'8
`
`_ 1—(2,4.6-trimethy1benzoyloxy)ethyl, pivaloyloxymethy1,_phenethy1,-
`
`phenpropyl, 2,2,2—trif1uoroethyl. 1- or 2—naphthyl,
`
`2,4—dimethylpheny1. aet—bucylphenyl and Snindanyl.
`
`'
`
`Of these a particularly preferred biolabi1e_estereforu1ng
`
`group is 5-indanyl.
`
`Compounfis:dffthe.fotmtlae'(E}“aufi-(Il)hwhereianoae or bath'of'
`
`R and R4 are-'Ci—C6 elkyl, particularly ethyl, or benzyl, are also
`active by virtue of their hydrolysis 1§_vivo,-and,
`in addition,
`
`are valuable intermediates for the preparation of the diacids
`
`wherein R and R4 are both 8.
`
`Particular examples of compounds of the'formula (I) wherein X
`
`'is a bridged cyclic group.include compounds wherein X is'a group
`
`of the formula:-
`
`
`
`COR
`
`'
`
`COR
`
`The above groups may be 2.5- or 2,6—linked, each attachment
`
`'being of either endo or one stereochemistrv._
`
`I!
`
`-u
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 601
`BIOCON P'HARMA' LTD (IPR2020-01263) Ex; 1015, p. 601
`
`
`
`_ 1—(2,4.6-trimethy1benzoyloxy)ethyl, pivaloyloxymethy1,_phenethy1,-
`
`phenpropyl, 2,2,2—trif1uoroethyl. 1- or 2—naphthyl,
`
`2,4—dimethylpheny1. aet—bucylphenyl and Snindanyl.
`
`'
`
`Of these a particularly preferred biolabi1e_estereforu1ng
`
`group is 5-indanyl.
`
`Compounfis:dffthe.fotmtlae'(E}“aufi-(Il)hwhereianoae or bath'of'
`
`R and R4 are-'Ci—C6 elkyl, particularly ethyl, or benzyl, are also
`active by virtue of their hydrolysis 1§_vivo,-and,
`in addition,
`
`are valuable intermediates for the preparation of the diacids
`
`wherein R and R4 are both 8.
`
`Particular examples of compounds of the'formula (I) wherein X
`
`'is a bridged cyclic group.include compounds wherein X is'a group
`
`of the formula:-
`
`
`
`COR
`
`'
`
`COR
`
`The above groups may be 2.5- or 2,6—linked, each attachment
`
`'being of either endo or one stereochemistrv._
`
`I!
`
`-u
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 601
`BIOCON P'HARMA' LTD (IPR2020-01263) Ex; 1015, p. 601
`
`
`
`9
`
`Examples of compounds uherein X is a bicyclic group include
`
`'in particular compounds wherein X is a group of the formula:
`
`
`
`The_group R5 is preferably 02—04 alkyl, 02—04 alkenyl, C
`
`2“C5
`
`alkynyl, CS-Cfi cycloalkyl, Cs—C6 cycloalkenyl,-Cl—C4I
`alkylsulphonamido, or tetrahydrofuranyl or wherein R5 is Cl-C3
`
`alkyl susbtituted by cl— alkoxy, cl—c6 alkoxymz—c4 alkoxy);
`Cs
`
`C3—C6 cycloalkyl. 4-pyridyl, 2—imidazolyl, C2-C4 alkanoyl. 02-04
`
`alkoxycarbonylamino, 01-64 slkylsulphonyl, 01-04 alkyl—
`
`sulphouamido, arylsulphonamido, heteroarylsulphonamido or
`
`benzdylamino. Thus Particular and preferred examples of R
`
`include methoxyethyl, 2—methoxyethoxymechyl, 44aminobutyl and_
`fi—merhylsulphonylethyl.
`I
`Particularly preferred individual compounds of the invention
`
`I.
`
`include 34$}—[6—endo—carboxybicyclo[212,2]octane—24exo—
`. carbamoyIJ—cyclopentylg eé-(meethoxyethyl)propanoic acid and
`3f{:l—[é—endo—carboxybicyclo{2,2,21octane-Z—exo—carbamoyl]—
`cyclopentyl} -2-(2-methoxyethoxymethyl)propanoic acid, especially
`
`the dextrorotatory diastereoisomer of the latter compound wherein
`
`-
`
`the bridged cyclic group X is resolved.
`
`BIOCON PHARMA LTD. (IPR2020-01263) EX. 1015, p. 602
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 602
`
`
`
`10
`
`The compounds of formula (I) are prepared by a number of
`different processes.
`fhe basic. procedure'involves-coupling a-
`partially prot_ected Spiro—substituted glutaric acid derivative
`to an amine to give the desired glutaramide. The carboxylic acid
`group in the amine. if free, or any reactive groups in R5, may'
`require protection during the coupling step end-sudn-protectimg
`groups are removed in the final stage of the process.
`
`The synthetic route is illustrated in scheme 1 wherein A,
`is
`I'
`as previously defined, R5
`is as defined for R5 with any reactive
`is as defined for
`group therein protected if necessary.
`(X)—COZR14
`11+
`13
`x except that R4 is R
`, and R
`andR14 are as defined for R and
`
`84 excluding H. or they are conventional carboxylic acid
`
`protecting groups:
`
`-;'
`
`Scheme-1
`
`n
`
`R5
`
`l%R
`
`A
`
`C
`
`\m} \
`130/
`I
`I
`
`H
`
`co2
`
`_
`
`+ H2N
`
`-
`
`(x)—coza
`
`I
`
`__
`
`(Iv)
`
`I
`
`.
