# US 4,201,770 A · Antigenic modification of polypeptides

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- Number: US 4,201,770 A
- Document ID: US4201770A
- Status: Granted
- Kind: A
- Application number: 05/936,876
- Filed: 1978-08-25
- Granted: 1980-05-06
- Earliest claimed: 1973-05-07
- Assignees: The Ohio State University
- Inventors: Vernon C. Stevens
- Web page: https://patentgrep.com/patent/US4201770A

## CPC

- C07K 14/59: Follicle-stimulating hormone [FSH]; Chorionic gonadotropins, e.g.hCG [human chorionic gonadotropin]; Luteinising hormone [LH]; Thyroid-stimulating hormone [TSH]
- A61K 39/0006: {Contraceptive vaccins; Vaccines against sex hormones}
- A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00)
- A61K 2039/5555: {Muramyl dipeptides}
- A61K 2039/6012: {Haptens, e.g. di- or trinitrophenyl (DNP, TNP)}
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)

## Abstract

Modified hormones or fragments of hormones are useful in producing antibodies when administered to an animal. Said antibodies in turn cause neutralization of endogenous natural protein hormones. The modification may be accomplished by attaching various kinds of modifying groups to the hormone or fragment. Modification may, for example, be achieved by chemically coupling diazosulfanilic acid groups to the hormone or fragment. The protein hormones to which this procedure can be applied are mammalian protein reproductive hormones such as, for example, Follicle Stimulating Hormone (FSH), or Human Chorionic Gonadotropin (HGG). These modified hormone or fragment may be administered to animals for the purpose of contraception, abortion, or treatment of hormone related disease states and disorders.

## Claims

### Claim 1

A method for controlling fertility in primate animals having naturally occurring endogenous Chroionic Gonadotropin hormone by neutralizing the biological action of the endogenous hormone, comprising the steps of:
administering to said primate animal an immunologically effective amount of the hormone, a subunit thereof, a peptide fragment of the subunit or a synthetically derived peptide having a sequence analogous to at least a portion of said subunit;
said hormone, subunit, fragment or synthetically derived peptide being modified by the coupling thereof with a non-endogenous material to effect the formation, following said administration, of antibodies having a specificity to endogenous Chorionic Gonadotropin;
thereby inhibiting the fertility of said primate animal by preventing one or more normal biological functions of the endogenous Chorionic Gonadotropin hormone.

### Claim 2 (depends on 1)

The method of claim 1 wherein said modified hormone, subunit, fragment or synthetically derived peptide is administered in combination with an adjuvant.

### Claim 3

The method of controlling fertility in primate animals having naturally occuring endogenous Chorionic Gonadotropin hormone comprising the steps of:
providing a quantity of the hormone, a subunit thereof, a peptide fragment thereof, or a synthetically derived peptide having a sequence analogous to at least a portion of said subunit and being non-antigenic within said primate animals;
modifying said hormone, subunit, fragment, or synthetically derived peptide by the coupling thereof with a non-endogenous material;
administering to said primate animal an immunologically effective amount of said modified hormone, subunit, fragment, or synthetically derived peptide;
thereby inhibiting the fertility of said primate animals by preventing one or more normal biological functions of endogenous Chorionic Gonadotropin.

### Claim 4 (depends on 3)

The method of claim 3 wherein said modified hormone, subunit, fragment or synthetically derived peptide is administered in combination with an adjuvant.

### Claim 5 (depends on 3)

The method of claim 3 wherein said non-endogenous material is coupled with said hormone, subunit, fragment or synthetically derived peptide through a sulfhydryl-group linkage.

### Claim 6 (depends on 3)

The method of claim 3 wherein said hormone, subunit, fragment or synthetically derived peptide is one having a sulfhydryl group therewithin, and said coupling is carried out by the treatment thereof with an activator of the structure ##STR10## wherein X represents a non-reacting connective entity so as to effect reaction of the maleiimide group of the activator with a said sulfhydryl group; and
treating the resultant activated hormone, subunit, fragment or synthetically derived peptide with said non-endogenous material.

### Claim 7 (depends on 6)

The method of claim 6 wherein said non-reacting connective entity, X, comprises an amino acid chain.

### Claim 8 (depends on 7)

The method of claim 7 wherein said amino acid chain is a Prolene spacer chain.

### Claim 9 (depends on 3)

The method of claim 3 wherein said non-endogenous material comprises sucrose copolymerized with epichlorohydrin.

### Claim 10 (depends on 3)

The method of claim 3 wherein said non-endogenous material comprises (poly)tyrosine, (poly)alanine, (poly)dextran, thyroglobulin, or a combination thereof.

### Claim 11 (depends on 6)

The method of claim 6 wherein said non-endogenous material comprises the protein, flagellin.

### Claim 12 (depends on 6)

The method of claim 6 wherein said non-endogenous material material comprises an influenza virus.

### Claim 13 (depends on 12)

The method of claim 12 in which said influenza virus is an influenza subunit incorporating substantially only the viral proteins Haemagglutin and Neuraminease.

### Claim 14 (depends on 6)

The method of claim 6 wherein said non-endogenous material is diptheria toxiod.

### Claim 15 (depends on 6)

The method of claim 6 wherein said non-endogenous material is a cholera organism.

### Claim 16 (depends on 6)

The method of claim 6 wherein said activated hormone, subunit, fragment or synthetically derived peptide is treated with said non-endogenous material at slightly alkaline pH.

