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`

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`-
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`u~ /14t>2• th us
`Annex tfs~. page I
`is·
`___________ P_C_T___..4.cant's Guide - Volume II ·- National Chaptc l
`~ SS HAIL: BB214759358US
`MAILED: 17 NOVEMBER 1993
`U S. DEPART~tth"T OF COMMERCE PATEto"T At-0 T RADEMARi; OFFICE ATTORtoEY"S ooci;ET NUMBER
`
`.
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`.
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`t1 ft...MPCt~ .. 17 NOV ~
`.._,U
`>O lllM rro.•J'JO
`( RliiV 11 -t ' j
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`lrl~
`
`TRANSMITTAL LETTER TO THE UNITED STATES
`DESIGNATED/ELECTED OFFICE (DO/EO/US)
`NTE!<NATIONl\L Al'l'L CATION NO.
`PCT/US92/05126
`TITLE Of INVENTION
`METHOD FOR MAKING HUMANIZED ANTIBODIES
`Al'PLICANT(S) FOR 00/EOIUS
`Paul J. CARTER Leonard G. PRESTA
`rrlit::1n1 hcrcwilh submils to the Uni1ed Slates Dcsigna1ed/Elcc1eJ Office (DO/'EOIUS) the following hems under 35 U.S.C. 371:
`I. cg This ellpress rcqucsl 10 immediately begin nalional examination procedures (35 U.S.C. 371(1)).
`2. ell The U.S. Na1ional Fee (3S U.S.C. 37l(c)(l)) and other fees as follows:
`(2) NUMBER FILED
`(3) NUMEIER EXTRA
`23
`3
`-20 -
`7
`10
`-3 -
`
`(4) RAT
`x $22.00
`X S74.00
`+ S230.00
`
`518.00
`230.00
`
`709Pl
`PRIORITY DATE CLAIMED
`14 June 1991 14.06.91
`
`JntematioMI preliminary ex.amination fee paid to USPTO (37 CFR 1.482)
`
`$830.00
`
`·sASIC NATIONAL FEE (37 CFR 1.492(&)(1)-(5))1
`, .. · 0 For filing with EPO or JPO search repon (37CFR I .492(a)(5)).....
`· 0
`111111
`N;·i:~;~·~;~~·i·~·~~i· ~~~i ;·,;;·i·,;~;·~~~~~·i·;;~;·i·~·.; .. i~~ ·~~id·;; ·0:s-P--r0·o1 cFR ~~:~2~0
`
`but intem:itional se:i.rch fee paid to USPTO (37 CFR l.445(a)(2})..
`
`$710.00
`
`Neither intema1ional preliminary exami11n1io11 fee (37 CFR 1.482) nor
`intemaiional search fee (37CFR I .445(a)(2)) paid to USPTO.......
`S950.00
`
`International preliminary examination fee p:i.id to USPTO ()7 CFR 1.482)
`and all claims satisfied provisions of PCT Anicles 33(2)-33(4)...
`$90.00
`
`Surcharge of SlJ0.00 for furnishing the N:1tional fee or oath or
`0 30 months from the earliest
`declaration later th:i.nO 20
`claimed priority date (37 CFR l.492(e)).
`
`950.00
`
`TOTAL OF AEIOVE CALCULATIONS = 1. 764.00
`
`Alrfidavit must be filed also.
`
`SUBTOTAL
`D 20 0 Jo
`
`+
`
`'
`TOTAL NATIONAL FEE Sl, 764.00
`
`+
`
`40.00
`
`TOTAL FEES ENCLOSED St,804.00
`
`a . 0 A check in the amount of S
`10 cover the above foes is enclosed.
`IKJ Please charge my Deposit Account No. 07-0630
`in the· amount of $ 1. 804. 00
`b.
`A duplicate copy of this sheet is enclosed.
`c. ~ The Commissioner is hereby authorized ''b~'a~jBY addi1ion:~I fees which may be required, or credit any
`overpayme11110 Deposit Account No.
`-
`. A duplicate copy of this sheet is enclosed .
`
`to cover the above fees.
`
`(Juuary 1993)
`
`6 of 1033
`
`BI Exhibit 1002
`
`

