Abstract:
Intra-vaginal delivery procedures including compositions and units therefor whereby an effective releasable amount of oestradiol 17β is released to achieve an efficacious effect insofar as oestrus expression is concerned. Preferably a cyclodextrin is utilized to enhance absorption.

Description:
TECHNICAL BACKGROUND  
         [0001]    The present invention relates to intra-vaginal delivery or dosage units, compositions suitable therefor, the use thereof and related means and methods.  
           [0002]    In our New Zealand patent specification numbers 207341, 286492 (PCT/NZ97/00052) and 314572/314175 (PCT/NZ98/00011) there are disclosed procedures applicable in a range of different animals insofar as the synchronisation of the onset of oestrus is concerned.  
           [0003]    In our EAZI-BREED™, CIDR™ “Controlled Breeding and Reproductive Management” publication there is disclosed the use of an oestradiol co-treatment (ie; CIDIROL™) with the use of our progesterone releasing CIDR™ intra-vaginal inserts in cattle for treatment of anoestrus or for oestrus synchrony. Current practice when selecting an oestradiol co-treatment, for use with a progesterone releasing intra-vaginal insert (such as our CIDR-B™ device), is to select oestradiol benzoate by intramuscular (i.m.) injection (eg; CIDIROL™).  
           [0004]    In the past attempts at vaginal administration of oestradiol benzoate by using a capsule has resulted in poor oestrus expression and poor fertility. This outcome has been notwithstanding doses typically 10 times higher than those usually administered by intramuscular injection.  
         BACKGROUND ART  
         [0005]    Our invention is directed towards a means whereby delivery of an active agent such as that to serve the role of previously used oestradiol benzoate is improved using and/or despite using a vaginal administration procedure.  
           [0006]    It is therefore an object of the present invention to provide intra-vaginal delivery systems, dosage units, compositions, methods of use thereof and related means and methods which will be useful (preferably in conjunction with a progesterone releasing intra-vaginal insert such as our CIDR-B™ range of intra-vaginal inserts) in animals.  
           [0007]    We have determined in cattle that the lack of performance when using the vaginal administration of oestradiol benzoate is not attributed to the dose, as typically doses are 10 times higher than those administered by intra-muscular injection, but to poor and variable absorption of oestradiol benzoate following vaginal administration. Typically peak plasma oestradiol concentrations following vaginal administration of 10 mg oestradiol benzoate range from 2 to 5 pg/ml. The peak in plasma levels is obtained between 2 and 48 hours following administration. In comparison peak plasma oestradiol concentrations following i.m. injection of 1 mg oestradiol benzoate range from 8 to 13 pg/ml. The peak in plasma level is obtained by approximately 2 hours following administration and maintained for up to 24 hours following treatment.  
           [0008]    We believe similar effects occur in other animal species, viz, buffalo, pig, goat, sheep and deer.  
           [0009]    As a result of our research in cattle we have determined that the natural oestradiol 17β rather than the synthetic analogue oestradiol benzoate can be effectively administered vaginally despite the fact that if given intramuscularly oestradiol 17β was known to be shorter acting than the intramuscularly efficacious oestradiol benzoate analogue thereof.  
           [0010]    Our research in cattle has established that irrespective of whether or not that it is oestradiol 17β or an analogue thereof (such as oestradiol benzoate) that is delivered intra-vaginally an efficacious delivery thereof is possible in conjunction with an appropriate agent.  
           [0011]    Our research has also established in cattle that intra-vaginal delivery of oestradiol 17β in preference to its analogues or the delivery of oestradiol 17β and/or its analogues in conjunction with at least one cyclodextrin can maintain serum levels of the active metabolite above normal for at least 24 hours.  
           [0012]    We believe each, any or all of these findings are also appropriate for other animal species, viz, buffalo, pig, goat, sheep and deer, where such a regime might from time to time be warranted.  
         DISCLOSURE OF THE INVENTION  
         [0013]    Accordingly in a first aspect the present invention consists in an intra-vaginal composition for intra vaginal administration into a mammal, said composition being or having, as a solids admixture, a active principle (eg; a steroid such as oestradiol benzoate, oestradiol 17β or a prodrug of either) in admixture with γ-cyclodextrin and/or hydroxypropyl β-cyclodextrin.  
