WO1997012038A1

WO1997012038A1 - Novel peptides useful in enhancement of fsh action

Info

Publication number: WO1997012038A1
Application number: PCT/GB1996/002365
Authority: WO
Inventors: Gabriele Bono; Luca Ferasin; James Beattie; Andrew Thomas Holder
Original assignee: Babraham Institute
Current assignee: Babraham Institute
Priority date: 1995-09-25
Filing date: 1996-09-25
Publication date: 1997-04-03
Anticipated expiration: 1998-03-25
Also published as: JP2000508293A; EP0858508A1; AU704010B2; AU7090096A; CA2232950A1; GB9519511D0

Abstract

Novel peptides useful in enhancing the activity of follicle stimulating hormone (FSH) are provided, together with their use in increasing fertility.

Description

NOVEL PEPTIDES USEFUL IN ENHANCEMENT OF FSH ACTION

The present invention relates to novel peptides useful in enhancement of follicle stimulating hormone (FSH) bioactivity in mammals. Particularly, the invention relates to such peptides and their use in stimulating FSH activity in, inter alia , humans, sheep, cattle, pigs, goats, cats, dogs, horses, camels, llamas and alpacas.

Follicle stimulating hormone (FSH) belongs to a family of glycoprotein hormones which includes lutenising hormone

(LH) , thyrotropin (TSH) and chorionic gonadotropin (CG) .

They are all composed of two different non-covalently bound subunits termed and jβ. Within a species the amino acid sequences of the ot subunits for these different hormones are identical. The hormone specific jβ subunits exhibit different amino acid sequences (for a review see Combarnous, Endocrine Reviews, 13:670-691 (1992)) .

FSH plays a key role in the control of folliculogenesis and spermatogenesis. The presence of FSH is fundamental to the recruitment of quiescent primary follicles and their development into antral follicles (for a review see Hillier, Human Reproduction, 9:188-191 (1994) ) . These biological effects are mediated by the binding of FSH to specific cellular receptors; such receptors bind to specific sites on the FSH molecule termed receptor recognition sites (Combarnous, supra) . Different animal models have been used to study the effects of FSH on follicular growth and ovulation. These include hypophysectomized mice (Wang and Greenwald,

Endocrinology 132: 2009-2016 (1993) ; Wang and Greenwald,

J. Reprod . Fert . 99: 403-413 (1993)) and hypogonadal mice

(Halpin & Charlton, J^". Reprod. Fert . 82: 393-400 (1988) ; Cattanach et al , Nature 269: 338-340 (1977) ; Mason et al, Science 234: 1366-1371 (1986)) . In particular, xn Snell and Ames dwarf mice, it has been observed that FSH supplementation can increase the total number of healthy follicles and uterine and ovarian weights in 24 day old animals (de Reviers, Acta . Endocrinologica . 106: 121-126 (1984) ) .

Animal models have also been used to look at the effects of antibodies on FSH bioactivity. Antibody action is normally considered in terms of neutralisation of biological activity. However, it has been found that a monoclonal antibody (MAb) can also enhance FSH activity

(Glencross et al, Journal of Endocrinology 136: R5-R7 (1993)) . In that study monoclonal antibodies prepared against bovine FSH (bFSH) were found to enhance the ability of bFSH to induce uterine growth in Snell dwarf mice. However, prolonged administration of murine MAbs to other species may have harmful consequences.

Thus, it would be preferable to generate site specific FSH enhancing antisera by administration of selected peptide sequences of FSH from the appropriate species. Given the commonality of one of the subunits with other glycoprotein hormones, administration of a complete hormone or antibodies raised against the complete hormone, may lead to some problems with respect to cross- reactivity effects on the bioactivity of other glycoprotein hormones. The aim, therefore, is to produce antibodies which are specific in that they stimulate the activity of FSH while having little or no effect on the bioactivity of other glycoprotein hormones which share some commonality of structure with FSH. The present inventors have now found a range of peptides which achieve this result. These peptides are comprised of amino acid residues 33-47 and 40-51 of the FSH molecule (see figure 1) . This in itself is surprising since this region has been implicated as a receptor recognition site. Thus, the binding of an antibody to this region of FSH should be incompatible with the binding of FSH to its receptor (Santa Coloma, et al, Biochemistry, 29:1194-1200 (1990) ; Santa Coloma and Reichert. J. Biol. Chem., 265:5037-5042 (1990) ; Lapthorn, et al , Nature, 369:455-461 (1994) ) .These peptides can be administered alone, or in combination, across a range of mammalian species to elicit an enhancement of FSH bioactivity.

