PCT
`
`WORLD INTELLECTUAL_ PROPERTY ORGANIZATION
`International Bureau
`
`
`
`INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT)
`
`(51) International Patent Classification 5 :
`A61K 31/495
`
`(11) International Publication Number:
`
`WO 99/18963
`
`(43) International Publication Date:
`
`22 April 1999 (2204.99)
`
`(21) International Application Number:
`
`PCT/IT98/00266
`
`(22) International Filing Date:
`
`6 October 1998 (06.10.98)
`
`(30) Priority Data:
`RM97A0OO6l3
`
`10 October 1997 (l0.10.97)
`
`IT
`
`(71) Applicant (for all designated States except US): MEDIVIS
`S.R.L. [IT/IT]; Via Marmolada, 4, l—20l00 Milano (IT).
`
`(81) Designated States: AL, AM, AT, AU, AZ, BA, BB, BG, BR,
`BY, CA, CH, CN, CU, CZ, DE, DK, EE, ES, FI, GB, GD,
`GE, GH, GM, HR, HU, ID, IL, IS, JP, KE, KG, KP, KR,
`KZ, LC, LK, LR, LS, LT, LU, LV, MD, MG, MK, MN,
`MW, MX, NO, NZ, PL, PT, RO, RU, SD, SE, SG, SI, SK,
`SL, TJ, TM, TR, T1‘, UA, UG, US, UZ, VN, YU, ZW,
`ARIPO patent (GH, GM, KE, LS, MW, SD, SZ, UG, ZW),
`Eurasian patent (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM),
`European patent (AT, BE, CH, CY, DE, DK, ES, FI, FR,
`GB, GR, IE, IT, LU, MC, NL, PT, SE), OAPI patent (BF,
`BJ, CF, CG, CI, CM, GA, GN, GW, ML, MR, NE, SN,
`TD, TG).
`
`(72) Inventor; and
`(75) Inventor/Applicant (for US only): LISI, Giuseppe [IT/IT];
`Medivis s.r.1., Via Marmolada, 4, I—20100 Milano (IT).
`
`Published
`
`(74) Agents: BANCHETTI, Marina et al.; Ing. Barzano & Zanardo
`Roma S.p.A., Via Piemonte, 26, I-00187 Roma (IT).
`
`With international search report.
`Before the expiration of the time limit for amending the
`claims and to be republished in the event of the receipt of
`amendments.
`
`(54) Title: USE OF FLUNARIZINE FOR THE TOPICAL TREATMENT OF GLAUCOMA
`
`(57) Abstract
`
`Use of flunarizine, a calcium channel blocking agent known for use as cerebral and peripheral vasodilator, in the treatment of glaucoma
`by topical administration. Differently from other calcium channel blockers already tested for use as antiglaucoma agents, flunarizine is highly
`active in lowering the intraocular pressure when administered by the topical ophthalmic route. The invention also comprises anti—glaucoma
`preparations containing flunarizine, or combinations of flunarizine with beta—blockers such as timolol.
`
`000001
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`ARGENTUM PHARM. 1012
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`ARGENTUM PHARM. 1012
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`000001
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`

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`
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`FOR THE PURPOSES OF INFORMATION ONLY
`
`
`Codes used to identify States party to the PCT on the front pages of pamphlets publishing international applications under the PCT.