`

`~ !\,""n1b· tr \
`- -
`Annex .U~. Vl,.page..2.· .
`
`1
`
`Applicant"s Guid~ - Volu~e 11 - Natio~'-r_-_u_s _________ _
`(f"e~~ -i .1:11 "' "' ....
`./" 7 Ji':; ·Y,.J.?.1 fS~ r(~I ~: 0
`\ .'~ ' •: r· TI'ORNEYSOOCl:ETNUlolHA· ' •
`- 709Pl
`'
`.
`
`j):t:\
`·I'
`"~ . ~,_,
`
`. l'l
`
`• .
`
`3.
`
`A copy of the International Application as filed (35 U .S.C. 371(c)(2})
`a. bl is transmitted herewith (required only if not transmitted by the International bureau).
`b. !l is not required, as the application was filed in the United States receiving Office (RO/US).
`c. D has been transmitted by the International Bureau.
`
`4 D A translation of the International Application into English (35 U.S.C. 371(c)(2)).
`
`5.
`
`Amendments to the claims of the International Application under PCT Anicle 19 (35 U.S.C. 371(c)(3))
`a D are transmitted herewith (required only if not transmitted by tll1e International Bureau).
`b. 0
`have been ITansmitted by the International Bureau.
`
`6. D A translation of the amendments to the claims under PCT Anicle 19 (3:5 U.S.C. 371(c)(3)).
`
`7. ~ An oath or declaration of the inventor (35 U .S.C. 371 (c)(4)).
`
`8. D A translation of the annexes to the International Prelimi'131Y Examinati1)n Report under PCT Article 36
`(35 U.S.C. 37 l(c)(5)).
`
`Other docwnent(s) or information included:
`9 . D An Information Disclosure Statement under 37 CFR 1.97 and 1.98.
`
`10. £]
`
`An assignment docwnent for recording.
`PLEASE MAIL THE RECORQED ASSIGNMENT DOCUMENT TO :
`a. Cl the person whose signature, name and address appears at the bottom of this page.
`b. D
`the following:
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`IL
`
`The above checked items are being transmited:
`a. D before the eighteenth (18) month publication.
`after publication or the Anicle 20 communication but before twenty (20) months ftom the priority date.
`b. D
`c. D
`after twenty (20) months but before twenty-two (22) months (surcharge and/or processing fee included).
`d . D
`after twenty-two (22) months (surcharge and /or processing fee included).
`NOTE: Petition to revive (37 CFR l.137(a) or (b)) is necessary irJS U.S.C. 371 requirements submitted
`after 22 months and NO pToper dcma11d foT lntcTflational p,~rliminary Examination was mode by 19 months
`fToM the earliest claimed prioritjl date.
`e. CD by thirty (30) months and a proper ctemand for International Prc:limina.ry Examination was made by tbe 19th
`month from the earliest claimed prio1ity date.
`after thiny (30) months bul before thiny-two (32) months and a1 proper demand for International Preliminary
`Examination was made by the 19th month from 1he earliest claimed priority dale (surcharge and/or processing
`fee included).
`g. D
`after thirty-two (32) months (surcharge and/or processing ree included).
`NOTE: Petition to revive (37 CFR 1.137(a) or (b)) is necessary irJS U.S.C. 371 requirements submitted
`after 32 months 1111d a pToper demand foT lntcmational Preliminary Examination was mode by 19 months
`/Tom the earliest claimed priority date.
`
`f. D
`
`12.
`
`At the time of transmittal, the time limit for amending claims under Article 19:
`lE has expired and no amendments were made.
`a.
`b. D
`has not yet expired.
`
`13. D
`
`I
`
`14 .
`
`Certain requirements under JS U.S.C. 371 were previously submitted by the applicant on - - - - - - - - (cid:173)
`namely:
`Submitted herewith are t Sequence Disk.eu:e. S.equence Listing. Preliminary
`Amendment:
`
`Janet E. Hasak
`"'AME
`ADDRESS Genentecb, Inc.
`
`$10 1"ATUl&
`
`lECISTRATIO"' "'UMll!I
`28,616
`
`E661 6 I i.~~:
`1 ~1 I '),: n.:r: J
`
`7 of 1033
`
`BI Exhibit 1002
`
`

