多晶型药物指定晶型筛选方案设计

Development of a Targeted Polymorph Screening Approach for a Complex Polymorphic and Highly Solvating APIANTHONY M.CAMPETA,BRIAN P.CHEKAL,YURIY A.ABRAMOV,PAUL A.MEENAN,MARK J.HENSON,BING SHI, ROBERT A.SINGER,KEITH R.HORSPOOLPharmaceutical Sciences,Pfizer Global Research&Development,Groton,Connecticut06340Received19February2010;revised21April2010;accepted22April2010Published online22June2010in Wiley InterScience().DOI10.1002/jps.22230ABSTRACT:Elucidation of the most stable form of an active pharmaceutical ingredient(API)isa critical step in the development process.Polymorph screening for an API with a complexpolymorphic profile can present a significant challenge.The presented case illustrates anextensively polymorphic compound with an additional propensity for forming stable solvates.In all,5anhydrous forms and66solvated forms have been discovered.After early polymorphscreening using common techniques yielded mostly solvates and failed to uncover several keyanhydrous forms,it became necessary to devise new approaches based on an advanced under-standing of crystal structure and conformational relationships between forms.With the aid ofthis analysis,two screening approaches were devised which targeted high-temperature deso-lvation as a means to increase conformational populations and enhance overall probability ofanhydrous form production.Application of these targeted approaches,comprising over100experiments,produced only the known anhydrous forms,without appearance of any new forms.The development of these screens was a critical and alternative approach to circumventsolvation issues associated with more conventional screening methods.The results providedconfidence that the current development form was the most stable polymorph,with a lowlikelihood for the existence of a more-stable anhydrous form.ß2010Wiley-Liss,Inc.and theAmerican Pharmacists Association J Pharm Sci99:3874–3886,2010Keywords:axitinib;polymorphism;polymorph screen;crystallization;desolvation;compu-tational chemistry;crystal structureINTRODUCTIONPolymorphism can be thought of as the state in which a solid chemical compound exists in more than one crystalline form1with only one polymorph being the thermodynamically most stable form at a specific temperature and pressure.This phenomenon is very common to most pharmaceutical APIs.It is well known that polymorphs of the same substance can have dramatic differences in pertinent pharmaceu-tical properties,such as solubility and stability that can often have a significant impact on bioavailability and overall drug product performance.A number of excellent texts on polymorphism and their influence on pharmaceutical development are available.2Thus,identifying the most appropriate solid form,typically the thermodynamically stable form,is a key element in the early developmental process for a new drug candidate.In regard to polymorph screening approaches,there are many common techniques employed that are designed to typically uncover all metastable and low-energy polymorphic forms.These include,for example,crystallizations through solvent evaporations,antisolvent crystallizations,slow and fast cooling of saturated API solutions to induce precipitation,and slurrying of solid API for extended periods of time.3A significant number of solvents and cosolvents of varying polarity and chemical composi-tion are usually employed,while variable tempera-tures are also incorporated in the design to assess enantiotropic behavior.These approaches have been incorporated into our practices for polymorph screen-ing and are typical throughout the pharmaceutical industry.In most cases,a thoroughly designed API form screen employing the approaches previously dis-cussed should typically identify the thermodynami-cally stable polymorph.Additional Supporting Information may be found in the online version of this article.Mark J.Henson’s present address is Molecular Biometrics,Inc., New Haven,CT06511.Correspondence to:Anthony M.Campeta(Telephone:860-441-5844;fax:860-715-2454;E-mail:anthony.m.campeta@pfi) Journal of Pharmaceutical Sciences,Vol.99,3874–3886(2010)ß2010Wiley-Liss,Inc.and the American Pharmacists Associationexceptions in the literature4,5that describe the appearance of a lower energy form at late stages in development.In these cases,common polymorph screening approaches were clearly unsuccessful,as unique physical and structural properties of the molecule hindered the anticipated cascade to the most stable form based on Ostwald’s rule.6In this article, we describe another such example.Axitinib(Fig.1)is an oncology