S22403S
`
`UNITED STATES DEPART.\IE~T OF COMMERCE
`United States Patent and Trademark Office
`
`March 14, 2022
`
`THIS IS TO CERTIFY THAT A~TEXED HERETO IS A TRUE COPY FROi\1
`THE RECORDS OF THIS OFFICE OF:
`
`Pre-Grant Publication Number: 2004/0126363
`Pre-Gr ant Publication Date: July 1, 200.:f.
`
`By Authority of the
`Under Secretary of Commerce for Intellec al Property
`and Director of the United States Patent d T ·ademark Office
`
`• /4
`C rt .'ing Office1· tJ
`
`y la o~
`
`Miltenyi Ex. 1007 Page 1
`
`

`

`1111111111111111 IIIIII IIIII 11111 1111111111 lllll lllll lllll 111111111111111 1111111111 111111111
`US 20040126363A I
`
`(19) Uni1ted States
`c12> Patent Application Publication
`Jensc10 et al.
`
`( JU) Pub. No.: US 2004/0126363 Al
`Jul. 1, 2004
`(4:1) Pub. Date:
`
`(54) C l>19-SPl~CfFJC REDIIU◄:CTlm IMMUNI<:
`CELLS
`
`(76)
`
`lnventors: Miclmcl C . .Jensen, Pa~adena. CA
`(US); Stephen Forman, San Marino,
`CA (US); Andrew Raubitschek, San
`Marino, CA (US)
`
`Correspondence /\ddrcf<.-;:
`ROTt-llWf+:LI,, FIGG, ERNST & MANHECK,
`P.C.
`1425 K STREE'J~ N. W.
`SUITE 800
`WAS.HINGTON, De 20005 (US)
`
`(21) Appl. No.:
`
`l0/416,01 I
`
`(22) PCT Filed:
`
`Nov. 7, 2001
`
`(86) PCl' Nlo.:
`
`PCT/US0J/42997
`
`Related U.S. Applicntion Data
`
`(60) Provisiional ~pplication No. 60/24G, I I 7, lilcd on Nov.
`7,2000.
`
`J'ublication Classification
`
`lnt. C t.7 ............................. A61K 48/00; Cl2N 5/08
`(51)
`(52) U.S. Cl .
`...................... 424/93.21; 424/93.7; 43.5/372
`
`(57)
`
`ABSTRACT
`
`Genetically engineered, CDl 9-spcdfic redirected [rnmuoe
`cells expressing s cell surface protcio baving an extracdlu lar
`domain comprising a receptor wbieh i.s specific for CD l 9. au
`iotraudlular signaling domain, am.I
`transmernbrane
`II
`domain. Use of sucb cells for cellu lar immunotherapy of
`CDJ 9+ malignaacies aad f0r abrogating any untoward B cell
`ruoctioo. To one embuclinu:nt, th.: immuo..: c.:ll is a Tcell and
`the cell surface protei n is a single chain scfvFc:t rec:eptor
`where scFv designates the VH and V t. chains of a i;ingle
`cbain monoclonal antibody to CD I 9, Fe represents al least
`par! of a coaslaol region of ao lgG,, and t rcprcscnL-; tbc
`ia1racellular signaling domain of lho zcia chain of bumao
`CD3. Tbe extracellular domain sc¥v11c and th.: intrac~,Uular
`domain t arc linked by a lransmcmbrane domain such ~s tbll
`lransmembrane domain of CD4. A metbocl of ma.king a
`redirected T cell expressing n chimeric T cell receptor by
`lllecLroporation u.sing naked DNA encoding Lhe receptor.
`
`Miltenyi Ex. 1007 Page 2
`
`