`

`.
`
`I
`
`• CERTIFICATION UNDER 37 CFR 1 .10
`
`HB214759358US : Express Mail Number
`
`17 NOVEMBER 1993 : Date of Deposit
`
`I hereby certify that this request to initial national processing, including: TRANSMITTAL
`LETTER, PRELIMINARY AMENDMENT, SEQUENCE LISTING & DISKETIE, COMBINED
`DECLARATION & POWER OF ATTORNEY, ASSIGNMENT and COPY OF PRELIMINARY
`EXAMINATION REPORT is being deposited with the United States Postal Service "Express
`Mail Post Office to Addressee" service under 37 CFR 1.10 on the date indicated above and
`is addressed to the Commissioner of Patents and Trademarks, Washington, D.C. 20231.
`
`Carol Koehler
`
`8 of 1033
`
`BI Exhibit 1002
`
`

`

`•
`
`405
`
`HU405
`
`50
`40
`30
`20
`10
`OIVMTQSHKFMSTSVGORVSITCJCASQDVNTAVAWYQQXPGHSPICLLIYSASFRYT
`I
`1111
`I
`I
`I
`I I
`IJ I
`I
`I I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`OIQMTQSPSSLSASVGDRVTITCRASQDVNTAVAWYQQXPGKAPKLLIYSASFLES
`I
`I I I
`I
`I
`I
`I I I
`I
`I
`._ ______ _
`OIQMTQSPSSLSASVGDRVTITCRASQDVSSYLAWYQQICPGXAPXLLIYAASSLES
`-------------
`
`405
`
`HU4D5
`
`100
`90
`80
`70
`60
`GVPDRFTGNRSGTOFTFTISSVQAEOLAVYYCQQHYTTPPTFGGGTKLEIKRA
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`I
`GVPSRFSGSRSGTDFTLTISSLQPEDFATYYCQQHYTTPPTFGQGTKVEIKRT
`I
`I I
`I
`I
`I
`I
`I
`I I
`I
`I
`GVPSRFSGSGSGTDFTLTISSLQPEOFATYYCQQYNSLPYTFGQGTKVEIJCRT
`
`9 of 1033
`
`BI Exhibit 1002
`
`

`

`...
`. t.
`
`.,
`
`•
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`'
`
`'-' 1 i Uv '1t:.1 u) I 2 ~
`t7f114&~~
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`•
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`405
`
`HU40S
`
`50 A
`40
`30
`20
`10
`EVQLQQSGPELVKPGASLKLSCTASGFNIXDTYIHWVJ<QRPEQGLEWIGRIYPTN
`II
`11
`I
`I
`I
`I
`I
`I
`II
`II
`I I
`I I
`I
`I
`I
`I
`I
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`I I
`I I
`EVQLVESGGGLVQPGGSLRLSCAASGFNIXDTYIHWVRQAPGKGLEWVARIYPTN
`111 1111
`I 1111
`t I
`I I I I
`I
`I I I I
`EVQLVESGGGLVQPGGSLRLSCAASGFTFSOYAMSWVRQAPGKGLEWVAVISENG
`
`~--------
`Va•CDR1
`
`405
`
`HU40S
`
`90
`100ABC
`80 ABC ,
`70
`60
`GYTRYOPXFQOKATITADTSSNTAYLQVSRLTSEDTAVYYCSRWGGDGFYAMDYW
`11111111
`I
`I
`I l l It
`I
`II 11111
`I
`I
`I l l II
`I
`GYTRYADSVXGRFTISADTSKNTAYLQMNSLRAEDTAVYYCSRWGGDGFYAMDVW
`II
`I
`I
`I
`I
`t
`II 111111
`I I 111111
`I I I
`I
`I
`I
`I
`SOTYYADSVXGRFTISRDOSJCNTLYLQMNSLRAEOTAVYYCARDRGGAVSYFOVW
`
`405
`
`HU405
`
`110
`GQGASVTVSS
`I I
`I
`I
`GQGTLVTVSS
`
`GQGTLVTVSS
`
`10 of 1033
`
`BI Exhibit 1002
`
`

`

`. ..
`
`..· .•
`
`•
`
`•
`
`/Uv 7 '-' v ./ ~ ·L... v
`
`01/1/.ft_, ~b
`
`Anneal huVL or huVH oligomers to pA.Kl template
`
`3·_,A__,,__ _ _,A__,A__,A_~ 5'
`.....
`--····
`
`1. Ligate
`_
`2. Isolate assembled oligomers
`3. Anneal to pAKl template (XhoJ-. StuJ+)
`4. Extend and ligate
`
`t
`
`Xhol
`
`1. Transform E. coli
`2. Isolate phagemid pool
`3. Enrich for hu'l and huVH(Xho J+, SruJ-)
`4. Sequence verify
`
`pAK2
`
`11 of 1033
`
`BI Exhibit 1002
`
`

