United States Patent
`
`[19]
`
`[11] Patent Number:
`
`4,537,883
`
`Alexander et al.
`
`[45] Date of Patent:
`
`Aug. 27, 1935
`
`trial Pharmacy” pp. 521-524 2d Ed., Lea And Febiger
`Publishers (1976) (Lachman et al.).
`“Remington’s Pharmaceutical Sciences” pp. 1483-1485
`Mack Publishing Co. (1975) (Remington).
`Brooke, et al., American Journal Of Hospital Phar-
`macy, 32:44-45 (1975).
`
`Primary Examiner-—Albert T. Meyers
`Assistant Examiner—Joyce L. Morrison
`Attorney, Agent, or Fz'rm—Richard P. Ryan; Robert H.
`Uloth
`
`[57]
`
`ABSTRACT
`
`A lyophilized pharmaceutical solid composition con-
`taining cyclophosphamide for reconstitution with water
`to provide a solution for oral or parenteral administra-
`tion. This lyophilized cyclophosphamide solid composi-
`tion demonstrates improved stability, solubility charac-
`teristics and enhanced appearance compared with cur-
`rently available dry powder pre-mix compositions of
`cyclophosphamide. The lyophilized solid composition
`contains about 20 parts by weight of cyclophospha-
`mide, about Ii-2 parts by weight of water and from
`about 10-85 parts by weight of excipient which is com-
`prised mainly of mannitol. Processes for making the
`composition are disclosed.
`
`[54] LYOPHILIZED CYCLOPHOSPHAMIDE
`
`[75]
`
`Inventors: Robert L. Alexander; Robert J.
`Bequette; Terry T. Kensler; Joseph A.
`Scott, all of Evansville, Ind.
`
`[73] Assignee: Mead Johnson & Company,
`Evansville, Ind.
`
`[21] Appl. No.: 589,202
`
`[22] Filed:
`
`Mar. 13, 1984
`
`Related U.S. Application Data
`
`[63]
`
`Continuation-in-part of Ser. No. 440,906, Nov. 12,
`1982, abandoned.
`
`Int. c1.3 ............................................ .. A61K 31/66
`[51]
`[52] U.S. c1. . . ..
`. .. .. 514/110; 514/960
`[58] Field of Search .......................... .. 424/209, ; 34/5
`
`
`
`[56]
`
`References Cited
`U.S. PATENT DOCUMENTS
`
`3,018,302
`3,732,340
`
`1/1962 Arnold et al. ....................... 260/936
`5/1973 Arnold et al. ....................... 260/936
`
`OTHER PUBLICATIONS
`
`Lachman et al., “The Theory And Practice Of Indus-
`
`16 Claims, No Drawings
`
`FRESENIUS KABI 1006-OOO1
`
`

`
`1
`
`LYOPHILIZED CYCLOPHOSPHAMIDE
`
`CROSS REFERENCE TO RELATED
`APPLICATION
`
`This is a continuation-in-part application of Ser. No.
`06/440,906 filed Nov. 12, 1982 now abandoned.
`BACKGROUND OF THE INVENTION
`
`Cyclophosphamide is the generic name for 2-[bis(2-
`chloroethyl)amino]tetrahydro-2H-1,3,2-oxazaphospho-
`rine-2-oxide monohydrate, a widely used antineoplastic
`drug chemically related to the nitrogen mustards. Cy-
`clophosphamide was one example of a group of novel
`cyclic phosphoric acid ester amides which were dis-
`closed and claimed in U.S. Pat. No. 3,018,302 granted
`Jan. 23, 1962 to H. Arnold, et al.
`A related series of compounds bearing substituents on
`the oxazaphosphorine ring nitrogen was disclosed and
`claimed in U.S. Pat. No. 3,732,340 granted May 8, 1973
`also to H. Arnold, et a1.
`Over the past 20 years a considerable amount of liter-
`ature concerning cyclophosphamide has accumulated.