`

`Patent Application Publication
`
`Jul. 1, 2004 Sheet 1 of 8
`
`US 2004/0126363 Al
`
`Xba!
`
`(l'JGMCSF s; .:;na l ;:ieptide-)
`s
`::
`i..
`!..
`'.'
`-:-
`!.
`L ~ !..
`L
`?
`£
`~
`ATCTCTAGAG CCGCCACCAT GCTTCTCCTG GiGACAAGCC TTCTGCTC".'G ,GAGTrAC::CA
`TAGAGATCTC GGCGGTGGTA CGAAGAGGAC CACfG17GGG AAGACGAGAC AC':-CAATGGT
`
`(V~ - l
`?
`::>
`-
`CCCAGACATC
`GGGTCTGTAG
`
`F L ~
`~ P A
`:l CACCCAGCAT TCCTCC~GAT
`G:GGGICGTA AGGAGGACTA
`
`0:-1~ Q 7 7 5 5~3
`CAGATGACAC AGACTACA~C C1CCCTG7C!
`GTCTACTGTG TCTGA:~TAG G~GGGACAGA
`s C
`., R A
`s
`T
`rt
`A S
`G 0
`i21 Gcc::'!'~~GG G_-,3ACAGAG7 ~ACCAIC;G: 7~=AGGGCM ~T:A~·~M:;~.-r -:.;G:;.,;....:.:-:.:..:(cid:173)
`CGGAuAi:iAC-'.: CTCTG7C':'CA :;TGGTAGTCA P.CGTCC':GTT CAGT::C7G7A A,CA:"!'TA,A.
`
`L N W Y Q Q K
`H ~ S
`L L ~ Y
`P D G T V K
`1B l TTM-.ATTGGT ATCAGGAGAA ACCAGATGGA ACTGTTAAAG TCCTGATCTJI CCJI .,.\CATCA
`AATTTAACCA TAGTCGTCTT TGGTCTACCT TGACAATTTG AGGAGTAGAT GGTATGTAGT
`s G s G s G T
`s R F
`s G V
`p
`R L H
`D
`5
`'{
`20 AGATTACACT CAGGAGTCCC ATCAAGGTTC' ?.GI'GGCAGTG GGTCTGGAAC AGATTATTCT
`TCTAATGTGA GTCCTCAGGG TAGTTCCAAG TCACCGTCAC CCAGACCTTG TCTAATAAGA
`
`L T
`s N L E
`y
`I
`1 A T
`Q E
`0
`3Cll C':'::ACCATTA GCAACCTGGA GCAAGAAGAT ATTGCCACTT ACTTTTGCCA ACAGGGTAA'!
`GAGTGGTAAT CGTTGGACCT GGTTCTTCTA ,AACGGTGAA TGAAAACGGT TGTCCCATTA
`
`r C Q " G N
`
`42:C
`
`:;
`s
`:<
`:'CCGGCA,;]:
`,l.~Gcc::;:-:-::G
`
`:;
`
`::;
`
`,Wr.itlow linker- )
`. G G G T
`Y r
`I T G 5
`T S G
`E
`-
`K
`T L
`?
`361 ACGCT~CCGT ACACGTTCGG AGGGGGGACT AAGT;GGAAA TAACAGGCTC CACCTCTGGA
`;GCGAAGGCA TG,GCAAGC: rc::ccccTGA ~-=:AACCTTT,ATTGTCCGAG GTGGAGACC7
`:·,~- :
`-
`C
`K
`::; S
`-
`~ ~ ~ E
`~
`:GAGGG;..rcc- Ac:.:..AGGGCG AG3TGAAACT GCAGGAGTCr.
`Gc:::c::ASG :GGT':'CCCGC TCCACTTTGA CGTCCTCAGT
`....
`..,
`s
`s V
`s
`T C
`s G
`'! V
`'I
`CTCAC,t,GAGC CTG7CCG7'C.A CA7G~ACTG7 -::TCAGGGG!'C
`GAGTG7CTCG GACAGGCAGT GTA.CGTGACA GAGTCCCCAG
`.
`-
`s
`::. E w
`:-;
`F.. K G
`p
`<'
`?. Q
`AAGCTGGATT CGCO.GCCT:: CACG/1.AAGGG TC':'GGAGTGG
`T'!'CGF>':C':'AA SCGG7CGGAG G7GC'!'TTCCC AGACCTCACC
`
`4 BJ,
`
`E' G
`G
`GGACCTGGCC
`ccr:;GACCGG
`
`;,
`!.,
`TGG!GGCGCC
`ACCACCGCGG
`
`541
`
`s L
`?
`TCATr;.. . ;.:G
`AGTAATGGGC
`
`y ~ ' /
`C
`-~C7AT~::.;-:'G":'
`!GATAcc;..:;..
`
`601
`
`L G
`·;
`CTGGGAG':'AA
`GACCCT:ATT
`
`.:;
`;;
`5
`TATGGGG':'A:;
`ATAcccc;;:-=::
`
`~
`
`.
`s A
`·,t N
`.
`E
`5 R L
`K
`J'GAAACCACA :'ACTATAATT CAGCTCTCAA ATCCAGACTG
`ACTTTGGTGT .ll,TGA:'ATTAA GTCGAGAGTT TAGGTCTGAC
`
`·.J
`
`Q T
`~ M ~ S
`~ ~ X 5
`7
`~ S Q
`~
`r
`- -rTT~T':'AA AAATGAACAG TC~GCAAACT
`661 ACCATCA1-A AGGACi....AC:'= ~AAGAGc~·-
`TGGTAGTAGT TCCTGTTGAG GTTCTCGG, T ~AAAAGAATT TTTACTTGTC AGACGTTTGA
`
`y A ~
`~ G G s
`~ A K
`-
`A
`y
`y
`~ ~ -
`~ V
`V
`721 GATGACACAS C~ATTTACTA C:GTGCCAAA CA~TAfTACT ACGGTGGTAG CTATGCTATG
`CTAC~GTGTC GGTA.nJI.TGA! GACACGGTTT ~,AA,AATGA TGCCACCATC GATACGATAC
`
`fIGURE. 1 lA
`
`Miltenyi Ex. 1007 Page 3
`
`