`

`•
`
`. . .
`
`100
`
`-= g _g 80
`§ t!
`.... ~
`o= -e
`5 c.. 60
`~= cf 8
`
`40
`
`huMAb4D5·8
`
`muMAb4D5
`
`4
`
`8
`12
`[MAb4D5 variant] µgtml
`
`16
`
`12 of 1033
`
`BI Exhibit 1002
`
`

`

`uruu 74! u.) 140
`- ~/
`• ' I /UV 7~ I
`,
`u .J .t L 0
`adv/465.20g
`'!/-'IC~~
`
`•
`
`\
`
`
`
`•
`
`•
`
`J
`
`13 0f1033
`
`BI Exhibit 1002
`
`13 of 1033
`
`BI Exhibit 1002
`
`

`

`(\(',·
`
`\ ·.
`'
`
`I
`
`•
`
`'
`
`•
`
`9''""'""' I ._ • .,, ., . • . '-...,
`o '-6/ J 4&, z...o l:,
`
`Vi.
`muxC03
`huxCD3vl
`huKI
`
`40
`10
`20
`30
`• • • • • •
`DIQMTQTTSSLSASLGDRVTISCRASQDIRNYLNWYQQKP
`**
`•
`*
`DIQMTQSPSSLSASVGDRVTITCRASQDIRNYLNWYQQKP
`# #
`#
`DIQMTQSPSSLSASVGDRVTITCRASQSISNYLAWYQQKP
`
`CDR-Ll
`
`muxCD3
`hwcC03vl
`huKI
`
`70
`60
`50
`80
`• • • • • •
`DGTVKLLIYYTSRLHSGVPSKFSGSGSGTDYSLTISNLEO
`*
`****
`*
`*
`• -~
`GKAPKLLIYYTSRLESGVPSRFSGSGSGTDYTLTISSLQP
`## '*
`#
`GKAPKLLIYAASSLESGVPSRFSGSGSGTDFTLTISSLQP
`
`CDR-L2
`
`muxCD3
`huxCD3vl
`huKI
`
`100
`90
`•••••••
`EDIATYFCQQGNTLPWTFAGGTKLEIK
`*
`•
`••
`•
`EDFATYYCQQGNTLPWTFGQGTKVEIK
`# #
`EDFATYYCQQYNSLPWTFGQGTKVEIK
`
`CDR-L3
`
`Va
`muxCD3
`huxCD3vl
`huIII
`
`40
`10
`20
`30
`• • • • •
`EVQLQQSGPELVKPGASMKISCKASGYSFTGYTMNWVKQS
`*
`•• **
`•
`• •••
`* *
`EVQLVESGGGLVQPGGSLRLSCAASGYSFTGYTMNWVRQA
`## ## # #
`EVQLVESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQA
`
`AAAA AAA
`
`CDR-Hl
`
`muxCD3
`huxCD3vl
`huIII
`
`50 a
`70
`60
`• • • • • • • • • •
`•
`HGKNLEWMGLINPYKGVSTYNOKFKDKATLTVDKSSSTAY
`* .;.. •• ••
`••
`••
`*
`*
`PGKGLEWVALINPYKGVTTYADSVKGRFTISVDKSKNTAY·
`## #### # #
`# #
`#
`PGKGLEWVSVISGrx;GSTYYADSVKGRFTISRDNSKNTLY
`
`CDR-H2
`
`muxCD3
`huxCD3vl
`huIII
`
`so abc
`110
`lOOabcde
`90
`••••••••
`MELLSLTSEDSAVYYCARSGYYGDSDWYFDVWGAGTTVTVSS
`•••• ••
`•
`•
`*
`LQMNSLRAEDTAVYYCARSGYYGDSDWYFDVWGQGTLVTVSS
`########### #
`LQMNSLRAEDTAVYYCARGRVGYSLSGLYDYWGQGTLVTVSS
`DE T
`S
`
`AAA.AA.AAA.AAA
`
`CDR-H3
`
`ftGuRf:.. 5
`
`14 of 1033
`
`BI Exhibit 1002
`
`