           [0014]    Preferably said composition is in the form of a tablet or as part of a capsule.  
           [0015]    Preferably said active principle is oestradiol 17β.  
           [0016]    Preferably said active principle is to achieve, in a target mammal, an efficacious effect insofar as oestrus expression is concerned or an efficacious effect insofar as oestrus synchronisation is concerned.  
           [0017]    Preferably said composition is provided as a dosage unit for a target mammal where the active principle is oestradiol 17β in an amount of between 0.72 and 7.2 mg.  
           [0018]    Preferably the active principle is oestradiol 17β and there is between 0.5 to 1.5 moles of γ-cyclodextrin and/or hydroxypropyl β-cyclodextrin per mole of oestradiol 17β.  
           [0019]    In another aspect the invention consists in the use of a composition of the present invention wherein, after an insertion of said composition as an intra-vaginal dosage unit in a target mammal, the plasma oestradiol concentration in the mammal 2 hours following the intra-vaginal administration is greater than 5 pg/ml and 24 hours following the intra-vaginal administration is less than 5 pg/ml.  
           [0020]    In yet another aspect the invention consists in, in a mammalian herd oestrus synchrony procedure which involves the insertion into and the subsequent withdrawal from each mammal of the herd of an intra vaginal device adapted to deliver progesterone, the use of an oestradiol co-treatment which involves a timely intra vaginal insertion into each such mammal of an intra vaginal dosage form having an active principal selected from the group consisting of (i) oestradiol benzoate, (ii) oestradiol 17β, (iii) prodrugs of (i) and, (iv) prodrugs of (ii) in admixture with a cyclodextrin selected from the group consisting of γ-cyclodextrin and hydroxypropyl β-cyclodextrin.  
           [0021]    In yet another aspect the invention consists in, in a mammalian oestrus expression procedure which involves the insertion into and the subsequent withdrawal from each mammal of the herd of an intra vaginal device adapted to deliver progesterone, the use of an oestradiol co-treatment which involves a timely intra vaginal insertion into each such mammal of an intra vaginal dosage form having an active principal selected from the group consisting of (i) oestradiol benzoate, (ii) oestradiol 17β, (iii) prodrugs of (i) and, (iv) prodrugs of (ii) in admixture with a cyclodextrin selected from the group consisting of γ-cyclodextrin and hydroxypropyl β-cyclodextrin.  
           [0022]    Preferably the active principal is oestradiol 17β.  
           [0023]    Preferably there are between 0.5 to 1.5 moles of γ-cyclodextrin and/or hydroxypropyl β-cyclodextrin per mole of oestradiol 17β.  
           [0024]    Preferably the oestradiol co-treatment is also a use.  
           [0025]    Preferably said oestradiol co-treatment intra vaginal dosage form is a tablet or capsule.  
           [0026]    Preferably each such tablet or capsule does not require active removal prior to, during or subsequent to the removal of said intra vaginal device adapted to deliver progesterone.  
           [0027]    In a further aspect the invention consists in the use of γ-cyclodextrin and/or hydroxypropyl β-cyclodextrin as absorption enhancers in the preparation of a pharmaceutical composition for the intra-vaginal administration of at least one active principle, said composition being a composition as aforesaid.  
           [0028]    Preferably said formula dosage unit is in the form of a capsule, tablet or similar product and may, for example, be associated with a delayed release mechanism of some intra-vaginal device adapted to release some preceding medicament.  
           [0029]    In another aspect the invention is a tablet or capsule (preferably capable in a target species recipient mammal of providing a plasma oestradiol concentration in the mammal two hours following intra vaginal administration of the table or capsule that is greater than 5 pg/ml and which 24 hours following the intra vaginal administration of the tablet or capsule will be less than 5 pg/ml) said tablet or capsule being formed of or having from 0.72 to 7.2 mg of oestradiol 17β in admixture with a cyclodextrin selected from one or both of γ-cyclodextrin and/or hydroxypropyl β-cyclodextrin, the mole ratio of the cyclodextrin(s) to oestradiol ranging from 0.5:1 to 1.5:1.  
