`
`.
`
`·•
`
`FIG. 7 ( contd.)
`T F P . A V L. Q S S G L Y S L S S V V . T V
`170
`600 ACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTG
`
`en
`
`C: I
`I
`
`P S S S L _G T Q T Y I C N V N H K P S· N
`190
`660 - CCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAAC
`
`T K V D K K V E P K S. C D K T H T C P P
`210
`720 ACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCG
`
`230
`C P A P E L L G G P S V F . L F P P K. P K
`780 TGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAG
`
`S R T P E V T C V V V D V S H
`I
`D T L M
`250
`840 GACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCAC
`
`~
`"' N .....
`ff:
`N t
`
`--= 8 C'l = "' -..... = -"' .... QO
`
`N ..... -w
`
`w
`
`189
`659
`
`209
`719
`
`229
`779
`
`249
`839
`
`269
`899
`
`BIOEPIS EX. 1002
`Page 1501
`
`
`
`•
`
`•
`
`t
`
`FIG . 7 ( contd.)
`E D P E V K F N W Y V D G V E V H N A K
`GAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAG
`
`270
`900
`
`289
`959
`
`~
`0
`\C
`
`N ....... = UI
`
`N ......
`A
`
`t'J) c::
`
`m I 290
`a, i
`
`960
`
`310
`1020
`
`T K P R E E Q Y N S· T Y R -V V S V L T V
`ACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTC
`
`L H Q D W L N G K E Y K C K V S N K A L
`CTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTC
`
`309
`1019
`
`1079
`
`329 -LI.I w
`
`N
`N
`
`330
`1080
`
`PAP IE KT IS KA KG Q P R .E P Q V
`CCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGA.ACCACAGGTG
`
`350
`1140
`
`Y T L P P S R D E L T K N Q V S L T C L
`TACACCCTGCCCCCATCCCGGGATGAGCTGACCAAGAACCAGGTCAGCCTGACCTGCCTG
`
`349
`1139
`
`369
`1199
`
`"'O .
`
`g
`
`C') = IC -....... = -VI
`
`......
`co
`
`BIOEPIS EX. 1002
`Page 1502
`
`
`
`'
`
`en
`C
`IJJ
`
`~
`"' Cl)
`ffl
`
`FIG. 7 (contd.)
`V K G F Y P S D I A V E .W E S N G Q P E
`GTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAG
`
`370
`1200
`
`390
`1260
`
`N N Y K T T P P V L D S D G S F F L Y S
`AACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCT~TACAGC
`
`410
`1320'
`
`K L T V D K S R W Q Q G N V F S C S V M
`AAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATG
`
`430
`1380
`
`H E A L H N H Y T Q K S L S L S P G K Trm
`CATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA
`
`Hin:lIII
`1440 GTGCGACGGCCCCAAGCTT
`
`N
`UJ
`.......
`UJ
`UJ
`
`389
`1259
`
`409
`1319
`
`429
`1379
`
`448
`1439
`
`1458
`
`~
`0
`\Cl
`N ......
`
`Ii N :.:!
`
`.,,
`
`8 Cl ' -~ -UI
`
`oi!
`
`BIOEPIS EX. 1002
`Page 1503
`
`
`
`. ~
`
`FIG.a
`Q V Q L V E s G G G V V Q
`CDR 1
`F s I s
`y A
`CDR 2
`A I I
`I w D
`
`p G R s L R L s C s s s G F
`
`I
`
`~WV
`
`R Q A p G K G L E w V
`
`(/J
`C
`m
`
`~
`C
`"'
`en
`ffl
`
`1
`
`14
`
`34
`
`54
`
`74
`
`94
`
`D G s D Q H y A D s V· K
`
`GI R
`
`F T
`
`I
`
`s R D
`
`N s K N T L F L Q M D s L R p E D T G V
`CDR 3
`y F C A RID G G H G F C s s A s C F G p
`
`114 ~w G Q G T p V T V s s
`
`"
`
`13
`
`33
`
`53
`
`73
`
`93
`
`113
`
`126
`
`-
`
`N
`+"
`UJ
`UJ
`
`\C
`
`< 0
`N ...... = UI
`N ......
`
`,I:..
`
`"'O g
`c:') = IC -......
`Q -I.II
`
`.....
`
`QC
`
`BIOEPIS EX. 1002
`Page 1504
`
`
`
`"
`
`. ~
`
`en
`
`C ;
`I
`
`Hin::lIII
`AAGCTTATGAATATGCAAATCCTCTGAATCTACATGGTAAATATAGGTTTGTCTATACC
`
`FIG. 9
`
`1
`
`59
`
`60 ACAAACAGAAAAACATGAGATCACAGTTCTCTCTACAGTTACTCAGCACACAGGACCTCA 119
`
`-5
`-19
`I L F L V A T A T
`M G W S C I
`120 CCATGGGATGGAGCTGTATCATCCTCTTCTTGGTAGCAACAGCTACAGGTAAGGGGCTCA 179
`
`180 CAGTAGCAGGCTTGAGGTCTGGACATATATATGGGTGACAATGACATCCACTTTGCCTTT 239
`
`~ -UJ
`
`UJ
`
`1 3
`· G V H S Q V Q L V E S G G G V V Q
`-4
`240 CTCTCCACAGGTGTCCACTCCCAGGTCCAACTGGTGGAGTCTGGTGGAGGCGTGGTGCAG 299
`CDRl
`I F S I N Y G
`33
`P G R S L R L S C S S S G F
`14
`300 CCTGGAAGGTCCCTGAGACTCTCCTGTTCCTCCTCTGGATTCATCTTCAGTAACTATGGC 359
`CDR2
`34 M A I W · V R Q A P G K G L E W V A I T
`53
`I
`S H
`360 ATGGCCTGGGTCCGCCAGGCTCCAGGCAAGGGGCTGGAGTGGGTCGCAACCATTAGTCAT 419
`
`~
`IC
`~
`~ ......