`

`Patent Application Publication Jul. 1, 2004 Sheet 2 of 8
`
`US 2004/012636~1 A l
`
`~ S
`3 V
`D Y W G Q ~
`7 8 'i GAC' AC-:'GGG ::;rcAAGG;.,;: :7CAGTCAC.::: :;: =::::c:-CAG
`CTGATGACCC :,;::;::~:::~ GAGTCAG7,G :AGAGGAG:C
`
`7 ;..GAA:::::A.z.
`.=-.!C7!G~~TT
`
`?
`_,,,.. ·---- _,.
`- ..;J\:~~<rJ:"".--
`
`.:::::ger,- .
`
`5 S
`
`:::;
`
`::
`
`'i
`
`p
`
`.. -
`
`~
`
`-r. - ..; . ..; ..:J\.:;-, ..,.
`
`--- ---· __ ,,..
`- . - .. ·""'""'- .
`
`---- -------
`
`- - - . . -:.--_ .. - ...:n
`
`::: 1
`.-... ..:.-.; ....... ---
`:<
`:(
`?
`-----,..,,.,. .... -
`... -..... - ...,.._,.,., ...
`... ..,.,..,op,_,...,..
`-,- .. ,,...,r-,-.-(cid:173)
`:;..1-_:...t..::c;_~_
`..:....;r. ... r1- - - - -
`GAGAAGGGGG GT':'T'!'G-GGT': CCTGTGGGAG :,;::-AGAGGG :c:GGG.j,\C, .:cAGTG,.:..:.::
`- N ~ Y V D G
`_ ~ K
`E D P
`~ V S
`V V V
`96ll GTGGTGGTGG ACGTGAGCCA CGAAGACCCT GAGGTCAAGT TCAACTGGTA ~GTGGACGGC
`CACCACCACC TGCACTCGGT GCTTCTGGGA CTCCAGTTG-. AGTTGACCAT GCACCTGCCG
`Y N s·- T
`'I R
`T K P
`R E E
`Q
`H N A K
`V E V
`CGGGAGGAGC AGTACAACAG CACGTACCGT
`1021. GTGGAGGTGC ATAATGCCAA GACAAAGCCG
`GCCCTCCTCG TCATGTTGTC GTGCATGGCA
`CACCTCCACG TATTACGGTT CTGTTTCGGC
`::; K E Y K C
`L H O O W ~ N
`V S V L T V
`lOBJ. GTGGTCAGCG TCCTCACCGT CCTGCACCAG GACTGGCTGA ATGGCAAGGA ~TACAAGTGC
`CACCAGTCGC AGGAGTGGCA GGACGTGGTC C:GACCGAC! TACCG!TCCT CATGT!CACG
`; : . . ! (G
`~SK
`-K
`t l i <A L PAP
`! <VS
`ll~i AAGGTC:CCA ACAA.~Gcc:r CCCAG:::cc::c ~r:~A~AA.~; :cA:CTCC~. ~GCCNt~G~G
`!TC:AGAGG! ~G7TT:~GGA GGG::~GCGG ~AG=~c~~: ~ ;~:AGAG5!7 TCGGr:~:c=
`C
`----,-------
`- · -'~• · -- - .:
`::
`
`?.
`
`;,20: CAG:C:.:-G.:i.:; ---------(cid:173)
`.. ..;..::: . ..; .. - __ -:;
`
`---· -.-. ..; _ __ :-____
`
`.... -. --· · --
`
`--------•;,,,
`::;
`... - -· . .
`.. -.
`..
`------- ---
`' .
`.,
`::;
`N
`',j
`E S
`~
`~
`v
`,_,..,,.-.-,,.. ---....
`GAACAAC':' ->.::
`1321 GAGAGC.l>,A'i':: ~GCrlGC:G:.:;A
`C'rTG~':'GA~:;
`CTCTCG':'TAC
`,_....,lo,!.~~.;,\,. ·- .
`~---~~T--- --,-.-----~-
`
`J
`5
`~J"-•---!...,..
`
`F
`_._,_ ___ ,M_r . .:,
`
`':AAG:::;,.:::
`GTTCGAr,7GG
`
`K T
`AAGACCACGC
`".';C:'GGTGCG
`
`.
`:::.
`5
`?
`' ~
`GGACTCCGAC
`C;CCCGTGCT
`CC:'GAGGCTG
`JAGGGCACGA
`
`:(
`)
`.3:'~~ACAAGA
`:.n.C.::'G~TC~
`
`'i
`5
`C
`5
`14-1: ~:--::i-r:::,:..~ Ge:-:::;:-:;;._:- 4~ATGAGGC: :7.jCA:AAc: M::i.:...:;..c~CA. GAAGAG:::cr=
`CAGAAGAG:A :GAG~:AC~A ::;TACTCCGA ~AC;:G7TG~ ~~A:'GTGC~T C~;CTCGGAG
`;_
`L
`:..
`_-.. G
`•
`~~:< .. A!...
`::;
`s
`CC7CCTGC::
`GCGT-:G::~G
`CC3G~~.A.Z...:.: GGCCC~GA~: :;:~:TGGGGG
`
`L,
`
`,,J
`
`5
`_ R V
`l
`•
`-
`I
`!
`~
`~
`T7~A77~GG~ :AGGcn:C~7 C!TCAGAG:~ AAUi!CAGCA
`.!\AG7AAC~CG A!CC~7A~AA GAAGTCTCAC :7:,;A~7~G:
`
`~c:cccc:;:::;
`~CGGGGGC~;
`
`E"IGURE : o
`
`Miltenyi Ex. 1007 Page 4
`
`