Thus, in a first aspect, the present invention provides a peptide having the following sequence:

(i) YTRDLVYRDPARPNI (ii) YTRDLVYRDPARPNI

(iii) YTRDLVYKDPARPKI

(iv) RDPARPNIQKTC;

(v) RDPARPKIQRTC;

(vi) KDPARPNIQKAC; or (vii) RDPARPNIQKTC

or a sequence substantially homologous thereto.

The peptides of the invention are useful in eliciting antibody responses in mammals, the antibody causing an enhancement of either endogenous or exogenous FSH activity. This in turn leading to enhanced ovulation or spermatogenesis and hence fertility. The peptide or peptides can be provided as a vaccine used to stimulate an immune response which recognises FSH in the subject mammal. Such a vaccine forms a second aspect of the invention.

In a third aspect, the present invention provides a nucleic acid sequence coding for a peptide as described above, or one substantially homologous thereto. Such nucleic acid sequences can either be DNA sequences or RNA sequences. Such nucleic acid sequences include all other nucleic acid sequences which, by virtue of the degeneracy of the genetic code, also code for the given amino acid sequence, or which are substantially homologous to such a sequence.

In the context of the present invention, substantial homology at the amino acid level refers to peptide sequences having a significant number of constituent amino acids homologous to the peptide sequences described above. For example, at least 40%, 50%, 60%, 70%, 80%, 90%, 95% or even 99%, in increasing order of preference, of the amino acids may be homologous .

As an alternative, the peptides of the present invention can be used to generate antibodies, these antibodies then being administered to a mammal to elicit enhanced bioactivity of either endogenous or exogenous FSH. Thus, in a further aspect, the present invention provides an antibody having specificity for one or more of the peptides of the present invention. In particular, the antibody will be a monoclonal antibody.

In one embodiment of this aspect of the invention, the antibody is provided as an antibody preparation, comprising one or more antibodies reactive with one or more of the peptides of the present invention.

In preferred embodiments of the above-noted aspects of the invention Peptides (i) or (iv) will be used to enhance FSH activity in sheep, or to raise antibodies to enhance FSH activity in sheep; Peptide (i) will be used to enhance FSH activity in cattle or pigs, or to raise antibodies to enhance FSH activity in cattle or pigs: Peptide (i) will be used to enhance FSH activity in horses or humans, or to raise antibodies to enhance FSH activity in horses or humans; Peptide (ii) will be used to enhance FSH activity in sheep or pigs, or to raise antibodies to enhance FS_H activity in sheep or pigs; Peptide (ii) will be used to enhance FSH activity in horses, humans or cattle, or to raise antibodies to enhance FSH activity in horses, humans or cattle; Peptide (iii) will be used to enhance FSH activity in horses or humans, or to raise antibodies to enhance FSH activity in horses or humans; Peptide (iii) will be used to enhance

FSH activity in cattle, sheep or pigs, or to raise antibodies to enhance FSH activity in cattle, sheep or pigs; peptide (iv) will be used to enhance FSH activity in cattle, or to raise antibodies to enhance FSH activity in cattle; Peptide (iv) will be used to enhance FSH activity in pigs, horses or humans, or to raise antibodies to enhance FSH activity in pigs, horses or humans; Peptide (v) will be used to enhance FSH activity in horses or humans, or to raise antibodies to enhance FSH activity in horses or humans; Peptide (v) will be used to enhance FSH activity in cattle, sheep or pigs, or to raise antibodies to enhance FSH activity in cattle, sheep or pigs; Peptide (vi) will be used to enhance FSH activity in sheep, or to raise antibodies to enhance FSH activity in sheep; Peptide (vi) will be used to enhance FSH activity in cattle, pigs, humans or horses, or to raise antibodies to enhance FSH activity in cattle, pigs, humans or horses; Peptide (vii) will be used to enhance FSH activity in pigs, or to raise antibodies to enhance FSH activity in pigs; Peptide (vii) will be used to enhance FSH activity in cattle, sheep, humans or horses, or to raise antibodies to enhance FSH activity in cattle, sheep, humans or horses.