`""
`
`~ tx:I
`IC -......
`c:i -!!l
`
`00
`
`BIOEPIS EX. 1002
`Page 1505
`
`
`
`. ..
`
`~
`
`"'
`
`FIG. 9(contd.)
`D G S D T Y F R D S V K G I R F T
`73
`S R D
`I
`54
`420 GATGGTAGTGACACTTACTTTCGAGACTCCGTGAAGGGCCGATTCACTATCTCCAGAGAT 479
`
`en
`
`C I ~ I
`
`93
`N S K N T L F L Q M D S L R · P E D T G V
`74
`480 AATAGCAAAAACACCCTATTCCTGCAAATGGACAGTCTGAGGCCCGAGGACACGGGCGTG 539
`CDR 3
`94 Y F C A R I Q G T
`I A G I R H I W G Q G T P
`113
`540 TATTTCTGTGCAAGACAAGGGACTATAGCAGGTATACGTCACTGGGGCCAAGGGACCCCC 599
`
`~ w
`
`w
`
`·. 118
`114 V T V S S
`600 GTCACCGTCTCCTCAGGTGAGTCCTTACAACCTCTCTCTTCTATTCAGCTTAAATAGATT 659
`
`660 TTACTGCATTTGTTGGGGGGGAAATGTGTGTATCTGAATTTCAGGTCATGAAGGACTAGG 719
`
`720 GACACCTTGGGAGTCAGAAAGGGTCATTGGGAGCCCGGGCTGATGCAGACAGACATCCTC
`
`779
`
`Ba.rrHI
`780 AGCTCCCAGACTTCATGGCCAGAGATTTATAGGGATCC
`
`817
`
`~
`~
`~
`,;!
`
`"1::1 8 Cl = '° -~ -UI
`
`iii!
`
`BIOEPIS EX. 1002
`Page 1506
`
`
`
`t
`
`-
`
`~
`
`..
`
`CJ)
`
`C: m
`~
`~
`I
`
`FIG .10
`
`I
`I L F
`M G W S C
`-19 HirrlIII
`1 AAGCTTTACAGTTACTCAGCACACAGGACCTCACCATGGGATGGAGCTGTATCATCCTCT
`
`-11
`60
`
`L V A T A T
`-10
`61 TCTTGGTAGCAACAGCTACAGGTAAGGGGCTCACAGTAGCAGGCTTGAGGTCTGGACATA
`
`-5
`120
`
`G V H S Q V
`-4
`121 TATATGGGTGACAATGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCACTCCCAGGTC
`
`2 N
`~
`180 ~
`
`Q L V E S G G G V V Q P G R S L R L S C
`3
`181 CAACTGGTGGAGTCTGGTGGAGGCGTGGTGCAGCCTGGAAGGTCCCTGAGACTCTCCTGT
`CDR 1
`I F S ! N Y G M A I W V R Q A P G
`S S S G F
`23
`241 TCCTCCTCTGGATTCATCTTCAGTAACTATGGCATGGCCTGGGTCCGCCAGGCTCCAGGC
`CDR 2
`K G L E W V A I T
`I
`S H D G S D T Y F R D
`43
`301 AAGGGGCTGGAGTGGGTCGCAACCATTAGTCATGATGGTAGTGACACTTACTTTCGAGAC
`
`22
`240
`
`42
`300
`
`62
`360
`
`. \C
`N
`
`~
`~
`
`C')
`
`"'d g
`! -~ ....
`
`!!,I
`l:IO
`
`BIOEPIS EX. 1002
`Page 1507
`
`
`
`. "
`
`,.
`
`..
`
`en
`
`C I
`I
`
`FIG . 10 ( contd.)
`S V K G ! R F T
`I S R D N S K N T L F L Q
`63
`361 TCCGTGAAGGGCCGATTCACTATCTCCAGAGATAATAGCAAAAACACCCTATTCCTGCAA
`CDR 3
`83 M D S L R P E D T G V Y F C A R I Q G T
`I
`421 ATGGACAGTCTGAGGCCCGAGGACACGGGCGTGTATTTCTGTGCAAGACAAGGGACTATA
`103 A G I R HI W G Q G T P V T V S S
`481 GCAGGTATACGTCACTGGGGCCAAGGGACCCCCGTCACCGTCTCCTCAGGTGAGTCCTTA
`
`82
`420
`
`102
`480
`
`122
`540
`
`N
`(X)
`
`OJ w
`
`541 CAACCTCTCTCTTCTATTCAGCTTAAATAGATTTTACTGCATTTGTTGGGGGGGAA.ATGT
`
`600
`
`601 GTGTATCTGAATTTCAGGTCATGAAGGACTAGGGACACCTTGGGAGTCAGAAAGGGTCAT
`
`660
`
`661 TGGGAGCCCGGGCTGATGCAGACAGACATCCTCAGCTCCCAGACTTCATGGCCAGAGATT
`
`720
`
`Ba.nHI
`721 TATAGGGATCC
`
`731
`
`~
`~
`~
`~
`
`.,,
`8 c:':I
`~ -......
`<:> ...
`~
`
`BIOEPIS EX. 1002
`Page 1508
`
`
`
`,.
`
`"
`
`,.
`
`..
`
`FIG .11
`HirxlIII
`AAGCTTATGAATATGCAAATCCTCTGAATCTACATGGTAAATATAGGTTTGTCTATACC
`
`1
`
`59 .