`Most of these references deal mainly with clinical appli-
`cations of this agent as an antineoplastic drug. Cyclo-
`phospharnide has been distributed for much of that time
`by Mead Johnson & Company under the proprietary
`name CYTOXAN ®. ENDOXAN ® and NEO-
`SAR ® are proprietary names for similar pharmaceuti-
`cal formulations of cyclophosphamide which are essen-
`tially comparable to CYTOXAN
`While cyclophosphamide comprises the monohy-
`drate drug form, which is the easiest to isolate and with
`which to work, the anhydrous form also exists. No
`other hydrate form has been reported. As used herein,
`the term “cyclophosphamide” refers generically to the
`drug substance (i.e., either the monohydrate or the
`anhydrous form) the term “cyclophosphamide mono-
`hydrate” refers specifically to the monohydrate and the
`term “anhydrous cyclophosphamide” refers to the an-
`hydrous form. The monohydrate form is preferred for
`pharmaceutical processing, as the anhydrous form
`readily picks up water to form the monohydrate when
`exposed to a relative humidity of about 20-30% or
`higher at about 25° C. While the monohydrate is stable,
`nonetheless, under dry conditions (relative humidities
`of about 20% or less) the monohydrate begins to lose
`this water of hydration which can cause problems in
`manufacture. Because of stability limitations which may
`be due in part to ready interconversion between the
`anhydrous and monohydrate forms, it is recommended
`that storage temperatures for cyclophosphamide prod-
`ucts not exceed 30° C. (86° F.), and preferably be stored
`at about 25° C. (77° F.).
`Currently, the parenteral dosage formulations of cy-
`clophosphamide consist of sterile packaged dry powder
`blend admixtures of cyclophosphamide monohydrate
`and sodium chloride. These premixes are dissolved in
`water prior to administration which can be oral as well
`as parenteral. It is intended that the solution itself be
`administered promptly after being prepared but it is
`satisfactory for use up to several hours after prepara-
`tion. During processing and/or storage of the present
`dry powder premix formulation, a glassiness and/or
`stickiness can be acquired by the premix composition
`giving an unattractive material with inferior solubility
`characteristics and decreased potency. This deteriora-
`tion is more pronounced as storage time is extended or
`
`10
`
`15
`
`20
`
`25
`
`30
`
`35
`
`40
`
`45
`
`50
`
`55
`
`60
`
`65
`
`4,537,883
`
`2
`if the upper limit of the storage temperature range is
`exceeded.
`
`A common practice used with constitution of sterile
`solids by a suitable aqueous vehicle consists of warming
`the container to expedite the dissolution process, espe-
`cially when the solids dissolve slowly. A study examin-
`ing the effect of briefly heating cyclophosphamide solu-
`tions was reported by D. Brooke, et al. in American
`Journal of Hospital Pharmacy, 32:44-45 (1975). This
`study concluded that warming vials of cyclophospha-
`mide in order to facilitate dissolution after adding an
`aqueous vehicle could decrease the potency of the final
`injectable product. In summary, these stability limita-
`tions and dissolution difficulties can often result in clini-
`cal usage of subpotent cyclophosphamide solutions.
`Therefore, the main objective of the project culmi-
`nating in the instant invention was to provide a cyclo-
`phosphamide dosage form with improved solubility
`characteristics and enhanced appearance, while main-
`taining stability comparable to the dry pre-mix compo-
`sition. Unexpectedly, the unique lyophilized solid com-
`position discovered has improved stability over the previ-
`ous dry pre-mix composition.
`Problems with unstable drug solutions have been
`handled previously by practitioners in the pharmaceuti-
`cal arts by applying lyophilization, cf: L. Lachman, et
`al., “The Theory and Practice of Industrial Pharmacy”,
`2nd Ed., Lea & Febiger, Philadelphia, PA., pp. 521-524
`(1976); “Remington’s Pharmaceutical Sciences”, 15th
`Ed., Mack Publishing Co., Easton, PA., pp. 1483-1485
`(1975). The technique known as lyophilization is often
`employed for injectable pharmaceuticals which exhibit
`poor stability in aqueous solution. This process involves
`freeze drying, whereby ice is sublimed from frozen
`solutions leaving only the solid, dried components of
`the original liquid.
`The particular advantages of lyophilization are that
`biologicals and pharmaceuticals which are unstable in
`aqueous solution yet relatively stable in the solid state
`can be processed and filled into dosage containers in
`solution, taking advantage of the relative ease of pro-
`cessing a liquid; dried without elevated temperatures,
`thereby eliminating adverse thermal effects; and then
`stored in the dry state in which there are relatively few
`stability problems. Additionally, freeze dried products
`are often more soluble and/or more rapidly solubilized,
`dispersions are stabilized, and products subject to deg-
`radation by oxidation or hydrolysis are protected.
`Pharmaceuticals to be freeze dried are usually in
`aqueous solution ranging from 0.01 to 40% in concen-
`tration of total solids. Final moisture content of the
`dried product is generally below 1.0% although some
`products, mainly biologicals, may have a final moisture
`content which could range as high as about 10%. Usu-
`ally, the improvement in stability of the lyophilizate,
`compared to the solution, is due to the absence of water
`in the pharmaceutical composition.