The peptides or antibodies of the invention may suitably be administered in the form of a pharmaceutically acceptable formulation. Thus, in a fifth aspect, the present invention provides a pharmaceutical formulation comprising one or more peptides of the present invention, or one or more antibodies or antibody preparations of the present invention, together with one or more pharmaceutically acceptable carriers, diluents or excipients.

In a sixth aspect, the present invention provides the use of a peptide or antibody of the present invention in enhancing FSH bioactivity in a mammal. In particular embodiments the bioactivity is enhanced in cattle, sheep, horses, pigs, humans, camels, goats, cats, dogs, llamas or alpacas.

In a seventh aspect, the present invention provides a method for enhancing FSH bioactivity in mammals, particularly cattle, sheep, horses, pigs, humans, camels, goats, cats, dogs, llamas or alpacas, which comprises the step of administering one or more peptides, or one or more antibodies, of the present invention to the mammal. The invention will now be described with reference to the following examples, which are not intended to be in any way limiting.

The examples refer to the figures in which:

FIGURE 1: is a comparison of the sequences of jβ subunits of FSH from various species;

FIGURE 2 : is a comparison between the titre of antibodies against whole bovine FSH (bFSH) in preimmune serum (bleed 0) and in antiserum after four administrations of peptide A (bleed 4) in sheep Al (2-a) and A2 (2-b) , and peptide B in sheep Bl (2- c) ;

FIGURE 3: shows the mean (± SEM) of uterine (3-a) and ovarian (3-b) weights and the keratinisation index (3-c) in groups of Snell dwarf mice given 3.3, 10, 30 and 90 μg/day of ovine FSH (oFSH) or phosphate buffered saline (hormone diluent, PBS) ;

FIGURE 4 : shows an estimation of antibody mediated enhancement of oFSH bioactivity by measurement of uterine (4-a) and ovarian (4-b) weights and the keratinisation index of vaginal smears (4-c) , in five different groups of Snell dwarf mice which received 20 μg/day oFSH (group FSH) , 20 μg/day oFSH + peptide A antiserum (group FSH + A) , 20 μg/day oFSH + preimmune serum (group FSH + p) , peptide A antiserum alone (group A) and PBS control (group C) ; all values are means ±SEM; and FIGURE 5: shows an estimation of antibody mediated enhancement of FSH oioactivity by measurement of uterine (5a) and ovarian (5b) weights and the keratinisation index of vaginal smears (5c) m five different groups of mice which received 20μg/day oFSH (group FSH) , 20μg/day oFSH+peptide B antiserum (group FSH+B) , 20ug/day oFSH+preimmune serum (group FSH+p) , Peptide B antiserum alone (group B) and PBS control group; all values are means ±SEM.

EXAMPLE 1

Preparation of Antiserum Against Peptide Sequences of bFSH

Peptides : different sequential peptides of FSH were synthesised by F_moc chemistry using an upgraded 431A automated peptide synthesiser (Applied Biosystems) . Peptides were synthesised with free amino and carboxytermmi . Peptide identity was confirmed by MALDI (matrix assisted laser desorption ionisation) mass spectroscopy and peptides were estimated at greater than 95% purity by analytical reverse phase chromatography.

KLH conj ugation : 20 ml of 10 mM sodium phosphate buffer pH 6.8 were added to a mixture composed by 5 mg of peptide and 5 mg of Reyhole Lympet Haemocyanin (KLH) ; this solution was stirred with the slow addition of 50 μl of 25% glutaraldehyde and the stirring was carried on for three hours. The solution was then aliquoted m 20 x 2 ml aliquots and stored at 20°C ready for use.

Animals and treatment ten mature female sheep were divided into five groups. Two sheep were immunised with eacn of peptides d) and (iv) (labelled A and B respectively in this study) , as well as the following peptides:

HADSLYTYPVAT (labelled C; bFSH sequence 69-80) ; HCSRCDSDSTDC (labelled D; bFSH sequence 83-94) ; and WCAGYCYTRDLVY (labelled E; bFSH sequence 27-39) .