`
`en
`C:
`m
`~
`I
`
`60 ACAAACAGAAAAACATGAGATCACAGTTCTCTCTACAGTTACTCAGCACACAGGACCTCA 119
`
`-19
`MG W.S CI IL FL VAT AT
`-5
`120 CCATGGGATGGAGCTGTATCATCCTCTTCTTGGTAGCAACAGCTACAGGTAAGGGGCTCA 179
`
`180 CAGTAGCAGGCT.TGAGGTCTGGACATATATATGGGTGACAATGACATCCACTTTGCCTTT 239
`
`G V H S Q V Q L V E S G G G V V Q
`13
`-4
`240 CTCTCCACAGGTGTCCACTCCCAGGTCCAACTGGTGGAGTCTGGTGGAGGCGTGGTGCAG 299
`CDR 1
`33
`P G R S L R L S C S S S G F I F SIN Y G·
`14
`300 CCTGGAAGGTCCCTGAGACTCTCCTGTTCCTCCTCTGGATTCATCTTCAGTAACTATGGC 359
`CDR 2
`3 4 M A I W V R 'Q A P G K G L E W V A I T
`5 3
`I S H
`· 360 ATGGCCTGGGTCCGCCAGGCTCCAGGCAAGGGGCTGGAGTGGGTCGCAACCATTAGTCAT 419
`
`a
`~ m w
`
`~
`
`N '° ......
`
`w
`I.JJ
`
`"'Cl g
`
`Cl
`
`I -c:i -~
`
`00
`
`BIOEPIS EX. 1002
`Page 1509
`
`
`
`t
`
`"
`
`..
`
`~
`
`FIG .11 (contd.)
`D G S D T Y F R D S V K GI R F T
`73
`S R D
`I
`54
`420 GATGGTAGTGACACTTACTTTCGAGACTCCGTGAAGGGCCGATTCACTATCTCCAGAGAT 479
`
`en
`C: m
`~
`~
`I
`
`93
`7 4 N S K N T L F L Q M D S L R P E D T G V
`480 AATAGCAAAAACACCCTATTCCTGCAAATGGACAGTCTGAGGCCCGAGGACACGGGCGTG 539
`CDR 3
`94 Y F C A R I Q G T I A G I R H j W . G Q G T T
`113
`540 TATTTCTGTGCAAGACAAGGGACTATAGCAGGTATACGTCACTGGGGCCAAGGGACCACG 599
`
`LU
`C)
`
`....... w
`w
`
`118
`114 V T V S S
`600 GTCACCGTCTCCTCAGGTGAGTCCTTACAACCTCTCTCTTCTATTCAGCTTAAATAGATT 659
`
`660 TTACTGCATTTGTTGGGGGGGAAATGTGTGTATCTGAATTTCAGGTCATGAAGGACTAGG 719
`
`720 GACACCTTGGGAGTCAGAAAGGGTCATTGGGAGCCCGGGCTGATGCAGACAGACATCCTC 779
`
`Ba.nHI
`780 AGCTCCCAGACTTCATGGCCAGAGATTTATAGGGATCC
`
`817
`
`\,C)
`
`~
`~ UI
`~
`
`.,,
`g
`
`<:':I = \Cl -C) -UI
`
`.....
`00
`
`BIOEPIS EX. 1002
`Page 1510
`
`
`
`t
`
`"'
`
`,,...
`
`~
`
`-19
`1
`
`FIG .12
`Hirx:1III
`I L F
`M G W S C I
`AAGCTTTACAGTTACTCAGCACACAGGACCTCACCATGGGATGGAGCTGTATCATCCTCT
`
`-10
`61
`
`L V A T A T
`TCTTGGTAGCAACAGCTACAGGTAAGGGGCTCACAGTAGCAGGCTTGAGGTCTGGACATA
`
`G V H S Q V
`-4
`121 TATATGGGTGACAATGACATCCACTTTGCCTTTCTCTCCACAGGTGTCCACTCCCAGGTC
`
`C/J
`
`C I
`I
`
`3 Q L V E S G G G V V Q P G R S L R L S C
`181 CAACTGGTGGAGTCTGGTGGAGGCGTGGTGCAGCCTGGAAGGTCCCTGAGACTCTCCTGT
`CDR 1
`I F S IN Y G M A I W V R Q A P G
`S S S. G . F
`TCCTCCTCTGGATTCATCTTCAGTAACTATGGCATGGCCTGGGT.CCGCCAGGCTCCAGGC
`CDR 2
`K G L E W V A I T
`I S H D G S D T Y F R D
`43
`301 AAGGGGCTGGAGTGGGTCGCAACCATTAGTCATGATGGTAGTGACACTTACTTTCGAGAC
`
`23
`241
`
`63
`361
`
`S V K GI R F T · I S R D N S K N T L F . L Q
`TCCGTGAAGGGCCGATTCACTATCTCCAGAGATAATAGCAAAAACACCCTATTCCTGCAA
`
`-11
`60
`
`-5
`120
`
`2
`180 ·
`
`-..:i.
`
`w
`.......
`22 ~
`240
`
`42
`300
`
`62
`360
`
`82
`420
`
`\C
`N
`
`~
`~
`i!
`
`.,,
`8 ~
`~ -;;;;;
`....
`~
`
`BIOEPIS EX. 1002
`Page 1511
`
`
`
`~
`
`"'
`
`.,.
`
`.,
`
`FIG .12 (contd.)
`
`CDR 3
`83 M D S L R P E D T G V Y F C A R IQ G T I
`421 ATGGACAGTCTGAGGCCCGAGGACACGGGCGTGTATTTCTGTGCAAGACAAGGGACTATA
`103 A G I R HI W G Q G T T V T V S S
`481 GCAGGTATACGTCACTGGGGCCAAGGGACCACGGTCACCGTCTCCTCAGGTGAGTCCTTA
`
`en
`
`541 CAACCTCTCTCTTCTATTCAGCTTAAATAGATTTTACTGCATTTGTTGGGGGGGAAATGT
`
`601 GTGTATCTGAATTTCAGGTCATGAAGGACTAGGGACACCTTGGGAGTCAGAAAGGGTCAT
`
`C I
`I
`
`102
`480
`
`122
`540
`
`600
`
`660
`
`l.JJ
`N
`.......