`

`Patent Application Publication Jul. 1, 2004 Sheet 3 of 8
`
`US 2004/012636~1 A l
`
`E s
`y N
`E L N
`'{ Q Q G
`L
`L G R R
`·Q N Q
`'(
`1621 TACCAGCAGG GCCAGAACCA GCTCTATAAC GAGCTCAATC TAGGACGAAG AGAGGA.GTAC
`ATGGTCGTCC CGGTCTTGGT CGAGATATTG CTCGAGTTAG ATCCTGCTTC TCTCCTCATG
`,,.. ..,
`p
`p E
`R R
`G R
`G
`:'I
`K
`i<
`D K R R
`D V
`L
`)
`1681 GATGTTTTGG ACAAGAGACG TGGCCGGGAC CCTGAGATGG GGGGAAAGCC GAGAAGGAAG
`CTACA.AAACC TGTTCTCTGC ACCGGCCCTG GGACTC'! ACC CCCCTTTCGG CTCTTCCTTC
`
`t
`A Y D
`-
`O K D K M A
`E G L Y N
`N ? Q
`174 ~ AACCCTCAGG AAGGCCTGTA CAATGAACTG CAGAAAGATA AGATGGCGGA GGCCTACAGT
`TTGGGAG'J'C::t: T',-...CGGACA! GTTAC'tTGi-..C G'!'.:TTTCTA7 !CTACCGCCT CCGGATGTCA
`
`G K G H D G L Y Q G
`R R R
`I G M K' G £
`E
`1801 GAGATTGGGA TGMAGGCGA GCGCCGGAGG GGCAAGGGGC ACGATGGCCT TTACCAGGGT
`CTCTAACCCT ACTTTCCGCT CGCGGCCTCC CCGTTCCCCG TGCTACCGGA AATGGTl.L:A
`
`P P R
`L S T A T K D T Y D A L H M Q A L
`1861 CTCAGTACAG CCACCAAGGA CACCTACGAC GCCCTTCACA TGCAGGCCCT GCCCCCTCGC
`GAGTCATGTC GGTGGTTCCT GTGGATGCTG CGGGAAGTGT ACGTCCGGGA CGGGGGAGCG
`
`Not I
`
`1921 TAAGCGGCCG C
`ATTCGCCGGC G
`
`FIGURE lC
`
`Miltenyi Ex. 1007 Page 5
`
`