Sheep were identified according to the peptide with which they were immunised (i.e. peptide A = sheep Al and A2) . A 2 ml aliquot of peptide/KLH conjugate, prepared as above, was diluted with 2 ml of complete Freund adjuvant and 1 ml was injected into each hindleg of each sheep. Three boosts were carried out at three week intervals with Freund incomplete adjuvant.

Radioimmunoassay: serum samples were taken from each sheep before the __first injection of peptide (preimmune) and ten days after each immunisation. Serial dilutions of each serum sample (1/10-1/6250) were tested for their ability to recognise iodinated bFSH in a liquid phase radioimmunoassay. In brief, 50 μl of serum was mixed with 100 μl of radioimmunoassay buffer (0.05 M phosphate buffer with 0.05% tween) plus 50 μl of iodinated bFSH (¹²⁵I; USDA bFSH-I-2 AFP 5318C; 20,000 CPM/tube, iodinated using iodogen by the method of Salacinski et al, Analyt . Biochem . 117: 136-146 (1981)) . Assay tubes were incubated at 4°C overnight and then 200 μl of 0.05 M Tris-HCl pH 8.5 containing 25% polyethylene glycol 6000 and 0.15% bovine gamma globulin were added. After a further incubation of lh at 4°C, tubes were centrifuged and the radioactivity in the pellet was counted using a gamma counter.

Measurement of FSH Bioactivity

Hormone : ovine-FSH (NIH-FSH-SI (Ovine R041212)) dissolved in PBS was used to measure the bioactivity of FSH .

Animals and trea tment : snell dwarf mice were bred, maintained and marked as previously described by Holder et al , J. Endocrinol . 85: 35-47 (1980) . Thirty 26 to 35 day old female dwarf mice were randomly allocated to five treatment groups, given 0, 3.3, 10, 30, 90 μg/day of oFSH using 0.05 ml of solution injected subcutaneously twice a day for five days. Fourteen hours after the last injection, the mice were sacrificed by decapitation under halothane/oxygen anaesthetic.

Ovarian and uterine weights : ovaries and uterus were removed, dissected free of fat under a stereo- microscope, fixed in formalin (3.7%) and weighed, after fixation, using a Cahn microbalance. Vaginal smears : vaginal cells were collected using a few drops of PBS introduced into the vagina via an ordinary pipette and then extracted by the pipette. The cells were transferred to a microscope slide, fixed in methanol and stained with Wright Giemsa. The smears were analysed under lOOx magnification and the keratinisation index (keratinised cells/total cells x 100) was calculated.

Statistical analysis : uterine and ovarian weights (expressed as mg/g body weight) and keratinisation index were used for the statistical analysis. Data from each group treated with hormone were compared with the PBS injected control group using Student's t_.-Test.

Enhancement of FSH Bioactivity Using Antiserum to Peptide A from Sheep Al

Hormone and purified antiserum: ovine FSH (NIH-FSH- SI) was dissolved in PBS. Preimmune serum or antiserum against peptide A was purified as follows: one volume of 27% sodium sulphate was mixed with 2 volumes of serum and incubated at 37°C for 3.5 h. The precipitated IG fraction was centrifuged and the pellet reconstituted to the volume of serum from which it was derived. This IG fraction was then extensively dialysed against PBS. Equal volumes of FSH and purified serum were combined to give a final concentration of 20 μg oFSH plus 50 μl of neat purified antiserum per 100 μl of injection solution. Where FSH was used alone, PBS was substituted for serum, and where antipeptide antisera alone was used, PBS was substituted for FSH. Control animals received PBS alone. Mice received two injections of 0.05 ml of the above solutions (as indicated) per day.

Animals and trea tment : dwarf mice were bred, maintained and marked as described previously. Thirty 31 to 40 days old female Snell dwarf mice were randomly allocated to five treatment groups:

Group 1 (FSH) : oFSH 20 μg/mouse/day dissolved in PBS;

Group 2 (oFSH + antiserum; FSH + A) : oFSH 20 μg plus purified peptide A antiserum prepared as above;

Group 3 (oFSH + preimmune serum; FSH + P) : oFSH 20 μg plus purified preimmune serum prepared as above;

Group 4 (antiserum alone; A) : purified antipeptide A antiserum prepared as above plus an equal volume of PBS:

Group 5 (control; PBS) : received PBS alone.