`w w
`
`\C
`
`~
`N ' ~
`~
`
`"ti 8 C) = \Cl -' C> -~
`
`661
`
`TGGGAGCCCGGGCTGATGCAGACAGACATCCTCAGCTCCCAGACTTCATGGCCAGAGATT
`
`720
`
`BanHI
`TATAGGGATCC
`
`721
`
`731
`
`BIOEPIS EX. 1002
`Page 1512
`
`
`
`~·
`
`"
`
`00/492
`0 .5500
`0.5000
`0 .4500
`0.4000
`0.3500
`0.3000
`0.2500
`0. 20 00
`0 .1500
`0.1000
`0 .0 500
`0 .0000
`
`/
`
`• V
`
`j~
`
`,,
`
`I I
`
`100
`
`~
`jj
`:.')
`-I
`::j
`.-
`::i
`rn
`en
`J:
`m
`Ill -~
`
`,>
`
`FIG .13
`
`_. -
`~ -
`~··
`~
`~
`
`-
`
`-
`
`/ '
`
`/
`
`/
`
`"
`
`/
`r". ~
`__.-1 ~
`
`0.39
`156
`6.25
`25
`ANTIBODY CONC. ug / ml
`AVIDITY EXPERIMENT
`• CD4VHKOL-THR113 A YNB46,1.8
`
`0.097
`
`0.024
`
`0.0061
`
`1.1.J
`
`uJ -l.lJ
`
`\JJ
`
`~
`~
`~
`
`.,,
`g
`c:"l = IQ ...
`~ .....
`lll
`~
`
`BIOEPIS EX. 1002
`Page 1513
`
`
`
`INTERNATIONAL SEARCH REPORT
`lnternatlOMI Application Ho PCT /GB 91/01578
`I. CLASSIFICATION OF SUBJECT MATTER (if 1evenl classlflcallon symbols apply, Indicate all)"
`According to lntarnallonal Patent Classlficatlon (IPC) or lo botb National Clmllication and IPC
`IPC5: C 12 P 21/08, C 12 N 15/13, A 61 K 39/395
`
`II. FIELDS SEARCHED
`
`Classiricallon Sy1tem
`
`Minimum Documentation Sean:hed7
`Clnslficalion Symbols
`
`!PCS
`
`C 12 P; C 12 N; A 61 K
`Documentation Searched olber lhsn Minimum Documentation
`ID Iha Extent that such Documents are Included in Fields Saarcltadl
`
`ea111gory •
`X
`
`Ill. DOCUMENTS CONSIDERED TO BE RELEVANT•
`Citation of Dacument,11 with Indication, wbere appropriate, or Iha ra1-nt pasaagas 12
`WO, Al, 9007861 (PROTEIN DESIGN LABS, INC.)
`26 July 1990, see page 5; page 10,
`line 25 - page 14; page 28 - page 30
`
`y
`
`X
`
`y
`
`y
`
`--
`Proc. Natl. Acad. Sci., vol. 86, December 1989,
`Cary Queen et al.: 11A humanized antibody that
`binds to the interleukin 2 receptor u
`'
`see pages 10029-10033, page 10031 right
`column-page 10033
`
`--
`Nature, vol. 341, October 1989, E. Sally Ward et
`al.: 11Binding activities of a repertoire of
`single immunoglobulin variable domains secreted
`from Escherichia coli II
`see page 544 -
`'
`page 546
`--
`
`Relevant to Claim Na.1'
`1-5
`
`1-9
`
`1-5
`
`1-9
`
`1-9
`
`i'o~~ ~~~r,t~·Jn~~t~a,r:i,IJ!l!gx t:\•
`.,.. ii:.tfri!~r:i:r: ~~~
`Flied Ip understand Iha principle or theory undertying tbe
`1nvent1on
`•x• document of particular re13vance, the claimed. Invention
`cannot ba c;ansadered nove DI' cannot be considered to
`involve an mvenlive step
`"Y· document ol particular rt>levance. the claimed invention
`0;,llt';,":~~g =:t!~:ri;:tCe?~: :,.:ra~r: .~·~e~.~~~ :~~;d
`cannot be FOnsidered ta involve an Inventive step when the
`~
`in the art.
`·a• document member of the same patent ramily
`
`1l
`
`Date.of Mailing or this International Search Report
`
`O !1ffl 199Z'
`.-
`Signature of Authorized~!.///:,, ~
`'
`/
`- - - -~E~
`.c--
`
`• Special categories of ciled documents: 10
`·A· =~r:1::ddf!lcl,88,'l:..ft~f~'r!f:~:JJhe art whlcb is not
`•£• ~a,tlar document but published an or arter the lnlem1tlona1
`ltlang date
`•L• doa;umpnl which ma{; throw doubts pn nnoriix clalm(sh,or
`wluch 1s cited to es ablish the pubhca 011 da e of ano er
`citation or other apecial reason (as specified)
`•o• document referring to an oral disclosure, use, exhibition or
`other means
`•p• document cublished prior to ll)e international filing dale but
`later than he priority date claimed
`IV. CERTIFICATION
`Date al the Actual Complellon of the International Search
`16th December 1991
`
`International Searching Authority
`EUROPEAN PATENT OFFICE
`orm ,,.,,.,,..1210 \second sneet) \January 1sB:.)
