NZ262435A - Kit for diagnosing an hcg fragment secreting cancer and immunotherapeutical means therefor - Google Patents
Kit for diagnosing an hcg fragment secreting cancer and immunotherapeutical means thereforInfo
- Publication number
- NZ262435A NZ262435A NZ262435A NZ26243594A NZ262435A NZ 262435 A NZ262435 A NZ 262435A NZ 262435 A NZ262435 A NZ 262435A NZ 26243594 A NZ26243594 A NZ 26243594A NZ 262435 A NZ262435 A NZ 262435A
- Authority
- NZ
- New Zealand
- Prior art keywords
- hcg
- polypeptides
- polypeptide
- primary sequence
- fragments
- Prior art date
Links
- 206010028980 Neoplasm Diseases 0.000 title claims description 38
- 239000012634 fragment Substances 0.000 title claims description 15
- 230000003248 secreting effect Effects 0.000 title claims description 14
- 201000011510 cancer Diseases 0.000 title claims description 13
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 74
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 72
- 229920001184 polypeptide Polymers 0.000 claims description 66
- 238000000034 method Methods 0.000 claims description 18
- 238000009169 immunotherapy Methods 0.000 claims description 11
- 238000002649 immunization Methods 0.000 claims description 10
- 210000002966 serum Anatomy 0.000 claims description 10
- 230000003053 immunization Effects 0.000 claims description 9
- 239000000203 mixture Substances 0.000 claims description 9
- 238000009007 Diagnostic Kit Methods 0.000 claims description 8
- 210000001744 T-lymphocyte Anatomy 0.000 claims description 8
- 231100000135 cytotoxicity Toxicity 0.000 claims description 6
- 150000001413 amino acids Chemical class 0.000 claims description 5
- 230000003013 cytotoxicity Effects 0.000 claims description 5
- 239000002671 adjuvant Substances 0.000 claims description 4
- 230000036039 immunity Effects 0.000 claims description 4
- 239000003795 chemical substances by application Substances 0.000 claims description 3
- 230000004044 response Effects 0.000 claims description 3
- 108010039918 Polylysine Proteins 0.000 claims description 2
- 210000004369 blood Anatomy 0.000 claims description 2
- 239000008280 blood Substances 0.000 claims description 2
- 238000003745 diagnosis Methods 0.000 claims description 2
- 238000004519 manufacturing process Methods 0.000 claims description 2
- 229920000656 polylysine Polymers 0.000 claims description 2
- 238000000338 in vitro Methods 0.000 claims 1
- 230000000638 stimulation Effects 0.000 claims 1
- 230000035755 proliferation Effects 0.000 description 11
- 108090000623 proteins and genes Proteins 0.000 description 10
- 208000007097 Urinary Bladder Neoplasms Diseases 0.000 description 8
- 210000004027 cell Anatomy 0.000 description 7
- 210000004698 lymphocyte Anatomy 0.000 description 6
- 239000005556 hormone Substances 0.000 description 5
- 229940088597 hormone Drugs 0.000 description 5
- 210000003819 peripheral blood mononuclear cell Anatomy 0.000 description 4
- 235000018102 proteins Nutrition 0.000 description 4
- 102000004169 proteins and genes Human genes 0.000 description 4
- 229960005486 vaccine Drugs 0.000 description 4
- 230000015572 biosynthetic process Effects 0.000 description 3
- 230000001573 trophoblastic effect Effects 0.000 description 3
- 241000193830 Bacillus <bacterium> Species 0.000 description 2
- 208000006332 Choriocarcinoma Diseases 0.000 description 2
- 102000011022 Chorionic Gonadotropin Human genes 0.000 description 2
- 108010062540 Chorionic Gonadotropin Proteins 0.000 description 2
- 229920001917 Ficoll Polymers 0.000 description 2
- 102000012673 Follicle Stimulating Hormone Human genes 0.000 description 2
- 108010079345 Follicle Stimulating Hormone Proteins 0.000 description 2
- 230000003509 anti-fertility effect Effects 0.000 description 2
- 239000000427 antigen Substances 0.000 description 2
- 108091007433 antigens Proteins 0.000 description 2
- 102000036639 antigens Human genes 0.000 description 2
- 210000001151 cytotoxic T lymphocyte Anatomy 0.000 description 2
- 229940028334 follicle stimulating hormone Drugs 0.000 description 2
- 230000002163 immunogen Effects 0.000 description 2
- 210000005087 mononuclear cell Anatomy 0.000 description 2
- 230000035935 pregnancy Effects 0.000 description 2