`
`(III)
`
`-
`
`I
`
`'
`
`A
`
`lR
`
`I
`(
`s
`\ c
`CHCH ”/// \\~‘~coNH—-&}CU R14
`"II
`
`2
`
`2
`
`/’///
`
`13'
`R 02c
`
`(v)
`
`' BIQCON PHARMA LTD (lfii2020—01263) EX. 1015,.p. 603
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 603
`
`
`
`ll
`
`- The reaction of the compounds of formula (III) and (IV) is
`achieved using conventional amide coupling techniques.
`Thus in
`one process the reaction is achieved with the reactants dissolved
`
`in an organic solvent. e.g. dichloromethane,_using a carhodiimide-
`condensing agent, for example l-ethyl—B-(dimethylaminopropyl)—
`carbodiimide, or N,N'-dicyclohexylcarbodiimide, advantageously in
`Ithe presence'of l—hydroxybenzotriazole and an organic_base such as-
`N—methylmorpholiner The reaction is generally complete aftEr a
`period of from 12 to 24 hours at room temperature and the product
`is then isolated by conventional procedures, i.e. hy washing with
`mater or filtration to remove the urea biproduct and evaporation
`of the solvent.
`The product may be further purified by
`
`crystallisation or chromatography, if necessary.
`'The compounds of
`Iformula (V)
`include compounds of formula (I) wherein R and Re are
`
`Cl—Cé alkyl or benzyl.
`In same cases the_coupled product,
`
`in protected form, may he
`
`subjected to conventional chemical
`
`transformation reactions to
`
`allow preparation'bf further compounds of formula (V). Thus_for
`example compounds of formula (V) wherein R5"is a bromoalkyl group
`may be reacted with sodium azide'and the product reduced hy
`
`catalytic hydrogenation to yield the corresponding derivative
`wherein R5
`is aminoalkyl. Similarly oxidation of compounds
`5' "
`'
`contains a sulphide group yields the corresponding
`
`hwerein R
`
`sulphoxide or sulphone derivative._ Such transformations are
`
`entirely conventional and appropriate conditions and reagents for
`
`-their performance will be well known to those skilled in the art
`
`as will other variations and possibilities.
`
`The diesters.of formula (V) may be further reacted to give
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 604
`the monoesterBIzOg-igggdPMRMALSTQ(¥BR%QQOCQ)1M}EQ& (116131 3,11). 604
`
`
`
`.both of R and R4 are H.
`
`'12
`
`_The conditions used will depend on'the'
`13
`14
`precise nature of the groups R -
`
`and B
`
`present in the compound
`
`- of formula (V) and a number of variations are possible; Thus for
`
`example when both of R13 and R14 are benzyl, hydrogenation of the
`
`product will yield the diacid of formula (I) wherein R-and Ra are
`. ..
`H
`:3.
`14
`and R
`-1s benzyl and the
`both E. Alternatively if one of R
`other is alkyl, hydrogenation will yield a monoester product;
`This can then be hydrolysed. if desired.
`to again yield_the diacid
`product. When one of R13 and-R14 is t-butyl,
`treatment of the
`I
`_ compound of formula (V) with tritluoroacetic acid yields the
`corresponding acid.
`The diester product wherein R13 and filé are
`_benzyl or lower alkyl can also be treated with trimethylsilyl
`iodide to_produce the dicarboxylic acid product. -If some other-
`carboxylic acid protecting group is used for R13 or R14 then
`clearly appropriate conditions for its removal.must be employed in H
`the final step to give the ester or diacid product of formula (I).
`In the case where the ring A or the substituent R5 is unsaturated,
`
`thus
`the deprotection must be effected by non-reductive methods,
`for example if either of R and R4 is benzYl,
`they may be removed:
`by treatment with.trimethylsilyl iodide;
`
`As an alternative to the above procedure the coupling '
`
`reaction is performed with an amine of the formula:-
`
`HZN-(X)-Ch203
`
`~—— (VI)
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 605
`' BIOCON PHARMA LTD (IPR2020-01263) EX. 1015, p. 605
`
`
`
`13
`
`The coupled product is deprotected as previously described
`
`and the product is then oxidised, for example by stirring with
`
`_p1atinum in the presence of oxygen.
`
`to yield the correspdnding
`
`acid of formula (I).
`
`As well as removing any protecting group which.mav be present
`I
`-
`.
`in R5 , a'npmber of chemical transformation reactions are possible_
`
`on the final mono-ester or diacid products as previously
`
`described.
`
`In each case the product_may be-obtained as the free
`
`_carboxylic acid or it may be neutralised with an-appropriate base
`
`and isolated in salt form.
`
`Compounds of the formula (I) wherein one or both R and 84 is
`
`Ia biolabile ester forming group are prepared following similar
`
`procedures to those described above. Thus,
`
`in one variant of the
`
`process outlined in Scheme 1; a compound of formula (III) wherein
`13
`
`is.a biolabile ester—forming_group is coupled to the
`
`R
`
`14 is a benzyli
`appropriate compound of formula (IV), wherein R'
`
`group, and the product is hydrogenated to give the compound of
`formula (I) wherein R is a biolabile estereforming group and R4 is
`H.