`
`SI
`SK
`SN
`SZ
`TD
`TG
`TJ
`TM
`TR
`TT
`UA
`UG
`US
`UZ
`VN
`YU
`ZW
`
`Slovenia
`Slovakia
`Senegal
`Swaziland
`Chad
`Togo
`Tajikistan
`Turkmenistan
`Turkey
`Trinidad and Tobago
`Ukraine
`Uganda
`United States of America
`Uzbekistan
`Viet Nam
`Yugoslavia
`Zimbabwe
`
`LS
`LT
`LU
`LV
`MC
`MD
`MG
`MK
`
`ML
`MN
`MR
`MW
`MX
`NE
`NL
`N0
`NZ
`PL
`PT
`R0
`RU
`SD
`SE
`SG
`
`Lesotho
`Lithuania
`Luxembourg
`Latvia
`Monaco
`Republic of Moldova
`Madagascar
`The fom1er Yugoslav
`Republic of Macedonia
`Mali
`Mongolia
`Mauritania
`Malawi
`Mexico
`Niger
`Netherlands
`Norway
`New Zealand
`Poland
`Portugal
`Romania
`Russian Federation
`Sudan
`Sweden
`Singapore
`
`000002
`
`Albania
`Annenia
`Austria
`Australia
`Azerbaijan
`Bosnia and Herzegovina
`Barbados
`Belgium
`Burkina Faso
`Bulgaria
`Benin
`Brazil
`Belarus
`Canada
`Central African Republic
`Congo
`Switzerland
`Cote d’Ivoire
`Cameroon
`China
`Cuba
`Czech Republic
`Germany
`Denmark
`Estonia
`
`ES
`FI
`FR
`GA
`GB
`GE
`GH
`GN
`GR
`I-IU
`IE
`IL
`IS
`IT
`JP
`KE
`KG
`KP
`
`KR
`KZ
`LC
`LI
`LK
`LR
`
`Spain
`Finland
`France
`Gabon
`United Kingdom
`Georgia
`Ghana
`Guinea
`Greece
`Hungary
`Ireland
`Israel
`Iceland
`Italy
`Japan
`Kenya
`Kyrgyzstan
`Democratic People's
`Republic of Korea
`Republic of Korea
`Kazakstan
`Saint Lucia
`Liechtenstein
`Sri Lanka
`Liberia
`
`AL
`AM
`AT
`AU
`AZ
`BA
`BB
`BE
`BF
`BG
`BJ
`BR
`BY
`CA
`CF
`CG
`CH
`CI
`CM
`CN
`CU
`CZ
`DE
`DK
`EE
`
`
`
`
`
`
`
`
`
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`000002
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`WO 99/18963
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`PCT/IT98/00266
`
`USE OF FLUNARIZINE FOR THE TOPICAL TREATMENT OF GLAUCOMIT
`
`SPECIFICATION
`
`The present invention concerns the use of flunarizine for the topical
`
`treatment of glaucoma. More specifically, this invention relates to the use of
`
`flunarizine, a calcium channel blocking agent known and employed as cere-
`
`bral and peripheral vasodilator, in a new indication as an antiglaucoma agent
`
`10
`
`for topical ophthalmic treatment.
`
`As it is known, glaucoma is a pathological ophthalmic condition the
`
`underlying causes of which are not well understood at present. This condition
`
`is usually shown by a progressive increase of the intraocular pressure, lead-
`
`ing to severe impairment of the eye structures, in particular to damage to the
`
`15
`
`optic nerve disc and to decrease in the visual field, finally resulting in optic
`
`atrophy. The disease is generally connected to an insufficient outflow of
`
`aqueous humour from the eye, although other causes, such as, e.g.,
`
`the
`
`production of aqueous humour and the episcleral veins pressure, take part in
`
`the regulation of the intraocular pressure.
`
`20
`
`The rationale of the pharmacological therapy presently in use is to
`
`lower the intraocular pressure. The drugs currently used to that aim, divided
`
`into classes according to their mechanism of action, are beta-blockers (such
`
`as timolol, betaxolol,
`
`levobunolol), sympathomimetics (such as epinephrine
`
`and dipivephrine), parasympathomimetics or miotics (such as pilocarpine and
`
`25
`
`acetylcholine) and carbonic anhydrase inhibitors (such as acetazolamide and
`
`dichlorphenamide). Besides the foregoing drugs well established in use, the
`
`search for agents having less side effects and longer lasting activity has lead
`
`to evaluate, more recently, the possibility of using for the treatment of glau-
`
`coma another class of drugs, i.e. the calcium blocking agents. The latter, also
`
`30
`
`known as “calcium entry blockers” or “calcium antagonists”, are currently
`
`used as vasodilators and in the treatment of cardiac affections. For such
`
`indications,
`
`the most widespread calcium antagonists are, e.g., nifedipine,
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`WO 99/18963
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`diltiazem and verapamil.
`
`The role of calcium in the dynamics of aqueous humour and in the
`
`control of intraocular pressure has not yet been entirely clarified, although it
`
`is known that the production and the outflow of aqueous are modulated also
`
`by calcium. As concerns the formation of aqueous,
`
`it is to be noted, firstly,
`
`that the hydrostatic component due to the arterial pressure and to the pres-
`
`sure of the vessels feeding the ciliary body is calcium-dependent, as it
`
`is
`
`confirmed by the known systemic vascular action of calcium antagonists.