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- 230000002381 testicular Effects 0.000 description 2
- 210000001550 testis Anatomy 0.000 description 2
- 238000002560 therapeutic procedure Methods 0.000 description 2
- IQFYYKKMVGJFEH-OFKYTIFKSA-N 1-[(2r,4s,5r)-4-hydroxy-5-(tritiooxymethyl)oxolan-2-yl]-5-methylpyrimidine-2,4-dione Chemical compound C1[C@H](O)[C@@H](CO[3H])O[C@H]1N1C(=O)NC(=O)C(C)=C1 IQFYYKKMVGJFEH-OFKYTIFKSA-N 0.000 description 1
- 108010042708 Acetylmuramyl-Alanyl-Isoglutamine Proteins 0.000 description 1
- 208000005623 Carcinogenesis Diseases 0.000 description 1
- VYZAMTAEIAYCRO-UHFFFAOYSA-N Chromium Chemical compound [Cr] VYZAMTAEIAYCRO-UHFFFAOYSA-N 0.000 description 1
- 108020004705 Codon Proteins 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 108010086677 Gonadotropins Proteins 0.000 description 1
- 101000957351 Homo sapiens Myc-associated zinc finger protein Proteins 0.000 description 1
- 241000701044 Human gammaherpesvirus 4 Species 0.000 description 1
- 208000006937 Hydatidiform mole Diseases 0.000 description 1
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 description 1
- 108700018351 Major Histocompatibility Complex Proteins 0.000 description 1
- 102100038750 Myc-associated zinc finger protein Human genes 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
- 108010033276 Peptide Fragments Proteins 0.000 description 1
- 102000007079 Peptide Fragments Human genes 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- 101800000874 Small capsid protein Proteins 0.000 description 1
- 101800000996 Small capsid protein precursor Proteins 0.000 description 1
- 230000024932 T cell mediated immunity Effects 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- 235000004279 alanine Nutrition 0.000 description 1
- 150000001294 alanine derivatives Chemical class 0.000 description 1
- 235000001014 amino acid Nutrition 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000000259 anti-tumor effect Effects 0.000 description 1
- 235000003704 aspartic acid Nutrition 0.000 description 1
- 230000002238 attenuated effect Effects 0.000 description 1
- 210000003719 b-lymphocyte Anatomy 0.000 description 1
- OQFSQFPPLPISGP-UHFFFAOYSA-N beta-carboxyaspartic acid Natural products OC(=O)C(N)C(C(O)=O)C(O)=O OQFSQFPPLPISGP-UHFFFAOYSA-N 0.000 description 1
- 230000036952 cancer formation Effects 0.000 description 1
- 231100000504 carcinogenesis Toxicity 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 229940015047 chorionic gonadotropin Drugs 0.000 description 1
- 229910052804 chromium Inorganic materials 0.000 description 1
- 239000011651 chromium Substances 0.000 description 1
- 210000000078 claw Anatomy 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 230000001472 cytotoxic effect Effects 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 239000000032 diagnostic agent Substances 0.000 description 1
- 229940039227 diagnostic agent Drugs 0.000 description 1
- 238000002405 diagnostic procedure Methods 0.000 description 1
- 238000010494 dissociation reaction Methods 0.000 description 1
- 230000005593 dissociations Effects 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 210000004696 endometrium Anatomy 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- 229960002743 glutamine Drugs 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 229940084986 human chorionic gonadotropin Drugs 0.000 description 1
- 238000002513 implantation Methods 0.000 description 1
- 238000010348 incorporation Methods 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 210000004072 lung Anatomy 0.000 description 1
- 230000001294 luteotrophic effect Effects 0.000 description 1
- 230000003211 malignant effect Effects 0.000 description 1
- 210000001161 mammalian embryo Anatomy 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- BSOQXXWZTUDTEL-ZUYCGGNHSA-N muramyl dipeptide Chemical compound OC(=O)CC[C@H](C(N)=O)NC(=O)[C@H](C)NC(=O)[C@@H](C)O[C@H]1[C@H](O)[C@@H](CO)O[C@@H](O)[C@@H]1NC(C)=O BSOQXXWZTUDTEL-ZUYCGGNHSA-N 0.000 description 1
- 239000002105 nanoparticle Substances 0.000 description 1
- 239000002077 nanosphere Substances 0.000 description 1
- 210000000496 pancreas Anatomy 0.000 description 1
- 229940049954 penicillin Drugs 0.000 description 1