candidate under development at Pfizer.This API targets the vascular endothelial growth factor(VEGF)to prevent the growth and proliferation of cancer cells via interrup-tion of tumor angiogenesis(formation of vascular supply tissue).7This compound has shown consider-able promise in the treatment of carcinomas in a number of target tissues and organs and is currently in late stage clinical development.8Understanding the polymorphism of axitinib has been a subject of considerable focus and effort,which we have initially reported.9In this work,polymorph investigations using the traditional approaches pre-viously mentioned,which incorporated well over 300experiments,identified a surprisingly high total of23unique solid forms.Distinction of these forms was assigned on the basis of powder X-ray diffraction (PXRD)patterns and thermal characteristics such as melting onset,melting enthalpy,and desolvation temperatures.This group of solid forms included three anhydrous forms with the remainder solvates (refer to Tab.1).The anhydrous form IV was characterized as a robust developmental form with acceptable solid-state properties and was advanced for early clinical studies.It was apparent that axitinib had a high tendency to form solvates.There was some question whether certain polymorph screening approaches may be challenged by this phenomenon,and the risk of not observing critical anhydrous forms(of which three had been already discovered)could exist.In parti-cular,axitinib had a propensity to form relatively stable solvated structures,as a majority of these solvates were characterized as possessing relatively high temperatures of desolvation(desolvation tem-peratures significantly higher than thenormalFigure1.Structure of axitinib.Table1.Summary of Axitinib Solid FormsName Form SolventForm I(1)Anhydrate—Form II(2)Hydrate WaterForm III(3)Solvate Ethyl acetate(EtOAc) Form IV(4)Anhydrate—Form V(5)No solid form designationForm VI(6)Anhydrate—Form VII(7)Solvate Isopropyl alcohol(IPA),IPA/waterForm VIII(8)Solvate Dioxane,tetrahydrofuran(THF) Form IX(9)Hydrate WaterForm X(10)Solvate Dimethylformamide(DMF),DMF/waterForm XI(11)Solvate THF/water,THFForm XII(12)Solvate Dichloromethane(DCM),ethanol(EtOH)Form XIII(13)Solvate Acetonitrile(ACN)Form XIV(14)Solvate Acetic acidForm XV(15)Solvate EtOHForm XVI(16)Solvate IPAForm XVII(17)Solvate AcetoneForm XVIII(18)Solvate Methylisobutyl ketone(MIBK) Form XIX(19)Solvate Methylethyl ketone(MEK) Form XX(20)Solvate Methyl benzoateForm XXI(21)Solvate2,2,2-CF3CH2OH/ether/hexane Form XXII(22)Solvate1-PentanolForm XXIII(23)Solvate PyridineForm XXIV(24)Solvate ChloroformForm XXV(25)Anhydrate—Form XXVI(26)Solvate THF/water,THFForm XXVII(27)Solvate Dimethylsulfoxide(DMSO) Form XXVIII(28)Solvate Benzyl alcoholForm XXIX(29)Solvate TrichloroethyleneForm XXX(30)Solvate Dimethylformamide(DMF)/octanol(1:1)Form XXXI(31)Solvate OctanolForm XXXII(32)Solvate MethanolForm XXXIII(33)Solvate1-ButanolForm XXXIV(34)Solvate3-Methyl-1-butanolForm XXXV(35)Solvate MEKForm XXXVI(36)Solvate Pyrrole/1-pentanolpyrrole/p-cymeneForm XXXVII(37)Solvate Allyl alcoholForm XXXVIII(38)Solvate Pyrrole allyl alcoholForm XXXIX(39)Solvate Acetic acidForm XL(40)Solvate EtOHForm XLI(41)Anhydrate—Form XLII(42)Solvate2-ButanolForm XLIII(43)Solvate2-Methyl THFForm XLIV(44)Solvate2-Methyl THFForm XLV(45)Solvate TolueneForm XLVI(46)Solvate N-Methylpyrrolidone Form XLVII(47)Solvate Isoamyl acetateForm XLVIII(48)Solvate MethylcyclohexaneForm XLIX(49)Solvate CyclohexanoneForm L(50)Solvate CyclohexanoneForm LI(51)Solvate1,2-DichloroethaneForm LII(52)Solvate Propionic acidForm LIII(53)Solvate Tert-butanolForm LIV(54)Solvate DimethoxymethaneForm LV(55)Solvate2-PentanoneForm LVI(56)Solvate Dimethyl acetate(DMA) Form LVII(57)Solvate NitromethaneForm LVIII(58)Solvate1,2,3,4-tetrahydronaphthalene Form LIX(59)Solvate Tetramethylene sulfone Form LX(60)Solvate Methyl acetateForm LXI(61)Solvate p-XyleneForm LXII(62)Solvate TrichloroethyleneForm LXIII(63)Solvate n-Butyl acetateForm LXIV(64)Solvate Isobutyl alcoholForm LXV(65)Solvate CyclohexanolForm LXVI(66)Solvate Isopropyl acetateForm LXVIII(67)Solvate p-Cymene/pyrrole(1:1) Form LXIX(68)Solvate t-Amyl alcoholForm LXX(69)Solvate4-Methyl-2-pentanone Form LXXI(70)Solvate CyclohexaneForm LXXII(71)Solvate1,2-Dichlorobenzene Form LXXIII(72)Solvate p-Cymene/acetone(1:1)TARGETED POLYMORPH SCREENING APPROACH DEVELOPMENT FOR AN API3875boiling point of the corresponding solvent),suggest-ing strong bonding within the crystal.These solvates may be anticipated to be thermodynamically stable in their corresponding mother liquor and may