`
`Further, the osmotic pressure due to ionic secretion at the level of the non-
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`10
`
`pigmented ciliary epithelium is likely to be modulated by calcium, as hy-
`
`pothesised by Abelson et al. (Abelson M.B., Gilbert C.M., Smith L.M., Sus-
`
`tained reduction of intraocular pressure in humans with the calcium channel
`
`blocker verapamil, Am. J. Ophthamol. 105; 155 (1988)).
`
`As far as the outflow of the aqueous humour is concerned, calcium
`
`15
`
`ions play a direct role in modulating the pressure of episcleral veins, and
`
`some studies suggest that calcium influences the outflow capacity, by main-
`
`taining the structural integrity of the trabecuale and of the exterior wall of the
`
`Schlemm’s canal.
`
`In spite of the foregoing suggestions several experimental works,
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`20
`
`both on animal models and clinical, and involving both systemic and topical
`
`administration, reported contradictory results about the activity of calcium
`
`channel blockers in the therapy of glaucoma. For instance, Monica et al.
`
`(Monica M.L., Hesse R.J., Messerll F.H., The effect of a calcium-channel
`blocking agent on intraocular pressure, Am. J. Ophthalmol. 96, 814 (1983))
`
`reports that the oral administration of nltrendipine to patients with moderate
`hypertension but with normal intraocular pressure slightly lowered the latter,
`while Beatty and co-workers (Beatty J.F., Krupin T., Nichols P.F., Elevation of
`intraocular pressure by calcium-channel blockers, Arch. Ophthalmol. 102;
`
`1072, (1984)) did not evidence any effect upon oral administration of vera-
`
`pamil to rabbits, and did even report an increase in the intraocular pressure
`upon topical administration. More recently,
`for instance, Payene and co-
`workers (Payene, L.J., Slagle T.M., Cheeks L.T., Effect of calcium-channel
`
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`30
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`_ 3 _
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`blockers on intraocular pressure, Ophthalmic Res. 22; 337, (1990)) obtained
`a reduction in the intraocular pressure upon systemic administration of vera-
`
`pamil or nifedipine to rabbits, but did not detect any significant effect upon
`topical administration of the same agents or of diltiazem by the topical route.
`In general, however, at least as far as verapamil is concerned, it may
`be said that the administration of this drug to man normally results in a reduc-
`
`tion of the intraocular pressure. A more consistent reduction upon topical
`administration has been explained,
`in particular, by a work of Ettl et al. (Ettl
`A., Daxer A., Hoffmann U., Calcium channel blockers in the management of
`low—tension and open-angle glaucoma, Am. J. Ophthalmol. 116; 778, (1993)).
`These authors have detected,
`in the rabbit eye, verapamil levels 200 times
`
`higher than the levels obtainable by systemic administration.
`Accordingly, the use of verapamil in the treatment of ocular hyper-
`tension is the object of the international PCT application No. WO 92/07563,
`filed by Abelson (i.e., the first author cited above) et al.. A later publication in
`the name of the same author is the international application No. W0 96/
`
`03986, concerning the treatment of a particular form of glaucoma, referred to
`as low—tension glaucoma. This pathology is characterised by an intraocular
`pressure which is almost normal,
`in spite of the fact that all of the other
`symptoms of glaucoma are present.
`In the latter document the therapeutic
`proposal is generically extended to all calcium-antagonists, many representa-
`tives of which are mentioned in a preliminary list. However, the only example
`
`of active agent disclosed in the document and supported by experimental
`
`10
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`15
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`20
`
`data is verapamil.
`Another calcium blocking agent that was specifically proposed for
`
`25
`
`in the treatment of intraocular hypertension is
`in a patent document,
`use,
`diltlazem (French patent No. 2593395, published in 1987), while a list of more
`than one hundred calcium antagonists is presented in the international PCT
`application No. WO 93/23082. The latter concerns, for use in the treatment of
`glaucoma, a combination of a compound which lowers the intraocular pres-
`sure (i.e., a conventional antiglaucoma agent) and a calcium channel block-
`ing agent. The disclosure does not contain any specific example of preferred
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`WO 99/18963
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`PCT/IT98/00266
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`combination, nor any experimental detail regarding the activity of any combi-
`
`nation.
`
`Some experimental trials on verapamil also allowed to ascertain that
`
`the ophthalmic use of the said agent causes an undesirable swelling of the
`
`cornea. (Green K., Cheeks L., Hull D.S., Effects of calcium channel blockers
`
`on rabbit corneal endothelial function, Curr. Eye Res. 13; 401-408, (1994)).