`I
`
`The amines of formula (IV) and (VI) are in many cases novel
`compounds but
`they may be prepared from known starting materials
`by conventional synthetic procedures in accordance.with literature
`
`precedents as illustrated in the Examples hereto.
`
`Thus for
`
`example the corresponding hydrosyfsubstituted bridged-cyclic
`carboxylate may be converted to the_amine by sulphonylation
`
`followed by aside displacement and reduction, or a bicyclic
`
`lactone may be.reduced by treatment with lithium aluminium hydride_
`
`and the resulting diol converted to an aminoalcohol
`
`in a similar
`
`fashion.
`
`BIOCION PHARMA LTD (IPR2020-01263) EX. 1015, p. 606
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 606
`
`
`
`1:.
`
`The starting spire-substituted glutaric acid mono esters of
`formula (III) may be prepared-by a number of processes as
`
`described in European patent application 8?310?84.1.
`
`As previously mentioned, the compounds of the invention are
`
`potent inhibitors of the neutral endopeptidase (E.C.3.4;24.11).
`
`This enzyme is involved in the breakdown of a number of peptide
`
`hormones and,
`in particular we have discovered that it is involved _
`in the breakdown of atrial natriuretic factor (ANF).I This_hormone
`
`consists of a family of related natriuretic peptides. secreted by
`the heart, of which the major circulating form in humans is knoun
`
`to be the 28 amino-acid peptide referred to as R.-hANP (See for
`
`_example G. A. Sagnella and G. A. MacGreggor, Nature, 1984, 309,
`
`666 and S. A, Atlas and others, Nature,'1984, 292' 717~?25);
`Thus,
`the compounds of the invention, by preventing the
`I
`
`degradation of ANF, by endopeptidase_E.C.3.4.24rll can potentiate
`its biological effects and the compounds are thus diuretic and
`
`natriuretic agents of utility in a number of disorders as
`
`.previously described.
`
`Activity against neutral endopeptidase E.C.3.4.24.11'isi
`
`assessed using a procedure based on the assay described by J. T.
`
`Gafford, R. A. Skidgel. E. G. Erdos and L. B. Hersh, Biochemistry,
`
`. 1983..§g. 3265—3271.
`
`The method involves_determining the
`
`_concentration of compound required to reduce by 502 the rate of
`
`release of radiolabelled hippuric acid from hippuryl—L—
`
`phenylalanyl-L—arginine by a neutral endopeptidase preparation
`
`from rat kidney.‘
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 607
`BIOCON PHARMA LTD (IPR2020-01263) EX. 1015, p. 607
`
`
`
`15
`
`. The'activity of the compounds as diuretic agents is
`determined_by measuring their-ability to increase urine output and
`sodium ion excretion in saline loaded conscious-miCe.
`In this
`
`.test, male mice (Charles River_CDl. 22-28_g) are acclimatised and
`«ctflrued onernigbrhin.merabowls...mhewmice43remdosed.intravenously
`Ivia the‘tail vein; with the test compound dissolved in a volume of
`saline solution equivalent to 2.5% of body weight. _Urine samples-
`are collected each hour for two hours-in:pre5weighed tubes and
`analysed for electrolyte concentration. Urine volume and sodium
`ion concentration from the test animals are compared to-a control
`
`group which received only saline.
`For administration to man_in the curative or prophylactic
`treatment of hypertension, congestive heart failure or renal
`insufficiency, oral doSages of the compounds will generally be in
`
`-the'range of from 10—1500 mg daily for an average adult patient
`I(70'kg).
`'Thus for a typical adult patient,
`individual tablets or
`capsules contain from 2 to 300 mg of active compound,
`in a '
`
`suitable pharmaceutically acceptable vehicle or carrier for
`administration singly, or in multiple doses; once or several times
`a day. Dosages for intravenous administration would typically be
`
`within the range 5 to 500 mg per single dose as required.
`§
`
`In
`
`- practice the physician will determine the actual dosage which will
`
`_be most suitable for an individual patient and it will vary with
`
`the age. Weight and response of the particular patient.
`
`The above
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 608
`BIOCON PHARMA LTD (IPR2020-01263) EX. 1015, p. 608
`
`
`
`16
`
`doSages are exemplary of the average case but there can, of
`
`'course,'be individual instances where higher or lower dosage
`
`ranges are merited, and such are within the scope of this
`
`invention.
`
`For human use.
`
`the compounds of the formula (I) can be
`
`‘- odefinistered alonecwbur.will generally be.adminis£ered in
`
`admixture with_a pharmaceutical carrier selected with regard to
`
`the intended route of administration and standard pharmaceutical
`
`practice. For example,
`
`they may be administered orally in the_
`
`form of tablets containing such excipients as starch or lactose,
`
`or in capSules or ovules-either alone or in admixture with
`
`excipients. or in the form of elixirs or suspensions containing-'
`
`flavouring or colouring agents.
`They may be injected
`parenterally, for example,
`intravenously,
`intramuscularly or
`subcutaneously.
`For parenteral administration,
`they are best used
`
`in the form of a sterile aqueous solution which may contain other
`Substances, for example. enough salts or glucose to make the
`
`solution isotonic with blood.'
`
`The compounds may be_administered alone_but may also be_
`
`administered together_with such other agents as the physician-
`
`Ishall direct to optimise control of blood pressure or to treat
`
`congestive heart failure, renal insufficiency or other disorders
`
`in any particular patient in accordance with established medical
`
`'practice. Thus the compounds can be co—administered with a
`
`variety of cardiovascular-agents, for example with an ACE
`
`.'