- 210000002826 placenta Anatomy 0.000 description 1
- 230000003169 placental effect Effects 0.000 description 1
- 238000004393 prognosis Methods 0.000 description 1
- 238000002271 resection Methods 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 238000004611 spectroscopical analysis Methods 0.000 description 1
- 239000003270 steroid hormone Substances 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 230000020382 suppression by virus of host antigen processing and presentation of peptide antigen via MHC class I Effects 0.000 description 1
- 230000001646 thyrotropic effect Effects 0.000 description 1
- 210000002993 trophoblast Anatomy 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/0005—Vertebrate antigens
- A61K39/0006—Contraceptive vaccins; Vaccines against sex hormones
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/0005—Vertebrate antigens
- A61K39/0011—Cancer antigens
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- 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
- C07K14/59—Follicle-stimulating hormone [FSH]; Chorionic gonadotropins, e.g.hCG [human chorionic gonadotropin]; Luteinising hormone [LH]; Thyroid-stimulating hormone [TSH]
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/74—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving hormones or other non-cytokine intercellular protein regulatory factors such as growth factors, including receptors to hormones and growth factors
- G01N33/76—Human chorionic gonadotropin including luteinising hormone, follicle stimulating hormone, thyroid stimulating hormone or their receptors
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Immunology (AREA)
- Molecular Biology (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Behavior & Ethology (AREA)
- Endocrinology (AREA)
- Public Health (AREA)
- Microbiology (AREA)
- Veterinary Medicine (AREA)
- Mycology (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Reproductive Health (AREA)
- Epidemiology (AREA)
- Biomedical Technology (AREA)
- Urology & Nephrology (AREA)
- Biochemistry (AREA)
- Hematology (AREA)
- Oncology (AREA)
- General Physics & Mathematics (AREA)
- Gastroenterology & Hepatology (AREA)
- Biophysics (AREA)
- Genetics & Genomics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Zoology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Toxicology (AREA)
- Pathology (AREA)
- Analytical Chemistry (AREA)
- Physics & Mathematics (AREA)
- Virology (AREA)
- Food Science & Technology (AREA)
- Cell Biology (AREA)
- Biotechnology (AREA)
Description
New Zealand No. 262435 International No. PCT/FR94/00265
Priority D«le(s):....ll|s.|3S Sjiate
Comply® Specification Fll»d:
Claw: (6) I.Ha«jay:..
P.O. Journal No: J.yd.Jf?..,...
NEW ZEALAND PATENTS ACT 1953 COMPLETE SPECIFICATION
Title of Invention:
Kit for diagnosing an HCG- or HCG fragment-secreting cancer and immunotherapeutical means therefor
K'amo, address and nationality of applicant(s) as in international application form:
LABORATOIRE L. LAFON, of 19, avenue du Professeur Cadiot, F-94701 Maisons Alfort, France
1 -
PCT/FR94/0(^^5^ 4 3 5
Diagnostic kit for a cancer secreting hCG or fragments of hCG and means intended for the immunotherapy of such a cancer.
The present invention relates to a method for 5 treating a cancer secreting hCG or fragments of hCG and to a means for implementing this method.
Human chorionic gonadotropin hormone (hCG) is an oligomeric glycoprotein consisting of the noncovalent association of two subunits a(hCG-a) and £(hCG-£). it 10 belongs to the family of gonadothyreotropic hormones also comprising the follicle-stimulating hormone (FSH), the luteotropic hormone (LH) and the thyrotropic hormone (TSH) . The hCG is secreted physiologically by the tropho-blast from the beginning of pregnancy and stimulates the 15 Corpus leuteum in order to maintain the synthesis of steroid hormones which is essential for the implantation of the embryo. Moreover, hCG and/or its subunits are also secreted by a majority of trophoblastic tumours and some nontrophoblastic tumours, thus constituting biological 20 markers which can help in the diagnosis or in the prognosis and in the monitoring of these tumours (Bellet et al. Rev. Prat. 40:1677-1990).