Each animal was injected and sacrificed with the same protocol as the previous experiment . Uterine and ovarian weights (expressed as mg/g body weight) plus the index of keratinisation were measured as described above. Differences between groups were assessed by Student's t_.-Test.

Enhancement of FSH Bioactivity Using Antiserum to Peptide B

Animals and trea tment : twenty-five 31-40 day old female Snell dwarf mice were randomly allocated to five treatment groups. Mice were bred and maintained as above.

Hormone and purified antiserum: hormone and sera were prepared and purified as described in the previous section.

Group 1 (FSH) : oFSH 20 μg/mouse/day dissolved in PBS;

Group 2 (oFSH + antiserum; FSH + B) : oFSH 20 μg plus purified peptide B antiserum;

Group 3 (oFSH + preimmune serum; FSH + P) : oFSH 20 μg plus purified preimmune serum;

Group 4 (antiserum alone; B) : purified antipeptide B antiserum alone:

Group 5 (control) : received PBS alone.

Each animal was injected and sacrificed with the same protocol as the previous experiment .

Uterine and ovarian weights (expressed as mg/g body weight) plus the index of keratinisation in vaginal smears were measured as described above. Differences between groups were assessed using Student's t.-Test.

RESULTS

Production of Antiserum Against Peptides A, B. C. D and

E

Figures 2a, b and c show the comparison between the titre of antibodies against bFSH in preimmune serum (bleed 0) , and in antiserum after four injections (bleed 4), in the two sheep injected with peptide A (Al and A2) and in one of the two sheep immunised against peptide B (Bl) . The second animal (B2) died before the end of the present trial. Of the two sheep injected with peptide A, sheep

Al had the highest titre of antiserum which recognised bFSH. Antisera from this sheep (Al) and from sheep Bl

(injected with peptide B) were investigated for their ability to enhance the biological actions of oFSH in hypopituitary Snell dwarf mice in vivo . Negligible titres of antisera were observed in animals immunised with peptides C, D or E.

Dose-response Curve of oFSH Bioactivity

Figure 3 illustrates the effects of various doses of oFSH alone, on uterine and ovarian weight (expressed as mg/g body weight; figs 3a and 3b) and on the index of keratinisation in vaginal smears (% keratinised cells; fig 3c) . Treatment with oFSH resulted in a significant dose-dependent increase in uterine weight (10 μg/day = 0.88 ± 0.19: P<0.05; 30 μg/day = 2.04 ± 0.34 m: P<0.01; 90 μg/day = 3.11 ± 0.18, P<0.001; significance values are versus the PBS control group 0.34 ± 0.05) . A similar dose dependent increase in ovarian weight was also observed, although ovarian weight was only significantly greater than that in the control group (0.36 ± 0.05) for the two highest doses of oFSH (30 μg/day = 0.60 ± 0.09, P<0.05; 90 μg/day = 1.18 ± 0.14, P<0.01) . The index of keratinisation showed a simlar pattern of dose dependency

(10 μg/day = 13.48 ± 2.95, P<0.05; 30 μg/day = 31.85 ±

6.12, P<0.05) ; 90 μg/day = 59.89 ± 5.23, P_<0.001; significance values are versus the PBS control group 2.40 ± 1.56) .

Enhancement of FSH Bioactivity bv Antiserum to Peptide A

Figure 4 illustrates the ability of antiserum to peptide A to enhance FSH activity, as determined by increased uterine (Fig 4a) and ovarian weight (Fig 4b) and an increased index of keratinisation in vaginal smears (fig 4c) in Snell dwarf mice.

For uterine weight (Fig 4a) , the response to FSH plus antiserum to peptide A (FSH + A) was significantly greater than that observed for either FSH alone (FSH; P<0.001) or FSH plus preimmune serum (FSH + P; P<0.01) . There was no significant difference between the response to FSH alone and to FSH plus preimmune serum. However, both responses were significantly greater than that observed for either antiserum to peptide A alone (A) or PBS injected controls (P<0.05 in all cases) .