           [0030]    In another aspect the invention is an intra vaginal table or capsule formed or having an analogue of oestradiol 17β in an amount equivalent to 0.72 to 7.2 mg of oestradiol 17β and from 6 to 150 mg cyclodextrin(s).  
           [0031]    Preferably the ratio of agent (eg; cyclodextrin) to enhance absorption to active material or pro-drug is less than 3:2 (agent:active) by molecular amount, that is 3 moles of cyclodextrin to every 2 moles of active.  
           [0032]    Preferably the intra-vaginal dosage unit has from 1.2 to 7.2 mg of oestradiol 17β [or an analogue equivalent amount, eg; from 10 to 30 mg if analogue is oestradiol benzoate] and from 6 to 150 mg cyclodextrin(s),  
           [0033]    and optionally other excipients, solid or liquid,  
           [0034]    and, preferably, if a capsule, is encased in a material such as gelatin that will release the capsule contents into vaginal fluids.  
           [0035]    A suitable source of cyclodextrins are the products BETA W7 HP hydroxypropyl γ-cyclodextrin, BETA W7 β-cyclodextrin and GAMMA W8 γ-cyclodextrin from Wacker Chemicals Australia, Victoria, Australia.  
           [0036]    A suitable source of oestradiol benzoate is from ICN Biomedical, Ohio, USA.  
           [0037]    A suitable source of oestradiol 17β is from Sigma Chemical Company, USA.  
           [0038]    The invention consists in the foregoing and also envisages constructions of which the following gives examples. 
       
    
    
     DETAILED DESCRIPTION OF THE INVENTION  
       [0039]    Preferred forms of the present invention will now be described with reference to the accompanying drawings in which:  
         [0040]    [0040]FIG. 1 is a plasma oestradiol concentrations following intramuscular injection of 0.72 mg (closed square) or vaginal administration of 7.2 mg (open square) of oestradiol 17β. Error bars are standard error means (n=3).  
         [0041]    [0041]FIG. 2 is a plasma oestradiol concentrations following vaginal administration of 10 mg oestradiol benzoate (open square), 10 mg oestradiol benzoate with 1:1 molar ratio β-cyclodextrin (open diamond), 10 mg oestradiol benzoate with 1:1 molar ratio hydroxypropyl β-cyclodextrin (open circle) or 10 mg oestradiol benzoate with 1:1 molar ratio γ-cyclodextrin (open triangle). Error bars are standard error means (n=4).  
         [0042]    [0042]FIG. 3 is a plasma oestradiol concentrations following vaginal administration of oestradiol 17β 7.2 mg (open square), 7.2 mg with 1:1 molar ratio β-cyclodextrin (open diamond), 7.2 mg with 1:1 molar ratio hydroxypropyl β-cyclodextrin (open circle) or 7.2 mg with 1:1 molar ratio γ-cyclodextrin (open triangle). Error bars are standard error means (n=4).  
         [0043]    [0043]FIG. 4 is a plasma oestradiol concentration following vaginal administration of 1.2 mg (closed diamond), 2.5 mg (closed square) or 7.2 mg (closed triangle) oestradiol 17β with 0.5:1 molar ratio of γ-cyclodextrin to oestradiol 17β. Error bars are standard error means (n=4).  
         [0044]    [0044]FIG. 5 is a plasma oestradiol concentration following vaginal administration of 1.2 mg (closed diamond), 2.5 mg (closed square) or 7.2 mg (closed triangle) oestradiol 17β with 1:1 molar ratio of γ-cyclodextrin to oestradiol 17β. Error bars are standard error means (n=4).  
         [0045]    [0045]FIG. 6 is a plasma oestradiol concentration following vaginal administration of 1.2 mg (closed diamond), 2.5 mg (closed square) or 7.2 mg (closed triangle) oestradiol 17β with 3:2 molar ratio of γ-cyclodextrin to oestradiol 17β. Error bars are standard error means (n=4).  