`

`., ...
`
`•
`
`··~ l/U~ 'J t. / ·U? :1. lb
`o~ /14~ 2.0ro
`
`PXGURB IA
`
`H52H4-160
`
`pH52-8.0
`
`H52.H4-160
`
`pHS.2-8.0
`
`H521H4-160
`
`pH5:2-8. 0
`
`H52H4-160
`
`pHS:?-8 . 0
`
`H52H4-160
`
`pH52-8 . 0
`
`H521il4-160
`
`pH52-8.0
`
`30
`20
`10
`QVQLQQSGPELVKPGASVKISCJCTSGYTFTE
`·*** ·** ** · ** · *·· · ** ********
`MGWSCIILFLVATATGVHSEVQLVESGGGLVQPGGSLRLSCATSGYTFTE
`so
`20
`30
`40
`10
`
`80
`70
`60
`50
`40
`YTMHWMl<QSHGKSLEWIGGFNPKNGGSSHNQRFMDKATLAVDKSTSTAYM
`******·* · **·***· · *· ******·********· *··**********
`YTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTISVDXSTSTAYM
`60
`70
`80
`90
`.
`100
`
`130
`120
`110
`100
`90
`ELRSLTSEDSGIYYCARWRGLNYGFDVRYFDVWGAGTTVTVSSASTKGPS
`.. ** ·**························· ** ************
`QMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTXGPS
`110
`120
`130
`140
`150
`
`180
`170
`160
`150
`140
`VFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVL
`****** *·*** · ************************************
`VFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVL
`160
`170
`180
`190
`200
`
`230
`220
`210
`200
`190
`QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDJCTH
`************** **··***** ***·********** ** * *
`QSSGLYSLSSVVTVTSSNFGTQTYTCNVDHKPSNTKVDKTVERJCCC---v
`210
`220
`230
`240
`
`280
`270
`260
`250
`240
`TCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVK
`*******
`·· ************************************* ·
`ECPPCPAPP-VAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQ
`250
`260
`270
`280
`290
`
`HS2.H4.-160·
`
`pH52-8.0
`
`330
`320
`310
`300
`290
`FNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCJ<VS
`*******·*************·*** · ********·***************
`FNWYVDGMEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVS
`300
`310
`320
`330
`340
`
`15 of 1033
`
`BI Exhibit 1002
`
`

`

`. l /Uv 7C..' VJ .L '- 0
`
`.0 8/ J 4to C.Ofo
`
`H52H4-160
`
`pHS2-8 . 0
`
`HS2H4-l60
`
`pH52-8.0
`
`H52H4-160
`
`pH52-8.0
`
`380
`370
`360
`350
`340
`NKALP~IE·KTISKAI<GQPREPQVYTLPPSREEMT~QVSLTCLVXGF'i'i'­
`** • *** ** * ** ** * · ** ****~~************•************•~
`NKGLPAPIEKTISKTI<GQPREPQVYTLPPSREEM'r.'ANQV;,LTCLVKGFYP
`350
`360
`370
`38U
`390
`
`430
`420
`410
`400
`390
`SDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFS
`·············~······························~·····
`SOIAVEWESNGQPENNYKTTPPMLDSOGSFFLYSKLTVDKSRWQOdN"'vFS
`400
`410
`420
`430
`440
`
`450
`440
`CSVMHEALHNHYTQKSLSLSPGK
`·····················~·
`CSVMHEALHNHYTQKSLSLSPGK
`450
`460
`
`16 of 1033
`
`BI Exhibit 1002
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`