For ovarian weight (fig 4B) a similar picture was obtained. FSH plus antiserum to peptide A (FSH + A) promoted a significantly greater increase in ovarian weight than that observed for either FSH alone (FSH; P<0.001) or FSH plus preimmune serum (FSH + P; P<0.01) . There was no significant difference between the response to FSH alone and FSH plus preimmune serum; however, both responses were significantly greater than those observed for either antisera to peptide A alone (A) or PBS injected controls (P<0.05 in all cases) .

Again, a similar picture was obtained for the keratinisation index of vaginal smears from these animals (fig 4c) . The response to FSH plus antiserum to peptide A (FSH +A) was significantly greater than that observed for either FSH alone (FSH; P<0.001) or FSH plus preimmune serum (FSH + P; P<0.001) . There was no significant differences between the response to FSH alone and FSH plus preimmune serum. fowever both responses were significantly greater than that observed for either antisera to peptide_ A alone (A) or PBS injected controls (P<0.05 in all cases) .

All of this data is summarised in Fig 4 plus associated Tables.

Enhancement of FSH Bioactivity by Antiserum to Peptide B

Figure 5 illustrates the ability of antiserum to peptide A to enhance FSH activity, as determined by increased uterine (Fig 5a) and ovarian weight (Fig 5b) and an increased index of keratinisation in vaginal smears (fig 3c) in Snell dwarf mice.

For uterine weight (Fig 5a) , the response to FSH plus antiserum to peptide B (FSH - B) was significantly greater than that observed for either FSH alone (FSH; P<0.01) or FSH plus preimmune serum (FSH + P; P<0.01) . There was no significant difference between the response to FSH alone and to FSH plus preimmune serum. However, both responses were significantly greater than that observed for either antiserum to peptide B alone (B) or PBS injected controls (P<0.01 in all cases) .

For ovarian weight (fig 5B) a similar picture was obtained. FSH plus antiserum to peptide B (FSH ₊ B) promoted a significantly greater increase in ovarian weight than that observed for either FSH alone (FSH; P<0.01) or FSH plus preimmune serum (FSH + P; P<0.01) . There was no significant difference between the response to FSH alone and FSH plus preimmune serum. The response to FSH alone or FSH plus preimmune serum was significantly greater than those observed for PBS injected controls (P<0.05 in all cases) .

Again, a similar picture was obtained for the keratinisation index of vaginal smears from these animals (fig 5c) . The response to FSH plus antiserum to peptide B (FSH + B) was significantly greater than that observed for either FSH alone (FSH; P<0.001) or FSH plus preimmune serum (FSH + P; P<0.001) . There was no significant differences between the response to FSH alone and FSH plus preimmune serum. The response to FSH alone was significantly greater than that observed for PBS injected controls (P<0.05) .

All of this data is summarised in Fig 5 plus associated Tables .

DISCUSSION

Immunisation of sheep with peptides A or B resulted in antisera capable of recognising bFSH, whereas peptides C, D and E did not . None of the sheep appeared to suffer any detrimental physiological effects due to peptide immunisation. The results clearly show that treatment of hypopituitary Snell dwarf mice with FSH mixed with antisera to either peptide A or peptide B induces a dramatic enhancement of FSH bioactivity as determined by increased uterine and ovarian weight and the keratinisation index of vaginal smears

The use of peptide vaccines to produce FSH enhancing antisera nas a number of advantages over alternative methods of controlling follicular development in commercially important species. Some of these advantages have been listed below.

1. This approach requires only transitory exposure of the animal to minute amounts of an antigen in order to produce relatively long lasting effects. Since antibodies persist in the circulation, it is not necessary accurately to predict surges in FSH production.

2. FSH enhancing antisera can be produced m a subject animal by active immunisation; alternatively antibody preparation from one animal may be administered to another animal of the same species by passive immunisation.

3. Using the known sequences of FSH from a variety of species, specific peptide vaccines can easily be designed to suit a variety of commercially important animals .

4 This technique can be used to enhance endogenous FSH activity; under these circumstances, circulating levels of FSH remain under pituitary control, thus enhancement only occurs when FSH is produced. This will serve to avoid hyperstimulation of the ovaries which is often caused by current superovulation treatments .