         [0046]    [0046]FIG. 7 is a area under the plasma oestradiol concentration against time curve (AUC) following vaginal administration of 1.2 mg, 2.5 mg or 7.2 mg oestradiol 17β with γ-cyclodextrin to oestradiol 17β molar ratio of 0.5 (closed diamond), 1 (closed square) or 1.5 (closed triangle). Error bars are standard error means (n=4).  
         [0047]    [0047]FIG. 8 is a time to maximum plasma concentration (tmax) following vaginal administration of 1.2 mg, 2.5 mg or 7.2 mg oestradiol 17β with γ-cyclodextrin to oestradiol 17β molar ratio of 0.5 (closed diamond), 1 (closed square) or 1.5 (closed triangle). Error bars are standard error means (n=4).  
         [0048]    [0048]FIG. 9 is a maximum plasma oestradiol concentration (Cmax) following vaginal administration of 1.2 mg, 2.5 mg or 7.2 mg oestradiol 17β with γ-cyclodextrin to oestradiol 17β, molar ratio of 0.5 (closed diamond), 1 (closed square) or 1.5 (closed triangle). Error bars are standard error means (n=4).  
         [0049]    [0049]FIG. 10 is a plasma oestradiol concentration at time=0 and time=24 hours post vaginal administration of various doses of oestradiol 17β (1.2, 2.5 or 7.2 mg) with various rations of γ-cyclodextrin (0.5, 1, 1.5 molar ration of γ-cyclodextrin to oestradiol 17β). Error bars are standard error means (n=4). * Denotes a significant difference between the plasma oestradiol concentration at time=0 and time=24 hours (p&lt;0.050). 
     
    
       [0050]    The use of oestradiol 17β and not the synthetic analogue oestradiol benzoate has not been firmly established due to the poor results with vaginally administered oestradiol benzoate and a perception that the shorter acting oestradiol 17β would not be as effications as the longer acting oestradiol benzoate. When oestradiol 17β in a dose equivalent to 1 mg oestradiol benzoate, i.e. 0.72 mg, is administered by i.m. injection in cattle the plasma oestradiol concentration rapidly rises to a maximum of approximately 100 pg/ml, followed by a rapid decline to pre injection levels by 24 hours following injection. See FIG. 1. Because of this rapid decline in plasma oestradiol levels an oestradiol 17β dose of 5 mg is commonly used to ensure adequate plasma oestradiol concentrations to achieve the same effect as 1 mg of oestradiol benzoate.  
         [0051]    We have found that unlike oestradiol benzoate vaginally administration oestradiol 17β is well absorbed, with a dose of 7.2 mg achieving a peak plasma concentration of between 10 and 20 pg/ml within four hours following administration, and the plasma oestradiol levels are elevated for at least 24 hours when oestradiol 17β is vaginally administered compared to the more rapidly cleared i.m. injection of oestradiol 17.  
         [0052]    We have found that the cyclodextrins improve the vaginal absorption of oestradiol benzoate. We have found that β or γ-cyclodextrin approximately double the plasma oestradiol concentration when vaginally administered with oestradiol benzoate (10 mg) compared with oestradiol benzoate administered without cyclodextrin. See FIG. 2.  
         [0053]    Furthermore we have found that the cyclodextrin hydroxypropyl β-cyclodextrin elevates plasma oestradiol concentrations approximately 6 fold following vaginal administration with oestradiol benzoate (10 mg) compared with oestradiol benzoate administered without cyclodextrin.  
         [0054]    We have also found that the cyclodextrins have indeed improved the vaginal absorption of oestradiol 17β. We have found that -cyclodextrin will approximately double the plasma oestradiol concentration when vaginally administered with oestradiol 17β (7.2 mg) compared with oestradiol 17β administered without cyclodextrin. See FIG. 3. Furthermore, we have found that the cyclodextrin hydroxypropyl β-cyclodextrin or γ-cyclodextrin elevates plasma oestradiol concentrations approximately 7 to 8 fold following vaginal administration with oestradiol (7.2 mg) compared with oestradiol 17β administered without cyclodextrin.  