`
`BIOEPIS EX. 1002
`Page 1514
`
`
`
`fCONTINUED FROM THE SECOND SHEET)
`Ill. DOCUMENTS CONSIDERED TO BE RELEVANT
`Citation of Docvmenl, with Indication. where apprapriale, of the relevant passages
`Catagory•
`
`Relevant to Claim No
`
`International Application Ho. PCT /GB 91/01578
`
`y
`
`X
`
`X
`
`A
`
`A
`
`A
`
`A
`
`Nature, vol. 332, March 1988, L Riechmann et
`al.: 11Reshaping human antibodies for therapy
`II , see page 323 - page 327
`.page 526 right column
`
`-
`EP, Al, 0328404 (MEDICAL RESEARCH COUNCIL)
`16 August 1989, see page 4; page 9,
`line 30; page 11, line 5
`--
`EP, A2, 0365209 (BECTON DICKINSON AND COMP~)
`25 April 1990,
`see in particular col. 3, lines 27-49 and
`columns 5-8
`
`--
`Proc.Natl.Acad.Sci., vol. 87, June 1990, J
`Sharon: 0 Structural correlates of high antibody
`affinity:Three engineered amino acid
`substitutions can increase the affinity of an
`anti-p-azophenylarsonate antibody 200-fold 11 ,
`see page 4814 - page 4817
`-
`Science, vol. 239, March 1988, M Verhoeyen et
`al.: 11Reshaping Human Antibodies: Grafting an
`Antilysozyme Activity 0
`, see page 1534 -
`page 1536
`
`-
`Nature, vol. 321, May 1986, PT Jones et
`al.: 11Replating the complementarity-determining
`regions in a humanantibody with those from a
`mouse 11
`, see page 522 - page 525
`page 525, left column
`
`--
`Nature, vol. 328, August 1987, S. Roberts et
`al.: 11Generation of an antibody with enhanced
`affinity and specificity for its antigen by
`protein engineering 0
`, see page 731 -
`page 734
`
`--
`
`1-9
`
`1-5
`
`1-5
`
`1
`
`1-9
`
`1
`
`1
`
`Fora PCT/ISA/ZlD Caxtra shaat) (January 198Sl
`
`BIOEPIS EX. 1002
`Page 1515
`
`
`
`ICONTINUED FROM THE SECOND SH~
`111. DOCUMENTS CONSIDERED TO BE RELEVANT
`Cllallon ol Document. with indication, Where appropriate, ol lhe relevant passages
`caieaory •
`
`Rel~t to Claim No
`
`lnlemational Application No. PCT /GB 91/01578
`
`P,X
`
`P,X
`
`P,X
`
`P,X
`
`WO, Al, 9109966 (ORTHO PHARMACEUTICAL CORPORATION)
`11 July 1991,
`see the whole document
`
`--
`WO, Al, 9107492 (CENTRAL BLOOD LABORATIORIES
`AUTHORITY) 30 May 1991,
`see page 3
`
`--
`EP, Al, 0403156.(GENZYME CORPORATION)
`19 December 1990,
`see example 12
`
`--
`WO, Al, 9109967 (CELLTECH LIMITED)
`11 July 1991,
`see the whole document
`
`-
`--------
`
`1-5
`
`1
`
`1-5
`
`1-9
`
`fora PCT.flSA/ZlO (extra shuU (January 198S)
`
`BIOEPIS EX. 1002
`Page 1516
`
`
`
`ANNEX TO THE INTERNATIONAL SEARCH REPORT
`ON INTERNATIONAL PATENT APPLICATION NO.PCT/GB 91/01578
`
`51310
`SA
`This annex lists the patent f.lmlly memban relating to the patent documents clled In the above-mentioned international surdl report.
`31/10/91
`TIie members ara aa contained In Iba Eurapaan Pa.tent Oflica l!DP file on
`The Europun Patent office Is In no way llable for lheseputlculars which are menily given ~or the purpose of Information.
`
`Patent document
`died In sun:11 rapart
`
`WO-Al- 9007861
`
`I
`
`Publlcatlon
`dale
`
`26/07/90
`
`I
`
`EP-Al- 0328404
`
`16/08/89
`
`EP-A2- 0365209
`
`25/04/90
`
`Patent f.lmlly
`member(sl
`
`I Publication
`
`dale
`
`i
`
`AU-0-
`CA-A-
`EP-A-
`AU-0-
`GB-A(cid:173)
`WO-A-
`
`JP-A(cid:173)
`
`5153290
`2006865
`0451216
`
`3062689
`2216126
`89/07452
`
`2238883
`
`13/08/90
`28/06/90
`16/10/91
`
`06/09/89
`04/10/89
`24/08/89
`
`21/09/90
`
`.
`
`.
`
`WO-Al- 9109966
`WO-A(cid:173)
`11/07/91
`11/07/91
`91/09967
`WO-A-
`91/09968
`11/07/91
`-----------------~--------------------------------------------------
`WO-Al- 9107492
`AU-0-
`13/06/91
`30/05/91
`6721490
`CA-A(cid:173)
`WO-A(cid:173)
`WO-A-·
`
`EP-Al- 0403156
`
`WO-Al- 9109967
`
`19/12/90
`
`11/07/91
`
`2018248
`
`91/09966
`91/09968
`
`07/12/90
`
`11/07/91
`11/07/91
`
`· For more details about lhis annex : sea Ofllcial Journal of the European patent Olllce, No. 12182
`
`EPO FORM P04711
`
`BIOEPIS EX. 1002
`Page 1517
`
`
`
`PCT
`
`WORLD INTELLECTUAL PROPERTY ORGANIZATION
`International Bureau
`
`INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCl)
`WO 92/15683
`
`(11) International Publication Number:
`
`Al
`
`(43) International Publication Date:
`
`17 September 1992 (17.09.92)
`
`(Sl) International Patent Oassification S :
`CtlN lS/13, C07K 13/00
`C12N 15/62, C12P 21/08
`A61K 39/395, 49/00
`
`(21) International Application Number:
`
`PCT /EP92/00480
`
`(22) International Filing Date:
`
`4 March 1992 (04.03.92)
`
`(30) Priority data:
`6 March 1991 (06.03.91)
`91 !03389.2
`(34) Countries for which the regional
`or intemational application
`was filed:
`
`EP
`
`(81) Designated States: AT (European patent), AU, BE (Euro(cid:173)
`pean patent), CA, CH (European patent), CS, DE (Eu(cid:173)
`ropean patent), DK (European patent), ES (European
`patent), FR (European patent), GB (European patent),
`GR (European patent), HU, IT (European patent), JP,
`KR, LU (European patent), MC (European patent}, NL
`(European patent), SE (European patent), US.