`
`inhibitor such as captopril or enalapril to facilitate the control
`
`of blood pressure in treatment of hypertension; or with digitalis,
`
`-or another cardiac stimulant or with an ACE inhibitor, for the
`
`:treatment of congestive heart failure. Other possibilities'
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 609
`BIOCON PHARMA LTD (IPR2020-01263) EX. 1015, p. 609
`
`
`
`l7
`
`include_co—administration mith a calcium antagonist (e.g.
`nifedipine or diltiazem) a beta—blocker (e.g. atenolol) or an
`
`alphaeblocher (e.g. prazosin) as shall be determined by the
`phpsician as appropriate for the treatment of the particular
`patient or condition'involved.
`
`In addition to the-above,
`
`the compounds may also be
`
`administered in conjunction with exogenous ANF, or a derivative
`
`thereof 0: related peptide or peptide fragment having
`
`diuretic/nattiuretic activity or with other ANF—gene related
`
`:peptides (e.g. as described by D. L._Vesely et a1, Biochem.
`
`Biophys. Res. Comm.l 1987. 1&3, 186).
`
`Thus in a further aspect
`
`the invention provides a
`
`pharmaceutical composition comprising a compound of the formula
`
`(I) or (II), or a pharmaceutically accéptable salt thereof or
`
`_bioprecursor thorefor,
`
`together with a pharmaceutically-acceptable.
`
`diluent or carrier.
`'The invention also includes a compounds of the formula (I) or
`(II), or a pharmaceuticallp acceptable salt thereof or
`hioprecursor therefOr._for use in medicine;
`in particular in the'
`
`treatment of hypertension, congestive heart failure or renal
`
`insufficiency in-a human being.
`I'The preparation of the compounds of the invention and of
`intermediates for use in their preparation is illustrated by the
`following Examples.-
`I
`
`Purity of compounds was routinely monitored by thin layer
`
`chromatography.
`
`H-N.M,R. Spectra of all products were recorded
`
`1
`
`using a Nicolet QE 300 spectrometer and were_in all cases
`
`consistent with the proposed structures.
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 610
`BIOCON PHARMA LTD.(IPR2020-.01263) EX. 1015, p. 610 '
`
`
`
`18
`
`EXAMPLE 1
`
`I Methil G-exo—aminobiczclOEZ,2,2]octane—2—endo—carboleate
`
`hxdrochloride
`
`
`
`A Solution of finendo~hydroxybicyclo[2,2,2]octane42—endo-
`I(a)
`carboxylic acid lactone (7;65 g; 50.3 mmole} in dioxau (100 ml)
`":'and 1N sodium hydrfikifie (200 El? was dllowed‘to_stand“at“ro0m
`Itemperature overnight. Host of the Solvent was evaporated under
`reduced pressure at 40°C, and the residue was acidified to nH 5
`
`with concentrated hydrochloric acid with ice cooling;
`
`The
`
`precipitate was collected by filtration and washed.with water,
`to
`
`give 6—endo—hydroxybicvcloI2,2,21octane—Euendo-carboxylic acid as
`
`a white solid (d.9_g. 802), m.pv 125—125.5‘C.
`I
`I
`I
`I
`(b) Cesium carbonate (7.88 g; 2d.2immolel
`in water t30'ml)
`was added to the above carboxylic acid (8.24 g: 48.4 mmole)
`in a
`1 1:1 mixture of methanol_and water (40 ml). The resulting clear
`solution was evaporated to dryness under vaCuum, and the residue
`was dried azeotropically with toluene giving the cesium salt as a
`
`white solid. Methyl iodide (5 ml:-81 mmole) was added at room
`
`temperature to-a stirred suspension of the cesium salt in dry
`dimethylfornamide (40 ml). Water was added after 2.5 hours.and
`the suspension was_extracted with ethyl acetate.
`The combined
`extracts ware washed with water, dried (MgSOa) and evaporated to
`give an oil (8.8 g) which solidified 0n standing.
`
`Recrystallisation from hexane gave methyl fi—endo-hydroxybicyclo-
`
`
`[2,2.2]octane—Z—endo—carhoxylate as a white solid (7.52 g, 842),
`
`m.p. 46—46.5°c.
`
`Found:
`
`c;55.09; H,8.83. c
`
`10H1603
`
`requires
`
`C,65.19;'H,8.?52.
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 611
`BIOCON PHARMA LTD (IPR2020-01263) EX. 1015, p. 611
`
`
`
`19
`
`(c) 'The above ester (7.11 g; 38.6-mmole) was added to an ice
`
`cold solution of.4—methylbenzenesulphonylchloride (11.04 g;
`
`I‘
`
`in dry pyridine (40_m1). After_0.5 hours the solution
`58 mmole)
`was allowed to warm.to room temperature and, after standing -
`
`overnight,
`
`the solvent was evaporated under vacuum.
`
`'The residue
`
`was partitioned between ethyl acatate and water. and the organic
`
`phase was washed in succession with 2N hydrochloric acid, water,
`
`:saturated aqueous sodium bicarbonate_solfltion and water.' Drying
`
`(M3804), followed by evaporation of the solvent and'
`
`
`-recrysta11isation from ethy1 acetate-hexane gave methyl o—endo—
`(demethylbenzenesulphonyloxy3bicyclo[2,2,2]octane+2-endo—
`
`carboxylate as'a white solid (11.65 g; 89%). m.p. 101-101.5°C.