         [0055]    We have found that the molar ratio of γ-cyclodextrin to oestradiol 17β influences the vaginal absorption of oestradiol 17β. Increasing the ratio of γ-cyclodextrin to oestradiol 17β from 0.5:1 to 1:1 has been found to increase the plasma oestradiol concentration. See FIGS. 4,5 and  6 .  
         [0056]    Further more the effect of the ratio of γ-cyclodextrin to oestradiol 17β upon the vaginal absorption of oestradiol 17β is more pronounced at higher doses (&gt;2.5 mg). See FIG. 7.  
         [0057]    We have found that vaginal administration of various amounts of oestradiol 17β (1.2, 2.5 and 7.2 mg) with various molar ratios of γ-cyclodextrin (0.5:1, 1:1 and 3:2) to oestradiol 17β has no significant effect upon the time to maximum plasma concentration (tmax) or the maximum plasma concentration (Cmax) of oestradiol. See FIGS. 8 and 9.  
         [0058]    We have found that the vaginal administration of a dose of oestradiol 17β greater than 2.5 mg with an amount of γ-cyclodextrin less than or equal to a molar ratio of 1:1 (γ-cyclodextrin to oestradiol 17β) results in plasma oestradiol concentrations 24 hours post administration significantly greater than those observed prior to administration. See FIG. 10.  
         [0059]    We have found vaginal administration of an oestradiol 17β dose of 5 mg and γ-cyclodextrin in a molar ratio of 0.5:1 (γ-cyclodextrin to oestradiol 17β) influences follicular dynamics in a similar manner to those observed following i.m. injection of 2 mg of oestradiol benzoate. See Table 1  
         [0060]    Table 1 tabulates a follicular dynamics and plasma oestradiol pharmacokinetics following vaginal administration of oestradiol 17β 2.5 mg or 5.0 mg and intramuscular administration of oestradiol benzoate 2 mg.  
                                                 TABLE 1                                   2.5 mg E-17   5.0 E-17   2 mg ODB                                    Follicular dynamics                   n   8   8   7       Day of oestrus   22.0 ± 0.0   22.0 ± 0.0   22.6 ± 0.2           (n = 7)   (n = 8)   (n = 7)       In oestrus by 48 h (n)   6   7   2       Follicle wave turnover   3   6   6       (i.e. DF3)       No follicle wave   5   2   1       turnover (i.e. no DF3)       Day DF2 emerged    9.9 ± 0.4 a     10.0 ± 0.6 a      9.6 ± 0.6 a         Diameter DF2 on    9.9 ± 0.5 a      9.3 ± 0.8 a      9.6 ± 1.0 a         Day 13 (mm)       Growth of DF2 from    2.4 ± 0.6 a      0.9 ± 0.8 ab     −0.6 ± 0.7 b         Day 13-17 (mm)       DF2 ovulated (n)   5   2   0       Age of ovulatory   12.0 ± 0.6   11.5 ± 0.5   —       DF2 (d)       Diameter of ovulatory   15.6 ± 0.7   17.0 ± 1.0   —       DF2 (mm)       Day DF3 emerged   18.3 ± 0.5   15.7 ± 0.6   17.0 ± 0.4       Interval, treatment to    5.3 ± 0.7    2.7 ± 0.6    4.0 ± 0.4       emergence of DF3 (d)   (4-6)   (1-5)   (3-5)       DF3 ovulated (n)   1   6   5       Age of ovulatory   6    7.0 ± 0.5    6.0 ± 0.5       DF3 (d)       Diameter of ovulatory   12   13.7 ± 0.7   14.2 ± 0.4       DF3 (mm)       Pharmacokinetics       Cmax (pg/ml)    8.8 ± 4.3 a      7.1 ± 4.1 a         Tmax (hours)    4.9 ± 7.8 a      5.9 ± 7.4 a         AUG (pg-hr/ml)   58.3 ± 25.8 a     97.8 ± 49.0 a         [oestradiol]pl (pg/ml)   0.17 ± 0.06 a1     0.89 ± 0.13 b1         at time = 0 hours       [oestradiol]pl (pg/ml)   0.15 ± 0.04 a1     1.26 ± 0.35 b2         at time = 24 hours