The amino acid sequence of hCG has been described by Morgan F.J. et al., The amino acid sequence of Human 25 chorionic gonadotropin, J. Biol. Chem. 250, 5247, 1975.
Within the normal population, very low serum levels of dimeric hCG are found in men (<0.05 ng/ml corresponding to the detection limit of the method used) and in nonmenopausal women (<0.1 ng/ml). The hCG is 30 mainly detectable 30% of women over 45 years old (from 0.1 to 1 ng/ml). Several studies suggest that the hCG present in the serum of healthy subjects might be produced by hypophyseal cells. The a-hCG is detectable in healthy men (<1 ng/ml) and in nonmenopausal women over 45 years old (<3 ng/ml) . In contrast, /S-hCG is not detectable the normal population.
In the case of trophoblastic tumours, it has been shown that benign (hydatidiform moles) and malignant (choriocarcinomas) placental tumours are characterized by
262 435
a high serum hCG//S-hCG ratio: the percentage of free hCG which is 0.05 to 1% during normal pregnancies increases to 1-5% in the event of mole and exceeds 5% in choriocarcinomas. This serum hCG//3-hCG ratio is used aB 5 control for the progression of testicular tumours: a percentage of /3-hCG which becomes less than 5% is often characteristic of a favourable progression of the tumour during the treatment. This ratio will rapidly increase in the event of an unfavourable progression or of relapse. 10 In the case of non trophoblastic tumours,
Marcillac et al., Cancer Res. 52:3901, 1992 have shown that jS-hCG, which is nondetectable . in healthy individuals, 'is found in numerous patients suffering from certain nontrophoblastio tumours (tumours of the bladder, 15 of the pancreas, of the neck of the womb, tumours without an established primary*cause# tumours of the endometrium and of the lungs).
Moreover, WO-A 90/02759 describes a product 20 termed UGP which is a mixture of a combination of three peptide fragments of which one is fi-hCG and the other two are said to be different from the peptides present in /3-hCG.
Under a very general definition, this document 25 defines diagnostic agents which may be constituted by UGP, whereas in practice it is the antibodies which are used.
EP-A-0,323,769 describes peptides comprising essentially a /3-hCG sequence coupled to an a-hCG sequence 30 in order to constitute an antifertility vaccine. These peptides are intended to induce the formation of antibodies .
US-A-4,201,770 also essentially describes an antifertility vaccine. This vaccine comprises a hormone 35 (such as CG) , its subunits or fragments of these subunits which have been modified by coupling with a nonendo-geneous material.
WQ-A-92/07272 simply describes a method for
N.Z. PATFNT OFFICE
13 MAS 1597
262 435
- 2a -
detecting, inter alia, a cancer and which consists in searching for the presence of "serum gonadotropin peptide complex" after dissociation.
The subject o£ the present invention is a method 5 for treating a cancer secreting hCG or fragments of hCG which consists in
- identifying, in the serum of a patient with the aid of a set of polypeptides covering at least part of the primary sequence of the hCG, a state of immunization
against one of these polypeptides,
- administering to the patient at least one polypeptide corresponding to the antigen responsible for the state of immunization.
262435
correspond to sequences which overlap.
There may be used for example a set of polypeptides corresponding to the following sequences:
a-hCG 1-20
15-35
-50 45-65 60-80 75-92
0-hCG 1-20
-35 30-50 45-65 60-80
75-95
90-110 105-125 120-145
At least a set of polypeptides which covers the
2 0 primary sequence of the jS subunit of hCG or at least part of this sequence will preferably be used.
It should be noted that the synthesis of the 0 subunit is dependent on a multigene group containing 7 genes (Talmadge et al., Nucl. Acids Res. 12, 8415,
1984) .
Analysis of the nucleotide sequences shows that three of these genes (1, 3, 5) have an identical structure, another four of these genes (2, 6, 7 and 8) encode a variant in codon 117 with substitution of alanine for
3 0 an aspartic acid. A study of the expression of the genes
(Bo et al., J. Biol. Chem. 267, 3179, 1992) shows that genes 1, 3 and 5 are expressed by the normal placenta.