`

`•
`
`r:IGtJRB 68
`
`HS2L6-158
`
`30
`20
`10
`DVQMTQTTSSLSASLGORVTINCRASQDINN
`*·****· ••••••·•••••• *********
`pH52-9.0 MGWSCIILFLVATATGVHSDIQMTQSPSSLSASVGORVTITCRASQOINN
`so
`20
`30
`40
`10
`so
`70
`60
`80
`40
`H52L6-158 YLNWYQQKPNGTVJ(LLIYYTSTLHSGVPSRFSGSGSGTDYSLTISNLDQE
`• ***************************·****·*· *
`*********
`YLNWYQQKPGKAPKLLIYYTSTLHSGVPSRFSGSGSGTDYTLTISSLQPE
`60
`70
`80
`90
`100
`
`pH52-9.0
`
`130
`120
`110
`100
`90
`H52L6-158 DIATYFCQQGNTLPPTFGGGTKVEIKRTVAAPSVFIFPPSOEQLKSGTAS
`*· ***·*******************************************
`OFATYYCQQGNTLPPTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
`110
`120
`130
`140
`150
`
`pHS2-9.0
`
`180
`170
`160
`150
`140
`H52L6-158 VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKOSTYSLSSTLTL
`**************************************************
`VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQOSKDSTYSLSSTLTL
`160
`170
`180
`190
`200
`
`pHS2-9.0
`
`HS2L6-158
`
`pHS2-9.0
`
`210
`200
`190
`SKADYEKHJ<VYACEVTHQGLSSPVTKSFNRGEC
`*********************************
`SKAOYEKHKVYACEVTHQGLSSPVTKSFNRGEC
`210
`220
`230
`
`17 of 1033
`
`BI Exhibit 1002
`
`

`

`••
`
`36,~d PCT/PTO 15JUN '92
`08/14G206
`
`I
`
`IMMUNOGLOBULIN VARIANTS
`
`--~--
`
`: 5
`
`10
`
`.. ... -..·
`
`· ~ .
`
`15
`
`20
`
`--·
`
`of the Invention
`
`:')./';~s invention relates to methods for the preparation and use of variant antibodies and
`ind~ application particularly in the fields of immunology and cancer diagnosis and therapy.
`'-
`
`~groynd of the Invention
`
`Naturally occurring antibodies (immunoglobulins) comprise two heavy chains linked
`
`together by disulfide bonds and two light chains, one light chain being linked to each of the
`heavy chains by disulfide bonds. Each heavy chain has at one end a variable domain (VH)
`.
`followed by a number of constant domains. Each light chain has a variable domain (VL) at one
`
`'
`
`end and a constant domain at its other end; the constant domain of the light chain is aligned
`with the first constant domain of the heavy chain, and the light chain variable domain is
`align•ed with the variable domain of the heavy chain. Particular amino acid residues are
`
`belie111ed to form an interface between the light and heavy chain variable domains, see e.g.
`Chotlhia eta/. , J . Mo/. Biol. 186:651-663 (1985); Novotny and Haber, Proc. Natl. Acad. Sci.
`
`USA 82:4592-4596 (1985).
`
`The constant domains are not involved directly in binding the antibody to an antigen,
`
`25
`
`but are involved in various effector functions, such as participation of the antibody in antibody-
`
`30
`
`depe1nd~nt cellular cvtotoxicity. The variable domains of each pair of light and heavy chains
`are involved directly in binding the antibody to the antigen. The do.mains of natural light and
`
`heav·~ chains have the same general structure, and each domain comprises four framework
`<FR) regio~s, whose sequences are somewhat conserved, connected by three hyper-variable
`or complementarity determining regions <CDRs) (see Kabat, E. A . et al. , Sequences of Proteins
`of lf!nmunological Interest, National Institutes of Health, Bethesda, MD, (1987)). The four
`
`framHwork regions largely adopt a P-sheet conformation and the CDRs form loops connecting,
`and in some cases forming part of, the P-sheet structure. The CD Rs in each chain are held in
`
`close proximity by the framework regions and, with the CDRs from the other chain, contribute
`
`18 of 1033
`
`BI Exhibit 1002
`
`