`
`This is particularly critical
`
`if one considers the use for the treatment of a
`
`chronic condition as is, actually, glaucoma.
`
`Although the entire class of calcium antagonists has already been
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`10
`
`considered for its potential use in the treatment of glaucoma, there does not
`
`seem to have been evidenced the particular activity, against this type of pa-
`
`thologies, of a specific agent belonging to the said class,
`
`i.e. flunarizine.
`
`It
`
`has now been found, and it is the subject-matter of this invention, that the
`
`specific calcium antagonist flunarizine, when administered through the topical
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`15
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`ocular route, is able to lower the intraocular pressure in a surprisingly more
`
`marked way than the other calcium antagonists so far proposed and tested for
`
`the therapy of glaucoma.
`
`Within the frame of the studies connected with this invention,
`
`it has
`
`also been found that some known receptors, referred to as c receptors, are
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`20
`
`localised in the ocular region,
`
`in particular in the ciliary body and in the iris,
`
`and that some specific “ligands”, having a cs-agonist activity, significantly
`
`lower the ocular pressure. Since it has been experimentally found that
`
`flunarizine shows a cs-agonist activity which is far higher than the activity of
`
`other calcium antagonists, this property may explain the unexpectedly greater
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`25
`
`activity of flunarizine in lowering the intraocular pressure, if it is hypothesised
`
`that such activity is exerted according to mechanisms of action that are at
`
`least partially different from the other calcium blocking agents.
`
`In order to identify the presence of 0 receptor sites in the eye the
`
`technique of “receptor binding” has been exploited. The latter has been car-
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`30
`
`ried out on cell membranes obtained from the irido—ciliary body complex. The
`
`irido—ciliary body complex had been explanted, after sacrifice, from male al-
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`bino rabbits of the New Zealand strain. The tissue was homogenised in buffer
`
`and a fraction rich in cell membrane proteins was isolated, obtained by cen-
`
`trifugation. The concentration of total proteins has been evaluated by the
`
`method of Lowry (Lowry, J. Biol. Chem. 193; 265 (1951)). Aliquots of the said
`
`fraction of the homogenate containing 300 pg of total proteins were incubated
`
`with scalar amounts of [3H](+)—pentazocine (which is used, for experimental
`
`purposes only, as a c: ligand). The reaction was carried out at 37°C for 150
`
`minutes and then, after filtering, the radioactivity left on the filters was meas-
`
`ured by liquid scintillation. The apparent dissociation constant (Kd) and the
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`10
`
`total number of receptors were determined, and it was thus ascertained that
`
`[3H](+)-pentazocine selectively binds to receptor sites present in the irido-
`
`ciliary body region of the rabbit. On the basis of the present scientific knowl-
`
`edge, the said receptors appear to be of the type cs-1.
`
`Further, “competitive binding” assays carried out with a constant
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`15
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`amount of [3H](+)—pentazocine and scalar amounts of (+)—N-allil—nor-meth-
`
`azocine (NANM) (which is used, for experimental purposes only, as a c li-
`
`gand), showed that the latter shift the radioactive ligands from the receptor
`
`sites. It has also been observed, by analysing the Hill coefficient, that NANM
`
`interacts with one only class of 0 receptor sites.
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`20
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`25
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`30
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`In the frame of the same research it has been found that cs-agonist
`
`agents show an ocular anti-hypertensive activity. A 1% preparation of NANM
`
`was administered (50 pl) in the conjunctival fornix of the right eye of male
`
`albino rabbits of the New Zealand strain, after measuring the (baseline) in-
`
`traocular pressure. Upon measuring again the intraocular pressure 60, 120,
`
`180 e 240 minutes after the instillation,
`
`it has been ascertained that the in-
`
`traocular pressure was significantly reduced (p<0.01) 60 minutes after the
`
`instillation, in comparison with the formulation containing the vehicle only.
`
`Lastly, as it was pointed out before, studies of receptor binding car-
`
`ried out with flunarizine (some of which are presented in the following) have
`
`shown that flunarizine has an affinity for 5-1 receptors which is not even com-
`
`parable to the affinity shown by the other calcium channel blocking agents
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`tested.