`The active constituent of many pharmaceutical prod-
`ucts, though, is present in such small quantity that if
`freeze-dried "alone, it may not give a composition of
`suitable bulk and, in some cases, its presence would be
`hard to detect visually. Therefore, excipients are often
`added to increase the amount of solids present. In most
`applications it is desirable for the dried product cake to
`occupy essentially the same volume as that of the origi-
`nal solution. To achieve this, the total solids content of
`the original solution is usually about 10 to 25%. Among
`
`FRESENIUS KABI 1006-0002
`
`

`
`4,537,883
`
`3
`substances found useful for this purpose, often in combi-
`nation, are sodium or potassium phosphates, citric acid,
`tartaric acid, gelatin, lactose and other carbohydrates
`such as dextrose, mannitol and dextran; and on occa-
`sion, preservatives. Various excipients contribute ap-
`pearance characteristics to the cake, such as whether
`dull and spongy or sparkling and crystalline, firm or
`friable, expanded or shrunken, and uniform or striated.
`Therefore, formulation of a composition to be freeze
`dried must include consideration not only of the nature
`and stability characteristics required during the liquid
`state, both freshly prepared and when reconstituted
`before use, but the characteristics desired in the final
`lyophilized cake. Additionally, for products to be re-
`constituted for parenteral usage, consideration must
`also be given to the pharmacological effects of excipi-
`ents chosen. In some instances,
`there may even be
`chemical interaction between the active ingredient and
`one or more of the excipients during processing. This
`could, of course, result in reduced potency of the fin-
`ished product.
`For all the above reasons, it becomes apparent that
`selection of a suitable excipient or excipients for a phar-
`maceutical product containing a reactive,
`thermally
`labile, and inherently unstable ingredient such as cyclo-
`phosphamide is not an obvious process. Considerable
`testing, including drug assay, would be required, as in
`the instant case, for the development of such a composi-
`tion. We surprisingly found that only the use of manni-
`tol as the primary excipient gave a far superior lyophili-
`zate compared to lyophilizates obtained using other
`excipients. Unexpectedly, we also discovered that a
`lyophilized cyclophosphamide solid composition con-
`taining about 4-% moisture gave a product with superior
`thermal stability, compared to currently available dry
`powder premixes, lyophilized cyclophosphamide solid
`compositions with moisture levels of about 1% or less,
`or even cyclophosphamide itself.
`SUMMARY OF THE INVENTION
`
`This invention concerns an improved solid pharma-
`ceutical composition and is based in part upon the dis-
`covery that a lyophilized cyclophosphamide-mannitol
`solid composition has improved thermal stability when
`it contains an amount of water approximately equimolar
`to the cyclophosphamide content taken as the anhy-
`dride. Of equal importance is the discovery that the
`desirable physical properties of the solid composition
`appear to be achieved only by using mannitol as the major
`excipient. Two processes have been developed which
`allow facile production of the solid composition in vials
`for packaging as unit dosage forms for reconstitution as
`sterile solutions.
`
`DETAILED DESCRIPTION OF THE
`INVENTION
`
`The therapeutically-active component of this inven-
`tion, cyclophosphamide,
`is a well known and widely
`used anticancer agent. Cyclophosphamide chemically is
`2-[bis-(2-chloroethyl)amino]tetrahydro-2H-l,3,2-oxaza-
`phosphorine-2-oxide monohydrate, shown in Formula I
`
`10
`
`15
`
`20
`
`25
`
`30
`
`35
`
`45
`
`50
`
`55
`
`60
`
`Cl
`
`I
`
`4
`
`21.1
`
`.l-I20
`
`It is appreciated by the practitioner that the degree of
`reactivity desired for efficacy in an agent of this sort
`necessarily acts as a limit to its inherent stability in
`aqueous solution. Mainly for this reason, cyclophospha-
`mide is compounded as a sterile dry powder mixture of
`cyclophosphamide monohydrate and NaCl for constitu-
`tion with sterile water just prior to administration.
`Some instability of cyclophosphamide dry powder
`mixes can be evidenced by change in physical proper-
`ties, e.g. decreased rate of dissolution, change in appear-
`ance, and the like, as well as loss of potency on storage.