resist further solvent-mediated transformation to an anhy-drous form.Solvent content was often found to be nonstoichiometric,suggesting nonsite-specific sol-vent incorporation.Also,many solvates formed from different solvent systems demonstrated isomorphic10 structures,as evidenced by very similar PXRD patterns,which made differentiation of solid forms complex.In addition,multiple polymorphs of solvates were observed from the same solvent,such as ethanolate forms XII and XV.During process development and refinement stu-dies with form IV,a new crystalline anhydrous form was discovered.9This new form,form XXV,was characterized as a thermodynamically more stable form than form IV,though it was not observed in any of the aforementioned polymorph screening studies. Crystal structure analysis would eventually show that the molecular conformation of most of the solvate structures was very similar to that of the anhydrous form IV.This coincides with the observation that the solvates would desolvate to form IV during DSC and variable temperature PXRD heating experiments. This suggested that the molecular conformation is mostly retained upon desolvation,resulting in the favored formation of form IV.Equally important was thefinding that the crystal packing and molecular conformation of the new form XXV were found to be different from those of form IV.In particular,the molecular arrangement of form XXV generally would be difficult to achieve through desolvation.Thus,it is believed the propensity to initially form stable solvates as well as structural properties that would favor desolvation to form IV contributed to the lack of appearance of form XXV in our polymorph screens.Though there was confidence that form XXV was the thermodynamically stable form of axitinib, another more-stable anhydrous form,form XLI, was later discovered during development.It became obvious that the polymorph environment of axitinib could not be confidently assessed through the use of conventional polymorph screening experiments.In this article,the discovery and characterization of this new form XLI are described,including a thorough evaluation and understanding of the crystal struc-ture,and its structural relationship relative to known polymorphs.Finally,with this detailed understand-ing of the structural properties,the development of two specific polymorph screening approaches were designed around desolvation to more completely assess the polymorphic environment of axitinib and improve confidence that form XLI was indeed the thermodynamically stable form.EXPERIMENTALMaterialsA sample of form XVI(isopropanol solvate)of axitinib (purity99.8%)was obtained from Pfizer Ireland Pharmaceuticals,Inc.,Little Island,CO.Cork,Ire-land.Samples of anhydrous forms IV,VI,XXV,and XLI were obtained from Pfizer Global Research& Development,Groton,CT.Full details on methods to prepare forms I,IV,and VI are presented by Ye et al.,11whereas methods to prepare forms XXV and XLI are described by Campeta et al.12Solvents used in this study were obtained from Sigma–Aldrich,St Louis,MO,Pharmco-AAPER,Brookfield,CT,or Mallinckrodt Baker,Phillipsburg,NJ.Solubility Measurements of Various Forms in Ethanol Solubility measurements of various forms in ethanol were completed using a100-mL jacketed glass vessel. When operated at temperatures above the normal boiling temperature of ethanol(788C),the vessel was pressurized to suppress boiling.During the solubility measurement,a measured amount of a particular form of axitinib was added to a measured amount (typically30–40g)of ethanol.The mixture was heated quickly to5–108C below the expected dissolu-tion temperature,and then heated slowly(0.25–0.58C/min)until all the solids were dissolved.The dissolution was monitored using a Mettler-Toledo Lasentec S400FBRM probe.Powder X-Ray DiffractionThe PXRD pattern measurement was carried out on a Bruker D5000diffractometer using copper radiation (CuK a,wavelength:1.54056A˚).The tube voltage and amperage were set to40kV and40mA,respectively. The divergence and scattering slits were set at1mm, and the receiving slit was set at0.6mm.Diffracted radiation was detected by a Kevex PSI detector.A theta-two theta continuous scan at2.48/min(1s/0.048 step)from 3.0to4082u was used.An alumina standard was analyzed to check the instrument alignment.Samples were prepared by placing them in a quartz holder.Single Crystal DiffractometryThe single crystal X-ray diffraction measurements were carried out at room temperature on either a Bruker SMART APEX CCD area detector system equipped with a graphite monochromator and