It is thought that in the tumorigenesis processes affecting the nontrophoblastic cells, the expression of
several genes leads both to the secretion of the normal protein and of a protein mutated in 117.
In the present invention, /S subunit therefore designates both the normal protein and the protein mutated in 117.
262 435
"Polypeptide corresponding to the antigen responsible for the state of immunization designates a polypeptide which has the same sequence as one of the polypeptides responsible for the state of immunization or 5 a sequence resulting from the combination of at least partial sequences of the polypeptides responsible for the state of immunization.
The state of immunization in relation to one of the polypeptides can be identified by a test of stimula-10 tion of the proliferation of the T lymphocytes by the polypeptides, or alternatively by a test of cytoxicity of the T lymphocytes in relation to the polypeptides.
In the case of cytotoxic T lymphocytes, the polypeptides recognized are formed by a chain of at least 15 9 amino acids corresponding to part of the primary sequence of the hCG. In order to be recognized by the cytotoxic T lymphocytes, the polypeptides interact with the class I molecules of the patient's major histocompatibility complex.
2 0 Each class I molecule interacts with peptides containing precise units. There may be used for example the polypeptides corresponding to the following sequences of the jS-hCG:
3 -
12
4 -
12
8 -
16
-
18
11 -
17 -
23 -
1-3
24 -
33
28 -
37
32 -
41
40 -
48
40 -
49
44 -
52
47 -
55
48 -
56
51 -
. 59
262435
55 - 62 61 - 69 68 - 76 71 - 80 5 72-80
75 - 84
77 - 86
78 - 86 83 - 92
100 - 108
119 - 128 125 - 134 134 - 143.
The possible presence of serum antibodies speci-15 fic for hCG or its fragments can also be detected. These antibodies can be detected especially by an imsiuno-enzymatic method such as an ELISA method.
The subject of the present invention is also a diagnostic kit for a cancer secreting hCG or fragments of 20 hCG comprising a set of polypeptides covering at least part of the primary sequence of hCG.
The subject of the present invention is also a composition intended for the immunotherapy of cancers secreting hCG or fragments of hCG comprising an effective 25 dose of a polypeptide corresponding to at least part of the primary sequence of hCG and especially a polypeptide corresponding to at least part of the primary sequence of the § subunit of hCG and a pharmaceutically acceptable vehicle.
The subject of the present invention is also the use of a polypeptide corresponding to at least part of the primary sequence of hCG and especially a polypeptide corresponding to at least part of the primary sequence of the /3 subunit of hCG for the manufacture of a composition 35 intended for the immunotherapy of cancers secreting hCG or fragments of hCG.
For therapy, the polypeptides can be used in free form or in a more immunogenic form, especially in a form in which the peptide is coupled to a polylysine matrix
262435
according to the MAP technique (Posnett et al. J. Biol. Chem. 263, 17, 1988) .
In addition, for therapy, the polypeptide is advantageously administered with an immunity adjuvant 5 agent. As examples of immunity adjuvants, there may be mentioned muramyldipeptide and SAF-1 (Allison et al., J. Immunol. Methods 45, 157, 1986).
The polypeptide can also be administered coupled to nanospheres or to nanoparticles such as those 10 described in EP 02 04 424.
Finally, the polypeptide can be protected at its NHj- and COOH-terminal ends in order to reduce the risk of degradation.
In the case of the treatment of certain cancers 15 and especially in the case of cancer of the bladder, BCG (Calmette-Guerin bacillus) vaccine consisting of attenuated live Btrains of tubercle bacillus can also be additionally administered concomitantly or as a mixture (see especially O. Lamm et al., New England Journal of 20 Medicine 325, 17, 1205, 1991).
The subject of the present invention is therefore, in addition, a composition intended for the immunotherapy of cancers secreting hCG or fragments of hCG comprising an effective dose of a polypeptide correspond-25 ing to at least part of the primary sequence of hCG and an effective dose of BCG in a pharmaceutically acceptable vehicle.