`

`Patent Application Publication
`Jul. 1, 2004 Sheet 4 of 8
`bGh'1JAn·'t ;:_ .. :c~f1 ColEl
`
`Pad(7Sl )
`.,
`
`US 2004/012636~1 Al
`
`SpAn
`
`C'M\I Promote r
`
`lntron A
`
`Miltenyi Ex. 1007 Page 6
`
`

`

`Patent Application Publication
`
`Jul. 1, 2004 Sheet 5 of 8
`
`US 2004/012636:1 Al
`
`hsp(FMC63)CD19R/pKEN
`09/27/00
`anti-CD3zeta - overnight
`· GAM-AP - 1 hour
`E;xposure - 1 hour
`
`t(?
`,-
`'c5
`...
`
`i
`
`.,..
`....
`N
`'l5
`l
`
`. a ~
`
`~ ~
`C7I N ♦
`ID
`.- -
`0 UI
`,-
`:t: ., X
`t: 0 &!
`~ 0 <
`
`~ -
`---
`
`Miltenyi Ex. 1007 Page 7
`
`

`

`Patent Application Publication
`
`Jul. 1, 2004 Sheet 6 of 8
`
`US 2004/01 2636~1 A l
`
`/11 vi"o Production of IFNy
`
`File· CD19>JD#8 of 12/6/99>CDl9(FMC63)pMG"·pac
`Elisa· 2/16/00
`IVS of2/8/00
`I O ~ I 06 each of responders or srimulato:s/well in 2 ml final. 24 well plate. 72 hr culture
`f .,·m;:t
`Rt!;ponders JD#B of 12/6/99 - CD 19 Clones: 86 (8T. W+. Fe-) and CI (8+.W + .. Fe-).
`Sthmulaton: Stimulator<. firTad to 8000 Rads) KS62,. ;)audi,. 1873-CRL\, JM-1 ,. DHL-4,. Parsons LCL,.
`SVP-815,, lona/PMA Media
`
`Samples
`
`I. 86/K562.
`2. 86/Daudi.
`l . B6/187J-CRL.,
`4. B6/ JM-1,
`.s. 8 6/ DHL-4.
`6 B6/ LCL.
`7. 86/ SUP-81.S,
`8. 86/1/P
`9. 86/Mcdio.
`10. CI/K562,
`II CI/Daudi,
`! 12. Cl/1873-CRL.
`' 13. Cl / JM-1.,
`1-1. Cl DHL--1
`IS. Cl/ LCL,.
`16. Cl/ SUP-81.S,
`17. Cl/ 1/P
`18. Cl/Media
`19. 1<562,
`-~-_!)audi,
`21. J 873-CRL.
`22. JM-1,
`23 DHL--1,
`24. LCL,
`' 25. SUP-815,
`I 26. 1/P
`27. Media
`
`d
`h. h
`Units = extrapolated a ove
`b
`I!{ est standar
`1 Diln
`n.,..1
`TNFO Dilo
`GM-CSF Diln
`( 0 2/m11 Plated
`(ne/ml) Plated
`(ne/ml)
`Plated
`Neat
`Neat
`0
`0
`Neat
`0
`1.10
`0
`Neat
`0
`Neat
`36
`1:10
`0
`Neat
`Neal
`0
`113
`1:10
`0
`Neat
`75
`Neat
`32
`1.10
`0
`Neat
`176
`Neat
`0
`1·10
`0
`Neat
`71
`Ne· t
`85
`I 10
`Neat
`0
`0
`Neat
`116
`1:10
`37.155
`1·20
`803
`Neat
`2231
`Neat
`0
`Neat
`Neat
`0
`0
`Neat
`34
`Neat
`Neai
`0
`0
`1:11)
`51
`60
`Neat
`Neut
`303
`!'JD
`}6
`Neat
`· 296
`611
`~·e.11
`u
`1· 10
`Neat
`Neal
`I)
`25
`~ .. ~\
`I Ill
`,.:~I
`"
`. (,9
`1:10
`0
`Neat
`350
`Near
`L:10
`0
`Neat
`Neat
`338
`ND
`29.892
`1·20
`I 10
`301
`Neat
`0
`Neat
`0
`Neal
`Neat
`0
`Neat
`0
`0
`Neat
`Neal
`0
`n
`Neal
`Neal
`0
`NeQI
`0
`Neat
`0
`Io
`Neat
`Neat
`II
`0
`Neat
`Neal
`ll
`0
`u
`1 Neat
`Neat
`Nellt
`0
`Neat
`0
`
`,,
`4D
`6.S
`5552
`0
`0
`0
`73
`n
`(I
`u
`,J
`0
`
`Diln
`Plated
`Neat
`Neat
`Neat
`Neat
`Neat
`Neat
`~-.:at
`1: 10
`Neat
`Neat
`Neat
`Neal
`Neal
`'.\.eJI
`Ne·
`Neat
`Neat
`Neal
`Neat
`Neat
`Neat
`Neat
`Neat
`Neat
`Neal
`Neat
`
`·1
`
`IFNy
`fno/ml\ 1
`25
`l
`1362
`967
`396
`645
`1!45
`946
`>10.000
`0
`0
`1679
`
`J
`
`631
`li~II
`• ~SJ
`1245
`
`0
`
`-
`
`~
`
`'
`!
`l
`I
`J
`1
`I
`I
`I
`I
`
`i
`!
`
`Miltenyi Ex. 1007 Page 8
`
`