`This deterioration becomes more pronounced on ex-
`tended storage or on exposure of the powder mix to
`temperatures above about 90° F., a temperature not
`uncommon during commercial transportation or ware-
`house storage. Freshly prepared solutions made up from
`this powder mix can exhibit lowered potency due not
`only to prior degradation of the active ingredient but
`also through incomplete dissolution of the powder mix
`as well.
`The instant invention has resulted from work under-
`
`taken to ascertain if physical properties, especially dis-
`solution rate, could be improved in a lyophilized com-
`position containing anhydrous cyclophosphamide.
`Cyclophosphamide was thoroughly tested with those
`excipients compatible with parenteral administration to
`determine if a suitable lyophilizate cake could be
`formed by freeze-drying. Following the lyophilization
`process, the resulting cake was evaluated visually on its
`physical appearance using as desired criteria: original
`shape, no shrinkage or meltback, good coloration, ho-
`mogeneity, firmness, and crystallinity. The dissolution
`rate was then tested and a global score of poor, fair, or
`good, based on the above criteria, was assigned, cf:
`Table 1.
`
`TABLE 1
`
`Excipient
`Lactose
`
`Dextrose
`Tartaric acid
`Urea
`L-arginine
`Polyvinylpyrro-
`lidone (PVP)
`KHZPO4
`K2]-IP04
`Na2CO3
`NaHCO3
`Nal-ICO3
`Mannitol
`
`Excipient Testing For Cake Formation
`Pre-lyophilization Solution:
`Lyophilization
`Concentration (% W/V)
`Result
`1.0, 2.0, 2.5, 3.3, 3.5,
`Poor cakes
`5.0, 6.7, 8.0
`2.0, 2.5
`2.0
`1.0, 2.0, 2.5
`0.5
`3.0
`
`Poor cakes
`Poor cake
`Poor cakes
`Poor cake
`Poor cake
`
`1.0
`1.0
`1.0
`1.0, 1.25, 1.5
`1.75, 2.0, 2.5
`1.7, 2.0, 2.5, 3.0, 4.0,
`5.0, 6.0, 7.0
`10.0, 15.0
`2.5, 3.0, 5.0
`
`Poor cake
`Poor cake
`Poor cake
`Poor to fair cakes
`Good cakes
`Good cakes
`
`Poor cakes
`Poor cakes
`
`Mannitol
`Cl-l3CO2K
`
`65
`
`Each solution for lyophilization contained 3.57%
`(W/V) cyclophosphamide monohydrate and an excipi-
`
`FRESENIUS KABI 1006-0003
`
`

`
`4,537,883
`
`6
`
`TABLE 2-continued
`
`5
`ent or excipients. The solutions were prepared using
`Water for Injection and were filtered through a prefilter
`and a 0.22 pm filter membrane. The solutions were
`filled into 30 mL serum vials with a fill volume of 15 mL .
`of solution per vial (the equivalent of 500 mg of anhy-
`drous cyclophosphamide).
`Prior to lyophilization, the filled vials were prechilled
`to 5° C. in a refrigerator. The vials were then frozen in
`the lyophilization chamber for approximately 2 hrs to a
`temperature of about -—35° C. After cooling the con-
`denser to about -60“ C., the lyophilizer chamber was
`evacuated to approximately 50 millitorr. An initial shelf
`temperature of 0° C. was maintained for the first 14-16
`hrs of the lyophilization cycle. The shelf temperature
`was then raised to 25° C. The cycle was completed
`when the product temperature stabilized near the shelf
`temperature, usually requiring 2-6 more hours, and the
`vacuum pressure reached equilibrium at a reading of
`approximately 100 millitorr. During lyophilization,
`product temperature was monitored using themocouple
`probes placed inside several vials.
`After the lyophilization process was completed, the
`material remaining in the vial was observed for color,
`appearance, texture, friability, and shrinkage from the
`original frozen volume. Also, each formulatin was
`tested for its moisture loss on drying (4 hrs at 70° C.
`under vacuum) and its dissolution characteristics upon
`reconstitution with 25 mL of Sterile Water for Injec-
`tion.
`
`5
`
`10
`
`15
`
`20
`
`25
`
`30
`
`Unexpectedly, only two excipients from this group
`(Table 1) exhibited favorable cake-forming ability in the
`presence of cyclophosphamide. These two prototype
`formulations (containing NaHCO3 or mannitol)
`re-
`tained their original shape, possessed suitable texture
`and appearance, and dissolved easily upon reconstitu-
`tion with water. Most of the other excipient formula-
`tions were unsuitable due to poor quality cakes due
`mainly to shrinkage and/or poor dissolution. As can be
`seen from Table 1, only mannitol, at concentrations of
`7% W/V or below, and sodium bicarbonate were ini- 40
`tially found to be acceptable excipients in the cyclo-
`phosphamide lyophilization process. The formulations
`containing these two excipients dissolved easily within
`a few seconds upon reconstitution with water.