`
`Another advantageous aspect distinguishing flunarizine from the
`
`other calcium channel blocking agents proposed so far for the topical treat-
`
`ment of glaucoma is, as it has now been found, that flunarizine does not show
`
`any side effect of corneal swelling.
`
`Therefore,
`
`the present
`
`invention specifically provides the use of
`
`flunarizine, optionally in the form of a pharmaceutically acceptable salt, for
`
`the topical treatment of glaucoma, i.e. the use of flunarizine, or of a pharma-
`
`ceutically acceptable salt thereof,
`
`in the manufacture of a topical ophthalmic
`
`medicament for the treatment and/or the prophylaxis of glaucoma. In general,
`
`the topical administration of flunarizine may take place by using a preparation
`
`in the form of an aqueous solution or suspension, or in the form a gel, an
`
`ointment or a cream in a pharmaceutically acceptable ophthalmic vehicle, or
`
`in the form of an erodible ocular insert or of a “reservoir“ system with a poly-
`
`mer membrane, to be placed in the conjunctival sac.
`
`The concentration of flunarizine in an ophthalmic vehicle may range
`
`from 10 pg/ml to 5 mg/ml,
`
`i.e. from 0.001 to 0.500% by weight. The optimal
`
`concentration is chosen firstly on the basis of the dosage to be administered:
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`10
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`15
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`in the case of use in eye-drop form, for instance, one drop should contain a
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`20
`
`sufficient amount of flunarizine for the drop to be effective as such or when
`
`instilled twice (i.e., two drops). Other criteria for the choice of the concentra-
`
`tion are the ocular tolerability (it should be considered that the conjunctival
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`sac,
`
`into which the ophthalmic preparation is to be instilled, has a limited
`
`capacity) and the stability of the active ingredient. The preferred concentra-
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`25
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`tion for an aqueous solution formulation (eye-drops) is 0.050% by weight, and
`
`preferably the product is present in the form of the corresponding hydrochlo-
`
`ride salt (optimal concentration of flunarizine hydrochloride: 0.052%).
`
`According to a particularly preferred embodiment of this invention, the
`
`anti-glaucoma activity of the proposed ophthalmic preparation is further en-
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`30
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`hanced by the presence,
`
`in combination with flunarizine, of an effective
`
`amount of a beta-blocking agent. The class of beta-blockers (or l3—adrenergic
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`blockers), referred to in the foregoing, represents to date the most wide-
`
`spread class of anti-glaucoma agents. These agents are used in the topical
`
`treatment of chronic open angle glaucoma and, more generally,
`
`in the treat-
`
`ment of intraocular hypertension. Their mechanism of action mainly consists
`
`in reducing the production of the aqueous humour, and therefore the unex-
`
`pected enhanced activity of the proposed combination of flunarizine (which
`
`has been found to be active in increasing the outflow of aqueous) with a beta-
`
`blocker may reasonably be explained in terms of a complementarity of the two
`
`actions.
`
`Preferably, the concentration of beta—blocking agent in the combina-
`
`tion according to the invention is from 0.1 to 2.5% by weight, and most pref-
`
`erably said beta—blocking agent is timolol or a pharmaceutically acceptable
`
`salt thereof.
`
`A vehicle that may be employed in an eye-drop preparation according
`
`to the invention is the simple physiological saline solution containing 0.9% by
`
`weight of sodium chloride. Such solution is isotonic with respect to the tear
`
`fluid, and therefore it is well tolerated by the eye. However, also hypotonic
`
`solutions or suspensions may be employed, as it is known that these prepa-
`
`rations are well tolerated by the ocular tissues.
`
`Other excipients may be added to the composition of the invention in
`
`order to adjust the tonicity of the solutions or suspensions, so as to stabilise
`
`the active ingredient(s) and to increase the tolerability of the preparation.
`
`Specifically, any buffers should maintain the pH into the range 4-8. For in-
`
`stance, the above saline solution may be buffered with any one of the buffers
`
`well known in the pharmaceutical art for ophthalmic use, such as, e.g., phos-
`
`phate buffer, or trizma buffer (i.e., tri-hydroxymethyl amino methane), so as to
`
`obtain a physiological pH,
`
`in the range of 7.0-7.4. Further, the solution may
`
`also have an osmolarity in the physiological range (295-305 mOsm/l). This
`
`allows to obtain a better ocular tolerability.