`
`Found:
`
`0.60.29; 3.6.57.
`
`5 requires c,60.33; H,6.ssz.
`
`C195205
`(d) The above hemethylhenzenesulphonate (11.7? g, 34.8
`mmole) was stirred under nitrogen with sodium azide (10 g; 150
`mmole)
`in dry dimethylformamide (30 ml) for five days at 75°C.
`Most of the solvent was evaporated under vacuum keeping the'
`
`temperature below 30°C.
`
`'The residue was partitioned between ethyl
`
`acetate and water and the organic extract washed with water.
`
`Drying (H3804) and evaporation of'the solvent gave an oil which
`
`was chromatographed on silica.' Elution with ethyl acetate, hexane
`(1:1) gave methyl—fi—exoLazidobicyclo[2,2,2]octane-2—endo—
`
`
`carhoxylate as an oil (4.28 g, 59%) contaminated with _
`
`
`approximately 10% of methyl bicycloI2,2,2]oct—5-ene—2—endo
`o
`
`carboxylate.
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 612
`BIOCON PHARMA LTD. (IPR2020-01263) EX. 1015," p. 612
`
`
`
`20
`
`(e) Therazide of example 1(d) above (3;2 g)
`
`in methanol
`
`(30 m1)'was'hydrogenated over 10% palladium on charcoal catalfst
`(300 mg) at room temperature and 50 p.s.i.
`(3.45 bar).' After fiue
`
`hours the mixture was filtered and evaporated to dryness.
`-The
`residue was chromatographed on silica, eluting with a mixture of_.
`dichloromethhnE'afid'methanol (9812}‘Ioiiowed by fiiéhloromethane,
`Imethanol, 0.88 aqueous ammonia (94:5fil)-to afford the reouired.
`amine as an 011.
`An etherial solution of the nroduct was treated-
`
`with 4N HCl in dioxan to give the title hydrochloride which was
`isolated as a white solid (2.55 g) after recrystallisation from-
`methanolediethylether,.m.p. 2004202°c,
`Found:
`0,54.13; H,8.47;
`
`N;6.17;
`
`c
`
`H N0'.HCl, 0.1 320 requires c,54.21;.a,8.2s;'N,6.32%;
`10 17
`2
`
`EXAMPLE 2
`
`- Mechl—G—exo-aminobiczclo[2,2,2]octane—Z—exo—carboxvlatg
`
`'hxdrochloride
`
`I
`
`(a)
`
`'Diethyl diazocarboxylate (0.63 ml, 4 mmole) and
`
`diphenylphorylaaide (0.86 ml; 4 mmole) were added simultaneously.
`
`
`to an ice cold stirred solution of methyl 6-endo-hydroxybicyclo—H
`
`[2,2,2]octane—Zfieggfcarboxylate (510 mg; 2.8 mmole) and triphenyl
`phosphine (1.05 g; 4 mmole)
`in dry tetrahydrofuran under nitrogen.
`After three hours the miuture was absorbed onto silica. Elution-
`Iwith a mixture of diethyl ether and hexane_(2:8) gave crude
`Inroduct which was rechromatographed eluting with diethy1_ethet_and
`
`hexane (1:9).
`
`The required methyl 6fg§gfazidobicyclo[2,2,2]-
`
`
`octane- 2—exo—carboxylate was obtained as a clear liquid (400 mg,
`
`692).
`
`IR (film) 2100 cmfl.
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 613
`BIOCON PHARMA LTD (IPR2020-01263) EX. 1015,‘ p. 613
`
`
`
`21
`
`(b)
`
`'The azide (390 mg, 1.8? mmole)
`
`from part
`
`(b) above was
`
`The
`reduced following the procedure described_in Example 1(e).
`hydrochloride salt was recrystalliSed from a mixture of methylene
`chloride and diethyl ether to give_methyl fifieggraminobicyclo—
`
`[2,2,2]octanefZ-exo—carboxylate hydrochloride as a white solid
`
`(230 fig. 562) m.p. 216-218°C.
`
`Found:
`
`'c.54.44: 3.8.63; s.6.32.
`
`c 'H no .HCl requires C,54.67; H.8.26; N.6.3?Z.
`10 l?
`
`EXAMPLE. 3
`Methxl 5—endo-aminobicxcloI2.2,2]octane—Z-endo-carhoxxlateI
`hzdrochloride.
`I
`I(a) Methzl 5-endo and 5—ero-hzdrorzbicxclo[2,2,2]octane-
`
`2 .
`
`2—endo-carboleate
`
`Sodium borohydride (824 mg; 21.8 mmole) was added to an_ice
`
`cold stirred solution of methyl 5—oxobicyclo[2,2.210ctane—2—
`
`
`'_endo—carboxylate (7.94 g; 43.6 mmole)
`
`in methanol
`
`(80 ml). After I
`
`2 hours the mixture was acidified to pH 4 with 2N hydrochloric
`
`acid and evaporated to a small volume under vacuum.
`
`The residue
`
`was partitioned between diethyl ether and water.
`
`The organic
`
`phase was washed in succession with 2N hydrochloric acid. water,
`saturated-aqueous sodium bicarbonate and water. Drying (Mg504)'
`and evaporation gave a crude mixture of isomers as a yellow oil
`
`(?.45 g).
`
`t.l.c. (silica;
`
`iso—propanol- methylene chloride (1:9)}
`
`Rf. 0.58 and 0.62.
