (iii) instructions for use with a prescription for reacting the ingredients present in the kit.
• Such a kit as described above can be delivered to the user.
• the user can easily perform the labelling of the chelated peptide with the lanthanide radionuclide by himself or herself in the clinical hospital or laboratory.
• the labelling procedure is simple and does not require complicated manipulations, so that the user is able to prepare the labelled composition from the kit ingredients by using the facilities that are at his or her disposal.
• the radionuclide is the only ingredient in the kit having a restricted shelf life. Therefore, if desired, the lanthanide radionuclide may be delivered separately and substituted after its expiration date.
• the DTPA-Octreotide kit formulated on basis of sodium acetate buffer with the final composition
• DTPA-Octreotide 0.5 mg is dissolved in 4 ml of acetic acid solution, and 5 ml of sodium acetate solution are added.
• Tb-161 For the separation of Tb-161, 5 ml of the Gd/Tb stock solution in 0.02N HCl is evaporated to dry and taken up in 200 ⁇ l of 0.02N HCl. This solution is loaded on a 0.8 x 12 cm column of SCX BioRad® 50 W-X8, 200-400 mesh, in NH 4 + form. As an eluent is used a 0.2 M solution of ⁇ -hydroxy- isobutyric acid, adjusted to pH 4.1 with ammonia. Fractions of 1 ml of eluate are collected for radiodnuclide identification. Combined fractions containing Tb-161 are made of 0.5N in HCl and run over a second small column of BioRed® 50 -X8 in H + form.
• Radionuclide purity determined by ⁇ spectrometry (ND 66 ⁇ spectrometer Ga/Li detector) : substantiallyl 100%; no other radionuclide detected.
• Radiochemical purity Thin layer chromatography -ITL SG (Gelman) plates, ascending, solvent, 1M sodium acetate pH 5. Result: single peak on front, 98.9% Tb-161 activity.
• kits of DTPA-Octreotide prepared according to Example 1 containing 10 or 50 ⁇ g DTPA-Octreotide, are labelled by addition of 0.5 ml of Tb-161 solution obtained under B. The mixture is incubated for 30 min. at room temperature.
• Tb-161-DTPA-Octreotide Rf ca 0.5-0.6 Free Tb-161 Rf ca 0.9-1.0
• the labelled solution is purified over a Sep- Pak R C18 cartridge, which after loading is washed with water (5 ml) and eluted with methanol (5 ml), the latter fraction containing Tb-161 Octreotide. Evaporation and dissolution of the residue in physiologic saline solution give after sterilization (membrane filtration) the desired injectable preparation.
• the obtained solution is withdrawn and trasferred to a small quartz beaker, the ampoule is washed with 3 x 1-ml portions of water and the washings are combined with the active solution.
• the solution of Yb-175 chloride is twice evaporated to dryness with concentrated HCl and the rest is taken up into 2 x 5-ml of 0.02N HCl, transferred to a 20 ml volumetric flask and diluted to the desired volume with 0.02N HCl.
• Example 1 Several kits containing 10 ⁇ g of DTPA-Octreotide prepared according to Example 1 are labelled by addition of 1 ml of the Yb-175 stock solution. The mixture is let to incubate 30 min. at room temperature. Samples for analysis are taken at time intervals indicated by the results. Used analytical methods are described in Example 1.
• Yb-175 Octreotide LY at 30 min. ITLC Rf 0.5-0.6 98.1%
• Yb-175-Octreotide can be used separately, in this case by administering Yb-175-Octreotide first to cause partial or deep tumour necrosis, followed by administration of DTPA-125-I-Tyr 3 -Octreotide to guide the tumours removal, or they can be administered simultaneously as a mixture in an appropriate ratio.
• a mixture is obtained by mixing both agents in the proper ratio.
• the difference in IPA of Yb-175 and 1-125, viz. 4.2 and 60.2 days respectively gives sufficient time for therapeutic effect while at the moment of surgery the background radiation, originating from Yb-175 (having also very low ⁇ abundance) , is already sufficiently low as not to diminish the sensitivity of the microprobe.
• Obtained Ho-166 stock solution (1 ml) has a specific activity 525 ⁇ Ci/ ⁇ g Ho-166, a radionuclide purity > 99.9% and a radiochemical purity > 99.9%, both values determined by the ⁇ ethods described in Example I.
• kits containing lO ⁇ g of DTPA-Octreotide prepared according to Example I., are labelled by addition of 0.5 or 1 ml of Ho-166 stock solution. The mixture is let to incubate 30 min. at room temperature. Samples for analysis at time intervals indicated by the results. Used analytical methods are described in Example I.
• Example II Similarly as described in Example II. is used a combination of both preparations, Ho-166-Octreotide and Tb-161-
• the favourable ratio of half-lives (Ho-166 26.9 h, Tb-161 d) guarantees that after one VA of Tb-161 no more than 1.3% of the originally bound Ho-166 activity remains at the site of the tumour so that the sensitivity of the microprobe cannot be influenced, particularly not since the range of gamma rays emitted by Ho-166 (48-80 keV) is comparable to that of Tb-161.

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Cited By (10)

Citations (2)

• 1993
  • 1993-03-24 JP JP5516829A patent/JPH07505621A/ja active Pending
  • 1993-03-24 EP EP93909165A patent/EP0636032A1/en not_active Withdrawn
  • 1993-03-24 WO PCT/US1993/002772 patent/WO1993018797A1/en not_active Application Discontinuation
  • 1993-03-24 AU AU39675/93A patent/AU3967593A/en not_active Abandoned
  • 1993-03-24 CA CA002131315A patent/CA2131315A1/en not_active Abandoned

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