`

`•
`
`•
`
`to the formation of the antigen binding site.
`Widespread use has been made of monoclonal antibodies, particularly those derived
`
`s
`
`10
`
`15
`
`from rodents including mice, however they are frequently antigenic in human clinical use. For
`
`example, a major limitation in the clinical use of rodent monoclonal antibodies is an
`
`anti-globulin response during therapy (Miller, R. A. eta/., 8/ood62:988·995 (1983); Schroff,
`
`R. W. et al. , Cancer Res. 45:879·885 (1985)).
`
`The art hes attempted to overcome this problem by constructing "chimeric" antibodies
`
`in which en animal antigen-binding variable domain is coupled to a human constant domain
`
`(Cabilly et al .• U.S. patent No. 4 ,816,567; Morrison, S. L. et al., Proc. Natl. Acad. Sci. USA
`81 :6851-6855 (1984); Boulianne, G. L. et al., Nature 312:643-646 (1984); Neuberger, M . S.
`
`et al., Nature 314:268-270 (1985)). The term "chimeric" antibody is used herein to describe
`
`a polypeptide comprising at least the antigen binding portion of an antibody molecule linked
`
`to at least part of another protein (typically an immunoglobulin constant domain).
`
`The isotype of the human constant domain may be selected to tailor the chimeric
`
`antibody
`for participation
`in antibody·dependent cellular cytotoxicity
`(ADCC> and
`complement-dependent cytotoxicity (see e.g. Bruggemann, M . et al., J . Exp. Med.
`166: 1 351 - 1361 ( 198 7); Riechmann, L. et al. , Nature 332:323-327 ( 1988); Love et al .•
`
`Methods in Enzymology 178:515-527 (1989); Bindon et al .. J . Exp. Med. 168:127-142
`
`(1988).
`
`20
`
`In the typical embodiment, such chimeric antibodies contain about one third rodent (or
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`other non·human species) sequence and thus are capable of eliciting a significant anti-globulin
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`response in humans. For example, in the case of the murine anti·CD3 antibody, OKT3, much
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`of the resulting anti-globulin response is directed against the variable region rather than the
`constant region (Jeffers, G. J . et al., Transplantation 41 :572-578 (1986)).
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`2S
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`In a further effort to resolve the antigen binding functions of antibodies and to minimize
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`the use of heterologous sequences in human antibodies, Winter and colleagues (Jones, P. T .
`et al., Nature 321 :522-525 (1986); Riechmann, L. et al., Nature 332:323-327 (1988);
`Verhoeven, M. et al. , Science 239: 1534-1536 {1988)) have substituted rodent CDRs or CDR
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`sequences for the corresponding segments of a human antibody. As used herein, the term
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`"humanized" antibody is an embodiment of chimeric antibodies wherein substantially less than
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`an intact human variable domain has been substituted by the corresponding sequence from a
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`non-human species. In practice, humanized antibodies are typically human antibodies in which
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`some CDR residues and possibly some FR residues are substituted by residues from analogous
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`sites in rodent antibodies.
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`The therapeutic promise of. this approach is supported by the clinical efficacy of a
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`s
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`humanized antibody specif ic for the CAMPATH-1 antigen with two non-Hodgkin lym phoma
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`patients, one of whom had previously developed an anti-globulin response to the parental rat
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`antibody (Riechmann, L. et al. , Nature 332:323-327 (1988); Hale, G. et al. , Lancet
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`i :1394-1399 (1988)). A murine antibody to the interleukin 2 receptor has also recently been
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`humanized (Queen, C. et al. , Proc. Natl. Acad. Sci. USA 86: 10029-10033 (1989)) as a
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`potential immunosuppressive reagent. Additional references related to humanization of
`antibodies include Co et al., Proc. Natl. Acad. Sci. USA 88:2869-2873 ( 1991 >; Gorman et al.,
`Proc. Natl. Acad. Sci. USA 88:4181 -4185 (1991 ); Daugherty et al., Nucleic Acids Research
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`10
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`19(9): 2471-2476 (1991 ); Brown et al. , Proc. Natl. Acad. Sci. USA 88 :2663-2667 (1991 );
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`Junghans eta/. , Cancer Research 50:1495-1502 (1 990).
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`In some cases, substituting CDRs from rodent antibodies for the human CDRs in human
`frameworks is sufficient to transfer high antigen binding aff inity (Jones, P. T . et al., Nature
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`ts
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`321 :522-525 ( 1986); Verhoeven, M . eta/. , Science 239:1534- 1536 (1988)), whereas in other
`cases "it has been necessary to additionally replace one (Riechmann, L. et al. , Nature