In practice, the immunotherapy can be carried out with polypeptide doses from 1 to 100 mg, administered 30 especially in the region of the tumour. For example, the polypeptides can be administered in the case of a cancer of the bladder, as is already done in the case of an immunotherapy using BCG, that is to say by intravesical instillation one week after the resection of the tumour, 3 5 then this administration is repeated each week for 6 weeks and additional intravesical administrations are performed 3, 6, 12, 18 and 24 months later. Administrations via the percutaneous route or via the general route are also possible.
262435
Description of the figures
Fig. 1 represents the proliferation of the lymphocytes obtained from a patient having cancer of the bladder in the presence of hCG, of the free a and /5 5 subunits and of different polypeptides.
Fig. 2 represents the proliferation of the lymphocytes obtained from another patient having cancer of the bladder in the presence of the free a and /S sub-units and of different polypeptides. 10 Fig. 3A represents the proliferation of the lymphocytes obtained from a patient having cancer of the bladder in the presence of different polypeptides.
Fig. 3B represents the proliferation of the lymphocytes obtained from a patient having cancer of the 15 testicle in the presence of different polypeptides.
An account of the diagnostic tests will be given below.
1) Isolation of the mononuclear cells (PBMC) bv Ficoll gradient:
Heparinized whole blood collected from cancer patients is deposited on a Ficoll-hypaque solution (1.077 g/1) in a conical tube. The tube is centrifuged for 30 min and 900 g, 18-20°C. The layer corresponding to the mononuclear cells is recovered with the aid of a. 25 pipette. The cells are washed in HANKs buffer.
2) Test of lvmphocvte proliferation
The PBMCs are cultured (2 x 10s cells per well) in RPMI 1640 supplemented with 10% human serum AB, 2 mM L-glut amine, 100 IU/ml of penicillin and 100 ^tg/ml of 30 streptomycin, in the presence of polypeptide at different concentrations. The cells are then incubated at 37°C with 5% C03 for 5 days. One fid of tritiated thymidine is then added to each well. The cells are collected 18 hours later and the incorporation of the radioactivity is 35 analysed by spectrometry in a scintillation liquid using a /3 counter.
The results of proliferation of the lymphocytes obtained from a patient having cancer of the bladder in the presence
262435
8 -
- of the culture medium (M)
- of the hCG and its a and 0 subunits
- of different polypeptides belonging to the a and £ subunits,
are given in Pig. 1.
The results reveal a significant proliferation with the polypeptide corresponding to the sequence 95-115 of the hCG jS subunit as well as with the polypeptide corresponding to the sequence 25-59 of the hCG a subunit. 10 The polypeptides 95-115 j3-hCG and 25-59 a-hCG can be used for an immunotherapy in the patient tested.
The same type of results on the lymphocytes from another patient having cancer of the bladder are given in Fig. 2. The results reveal a significant proliferation 15 with the polypeptide corresponding to the sequence 31-50 of the hCG /? subunit as well as with the polypeptide corresponding to the sequence 41-61 of the hCG a stibunit. In this patient, the polypeptides 31-50 of the /3-hCG and 41-61 of the a-hCG can be used for an immunotherapy. 20 The results of tests of proliferation of PBMC
from another patient having cancer of the bladder secreting the hCG fi subunit in the presence of different polypeptides covering the entire hCG /5 subunit (50 fan) are given in Fig. 3A.
The results show that for this patient, the poly peptide corresponding to the /J region (2 0-47) of hCG is most effective for inducing proliferation of the T cells.
Moreover, other tumours (pancreatic and testicular) were studied. The T cells of a patient, having 30 cancer of the testicles, secreting 0-hCG, were proliferated in the presence of the /S-hCG peptide (20-47) (Fig. 3B).
These results show that the peptides corresponding to part of the /3-hCG sequence are immunogenic in 35 patients having a j8-hCG-producing tumour and can thus be used to develop an antitumour immunotherapy.
In contrast, no response against hCG, its sub-units or peptides of analogous structure was observed in 21 pregnant women.
4 65
The results thus obtained will be given below. Cellular immune response against hCG or its subunits.