`
`IChromatography on silica and eluting with
`
`diethyl ether and methylene chloride (1:9) gave an initial
`
`to he 96% pure 5—endo isomer.
`
`shown by g.1.c.
`
`fraction (2.6 g),
`1
`
`H—N.M'.R.
`
`(00013) 53.80 (m fl—C5), 3.72 (s £02933).
`
`Following a
`
`BIOCON PHARMA LTD (IPR2020-01263) EX. 1015, p. 614
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 614
`
`
`
`22
`
`mired fraction. a third fraction (1.64 g) was obtained which was
`shown by g.1.e.
`to be 94% pure Siéifl isomer.lH—N.M.R.'(CDC13)A?
`I 3.99 (m, H-CS) 3. 68 (5, —CO
`20KB).
`I
`I
`I
`(b) The 5fgggjisomer from part
`(a) above (1.95 g;.10.58
`
`mmole} was treated with 4-methy1benzenesu1phonyl chloride in
`pyridine as def-curibed in EnaE-ple- l{:t).
`The crudutproduot was
`
`chromatographed on silica, eluting with diethyl ether and hexane
`
`(4:6) to give methyl 5—exo—(4—methy1benzenesulphonyloxy)hicyclo—
`
`
`[2,2,2]0ctane-2-endo.carboxylate as an'oil (3.21 g, 90%).
`C,60.29; H,6.53Z.
`c17H22053 requires c,6u.33: 3.6.552.
`
`(c) The above S—exo—methylbenzenesulphonate (3.18 g. 9.4
`
`Found:
`
`mmole) was stirred under nitrogen with sodium azide (3.05 g; 4?
`
`mmole)
`
`in dry dinethylformamide (10 ml) at 55°C for eighteen hours
`
`-and then at'75°C for forty-eight hours. Work up as described in
`
`Example 1(6) gave a pale yellow oil (1.95 g) which was
`
`chromatographed on silica. Elution with diethyl ether and hexane
`
`
`(5:95) gave methyl 5—endo—azidobicyc1oI2,2,2]octane—2—endo
`
`carboxylate as a clear liquid (950 mg; 48%).
`
`Found:
`
`0.57.15;
`
`requires c. 57. 40; s, 7. 23; u, 20. 082.
`
`H,7.4o;_w.20.22.c
`c10H15N302
`IThe above azide (940 mg; 4.49 mmole) was reduced as
`(d)
`described in Example 1(e). The crude hydrochloride was.
`recrystallised from methanolkdiethyl ether to give the title amine
`
`as e white solid (430 mg, 442), e... 156-7°C.
`
`Found:
`
`c, 54.79;
`
`3,8.36; N,6.3?.
`
`C10H17N02..HC1 requires C, 54. 67; H, 8. 26; N, 6. 372.
`
`BIOCON PHARMA LTD (IPR2020-01263) EX. "1015, p. .615
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 615
`
`
`
`23
`
`EXAMPLE 4
`
`I
`
`'Methxl—S—exoéaminobicxcloI2;2,2Ioctane—Z—endo—oarboleate
`5 hydrochloride
`I
`I
`I
`(a) The Seengg—hydroxy compound from Example 3(a)_(3.1'g;--I
`16.82 mmole) was treated with 4—methylbenzenesulphony1-chloride
`
`(£;81 g; 25L2 nmole) in dry pyridine as described in Example 1(c).
`
`The erude product was chromatographed on silica, eluting with
`diethyl ether and hexane-(4:6) to give methyl-5—gnfig}(4—methyl-
`
`
`benzenesulphonyloxy)bicyclo[2,2,2]octane-Z—endo—carboxylate as an
`
`011 (5.20 g, 912).
`
`Found: C,60.29; H;6.54.
`
`C17H22°5
`
`s requires-
`
`c,50.33; H,6.552.
`
`
`(b) The above S-endo—methylbenzenesulphonate (5.15 g; 15.?
`
`mmole) was stirred under nitrogen with sodium aside (4.94 g; ?6
`mmole) in dry dimethylformamide (15 m1) at 55°C for eighteen hours
`
`L and then at 75°C for thirty hours. Work_up as described in
`
`'Example 1(d) gave a pale yellow liquid which was chromatographed
`
`on silica. Elution with diethyl ether and hexane (5:95) gave
`
`
`methyl—SfigggfazidobicyCloI:,2,2]octane—Z-endo-carboxylate, as a
`
`clear liquid (1.9 g, 612).1 Found:
`c;5?.17;-H,?.26; n.2o.oz.
`I¢10315N302 requires C.57.40; H,l.23; N.20.08%.
`(c) The above aside (1.9 g; 9.1 whole) was reduced as
`described in Example 1(e3.
`The crude hydrochloride was
`recrystallised from nethanol/diethyl ether to give the title amine
`
`as a white solid (840 mg. 42%); m.p. 228—229°C.
`
`Found: C.54.64;
`
`' H,8.45; N,6.31. C10H1?302.HC1 requires C,54.6?; H.8f26; N,6.3?Z.
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 616
`BIOCON PHARMA LTD (IPR2020-01263) EX. 1015, p. 616
`
`
`
`'24.