`332:323-327 (1988)) or several
`(Queen, C. et al. , Proc. Natl. Acad. Sci. USA
`
`86: 10029- 10033 < 1989)) framework region (FR) residues. See also Co et al., supra.
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`For a given antibody a small number of FR residues are anticipated to be important for
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`antigen binding. Firstly for example, certain antibodies have been shown to contain a few FR
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`residues which directly contact antigen in crystal structures of antibody-antigen complexes
`
`(e.g., reviewed in Davies, D. R. et al., Ann. Rev. Biochem. 59:439-473 (1990)). Secondly,
`a number of FR residues have been proposed by Chothia, Lask and colleagues (Chothia, C. &
`Lask, A. M ., J . Mo/. Biol. 196:~01-917 (1987); Chothia, C. et al. , Nature 342:877-883
`
`(1989); Tramontano, A . et al., J . Mo/. Biol. 215: 175-1 $2 (1990)) as critically affecting the
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`conformation of particular CORs and thus their contribution to antigen binding. See also
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`30
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`Margolies et al., Proc. Natl. Acad. Sci. USA 72: 2180-2184 ( 1975).
`
`It is also known that, in a few instances, an antibody variable domain (either VH or VL)
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`may contain glycosylation sites, and that this glycosylation may improve or abolish antigen
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`binding, Pluckthun, Biotechnology 9 : 545-51 (1991 ); Spiegelberg et al. , Biochemistry 9 :4217-
`4223 (1970); Wallie eta/., J. Exp. Med. 168:1099- 1109 (19881; Sox eta/., Proc. Natl. Acad.
`Sci. USA 66:975-982 (1970); Margni et al., Ann. Rev. lmmunol. 6 :535-554 (1988) .
`Ordinarily, however, glycosylation has no influence on the antigen-binding properties of an
`antibody, Pluckthun, supra, (1991 ).
`
`The three-dimensional structure of immunoglobulin chains has been studied, and crystal
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`structures for intact immunoglobulins, for a variety of immunoglobulin fragments, and for
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`antibody-antigen complexes have been published (see e.g., Saul et al., Journal of Biological
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`Chemistry 25:585-97 (1978); Sheriff et al. , Proc. Natl. Acad. Sci. USA 84:8075-79 (1987);
`
`Segal et al., Proc. Natl. Acad. Sci. USA 71 :4298-4302 (1974); Epp et al., Biochemistry
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`14(22):4943-4952 (1975); Marquart etal. , J . Mo/. Biol. 141 :369-391 (1980); Furey eta/.,
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`J . Mo/. Biol. 167:661-692 < 1983); Snow and Amzel, Protein: Structure, Function, and
`Genetics 1 : 267-279, Alan R. Liss. Inc. pubs. ( 1986); Chothia and Lask, J. Mo/. Biol. 196:901 -
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`917 (1987); Chothia eta/. , Nature 342:877-883 (1989); Chothia eta/., Science 233:755-58
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`(1986); Huber et al., Nature 264:415-420 (1976); Bruccoleri et al., Nature 335:564-568
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`( 1988) and Nature 336:266 ( 1988); Sherman et al., Journal of Biological Chemistry 263:4064-
`
`4074 (1988); Amzel and Poljak, Ann. Rev. Biochem. 48 :961-67 (1979); Silverton et al., Proc.
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`Natl. Acad. Sci. USA 74:5140-5144 (1977); and Gregory et al., Molecular Immunology
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`24:821-829 (1987>. It is known that the function of an antibody is dependent on its three
`
`dimensional structure, and that amino acid substitutions can change the three-dimensional
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`structure of an antibody, Snow and Amzel, supra. It has previously been shown that the
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`antigen binding affinity of a humanized antibody can be increased by mutagenesis based upon
`molecular modelling (Riechmann, L. et al. , Nature 332:323-327 ( 1988); Queen, C. et al., Proc.
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`Natl. Acad. Sci. USA 86: 10029-10033 (1989)).
`
`Humanizing an antibody with retention of high affinity for antigen and other desired
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`biological activities is at present difficult to achieve using currently available procedures.
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`Methods are needed for rationalizing the selection of sites for substitution in preparing such
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`antibodies and thereby increasing the efficiency of antibody humanization.
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`The proto-oncogene HER2 (human epidermal growth factor receptor 2) encodes a
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`protein tyrosine kinase (p195HER2) that is related to and somewhat homologous to the human
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`epidermal growth factor receptor (see Coussens, L. et al., Science 230: 1132-1139 (1985);
`
`Yamamoto, T. et al., Nature 319:230-234 (1986); King, C. R. et al., Science 229:974-976
`
`(1985))~ HER2 is also known in the field as c-erbB-2, and sometimes by the name of the rat
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`homolog, neu. Amplification and/or overexpression of HER2 is as