### Claim 17 (depends on 3, 6)

The method of claim 3 or 6 wherein said fragment which is modified is of the chemical configuration:
Thr-Cys-Asp-Asp-Pro-Arg-Phe-Gln-Asp-Ser-Ser-Ser-Ser-Lys-Ala-Pro-Pro-Pro-Ser -Leu-Pro-Ser-Pro-Ser-Arg-Leu-Pro-Gly-Pro-Ser-Asp-Thr-Pro-Ile-Leu-Pro-Gln;
Asp-Asp-Pro-Arg-Phe-Gln-Asp-Ser-Pro-Pro-Pro-Cys-Pro-Pro-Pro-Ser-Asp-Thr-Pro -Ile-Leu-Pro-Gln;
Asp-Asp-Pro-Arg-Phe-Gln-Asp-Ser-Pro-Pro-Pro-Pro-Pro-Pro-Cys-Pro-Pro-Pro-Pro -Pro-Pro-Ser-Asp-Thr-Pro-Ile-Leu-Pro-Gln;
Asp-His-Pro-Leu-Thr-Aba-Asp-Asp-Pro-Arg-Phe-Gln-Asp-Ser-Ser-Ser-Ser-Lys-Als -Pro-Pro-Pro-Ser-Leu-Pro-Ser-Pro-Ser-Arg-Leu-Pro-Gly-Pro-Ser-Asp-Thr-Pro-Il e-Leu-Pro-Gln-Pro-Pro-Pro-Pro-Pro-Pro-Cys;
Asp-His-Pro-Leu-Thr-Aba-Asp-Asp-Pro-Arg-Phe-Gln-Asp-Ser-Ser-Ser-Ser-Lys-Ala -Pro-Pro-Pro-Ser-Leu-Pro-Ser-Pro-Ser-Arg-Leu-Pro-Gly-Pro-Ser-Asp-Thr-Pro-Il e-Leu-Pro-Gln-Cys;
or
Cys-Pro-Pro-Pro-Pro-Pro-Pro-Asp-Asp-Pro-Arg-Phe-Gln-Asp-Ser-Ser-Ser-Ser-Lys -Ala-Pro-Pro-Pro-Ser-Leu-Pro-Ser-Pro-Ser-Arg-Leu-Pro-Gly-Pro-Ser-Asp-Thr-Pr o-Ile-Leu-Pro-Gln.

### Claim 18 (depends on 3)

The method of claim 3 wherein said fragment which is modified is of the chemical configuration:
Asp-Asp-Pro-Arg-Phe-Gln-Asp-Ser-Ser-Ser-Ser-Lys-Ala-Pro-Pro-Pro-Ser-Leu-Pro -Ser-Pro-Ser-Arg-Leu-Pro-Gly-Pro-Ser-Asp-Thr-Pro-Ile-Leu-Pro-Gln;
Gln-Asp-Ser-Ser-Ser-Ser-Lys-Ala-Pro-Pro-Pro-Ser-Leu-Pro-Ser-Pro-Ser-Arg-Leu -Pro-Gly-Pro-Ser-Asp-Thr-Pro-Ile-Leu-Pro-Gln,
Phe-Gln-Asp-Ser-Ser-Ser-Ser-Lys-Ala-Pro-Pro-Pro-Ser-Leu-Pro-Ser-Pro-Ser-Arg -Leu-Pro-Gly-Pro-Ser-Asp-Thr-Pro-Ile-Leu-Pro-Gln;
or
Asp-Asp-Pro-Arg-Phe-Gln-Asp-Ser-Ser-Ser-Ser-Lys-Als-Pro-Pro-Pro-Ser-Leu-Pro -Ser.

### Claim 19 (depends on 3)

The method of claim 3 wherein said non-endogenous material is one having an amino group and said coupling is carried out by activating said non-endogenous material with an activator of the structure: ##STR11## where X represents a non-reacting connective entity so as to effect reaction of the activator with said amino group; and
treating the activated non-endogenous material with said hormone, subunit, fragment or synthetically derived peptide which has a sulfhydryl group.

### Claim 20 (depends on 19)

The method of claim 19 wherein said activation is carried out under neutral or acid conditions.

### Claim 21 (depends on 3)

The method of claim 3 wherein said modification is carried out to effect the constitution of two or more immunological determinants effective to elicit antibody response to the endogenous hormone, Chorionic Gonadotropin.

### Claim 22 (depends on 3)

The method of claim 3 wherein:
said fragment or synthetically derived peptide which is modified is of the chemical configuration:
Cys-Pro-Pro-Pro-Pro-Pro-Pro-Ser-Asp-Thr-Pro-Ile-Leu-Pro-Gln
and said administration thereof is in combination with another, different modified Chorionic Gonadotropin hormine, subunit thereof, peptide fragment of the said subunit, or a synthetically derived peptide having a sequence at least analogous to at least a portion of the fragment, the said modification of which effects the constitution of at least one immunological determinant.

### Claim 23 (depends on 3)

The method of claim 3 wherein:
said fragment or synthetically derived peptide which is modified is of the chemical configuration:
Asp-Asp-Pro-Arg-Phe-Gln-Asp-Ser-Pro-Pro-Pro-Pro-Pro-Pro-Cys
and said administration thereof is in combination with another, different modified Chorionic Gonadotropin hormine, subunit thereof, peptide fragment of the said subunit, or a synthetically derived peptide having a sequence at least analogous to at least a portion of the fragment, the said modification of which effects the constitution of at least one immunological determinant.

### Claim 24 (depends on 1)

The method of claim 1 wherein said animal is human and said hormone is human Chorionic Gonadoptropin.