Populations tested
Total
Responses
Vesical tumours
21
/5-hCG+ tumours
14
/S-hCG- tumours
7
0
Pregnant women
21
0
Healthy subjects
13
0
3) Test of cytotoxicity
The PBMCs isolated by Ficoll gradient are tested for their cytotoxic action against autologous targets (B lymphocytes from the patient which are immortalized by the Epstein-Barr virus) labelled with "chromium and 15 serving as the hCG polypeptide-presenting cells. The cytotoxicity is evaluated by counting the radioactivity due to the release of sxCr from the lysed target.
2624
Claims (25)
1. CLAIMS I. Diagnostic kit for a cancer secreting hCG or fragments of hCG comprising a set of polypeptides covering at least part of the primary sequence of hCG./ 5
2. Diagnostic kit according to Claim 1, comprising a set of polypeptides which covers the primary sequence of the /3 subunit of hCG (/3-hCG or /3-hCG-Ala 117) . v
3. Diagnostic kit according to Claim 1, comprising a set of polypeptides which covers at least part of the 10 primary sequence of the /3 subunit of hCG (/3-hCG or jS-hCG-Ala 117) . /
4. Diagnostic kit according to any one of Claims 1 to 3, comprising polypeptides having from 9 to 40 amino acids. j 15
5. Diagnostic kit according to any one of Claims 1 to 4, comprising polypeptides whose sequences overlap,
6. Use of a polypeptide corresponding to at least part of the primary sequence of hCG for the manufacture of a composition intended for the immunotherapy of 2 0 cancers secreting hCG or fragments of hCG. /
7. Use according Co Claim 6, in which the polypeptide corresponds to at least part of the primary sequence of the /3 subunit of hCG (/5-hCG or /5-hCG-Ala 117)./ 25 8.
Use according to Claim 6 or Claim 7, in which the polypeptide(s) have from 9 to 40 amino acids./ 9.
Use according to any one of Claims 6 to 8/ in which the polypeptide is coupled to a polylysine template .
J 3 0 10.
Use according to any one of Claims 6 to 9, comprising in addition an immunity adjuvant agent.y II.
Use according to any one of Claims 6 to 10, in which the polypeptide is protected at the NH2- and COOH-terminal ends. / 35 12.
Composition intended for the immunotherapy of cancers secreting hCG or fragments of hCG comprising an effective dose of a polypeptide corresponding to at least part of the primary sequence of hCG and an effective dose AMENDED SHEET -ii- 262 435 in identifying, in the circulating blood of the patient with the aid of a set of polypeptides covering at least part of the primary sequence of hCG, a state of immunization in against one of these polypeptides. 5
14. Method according to Claim 13, in which the state of immunization in relation to one of the polypeptides is identified by a test of stimulation of the T lymphocytes by the polypeptides.
15. Method according to Claim 13, in which the pos-10 sible presence of serum antibodies specific for hCG or for its fragments is detected.
16. Method according to Claim 13, in which the state of immunization in relation to one of the polypeptides is identified by a test of cytotoxicity of the T lymphocytes 15 against the polypeptides.
17. Method according to claim 13, in which the state of immunisation against one of the polypeptides is identified by a test of cytotoxicity of the T lymphocytes in relation to the polypeptides. 20
18. A diagnostic kit as claimed in any one of claims 1-5, substantially as herein described and with reference to any one of the figures. 25
19. A use of a polypeptide, as claimed in any one of claims 6-11, substantially as herein described.
20. A composition as claimed in claim 12, substantially as herein described. 30
21. A method of in vitro diagnosis, as claimed in any one of claims 13-17, substantially as herein described. 35 262435 - 12
22. Method according to any one of Claims p. to 20, in which the possible presence of serum antibodies specific for hCG or for its fragments is detected.
23. Method according to Claim 13, in which the immune 5 response in relation to one of the polypeptides is identified by a test of cytotoxicity of the T lymphocytes in relation to the polypeptides.