sealed tube Cu radiation(1.54178A˚)source or a Bruker FR591rotating anode with Motel Multilayer Optics, Cu radiation(1.54178A˚),equipped with an APEX II detector.Single crystal X-ray diffraction data were collected at room temperature in order to provide a calculated powder pattern that best matches3876CAMPETA ET AL.experimental powder patterns for API samples, which are measured at room temperature.Tempera-ture changes may result in the contraction or elongation of unit cell dimensions to such an extent that the calculated powder patterns at100K and room temperature may be significantly different.All crystallographic calculations were facilitated by the SHELXTL software suite system.In general,hydro-gens bonded to heteroatoms were located by differ-ence Fourier techniques.The remaining hydrogen atoms were placed in idealized positions.The hydro-gen parameters were added to the structure factor calculations but were not refined.Crystals suitable for this analysis were grown through crystallization techniques including evaporations of saturated solu-tions,vapor and liquid diffusion,and antisolvent crystallization.Thermal AnalysisDifferential scanning calorimetry(DSC)was carried out on a Mettler Instruments DSC821e,Columbus, OH.The instrument was calibrated for cell constant and heat capacity using indium and zinc.Samples were prepared by weighing about3mg of into a40m L aluminum pan which was then covered with an aluminum lid.The lid is pierced prior to measure-ment.Data were analyzed using STAR e SW8.10.The experiments started at ambient temperature and heated the sample at58C/min to3008C under a nitrogen gas purge(flow rate was60mL/min). Thermogravimetric analysis(TGA)was carried out on a TA Instruments,New Castle,DE,DSC Q500 V6.7Build203.The instrument was weight cali-brated with200and1000mg weights.Samples were prepared by weighing about6mg of sample into a 50m L aluminum pan.Data were analyzed using Universal Analysis2000for Windows2000/XP version4.2E,Build4.2.0.38.The experiments started at ambient temperature and heated the sample at 108C/min to3008C under a nitrogen gas purge(flow rate was50mL/min).Conventional Polymorph Screen With Form XXVThe polymorph screen consisted of generating solid samples using a variety of techniques and solvents.A total of12solvents and8aqueous cosolvent mixtures were used.Techniques for generating samples included fast evaporations,slow cools of saturated solutions,and slurries consisting of excess solid in saturated solutions at different temperatures.Resul-tant solids were examined under a microscope for birefringence and morphology,and analyzed at a minimum by PXRD.Selected samples were also assessed by thermal methods.Approximately70total experiments were performed.Fast evaporation experiments:Solutions were prepared in various solvents and sonicated to assist in dissolution.The final solutions were typicallyfiltered through a 0.2-m m nylonfilter.The solutions were allowed to evaporate at ambient(20–258C)in a vial.The solids that formed were isolated and analyzed.Slow cooling experiments:Saturated solutions were prepared in various solvents at elevated temperatures(approxi-mately608C)and rapidlyfiltered through a0.2-m m nylonfilter into open vials.The vials were covered and allowed to cool slowly to room temperature.The presence or absence of solids was noted.If there were no solids present,or if the amount of solids was judged too small for PXRD analysis,the vial was placed in a refrigerator overnight.Again,the presence or absence of solids was noted and if there were none, the vial was placed in a freezer overnight.Solids that formed were isolated byfiltration and allowed to dry prior to analysis.Slurry experiments:Solutions were prepared by adding enough solids to a given solvent so that excess solids were present.The mixtures were then agitated in sealed vials at either ambient or an elevated temperature,typically608C.After a given amount of time(typically7days for ambient and1day for608C),the solids were isolated by vacuum filtration.Preparation of Solvates and Three-Component Desolvation ScreenThe solvate–desolvation polymorph screen was run byfirst preparing a variety of axitinib solvates,and then desolvating using multiple methods.The solvate samples were prepared on a1g scale starting from form XVI(isopropanol solvate)and were formed by reslurrying in a single solvent or mixed solvent system for3days at508C.The isolated solids were initially characterized by PXRD to determine if a unique solid form had been generated.Additional characterization utilized DSC and TGA to confirm