`
`In addition, the formulation may
`
`advantageously contain an antioxidant, such as, e.g., gallates, ascorbic acid,
`
`superoxide dismutase (SOD), BHT, sodium metabisulphite, tocopherols, BHA,
`
`nordihydroguaiaretic acid, ascorbic acid esters, dimethylthiourea and the like.
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`The tolerability may be further enhanced by means of other excipients
`
`such as cyclodextrins, polysorbate 80 (or Tween 80), dextrane (e.g., dextrane
`
`70), polyethylene glycol (e.g. PEG 400), poloxamers and other similar agents.
`
`The formulation may include viscosifying/thickening agents such as methyl-
`
`cellulose, polyvinyl alcohol, glucosamine glucans, polyvinyl pyrrolidone and
`
`the like,
`
`in order to increase the ocular bioavailability, the stability and the
`
`tolerability of the active ingredient(s).
`
`The ocular bioavailability of flunarizine may be further enhanced by
`
`the addition of substances which increase the corneal permeation of the drug,
`
`such as, e.g., dimethyl sulphoxide, taurocholates, membrane phospholipides,
`
`benzalkonium chloride and other surface active agents for ophthalmic use
`
`(such as disodium lauryl sulphosuccinate).
`
`Lastly,
`
`in the preparations to be packaged in multidose bottles com-
`
`positions a preservative with antimicrobial activity will have to be added,
`
`in
`
`order to prevent contamination of the product. Such agent may be chosen
`
`among the preservative agents well known for this use in the pharmaceutical
`
`art.
`
`Products to be administered in the form of suspensions should con-
`
`tain suitable agents such as carboxymethyl cellulose and the like. In the event
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`that the preparation is to be employed in the form of an ointment, a gel or a
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`cream for ophthalmic use, flunarizine will be admixed with carriers such as
`
`polyethylene glycols, polyacrylates, polyethylene oxides, fatty acids and alco-
`
`hols or lanolin, paraffin and other similar products. Suitable ingredients for the
`
`production of emulsions or microemulsions may be chosen among the follow-
`
`ing: diethylene glycol—monobutyl ether, di(ethylene glycol) buthyl ether, ca-
`
`prylic acid ethyl ester, oleic acid ethyl ester, soybean oil, hexadecane, tribu-
`
`tyrin, ethylene glycol—monobutyl ether, 1-hexadecane, n-heptane, 1-heptene,
`
`Tween 80, PEG, poloxamers, polyoxyethylene ethers.
`
`The dosage of the main active ingredient of the invention, to be ad-
`
`ministered by the topical route, may vary from about 20 pg to about 200 pg
`
`per day for each eye. The prescription dosage of the ophthalmic preparations
`
`based on flunarizine will depend on the daily dose that will be necessary to
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`achieve the therapeutic effect and, obviously, on the specific formulation
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`employed. Ophthalmic solutions or suspensions will require from 1 to 4 instil-
`
`lations per day; ointments, gels and creams will require 1 or 2 applications;
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`solid inserts with polymeric matrix, either biodegradable or not, will require
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`one only administration per day.
`
`The present invention further concerns compositions which allow the
`
`administration of flunarizine through the topical ophthalmic route, and specific
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`ophthalmic compositions for use in the treatment and/or in the prophylaxis of
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`glaucoma comprising, as an active ingredient, a therapeutically effective
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`amount of flunarizine. A group of preferred compositions have the following
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`formulation (wherein all percentages are by weight):
`
`flunarizine hydrochloride
`
`0.059
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`%
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`(corresponding to 0.05% flunarizine)
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`sodium chloride
`
`0.10-0.80 %
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`trizma buffer
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`PEG 400
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`Tween 80
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`0.02-0.20 %
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`1.00-6.00 %
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`2.00-12.00 %
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`sodium metabisulphite
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`0.01-0.20 %
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`propyl gallate
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`20
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`EDTA
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`purified water
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`0.01-0.50 %
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`0005-020 %
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`q.s. to 100 %
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`optionally comprising further pharmaceutically acceptable ingredients.
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`In a particularly preferred embodiment of this invention, the composi-
`
`tions for use in the treatment and/or in the prophylaxis of glaucoma further
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`contain from 0.1 to 2.5% by weight of a beta-blocking agent, the latter being
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`by preference timolol or a pharmaceutically acceptable salt thereof, such as
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`timolol maleate.