`
`AT et al. Published
`With international search report.
`
`(71) Appllcant (for all designated States except US): ME.~CK PA(cid:173)
`TENT GESELLSCHAFr MIT BESCHRANKTER
`HAFrUNG{DE/DE]; Frankfurter Str. 250, D-6100
`Darmstadt (DE).
`
`(72) Inventors; and
`(75) Inventors/ Applicants (for US only) : BENDIG, Mary, M.
`· [US/GB}; 64 Solent Road, West Hamstead, London
`NW6 ITX (GB). KETTLEBOROUGH, Catherine, A.
`[GB/GB); 28 Milton St., Watford, Herts, WD2 5W (GB).
`SALDANHA, Jose [GB/GB]; 22 Lincoln Way, Enfield,
`Middlesex ENI !TE (GB).
`
`(54) Title: HUMANIZED AND CHIMERIC MONOCLONAL ANTIBODIES
`
`(S7) Abstract
`
`The invention relates to new humanized monoclonal antibody comprising an artificial modified consensus sequence at
`least of the FRs of the heavy chain variable region of a human immunoglobulin. The invention relates, furthermore, to corre(cid:173)
`sponding humanized and chimeric monoclonal antibodies which are binding to epitopes of the Epidermal Growth Factor wher(cid:173)
`ein the responsible hypervariable regions have the following amino acid sequence: light chain:CDR-1: -Ser-Ala~Ser-Ser-SeraVal(cid:173)
`Thr-Tyr-Met-Tyr-; CDR-2: -Asp-Thr-Ser-Asn-Leu-Ala-Ser-; CDR-3: -Gln-Gln~Trp-Ser-Ser-His-Jle-Phe-Thr~; heavy chain:
`CDR-1: -Ser-His-Trp-Met-His,; CDR-2:. -Glu-Phe-Asn-Pro-Ser-Asn-Gly-Arg-Thr-Asn-Glu-Lys-Phe-Lys-Ser~;. CDR-3: -Arg(cid:173)
`Asp-Tyr-Asp-Tyr-Asp-Gly-Arg-Tyr-Phe~Asp-Tyr-. The antibodies can be used for therapeutical and diagnostic purposes.
`
`:·,:~.·
`...
`
`4
`
`BIOEPIS EX. 1002
`Page 1518
`
`
`
`...
`
`• -,
`•
`
`FOR THE: PURPOSES OF INFORMATION ONLY
`
`Cude:; uSt:d lo identify S1ali:~ 11any lo the PCf on the f1onl pages or pam1>hlets publishing international
`a1J11lications under the PCT.
`
`AT
`All
`HB
`BE
`BF
`BG
`BJ
`HR.
`CA
`CF
`·("G
`CH
`(:'I
`CM
`cs
`OE
`UK
`ES
`
`Au:.tria
`Australia
`H.irhildo,
`Hi:li;ium
`Burkina f..ct11
`· Bulgaria
`e(...,lin
`B~a,il
`("anada
`C,mtral African Kc:11uhlii:
`(uni.~•
`Swil7cda11d
`( 'olc ,11,uirc
`,·anu:ruon
`C1c:chu:1luvitLia
`(jurn,any
`Dc111n.ark
`
`s, ... ; ..
`
`,..
`
`, ...
`
`FR
`GA
`GB
`GN
`CR
`HU
`IE
`11'
`JP
`kP
`
`KR
`u
`LK
`LU
`MC
`MG
`
`Vinland
`r,aiu:a:
`UJhun
`Uuiri:d t.:i11L~lum
`(.i-oinc.:a
`Urccc.:
`HUIIJ:ilr)
`Ireland
`llid)
`Japan
`l.lcmoo:rJlic Pcu11h:'i, Ki:public
`. ur Kurca
`K<.11ublic: ur Korea
`l.ii:ch1c:n,1cin
`Sri l.anl.r
`l.u•cmhuurg
`Mun.ico
`M.nt.,g=Jr
`
`Ml.
`MN
`MR
`MW
`NI.
`NO
`PL
`RO
`RU
`SD
`SE
`SN
`SU
`TD
`TG
`us
`
`Mali
`Moni.'Ulia
`Mauritani.&
`M.dawi
`Nc:rhr:rland,
`Norwa)
`PolJnJ
`Roniania
`KU>Sian Fi:Jcration
`Sudan
`Sweden
`Senegal
`Sovii:t Uniun
`C'had
`Tugo
`Unh.:d Scales of An,cri<;a
`
`BIOEPIS EX. 1002
`Page 1519
`
`
`
`W092/15683
`
`PCT/EP92/00480
`
`1
`
`· Humanized and Chimeric Monoclonal Antibodies
`
`,.
`
`5
`
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`15
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`
`25
`
`30
`
`•
`
`TECHNICAL FIELD OF THE INVENTION
`
`The invention relates to new humanized monoclonal antibodies
`comprising·an artificial modified consensus sequence at least
`of the FRs in the variable region of the heavy chain of human
`immunoglobulins.
`
`The invention relates, furthermore, to humanized and chimeric
`monoclonal antibodies which are binding to epitopes of the
`Epidermal Growth Factor. The invention disclQSes the amino
`acid sequences of the responding antigen-binding site for
`this receptor.
`
`The invention relates to pharmaceutical compositions·compris(cid:173)
`ing the said antibodies for the purposes of treating tumors
`like melanoma, glioma or carcinoma. The said.antibodies can
`be used also for diagnostic applications regarding locating
`and assessing the said tumors in vitro· or in vivo.