`Further experimentation utilizing the same experi-
`mental procedure to investigate lyophilization results
`using mixed excipients demonstrated the surprising
`finding that fair to good lyophilizate cakes could be
`obtained only with mannitol compared to lactose or
`sodium bicarbonate as a primary excipient; cf: Table 2. 5°
`
`35
`
`45
`
`Combined Excipient Testing
`
`Na citrate (0.5)
`
`Mannitol (2.0)
`
`Na ascorbate (0.33)
`
`Na acetate (0.33)
`
`Na citrate (0.33)
`
`Na2CO3 (0.5)
`Na}-[CO3 (0.5)
`PVP (1.0)
`
`Primary Excipient
`(Concentration
`Secondary Excipient
`Lyophilization
`
`% W/V)
`(Concentration % W/V)
`Result
`Nal-[CO3 (2.0)
`Glycine (0.5)
`Poor cake,
`very fragile
`poor cake,
`very fragile
`Poor cake,
`shrinkage
`Poor cake,
`slow dissolution
`Poor cake,
`shrinkage
`No cake
`No cake
`Fair cake,
`sl. shrinkage
`Poor cakes,
`shrinkage
`Poor cake,
`slow dissolution
`Poor cakes,
`unhomogenous
`Fair to good
`cakes
`Fair cake,
`sl. shrinkage
`Na citrate (0.33) Good cake
`
`Mannitol (2.5)
`
`Na citrate (0.33, 0.5, 0.67)
`
`L-arginine (0.5)
`
`Glycine (0.5, 1.0)
`
`PVP (0.5, 1.0)
`
`Mannitol (3.0)
`
`Glycine (0.5)
`
`
`
`Most of the compositions listed in Table 2 exhibited
`some ability to form lyophilizate cakes except those in
`which lactose was the primary excipient; however,
`most of the resulting cakes were of poor quality due to
`friability, shrinkage and/or poor dissolution. The lyo-
`philizates containing various secondary excipients with
`sodium bicarbonate were of poorer quality than the
`lyophilizates with sodium bicarbonate as the only excip-
`ient. One formulation containing mannitol as the pri-
`mary excipient and sodium citrate and a second formu-
`lation containing mannitol as the primary excipient and
`polyvinylpyrrolidone (PVP) formed acceptable lyo-
`philizates thate were firm, homogenous cakes with min-
`imum friability and shrinkage. These two combination
`excipient products dissolved easily in a few seconds
`upon reconstitution with water. These selected lyophi-
`lized solid compositions all had low moisture levels, as
`measured by loss on drying, on the order of about 2%
`or less. While considerable improvement of the dissolu-
`tion properties and physical appearance was achieved
`with these low moisture cyclophosphamide lyophili-
`zates, six-week stability testing indicated that these ly-
`ophilized solid compositions were heat sensitive and, in
`fact, much less stable than the existing sodium chloride/-
`cyclophosphamide monohydrate dry powder premix-
`ture.
`
`
`
`TABLE 2
`
`Combined Excipient Testing
`
`Nal-lCO3 (1.5)
`
`Primary Excipient
`(Concentration
`Secondary Excipient
`Lyophilimtion
`
`% W/V)
`(Concentration % W/V)
`Result
`Lactose (2.0)
`Mannitol (1.0, 1.5)
`No cakes
`Lactose (3.3)
`Citric acid, anhyd. (1.0)
`No cake
`Na2CO3 (1.0)
`No cake
`Na}-ICO3 (1.25)
`No cake
`NaCl (0.25)
`Poor cake,
`shrinkage
`Poor cakes,
`shrinkage
`Poor cake,
`shrinkage
`Poor cake,
`shrinkage
`Poor cakes,
`very fragile
`
`Glycine (0.25; 0.5)
`
`K2HPO4 (0.5)
`
`Citric acid, anhyd. (0.5)
`
`Lactose (0.5, 1.0, 1.5)
`
`55
`
`60
`
`55
`
`It was unexpectedly discovered, however, that hu-
`midification of these low moisture lyophilized cyclo-
`phosphamide solid compositions (“dry” lyophilizates)
`could be easily and reproducibly effected by subjecting
`them to an air atmosphere of about 60 to 80% relative
`humidity at about 25° C. in a closed container for 1-2
`days. Under these conditions, a state of equilibrium
`could be achieved in which the moisture gained on a
`molar basis was approximately equivalent to the anhy-
`drous cyclophosphamide content of the lyophilizate.