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`Some specific embodiments of the invention are described below for
`
`merely illustrative purposes,
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`together with the results of the experimental
`
`studies carried out on the proposed anti-glaucoma agent, including compara-
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`tive tests with other calcium-blocking agents.
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`EXAMPLE 1
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`Ophthalmic solution based on flunarizine
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`A composition according to the invention that turned out to be particu-
`
`larly effective (the performance of which was experimentally evaluated as it is
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`5
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`partly reported further on) has the following composition (the percentages
`
`being given by weight):
`
`flunarizine hydrochloride
`
`0.059 %
`
`(corresponding to 0.050% flunarizine)
`
`sodium chloride
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`10
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`trizma buffer
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`PEG 400
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`Tween 80
`
`0.485 %
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`0.100 %
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`2.500 %
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`5.000 %
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`sodium metabisulphite
`
`0.050 %
`
`propyl gallate
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`15
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`EDTA
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`0.050 %
`
`0.010 %
`
`purified water
`
`q.s. to 100 %
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`20
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`25
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`30
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`The above composition is suitable for being packaged in single dose
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`containers; in the event that a multidose packaging is desired, a preservative
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`(such as, e.g., benzalkonium chloride) will have to be added in order to
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`maintain the sterility of the product for the whole period of use.
`
`EXAMPLE 2
`
`Ophthalmic microemulsion based on flunarizine
`
`A composition suitable for use as an ophthalmic ointment was pre-
`
`pared according to the formulation given below (weight percentages) :
`
`flunarizine hydrochloride
`
`0.059 %
`
`(corresponding to 0.050% flunarizine)
`
`trizma buffer (to pH 7.20)
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`0.100 %
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`PEG 400
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`soybean oil
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`Tween 80
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`sodium metabisulphite
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`sorbitol
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`10.000 %
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`2.00
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`%
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`20.000 %
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`0.050 %
`
`2.057 %
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`propyl gallate
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`purified water
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`0.050 %
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`q.s. to 100 %
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`As a tonicity adjusting agent, 455 mg of sodium chloride per 100 ml
`
`(i.e. 0.455 wt. %) may be used in place of the above amount of sorbitol.
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`EXAMPLE 3
`
`Ophthalmic emulsion based on flunarizine
`
`An ophthalmic product similar to that shown in the previous example,
`
`but having a coarser size of the drops of the dispersed phase, was obtained
`
`excluding the soybean oil from the composition, according to the following
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`formulation (weight percentages) :
`
`flunarizine hydrochloride
`
`0.059 %
`
`(corresponding to 0.050% flunarizine)
`
`trizma buffer (to pH 7.20)
`
`0.100 %
`
`PEG 400
`
`15
`
`Tween 80
`
`sodium metabisulphite
`
`sorbitol
`
`propyl gallate
`
`purified water
`
`2.000 %
`
`7.000 %
`
`0.050 %
`
`2.014 %
`
`0.050 %
`
`q.s. to 100 %
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`As an alternative to sorbitol as a tonicity adjusting agent, the compo-
`
`sition may include 433 mg of sodium chloride per 100 ml (i.e. 0.433 wt. %).
`
`EXAMPLE 4
`
`Ophthalmic solution based on a combination of flunarizine and timolol
`
`A particularly preferred composition according to the invention was
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`obtained by adding to the formulation of Example 1 a sufficient amount of
`
`timolol maleate to achieve a concentration of 0.5% by weight of timolol in the
`
`overall composition (corresponding to about 0.68% by weight of timolol
`
`maleate). The concentrations of the other ingredients were the same as
`
`specified above for Example 1.
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`Similarly, also the formulations given in Examples 2 and 3 can be
`
`modified with the addition of a proper amount of timolol maleate. Also in this
`
`case, it is preferred to obtain a concentration of 0.5% by weight of timolol in
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`the overall composition.