`
`EXAMPLE 5
`
`1H—ZE—Amino—2 3. 3ao<,4, S,6,?, 7aoL—octahxdroinden1e—awcarboleic
`
`acid ethzl ester
`
`(3)
`
`2H—2*0xo—1,3.3ao(,4,5,6.7,?aod-octahydroindene-3aod-'
`
`carboxylic acid ethyl ester (2.9 g; 13.8 mmole)
`
`(W. Dauben et al.,
`
`J. org.'Chem., 1961, gg, 29?}-is-teiiarydiofuras-(30*m13-wasjaaaaa‘“"r'
`
`dropwise under nitrogen to a stirred 1M solution of lithium
`
`trisiamylborohydride in tetrahydrofuran (15.2 ml; 15.2_mmole) at
`
`—?0 to -60°C. _After two hours the mixture was allowed to warm to '
`
`ambient
`
`temperature and left to stand for eighteen hours.
`
`The
`
`-mixture=was then cooled to-10°C.- water (1 ml), ethanol (3.é_mlj.
`
`I 6N aqueous sodium hydroxide (2.3 m1) and 302 aqueous hydrogen
`
`peroxide (3.4 ml) were added in succession. After five minutes
`
`the aqueous phase was saturated with potassium carbonate, diethyl
`
`ether and saturated sodium chloride solution were added and the
`
`organic layer was separated off. The aqueous phase was
`Ire—extracted with diathyl ether and the combined extracts were
`
`washed with water, dried.(Mgsoé) and evaporated to give an oil
`which was chromatographed on silica. Gradient elution using
`
`hexane and ethyl acetate (3:? to 1:1) gaVe 1H-2fi-hydroxy-2,3,3aoc- I
`
`I 4.5.6.?,7aoéroctahydroeindene—Saocrcarbonyl acid ethyl ester as an _
`
`oil (2.52 g; 862).
`
`Found: C, 67. _70; H, 9. 67. C
`12B2003
`
`requires'
`
`0.67.89; H.9.SOZ.
`
`(b) Diethyl azodicarboxylate (2.47-m1; 15.?‘mmole) in
`
`tetrahydrofuran (15 ml) was added dropwise at 10°C to a_ stirred
`
`(2. 22 g; 10.5 mmole),
`(a)
`solution of the hydroxy acid from part
`methyl 4—methylhenzenesulphonate (2.43; 13.1 mmole) and triphenyl
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 617
`BIOCON PHARMA LTD (IPR2020-01263) EX. '1015, p. "617
`
`
`
`----1.—--~
`
`25
`
`phosphine (3. 43 g; 13.1 mmole)
`
`in tetrahydrofuran (25 ml). I After
`
`the mixture was
`stirring for 20 hours at ambient temperature,
`_evaporated to dryness, absorbed onto silica and chromatographed on
`silica. Gradient elution starting from hexane'and methylene
`
`to neat methylene chloride gave 1H-zac—(4-methyl-
`chloride (3: 7)
`benzensulphonyloxy)~2, 3 ,3aG¢,4, 5, 6, 7, 7a0<roctahydroindene—3aa<9
`
`carboxylic.acid ethyl ester as a clear 011 (2.01 g; 582).
`
`Found:
`
`c19H26055 requires 0.62.27; 3.7.152
`_C,62.2?; 3.7.27.
`(c) The product from part
`(b) above (1.99 g) mas treated
`_ with sodium azide and reduced as described in Example 4 to give
`
`'
`
`the crude amine which was chromatographed on silica. Gradient
`
`elation with increasing proportions of ethanol in methylene:
`chloride (0 to 20%) gave pure title product as an oil (770 mg; 62%
`overall).
`'Found:
`0.65.85; H,9.74; N,6.32.
`c12321N°2,0 5 H20
`
`requires 0.65.42; H,10.07; N,6.362.
`
`
`EXAMPLE 6'
`
`_
`Suzi[6—endo—methoxzcarbonglbicxclof2, 2. 2]octane—2—exo—
`carbamozlkxclogentxlg-2—(HZ—methoxzethzlmroganoic acid [:31le
`I ester
`
`l—Ethy1—3—(3-dimethylaminopropyl)carbodiimide'hydrochloride
`
`(958 mg; 5 mmole) was added to an ice cold stirred mixture of _
`3-(l—carboxycyélopentfl)—2r(2"m3th03YEthyl)PfOPanOic acid benzyl
`ester (836 mg; 2.5 mmole). methyl 6sefig—aminobicyclof2.2,2]—
`octane—2—Eggg carboxylate hydrochloride (550 mg; 2.5 mmole),
`l—hydroxybenzotriazole-(33% mg; 2.5 mmole) and N—methylmornholine
`
`(834 mg; 8.25 mmole)
`
`in dry methylene Chloride (10 ml). After 0.5 -
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 618
`BIOCON PHARMA LTD (IPR2020-01263) EX. 1015, p. 618
`
`
`
`26 _
`
`hours the mixture was allowed to attain room temperature and
`
`stirred for a further eighteen hours. The mixture_was diluted
`
`'with methylene chloride. washed in succession with water, 1N
`
`'hydroohloric acid, saturated aqueous sodium bicarbonate and water,
`
`and dried (Hgsoé). Evaporation gave an oil which_was.
`chromatographed'on-sdlioa.'~Eluticc.with ethylpaoetate and hexane
`
`(3:?) gave the title diester as a gum (1.08 g; 852).
`
`Found:
`
`6.69.42; H,8.29; N,2.7§.
`0.69.?1; H,8.27; N,2.802.