the presence of desolvation events.Once the solvated forms were characterized, parallel desolvation experiments were run on sam-ples of the solvated forms.The desolvation techniques utilized were a)vacuum drying at1008C,b)reslurry in ethanol at908C in a sealed vial,and c)reslurry in heptane at1058C in a sealed vial.Desolvation studies were carried-out on a100mg scale.Samples exhibit-ing the initial solvate PXRD powder pattern were exposed again to the same desolvation conditions.If a unique powder pattern was observed,additional thermal analysis(DSC and TGA)was conducted to characterize the material.Raman SpectroscopyThe solid form of larger scale reslurry studies was monitored via Raman spectroscopy with a Kaiser Optical Systems HoloLab5000equipped with a 785nm laser.Afiber optic cable linked the instru-ment to an MkII probe headfitted with a Hastelloy1TARGETED POLYMORPH SCREENING APPROACH DEVELOPMENT FOR AN API38771.3cm diameter immersion probe.The probe incor-porated a short focal length sapphire optic.Data collection was controlled via Kaiser’s HoloReact software(v.4.0.0233).The software resampled experi-mental data to a spectral resolution of0.6cmÀ1. Wavelength calibration was performed using cyclo-hexane.Ten second spectral accumulation times were found to be sufficient to achieve slurry signal levels within the recommended segment of the detector dynamic range.Dark background spectra were acquired prior to each spectrum,and automatic cosmic ray rejection was employed.Utilization of these two options extended the total acquisition time for each spectrum to40s.During the experiments, spectra were acquired at5-min intervals.As Raman spectra were being acquired,the univariate intensity trending of key spectral bands in real time was performed using the HoloReact package based on MatLab.Subsequent offline spectral data analysis was performed using Delight v.3.2.1(DSquared Development,Inc.,LaGrande,OR).High Boiling Solvent Polymorph Screen Approximately40axitinib solvates werefirst pre-pared on a1g scale starting from form XVI (isopropanol solvate)and were formed by reslurrying in a single solvent or mixed solvent system for3days at508C.These samples were then reslurried at1508C in p-cymene for at least7days before isolating.The isolated solids were initially characterized by PXRD to determine if a unique solid form had been generated.If needed,thermal analysis was completed by DSC and TGA.Additional high-temperature reslurry experiments in p-cymene at1508C were completed on a50mL scale using the Biotage AS2350reactor system starting from the anhydrous forms XLI and XVI with the solid form monitored in situ by Raman spectroscopy.An additional set of1308C reslurry experiments were completed in3:1p-cymene/benzyl alcohol(v/v) mixture and3:1p-cymene/o-cresol(v/v)mixture starting with form XVI.Isolated material was characterized by PXRD.Computational Chemistry MethodsCrystallographic and generated conformations of axitinib were optimized using aqueous media at the PBE/DNP/COSMO level of theory(defined by theo-retical method and a basis set)as implemented in DMol3.13This utilizes density functional theory (DFT)PBE approximation14with all-electron dou-ble-numeric-polarized basis set.The effect of bulk water is estimated by conductor-like screening model (COSMO)as implemented in DMol3.15The generated cosmofiles by the PBE/DNP/COSMO calculations for each conformer are further used for the prediction of solubility and conformational distribution in different solvents adopting COSMO-RS theory16as implemen-ted in COSMOTherm software.17RESULTS AND DISCUSSIONConventional Polymorph Screen of Form XXVThe properties and characteristics of the newly discovered anhydrous form XXV have been recently described,which demonstrated that it was a more stable polymorph at room temperature than the early development form IV.9After discovery,an additional polymorph screen with this new form was performed to reassess the polymorph environment of axitinib. The screening approaches were similar to those used for polymorph screening of form IV,which comprised well-precedented techniques previously mentioned. Solubility assessment showed that form XXV had solubilities roughly2Âlower than form IV in most solvent and aqueous cosolvent mixtures.It was acknowledged that the lower solubilities may hinder solvent-mediated transformations in slurry-type experiments.At this point in time,extensive crystal structure information of axitinib solid forms had not been discerned,including those of forms XXV and IV. Thus,though it was unclear as to definitive reasons for