`The improved dissolution characteristics found in the
`“dry” cyclophosphamide lyophilizates remained essen-
`tially unchanged upon humidification. Stability testing
`of the humidified solid compositions (“hydrated” lyo-
`philizates) demonstrated that those compositions con-
`
`FRESENIUS KABI 1006-OOO4
`
`

`
`7
`taining mainly mannitol as the excipient were unexpect-
`edly superior, compared with the sodium bicarbonate
`excipient compositions in maintaining minimal dissolu-
`tion times, cf: Table 3.
`TABLE 3
`Comparative Stability Testing of “I-lydrated" Solid
`Lyophilizates: Mannitol as excipient vs Nal-[CO3 as excipient
`Storage
`Time
`Dissolution Times (sec)
`Temperature
`(Wks)
`Mannitol
`Nal-[CO3
`40°
`3
`~ 20
`> 180
`6
`~ 60
`> 180
`12
`- 60
`~ 120
`3
`~20
`> 180
`6
`~ 60
`> 180
`12
`~ 60
`~ 120
`6
`~ 60
`> 180
`12
`~ 60
`> 180
`
`35°
`
`25'’
`
`Preferred solid compositions of the instant invention
`are comprised of about 20 parts by weight of cyclophos-
`phamide, taken as the anhydrous form, about 15-2 parts
`by weight of water and from about 10-40 parts by
`weight of excipient which is primarily mannitol. Most
`preferred compositions are comprised of about 20 parts
`by weight of cyclophosphamide,
`taken as the anhy-
`dride, about 1% parts by weight of water and about 15
`parts by weight of excipient consisting essentially of
`mannitol.
`These solid compositions may be prepared by either
`of two methods. The first method, of’ which one em-
`bodiment as mentioned hereinabove, involves lyophili-
`zation and humidification. Common to both methods is
`preparation of a sterile solution for lyophilization. A
`typical solution is composed of about 1 part by weight
`1 excipient which is mainly mannitol, about 1.5 parts by
`weight cyclophosphamide (as the monohydrate), and
`about 40 parts water. In order to obtain optimal results
`with the lyophilizate composition this solution should
`not contain much more than about 4% cyclophospha-
`" mide content. Continuing with the first method,
`this
`solution is aseptically filled into suitable containers and
`then lyophilized to a “dry” lyophilizate cake which has
`a moisture content on the order of about 2% or less.
`This lyophilization is done in a short period of time
`(~24 hrs) at a lyophilization chamber pressure of about
`100 to 500 millitorr using a very cold condenser temper-
`ature (about —-60° C.) and a warm shelf temperature
`(about 20°—25° C.). This typical “dry” lyophilizate is
`then humidified to give a “hydrated” lyophilizate con-
`taining approximately 4% moisture. With humidif1ca-
`tion at atmospheric pressure, further water pick-up
`essentially stops at this point giving the stable “hy-
`drated” lyophilizate solid composition of the instant
`invention. The amount of water taken up is approxi-
`mately equimolar ranging to about 30% excess com-
`pared with the cyclophosphamide (on an anhydrous
`basis) present in the cake. While the humidification can
`be done at approximately atmospheric pressure,
`the
`time required for complete humidification is considera-
`bly shortened (to at least a third or less) when done
`under reduced pressure. By reduced pressure is meant a
`pressure of about 0.1 to 25 torr. A unit of 1 torr is equiv-
`alent to 1/760 standard atmospheric pressure.
`A second method of obtaining the desired lyophilized
`solid composition involves lyophilizing the sterile solu-
`tion directly using a longer lyophilization cycle (~48
`hrs). This method, done more slowly under milder con-
`ditions, requires adjustment of certain parameters of the
`lyophilization process such as shelf temperature, and
`
`10
`
`15
`
`20
`
`25
`
`30
`
`35
`
`45
`
`50
`
`55
`
`60
`
`65
`
`4,537,883
`
`8
`condenser temperature during the process of drying.