24. Method according to any one of Claims 17 to 23, in which an immunity adjuvant agent ie administered with 10 the polypeptide .^/
25. Method according to any one of Claims 17 to 24, in which BCG is administered concomitantly with the administration of the polypeptide or as a mixture with the polypeptide. ^ AMENDED SHEET
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| FR9302820A FR2702494B1 (en) | 1993-03-11 | 1993-03-11 | Diagnostic kit for cancer secreting hCG or fragments of hCG and vaccine intended for immunotherapy of such cancer. |
| FR9312039A FR2710845B1 (en) | 1993-10-08 | 1993-10-08 | Composition intended for the immunotherapy of a cancer secreting hCG or fragments of hCG. |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| NZ262435A true NZ262435A (en) | 1997-05-26 |
Family
ID=26230164
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| NZ262435A NZ262435A (en) | 1993-03-11 | 1994-03-10 | Kit for diagnosing an hcg fragment secreting cancer and immunotherapeutical means therefor |
Country Status (6)
| Country | Link |
|---|---|
| EP (1) | EP0688432A1 (en) |
| JP (1) | JPH08507604A (en) |
| AU (1) | AU6211294A (en) |
| CA (1) | CA2157868A1 (en) |
| NZ (1) | NZ262435A (en) |
| WO (1) | WO1994020859A1 (en) |
Families Citing this family (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5677275A (en) * | 1994-08-05 | 1997-10-14 | The United States Of America As Represented By The Department Of Health And Human Services | Treatment of cancer with human chorionic gonadotropin |
| FR2745638A1 (en) * | 1996-03-04 | 1997-09-05 | Roussy Inst Gustave | METHOD FOR DETECTION OF MALIGNANT TRANSFORMATION OF HUMAN CELLS |
| US5997871A (en) * | 1996-06-24 | 1999-12-07 | University Of Maryland Biotechnology Insitute | Treatment and prevention of cancer by administration of derivatives of human chorionic gonadotropin |
| US6319504B1 (en) | 1996-06-24 | 2001-11-20 | University Of Maryland Biotechnology Institute | Treatment and prevention of HIV infection by administration of derivatives of human chorionic gonadotropin |
| US5968513A (en) * | 1996-06-24 | 1999-10-19 | University Of Maryland Biotechnology Institute | Method of promoting hematopoiesis using derivatives of human chorionic gonadotropin |
| US6583109B1 (en) | 1997-06-24 | 2003-06-24 | Robert C. Gallo | Therapeutic polypeptides from β-hCG and derivatives |
| US7994278B1 (en) * | 1999-08-06 | 2011-08-09 | Nobel Biosciences Llc | Biologically active polypeptides derived from a novel early stage pregnancy factor designated maternin (MA) |
Family Cites Families (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4201770A (en) * | 1973-05-07 | 1980-05-06 | The Ohio State University | Antigenic modification of polypeptides |
| US4234561A (en) * | 1978-02-06 | 1980-11-18 | Research Corporation | Antigen for early pregnancy test and contraceptive vaccine |
| US4713366A (en) * | 1985-12-04 | 1987-12-15 | The Ohio State University Research Foundation | Antigenic modification of polypeptides |
| FR2623715B1 (en) * | 1987-11-26 | 1990-12-21 | Lafon Labor | PEPTIDE STRUCTURES, IMMUNOGENS CONTAINING THEM AND THEIR APPLICATIONS IN FERTILITY CONTROL |
| GB8821507D0 (en) * | 1988-09-14 | 1988-10-12 | Cancer Res Campaign Tech | Improvements relating to peptides |
| JPH06502494A (en) * | 1990-10-17 | 1994-03-17 | コール，ローレンス エイ | How to detect and follow the course of cancer, pregnancy and trophic cell diseases |
-
1994
- 1994-03-10 CA CA002157868A patent/CA2157868A1/en not_active Abandoned
- 1994-03-10 JP JP6519675A patent/JPH08507604A/en active Pending
- 1994-03-10 EP EP94909172A patent/EP0688432A1/en not_active Withdrawn
- 1994-03-10 AU AU62112/94A patent/AU6211294A/en not_active Abandoned
- 1994-03-10 NZ NZ262435A patent/NZ262435A/en unknown
- 1994-03-10 WO PCT/FR1994/000265 patent/WO1994020859A1/en not_active Application Discontinuation
Also Published As
| Publication number | Publication date |
|---|---|
| CA2157868A1 (en) | 1994-09-15 |
| AU6211294A (en) | 1994-09-26 |
| WO1994020859A1 (en) | 1994-09-15 |
| EP0688432A1 (en) | 1995-12-27 |
| JPH08507604A (en) | 1996-08-13 |
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