`
`C29H41N96, 0.11 CH3002d2H5
`
`requires
`
`EXAMPLE ?-
`
`{h-EJrlfi—endo—carhoxybiczcloI2,2,2]octane—Z—exo-carbamole-
`
`czclopentzlg —2—(2—methox2ethzl)propanoic acid'
`in methanol
`The diester from Example 6 (980 mg; 1.96 mmole)
`(18 ml)'and water (12 ml) was hydrogenated over 5% palladium on
`
`charcoal'catalyst
`(100 mg) at 50 p.s.i..(3.45 bar).' After four
`hours the mixture was filtered through avicel, washing with
`methanol, and evaporated to dryness.
`The residue was partitioned
`
`between diethyl ether'and 1N sodium hydroxide (4 ml).
`The aquebus
`phase was separated and the ether phase was again extracted with
`1N sodium hydroxide (4 ml).
`The combined aqueous extracts were.
`
`allowed to stand at room temperature for eighteen hours and then
`
`acidified with 2N hydrochloric acid.
`
`The suspenSion was extracted -
`
`I with methylene chloride,
`
`the extract washed with saturated aqueoUS'
`
`sodium chloride solution and dried (M3804).' Evaporation of the
`
`solvent gave the required diacid as a white foam (705 mg. 91%).
`
`Found:
`
`.C,63.50; H,8.6l; N,3.31.
`
`C21H33N06 requires 0.63.77;
`
`H.8.41; N,3.saz;
`
`'
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 619
`BIOCON PHARMA LTD (IPR2020-01263) EX. '1015, p. 619
`
`
`
`27
`
`EXAMPLES 8 - .15'
`The-following compounds-were prepared- from the aopropriate
`
`amine of Examples 2 to 5 following the coupling and deprote'ction
`
`procedures of Examples 6_and ?.
`
`' ;;
`ca30(C32)2
`-\
`/CHCH2
`ROZC/
`
`_
`
`CONE—X _
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 620
`BIOCO'N PHARMA LTD _(IPR2020-01263_) EX. 1015, p. 620
`
`
`
`28
`
`N29
`
`:53
`
`.Enw
`
`Han
`
`
`
`noun«.09
`
`Ewen
`
`m00.
`
`
`
`1...n
`
`.3
`
`
`
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 621
`BIOCON PHARMA LTD (IPR2020-01263.) EX. "1015, p. 621
`
`2monmuwxuwuncwHmUHuuuowphvNmfimhfimp¢
`umumfiowhEhom1NIEZI
`
`.l:llllllIIllllIl-Irtailrl
`
`I'Illlllllllllllllniillllullllllo.
`
`wHamem
`
`
`
`
`
`
`
`
`
`
`
`
`nmuwxuwunufinfiUfiuwuouzhvNmamhamam.
`
`2..m_.o.
`
`
`
`uuumHOmHSham
`
`.uamfimwm
`
`29
`
`
`
`
`
`cm.m.mm.m....m¢.mm
`
`
`
`.nmm.m.no.mmn.meU
`
`houmq.ou
`
`.Em0m
`
`.36
`
`nonmm.ov
`
`EmOm
`
`Ill-I-llllllllllllllilllqlll.II
`
`BIOCON PHARMA LTD (IPR2020-01263) Ex. 1015, p. 622
`BIOCON PHARMA LTD. (IPR2020-01263) EX. 1015, p. 622
`
`
`
`
`
`30
`
`EXAMPLE 16
`
`
`6-endoeflxdroxzmethxlbiczclo[2,2,1]heptan-2—exo—amine hydrochloride _
`
`
`(a)
`A solution of S—endo-hydroxybicyclo[2,2,1]heptane-2—
` endo-carboxylic acid lactone (11.0 g;_?9.? mmole)
`in diethfl ether
`(150 ml) was added dropwise over 0.5 hours under nitrogen to a
`stirred ice couled suspension 6f lithium aluminium hydride
`(3. 02 g; 79. ? mmole)
`in dry diethyl ether (50 m1), keeping the
`temperature between 10 and 20°C.
`The mixture was then stirred at
`_room temperature for 2 hours. saturated aqueous ammonium chloride
`(20 ml) was carefully added followed by solid ammonium chloride
`(15 g) and magnesium sulphate to produce a granular suspension.-
`Ifhe mixture was filtered, dried (M3804) and the solvent evaporated
`under reduced pressure to give a white solid (11.15 g).
`
`Chromatography on silica, eluting with diethyl ether and
`
`recrystallisation from ether and hexane gave 6—endo~hydroxy4
`
`
`methylbicyclo[2,2,l]heptan-2-endo-ol as a white solid (9.49; 84%).
`An analytical sample had m.p. 118—120°C.
`Found:
`0.67.87;
`.
`H.10.24.' C8E1402 requires C, 67. 5?; H. 9. 93K.
`(b)
`(1, l—Dimethylethyl)dimethylsilylchloride (10. 98 g; 72r8
`uncle)
`in dry methylene chloride (45 ml) was added with ice
`cooling to a stirred solution of the abowe dio]
`(9.42 g; 6622
`mmole),
`triethylamine (1.37 g; 72.82 mmole) and
`_A-dimethylaminopyridine (325 mg, 2.65 mmole)
`in methylene chloride'
`(55 ml). After stirring at room temperature.for'one and a half
`I
`hours the solution was washed in succession with water, saturated
`
`aqueous ammonium-chloride and water. Drying (M3804) and .
`t .
`.
`
`evaporation
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