`This second lyophilization process comprises 2 stages of
`freeze-drying done under a 500 millitorr chamber pres-
`sure. The first stage of freeze-drying is done with a low
`condenser temperature of about -60“ C., and a shelf
`temperature at about 10° C. The second stage of freeze-
`drying requires raising the condenser temperature to
`about —30° C. and lowering the shelf temperature to
`about — 10° C.
`
`These lyophilized cyclophosphamide solid composi-
`tions may be provided in single dose container forms by
`aseptically filling suitable containers with the sterile
`pre-lyophilization solution to a prescribed cyclophos-
`phamide content; preparing the desired lyophilized
`solid composition, using either of the two methods de-
`scribed hereinabove; and then hermetically sealing the
`single dose container. It is intended that these filled
`containers will allow rapid dissolution of the solid com-
`position upon reconstitution with water in situ giving an
`appropriate sterile solution of desired cyclophospha-
`mide concentration for administration. By suitable con-
`tainers is meant a container capable of maintaining a
`sterile environment such as a vial capable of being her-
`metically sealed by a stopper means. Additionally, suit-
`able containers implies appropriateness of size, consid-
`ering the volume of solution to be held upon reconstitu-
`tion of the lyophilized composition; and appropriate-
`ness of container material, generally Type I glass. The
`stopper means employed, e. g. sterile rubber closures or
`an equivalent, should be understood to be that which
`provides the aforementioned seal but which also allows
`entry for the purpose of introduction of diluent, e.g.
`sterile water, the reconstitution of the desired cyclo-
`phosphamide solution. These and other aspects of the
`suitability of containers for pharmaceutical products
`such as those of the instant invention are well known to
`those skilled in the practice of pharmaceutical arts.
`While the physical properties, such as appearance
`and particularly dissolution time, were improved in the
`instant solid compositions, thereby achieving one objec-
`tive of the invention, we unexpectedly found that these
`instant solid compositions also possessed improved ther-
`mal stability compared with currently known dry pow-
`der premix formulations as well as cyclophosphamide
`itself.
`In practice, expectation for enhancement of
`chemical stability by lyophilization relates to a compari-
`son of the lyophilizate solid with the solution form of
`the pharmaceutical composition. In contrast, the instant
`compositions demonstrate enhanced chemical stability
`between solid dosage forms, cf: Table 4.
`TABLE 4
`
`Comparative Chemical Stabilities for “Hydrated"
`Cyclophosphamide/Mannitol Lyophilizate and
`CYTOXAN Q for Injection {Powder Premixg
`Storage
`Time
`Potency 1% of Zero-time)
`Temperature
`(Wks)
`Lyophilizate
`CYTOXAN ®
`50°
`1
`31
`31
`45°
`1
`94
`80
`2
`76
`42
`1
`100
`98
`3
`101
`85
`6
`99
`33
`I2
`98
`2
`3
`101
`105
`6
`100
`98
`12
`100
`98
`26
`98
`85
`6
`100
`101
`12
`100
`101
`
`35°
`
`25°
`
`40°
`
`FRESENIUS KABI 1006-OOO5
`
`

`
`4,537,883
`
`9
`
`TABLE 4-continued
`Comparative Chemical Stabilities for “Hydrated”
`Cyclophosphamide/Mannitol Lyophilizate and
`CYTOXAN Q for Injection (Powder Premixg
`Storage
`Time
`Potency 5% of Zero-time}
`Temperature
`(Wks)
`Lyophilizate
`CYTOXAN ®
`26
`100
`103
`
`
`
`5
`
`10
`
`I5
`
`20
`
`25
`
`30
`
`35
`
`45
`
`50
`
`55
`
`60
`
`65
`
`10
`final volume of this solution to 15 liters. The solution is
`first passed through a sterile prefilter and then a sterile
`0.22 pm pore size membrane filter, following which the
`sterile solution is then aseptically filled into sterile 30
`mL glass vials. Each vial is filled with 15.0 mL solution
`(containing the equivalent of 500 mg anhydrous cyclo-
`phosphamide) and then sterile rubber closures are asep-
`tically inserted in the lyophilization position. These
`filled vials are placed in suitable lyophilization equip-
`ment and cooled as quickly as possible to a temperature
`of about -35” C.
`The lyophilizer condenser is then cooled to about
`-60” C. and the lyophilization chamber is evacuated to
`a pressure of approximately 500 millitorr. Shelf heat is
`set at +20° C. to begin the drying process. Following
`about 15-20 hours of primary drying, the shelf tempera-
`ture is raised in order to bring the product temperatures
`to near +25° C. provided that the product temperature
`thermocouples have reached + 15° to + 18° C. and the
`lyophilization chamber pressure is not more than 500
`millitorr at that point. Lyophilizatin is then continued
`until a final product temperature of +22° to +24° C.