5. Both of the active component, namely antisera and FSH, are produced by the animal.

Claims

CLAIMS :

1. A peptide having the sequence:

(i) YTRDLVYRDPARPNI

(ii) YTRDLVYRDPARPNI ,

(iii) YTRD VYKDPARPRI

(iv) RDPARPNIQKTC;

(v) RDPARPRIQRTC; (vi) KDPARPNIQKAC; or

(vii) RDPARPNIQKTC or a sequence substantially homologous thereto.

2. A vaccine comprising one or more peptides as defined in claim 1.

3. A vaccine as claimed in claim 2 which is for administration to cattle, sheep, pigs, horses, humans, goats, camels, cats, dogs, llamas or alpacas.

4. A vaccine as claimed in claim 3 which is for administration to humans.

5. A nucleic acid sequence coding for a peptide as defined in claim 1 or a nucleic acid sequence substantially homologous thereto.

6. A nucleic acid sequence as claimed in claim 5 which is a DNA sequence.

7. An antibody having specificity for one or more of the peptides defined in claim 1.

8. An antibody as claimed in claim 7 which is a monoclonal antibody.

9. An antibody preparation comprising antibodies as defined in claim 7 or claim 8.

10. A peptide as defined in claim 1 for use in enhancing FSH activity in mammals.

11. A peptide as claimed in claim 10 wherein the mammals are sheep, cattle, horses, pigs, humans, camels, goats, cats, dogs, llamas or alpacas.

12. A peptide as claimed in claim 11 wherein the mammal is a human.

13. Peptide (i) , as defined in claim 1, for use in enhancing FSH activity in sheep.

14. Peptide (iv) , as defined in claim 1, for use in enhancing FSH activity in sheep.

15. Peptide (i) , as defined in claim 1, for use in enhancing FSH activity in cattle or pigs.

16. Peptide (i) , as defined in claim 1, for use in enhancing FSH activity in horses or humans.

17. Peptide (ii) , as defined in claim 1, for use in enhancing FSH activity in sheep or pigs.

18. Peptide (ii) , as defined in claim 1, for use in enhancing FSH activity in horses, humans or cattle.

19. Peptide (iii) , as defined in claim 1, for use in enhancing FSH activity in horses or humans.

20. Peptide (iii) , as defined in claim 1, for use in enhancing FSH activity in cattle, sheep or pigs.

21. Peptide (iv) , as defined in claim 1, for use in enhancing FSH activity in cattle.

22. Peptide (iv) , as defined in claim 1, for use in enhancing FSH activity in pigs, horses or humans.

23. Peptide (v) , as defined in claim 1, for use in enhancing FSH activity in horses or humans.

24. Peptide (v) , as defined in claim 1, for use in enhancing FSH activity in cattle, sheep or pigs.

25. Peptide (vi) , as defined in claim 1, for use in enhancing FSH activity in sheep.

26. Peptide (vi) , as defined in claim 1, for use in enhancing FSH activity in cattle, pigs, humans or horses.

27. Peptide (vii) , as defined in claim 1, for use in enhancing FSH activity in pigs.

28. Peptide (vii) , as defined in claim 1, for use in enhancing FSH activity in cattle, sheep, humans or horses .

29. A pharmaceutical formulation comprising at least one peptide as defined in claim 1, together with one or more pharmaceutically acceptable carriers, excipients or diluents .

30. A method for enhancing FSH activity in mammals comprising administering to the mammals a peptide as defined in any one of claims 1 or 10 to 28, an antibody as defined in claim 7 or claim 8, an antibody preparation as claimed in claim 9, or a pharmaceutical formulation as claimed in claim 29.

31. A method for increasing fertility in mammals comprising administering to the mammals a peptide as defined in any one of claims 1 or 10 to 28, an antibody as defined in claim 7 or claim 8, an antibody preparation as claimed in claim 9, or a pharmaceutical formulation as claimed in claim 29.

32. A method as claimed in claim 30 or claim 31 wherein the mammals are sheep, cattle, horses, pigs, humans, camels, goats, cats, dogs, llamas or alpacas.

33. A method as claimed in claim 32 wherein the mammals are humans.

34. The use of a peptide as defined in claim 1, or in any one of claims 10 to 28, an antibody as defined in claim 7 or claim 8, or an antibody preparation as defined in claim 9, in the preparation of a medicament to enhance FSH activity.