the lack of appearance of form XXV in previous form screens,conducting another traditional screen in the same manner as with form IV was considered the bestfirst approach for the screening of form XXV. These experiments discovered no new anhydrous forms.Form XXV was the predominant anhydrous form,while a number of new solvates were found through the use of some solvents not evaluated in previous studies.As a result,a total of39forms of axitinib had been found,see Table1.Discovery of Form XLIOnce form XXV was identified as the lead develop-ment form,chemical process development studies identified facile conditions to isolate the desired form. The process utilized a two-step reslurry process to convert the crude API to thefinal form.An initial reslurry in isopropanol converted the crude API form to an isopropanol solvate,which was subsequently converted to the desiredfinal form via a reslurry in ethanol with the addition of form XXV seed crystals to control the polymorph conversion.In the initial manufacturing campaign for producing form XXV, the conversion of the isopropanol solvate to form XXV required extended processing time possibly due to poor dispersion of the form XXV seed crystals in the crystallization vessel attributed to mixing.Subse-quent lab-scale development studies demonstrated that the seeded ethanol reslurry process conducted at reflux with a high jacket temperature resulted in a3878CAMPETA ET AL.faster and more robust conversion to form XXV.The rationale for the faster form conversion was not determined but may have been due to the higher solubility difference between the solvate form and form XXV at higher temperatures,and/or it may be due to the improved solid –liquid mixing in the crystallization vessel due to the re fluxing solvent.The lab-scale process was modi fied to utilize the higher jacket temperature for the subsequent man-ufacturing campaign.During the final isolation,an in process control sample was pulled and analyzed by PXRD to determine if the conversion to the desired form XXV was complete.PXRD analysis indicated a previously unknown powder pattern.The slurry was seeded with form XXV a second time,and the process was allowed to reslurry at re flux for 12h.Analysis of a subsequent sample indicated the same unknown powder pattern with no indication of form XXV,which suggested that the unknown form (form XLI)was thermodynamically favored versus form XXV.Characterization of Form XLI and Comparison to Known Anhydrous FormsWith the discovery of form XLI,an intensive evaluation of its physical properties was conducted to characterize this new form and understand its relationship to the other known anhydrous forms.DSC experiments showed form XLI possessing a single endotherm,with a melting temperature and heat of fusion value higher than other known anhydrous forms,see Table 2.Together,a total of five anhydrous polymorphs of axitinib were now known,which presents an unusual case.On the basis of the DSC data and Burger ’s rule,18form XLI is shown to be monotropically related to all otheranhydrous forms and is thus expected to be the most stable solid form at all temperatures.Form XLI also has the lowest solubility of all the anhydrous forms,providing further support for its designation as the lowest energy form.Figure 2illustrates the signi fi-cantly lower solubility of form XLI in ethanol.Table 2provides the rank order of the relative stabilities of the anhydrous forms.The calculated density values for all forms do not correlate with relative stability,suggesting this parameter is not a reliable measure of stability,as is often the case.18As will be described later,the various forms have different conformations,which makes comparisons based on the density rule invalid.In regard to relationships between the other forms,forms XXV and VI are enantiotropically related and very close in energies.The average heat of fusion of form VI is slightly higher,though the range of values from multiple determinations of each form is overlapping and thus may be within the experimental sensitivity of the instrument.Table 2.Thermal Properties and Calculated Density of Axitinib Anhydrous FormsMelt Onset (8C)D H f (J/g)Density (g/mL)aForm XLI 225.9153 1.369Form XXV 217.2129–132b 1.382Form VI 211.6132–136b1.330Form IV 218.7122 1.293FormI 210.51151.333a Calculated from single-crystal structure information.bRange of values from multiple DSC (5–10)determinations at 58C/min heatingrate.Figure 2.Solubility in ethanol of ethanol solvate,forms IV,VI,XXV,and XLI.TARGETED POLYMORPH SCREENING APPROACH DEVELOPMENT FOR AN API 3879。