`with a chamber pressure of not more than about 200
`millitorr, is reached. Depending on the capacity of the
`lyophilization equipment and the product batch size, the
`total lyophilization time will vary but is generally in the
`range of 24-36 hours. Following completion of the
`lyophilization process, the high vacuum is relieved by
`the aseptic introduction of sterile air and/or nitrogen
`having 80% relative humidity at room temperature. An
`aseptic environment is maintained during this rehydra-
`tion process which is complete within approximately
`18-24 hours at atmospheric pressure. At this point, the
`vials are closed by aseptically seating the lyophilization
`stoppers into the vials by mechanical collapse of the
`shelves.
`
`A preferred method of humidification involves intro-
`ducing water vapor into the lyophilization chamber
`while it remains under reduced pressure. With the
`chamber at about 25° C. and with a closed vacuum
`(pump off) of about 100-200 millitorr, a quantity of
`sterile water is vaporized into the chamber during
`which time the pressure may increase to about 25 torr.
`The quantity of sterile water used depends on the size of
`the lyophilization equipment being used as well as the
`product batch size. Typically, for 180 vials of the size
`and content described hereinabove and with a lyophili-
`zation chamber volume of approximately 125 liters, the
`amount of water used is on the order of 9 mL. The
`introduction of water is accomplished by establishing a
`closed system and bleeding in the water vapor issuing
`from a reservoir of liquid water kept at about 60°—80° by
`external heating. Boiling chips are used to prevent
`bumping and the introduction of water is completed
`within approximately 45 minutes. The internal chamber
`pressure during this “vacuum humidification” process
`varies from about 0.1 to 25 torr with the introduction of
`
`the water vapor. Total time of humidification using this
`method generally requires about 2-6 hours, again de-
`pending on lyophilization equipment size and product
`batch size. After humidification, the remaining vacuum
`is relieved by introduction of sterile air and/or nitrogen
`and the vials are closed by aseptically seating the lyoph-
`ilization stoppers into the vials by mechanical collapse
`of the shelves.
`
`FRESENIUS KABI 1006-0006
`
`It is recognized by practitioners in the pharmaceuti-
`cal arts that the use of elevated storage temperatures for
`shorter time periods is standard practice in obtaining
`comparative stability data. As can be seen in Table 4,
`the “hydrated” cyclophosphamide/mannitol
`lyophi-
`lized solid composition possesses greater thermal stabil-
`ity than the current CYTOXAN® for Injection dry
`powder formulation.
`Short term comparative stability studies have also
`been made using the “hydrated” cyclophosphamide/-
`mannitol lyophilized solid composition, a cyclophos-
`phamide monohydrate/mannitol dry powder blend (not
`lyophilized), CYTOXAN ® for Injection dry powder
`premix, and pure cyclophosphamide monohydrate.
`These studies at a storage temperature of 40° C. gave
`the results summarized in Table 5.
`
`TABLE 5
`Comparative Stabilities at 40° C. for:
`“Hydrated” Cyclophosphamide/Mannitol Lyophilizate
`Cyclophosphamide Monohydrate/Mannitol Powder Blend
`CYTOXAN ® for Injection (Powder Premix)
`. Cyclophosphamide Monohydrate
`Potency
`Physical Appearance
`g% of Zero-time!
`B
`C
`D
`A
`B C D A
`s1.
`s1.
`un-
`104
`96
`93
`95 un-
`changed melted melted
`changed
`un-
`melted melted melted
`changed
`un-
`melted melted
`4 —
`42
`101
`12
`
`changed
`
`102
`
`57
`
`39
`
`37
`
`UOP’?
`
`Time
`(Wks)
`3
`
` 6
`
`As these tabular results show, the “hydrated” cyclo-
`phosphamide/marmitol
`lyophilized solid composition
`demonstrates superior thermal stability as well as main-
`tenance of initial physical appearance. In summary, the
`“hydrated” cyclophosphamide/mannitol
`lyophilized
`solid composition shows unexpected improved stability,
`superior solubility characteristics and enhanced appear-
`ance when compared with other powder premix formu-
`lations of cyclophosphamide monohydrate and cyclo-
`phosphamide monohydrate alone. These improved
`characteristics are indicative of a superior pharmaceuti-
`cal solid composition.
`The following examples describe in detail methods
`for prep