`
`The specification relates to several technical terms which
`are here defined as follows.:
`
`BIOEPIS EX. 1002
`Page 1520
`
`
`
`W092/15683
`
`PCT/EP92/00480
`
`"Humanized" antibodies mean antibodies comprising FRs of the
`variable regions and constant regions of amino acids located
`in the light and heavy chain which derive from human sources
`.whereas the hypervariable regions derive from non-human
`sources.
`
`•· ..
`
`"Chimeric" antibodies mean antibodies comprising variable and
`hype+Variable regions which derive from non-human sources
`whereas the constant regions derive from human origin.
`
`11FRs 11 ·mean the framework regions of an antibody and are found
`within the variable regions. In these regions a certain
`alteration of amino acids occurs.
`
`"CDRs" mean the complementarity determining or "hypervari(cid:173)
`able" regions of an antibody and are found within the vari(cid:173)
`able regions. These.regions represent the specific antigen(cid:173)
`binding site and show an immense exchange of amino acids.
`CDRs are primarily responsible for the binding affinity of
`the antigen.
`
`"Consensus sequence 11 means a non-naturally occurring amino
`acid sequence as light or heavy chain variable regions and is
`used as substitute for the originally present non-human heavy
`or light chain variable regions. The consensus sequences is
`synthetic and therefore an artificial sequence of the most
`common amino acids of a distinct class or subclass or sub(cid:173)
`group of heavy or light chains of human immunoglobluins.
`
`5
`
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`
`"EGF" and "EGFR" mean the Epidermal Growth Factor and its
`receptor.
`
`•
`
`BIOEPIS EX. 1002
`Page 1521
`
`
`
`W092/JS683
`
`PCT/EP92/00480.
`
`- 3 -
`
`"VL" regions mean light chain variable regions.
`
`"VH" regions mean heavy chain variable regions.
`
`5
`
`BACKGROUNDOF THE INVENTION
`
`(MAb 425) was raised
`The murine monoclonal antibody 425
`against the human A431 carcinoma cell line and found to bind
`. to a polypeptide epitope on the external domain of the human
`epidermal growth factor receptor (EGFR). It was found to
`inhibit the binding of epidermal growth factor (EGF) at both
`low and.high affinity EGFR sites (Murthy et al., 1987),
`Enhanced expression of EGFR is found to occur on malignant
`tissue from a variety of sources thus making MAb 425 a possi(cid:173)
`ble agent for the diagnosis and therapeutic treatment of
`human tumors. Indeed, MAb 425 was found to mediate tumor
`cytotoxicity in vitro and to suppress tumor cell growth of
`epidermoid and colorectal carcinoma-derived cell lines in
`vitro (Rodeck et al., 1987). Radiolabelled MAb 425 has also
`been shown to bind to xenografts of human malignant gliomas
`in mice (Takahashi et al., 1987).
`
`EGF is a polypeptide hormone which is mitogenic for epidermal
`and epithelial cells. When EGF interacts with sensitive
`cells, it binds to membrane receptors; the receptor EGF
`complexes cluster and then are internalized in endocytotic
`vesicles. This is responsible for the phenomenon.of "down(cid:173)
`regulation". EGF binding induces a tyrosine kinase activity
`of the receptor molecule and induces synthesis of DNA.
`
`10
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`
`BIOEPIS EX. 1002
`Page 1522
`
`
`
`W092/15683
`
`PCT/EP92/00480
`
`- 4 -
`
`The EGF~receptor is a transmembrane glycoprotein of about
`170,000 Daltons (Cohen, 1982). It is the gene product of the
`c-erb-B proto-oncogene (Downward et al., Nature. Vol. 307,
`pp. 521-527, 1984). The receptor exists in two kinetic forms:
`so-called low affinity and high-affinity receptors.
`
`..
`
`The A431 carcinoma cell line expresses abundant EGF-receptors
`on its cell surfaces, and thus has been used in many studies
`to generate anti-EGF-receptor antibodies. However, the recep-
`tors on A431 differ from those of other cell types in the
`carbohydrate moieties attached to the polypeptide. Thus many·
`antibodies raised against A431 membranes are directed against
`carb~hydrates which are not common to all forms of the recep(cid:173)
`tor molecule (e.g. Schreiber, 1983).
`
`I
`
`Other monoclonal antibodies are reactive with the protein
`moiety of EGF-receptors. These antibodies display a variety
`of properties upon binding to EGF-receptors, presumably
`dependent on the particular portion of the receptor molecule
`bound, and the isotype of the antibody. Some antibodies mimic
`some of the effects of EGF {agonists} and some inhibit the
`effects (antagonists).
`
`Expression of EGF-receptors has been implicated in the pro-
`gression of tumor growth. The gene for the receptors has been
`found to be the cellular analogue of the avian viral oncogene
`v-erb-B (Ulrich, 1984). In addition an association has been
`detected between late stages of melanoma devel,opment and
`extra copies of the chromosome carrying the receptor gene
`(Koprowski et al., Somatic Cell and Molecular Genetics, Vol.
`11, pp. 297-302, 1985).
`
`•
`
`5
`
`10
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`15
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`20
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`25
`
`30··
`
`BIOEPIS EX. 1002
`Page 1523
`
`
`
`•
`••
`
`W092/15683
`
`PCT/EP92/00480.
`
`- 5 -
`
`Because of EGF-receptors are expressed on a wide variety of
`solid tumors they provide a suitable target for anti-tumor
`therapy. However, there is a need in the art for a suitable
`anti-receptor antibody. Many of the known antibodies have
`properties which would be deleterious if used as anti-tumor
`agents. For example, antibodies which mimic the effects of
`EGF could stimulate the progression of the tumor rather than
`arresting it. Other antibodies which only bind to high or low
`affinity receptors could be less than optimally effective
`because EGF could still exert its effect through the unbound
`receptors. Still other antibodies convert low affinity recep(cid:173)
`tors to high affinity receptors, which could exacerbate tumor
`gro~h rather than inhibiting it. Thus there is a need in the
`art for an anti-EGF-receptor antibody which would be suitable
`for anti-tumor therapy.