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`Experimental results
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`An isotonic solution, buffered and viscosified according to the formu-
`
`lation of Example 1, but having variable concentrations of flunarizine (ranging
`
`from 0.01% to 0.1% by weight), was generally referred to as MEG O1 in the
`
`experimental work the results of which are set forth below. The experimenta-
`
`tion also considered combinations of flunarizine and beta-blocking agents
`
`formulated as shown for timolol in Example 4. The combination of flunarizine
`
`and timolol was referred to as MEG 02. Some of the said experimental results
`
`are also shown in the graphs of the accompanying drawings, wherein:
`
`Figure 1 shows the percent reduction in the intraocular pressure
`
`obtained upon instillation of flunarizine in the eyes of rabbits with hyperten-
`
`sion, in comparison with the corresponding reduction obtained with the instil-
`
`lation of placebo and with the instillation of other calcium antagonists;
`
`Figure 2 shows the percent reduction in the intraocular pressure
`
`obtained upon instillation,
`
`in the eyes of rabbits with ocular hypertension, of
`
`flunarizine at various concentrations; and
`
`Figure 3 shows the percent reduction in the intraocular pressure
`
`obtained upon instillation,
`
`in the eyes of rabbits with ocular hypertension, of
`
`flunarizine in combination with various beta-blocking agents.
`
`Pharmacodynamic studies
`
`a. Study on rabbits with normal intraocular gressure
`
`The effects of the agent of the invention on the intraocular pressure of
`
`rabbits showing normal baseline intraocular pressure were evaluated in com-
`
`parison with the action of a placebo, and with that of various other calcium
`
`channel blocking agents. Female pigmented rabbits of the Vienna Blue strain
`
`were used (supplied by Charles River ltaliana, of Calco (C0)). The age of the
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`animals at the time of starting the experimentation was 9 weeks, and their
`
`weight was 2.0-2.5 kg.
`
`The choice of a species with pigmented iris is due to the fact that the
`
`latter represents a reliable model for the evaluation of possible modifications
`
`of the intraocular pressure caused by the products under test. The strain g
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`chosen is genetically defined, so as to limit to a minimum the variability of the
`
`biological characteristics between one animal and the other.
`
`The animals were kept in rooms maintained under constant and con-
`
`trolled conditions of temperature and humidity, illuminated for 12 hours a day
`
`with artificial light and with continuous renovation of the air. The feed con-
`
`sisted of a standard diet having a constant and known composition, and both
`
`feed and water were available ad libitum during the whole period of the test.
`
`The rabbits were stabled for 21 days before starting the test, so as to allow a
`
`sufficient acclimatation and to suitably evaluate the health conditions of the
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`rabbits. Each experimental group consisted of 4 animals, which were allotted
`
`to the treatment groups in a randomised way.
`
`Each different group of animals received, by instillation in the right
`
`conjunctival fornix, 50 ul of the following products:
`
`a) eye-drops of MEG 01, containing 0.050 wt. % flunarizine (0.052 wt. %
`
`15
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`flunarizine hydrochloride);
`
`b) placebo solution (i.e., the vehicle of MEG 01);
`
`c) eye-drops containing 0.056 wt. % verapamil in the vehicle of MEG 01;
`
`d) eye-drops containing 0.051 wt. % diltiazem in the vehicle of MEG 01;
`
`e) eye-drops containing 0.043 wt. % nifedipine in the vehicle of MEG 01.
`
`The weight concentrations of the various agents under test are cho-
`
`sen so as to correspond to the same molar concentration.
`
`The pressure in the treated eye was measured by flattening tonome—
`
`ter (TonopenXL®, Mentor), 15 minutes before the instillation of the eye-drops
`
`(time 0) and then 30, 60, 90, 120, 180 and 240 minutes after. As a local an-
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`aesthetic, 5 minutes before carrying out each measurement 25 pl of a com-
`
`mercial ophthalmic solution containing 0.4% oxybuprocaine hydrochloride
`
`(Novesine®, Sandoz) was instilled. To carry out the measurement the rabbits
`
`were placed in a suitably designed cage, that prevents any sudden movement
`
`of the animal under test.
`
`For each animal and at each of the times listed above the average of
`
`three subsequent measurements was calculated and recorded, each one said
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`measurements being made after 1 minute from the previous one. The in-
`
`traocular pressure values at the various times were compared with the values
`
`obtained before the treatment, by means of the Student’s “t” test. The com-
`
`parisons between different groups were made by processing the data by the
`
`variance analysis (ANOVA) and, where possible, by the Student’s “t” test for
`
`the comparison of two different experimental groups. Values of p < 0.05 were
`
`considered to be statistically significant.
`
`The following table shows the values of intraocular pressure deter-
`
`mined on each one of the animals treated, as well as the average values for
`
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`each test group (i standard deviation).
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`TABLE 1
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`PCT/IT98/00266
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`_
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`lntraocular pressure in rabbits with normal pressure treated with the tested agents
`
`
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`Rabbitwo.
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`