`
`Although murine MAbs have been used for therapeutic treatme~t
`in humans, ·they have elicited an immune response (Giorgi et
`al., 1983; Jaffers et al., 1986) . To overcome this problem,
`several groups.have tried to "humanize" murine antibodies.
`This can involve one of two approaches. Firstly, the murine
`constant region domains for both the light and heavy chain
`can be replaced with human constant regions; Such "chimeric"
`murine-human antibodies have been successfully constructed.
`from several murine antibodies directed against human tumor-.
`associated antigens (Sun et al., 1987; Whittle et al., 1987;
`Liu et al., 1987; Gillies and Wesoiowski, 1990). This
`approach totally conserves the antigen-binding site of the
`murine antibody,. and hence the antigen affinity, while .. con-
`ferring the human isotype and effector functions. In the
`second approach only the complementarity determining regions
`
`5
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`
`BIOEPIS EX. 1002
`Page 1524
`
`
`
`W092/15683
`
`PCT/EP92/00480
`
`-6-
`
`(CDRs) from the mouse variable regions are grafted together
`with
`human framework regions (FRs) of both the light and
`heavy chain variable domains (VL and VH). It is reasoned that
`
`this technique will transfer the critical and major portion
`of the antigen-binding site to the human antibody (Jones et
`al., 1986).
`
`•
`
`CDR grafting has been carried out for several rodent mono(cid:173)
`clonals (Jones et al., 1986; Reichmann et al., 1988; Verhoe-
`yen et al.; 1988; Queen et al.; 1989; Co et al., 1991; Gorman
`et al., 1991; Maeda et al., 1991; Temptest et al., 1991). All.
`retained their capacity to bind antigen, although the affin(cid:173)
`ity was usually diminished. In most cases it was deemed
`necessary to. alter certain amino acids in the human framework
`residues (FRs). Both chimeric and CDR grafted antibodies have
`proved superior to the mouse antibodies in the clinic (Hale
`et al., 1988; LoBuglio et al., 1989; Mathieson et al., 1990).
`However, a general teaching of which amino acids have to be
`changed, is not known and not completely predictable in any
`case.
`
`EP 088 994 proposes the construction of recombinant D~
`vectors comprising of a DNA sequence which codes for a vari(cid:173)
`able domain of a light or a heavy chain of an immunoglobulin
`specific for a predetermined ligand. The application does not
`contemplate variations in the sequence of the variable
`domain.
`
`EP 102 634 describes ~he cloning and expression in bacterial
`host organisms of genes coding for the whole or a part of
`human IgG heavy chain polypeptide, but does not contemplate
`variations in the sequence of the polypeptide.
`
`5
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`
`BIOEPIS EX. 1002
`Page 1525
`
`
`
`W092/l5683
`
`PCTiEP92/00480
`
`..
`
`EP 239 400 proposes that humanized antibodies cart be obtained
`by replacing the antigen-binding site (hypervariable regions)
`of any human antibody by an antigen-binding site of a non-hu(cid:173)
`man, for example of a mouse or a rat antibody by genetechno(cid:173)
`logical met"hods.
`
`Thus, following this teaching, human or humanized antibodies
`can be manufactured having specific antigen-binding sites
`which were not available up to now in antibodies originating
`from humans.
`
`Chimeric antibodies can be obtained by replacing not only the
`CDRs but the whole variable regions of the light and heavy
`chains. Chimeric antibodies, however, can still be imrnuno-
`genie. Chimeric antibodies are, however, very useful for
`diagnostic purposes and optimizing humanized antibodies.
`
`It could be shown that the affinity of the antigen-binding
`sites can be influenced by selective exchange of some sing+e
`amino acids within the variable regions which are not
`directly part of the CORs (Reichmann et al., 1988).
`
`As consequence in the worst case, the binding affinity of the
`antigen can be completely lost if one works according to the
`teaching of the EP 239 400. This fact could be demonstrated
`by the inventors of the instant invention, who failed in
`constructing a correspondingly humanized antibody which was
`directed to epitopes of the EGF-receptor.
`
`Therefore, it ~st be considered that the success of such a
`humanization depends on the constitution and conformation of
`the used variable regions and their interactions with the
`
`5
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`
`BIOEPIS EX. 1002
`Page 1526
`
`
`
`W092/15683
`
`PCT/EP92/00480
`
`- 8 -
`
`.corresponding antigen-binding site. Thus, it is not com(cid:173)
`pletely predictable whether or which modifications within the
`variable domains of the antibody are necessary in order to
`obtain or to improve the binding of the antigen to the anti(cid:173)
`body.
`
`..
`
`•
`
`SUMMARY OF THE INVENTION
`
`Thus, the invention has the object of providing a humanized
`monoclonal antibody which is, in particular, directed to the
`EGF-receptor, comprising an antigen-binding site of non-human
`sources and the FRs of the variable regions and constant
`regions of human origins, which are, if necessary, modified
`in a way that the specificity of the binding site can be
`conserved or restored.
`
`In particular, the invention has the object of characterizing
`the hypervariable regions of the antigen-binding site of an
`antibody against the EGF-receptor and providing these CDRs
`within a humanized monoclonal antibody defined as above.
`
`This antibody and its chimeric variant can play an important
`role.as a therapeutic or diagnostic agent in order to combat
`tumors, as melanoma, glioma or carcinoma.
`
`It has been found, that effective and specific humanized
`monoclonal antibodies can be easily obtained by using a
`consensus sequence of at least the heavy chain variable
`regions of human inununoglobulins. In particular, all those
`consensus· ·sequences are suitable which have a good Cat least
`60-70 %, particularl