`

`Pa1c1Jt Application Publication
`
`Jul. l , 2004 Sheet 7 of' 8
`
`lJS 2004/0l2636~1 Al
`
`0
`0
`(0
`~
`
`0
`0
`'V
`
`~
`
`~ z
`LL -
`
`"-
`0
`N
`
`r -u E ·
`
`0
`0
`co
`
`IV)
`
`.._
`0
`C>I
`0
`~ C.,
`.._,,,,
`ffl
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`I
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`0 -
`
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`0
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`
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`0
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`
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`
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`...J
`
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`
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`I a.
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`
`Miltenyi Ex. 1007 Page 9
`
`

`

`Patent Application Publication
`
`Jul. 1, 2004 Sheet 8 of 8
`
`US 2004/01263621 Al
`
`Tuaet
`187lCIU.
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`!dl 9 51 c r . xl5
`
`Miltenyi Ex. 1007 Page 10
`
`

`

`US 2004/0126363 Al
`
`J ul. 1, 2004
`
`CD19-SPE◄ClFlC RIWrRECT ED IMMUNE CELLS
`
`CROSS-REFERENCE 1'0 RELATED
`APPLICATION
`
`[<H)0l] This applicalioa claims priority 10 Provisional
`t\pplil:a1ioa Seriru No. 60/246, 11 7, fileJ Nov. 7, 2000, the
`disclosure of which is incorporated by reforeace.
`
`BACKGROUND OF THE [NVENTION
`
`[0002] Tb.is invention rclalcs to the field of gcnclically
`engim:ercd, redircc!t:!d immune cells and to the field of
`cellular immunorbcrapy of 8 -cell malignancies. B-cell lym(cid:173)
`phoproliferative syadromes and B-cell mediaced au1oim(cid:173)
`mune disei1$c,s. The publications and other materials used
`herein to il111min3k the background of the invenl ion or
`provide additional details respecting the practice arc incor(cid:173)
`porated by reference.
`[0003] Approximately half of all bcmatopoictic stem t:cll
`traosplanlatioo (HSC') procedures performed io tbt' Uoited
`States are for the trna1men1 of hematologic rnn.lignaocy [1 ].
`Tbc initial abstadl,'S for succ~ssflll HSC transplantation
`were ia lnrge part due to inadequate lrealmenl modalities for
`amclirm1ti11g regimen-related toxicities and for cootrollin_g
`infections and gra(t-versus-lmJ;I disease
`opportUJJistic
`(GVHD) [2-5]. /\s supportive care measures have improved
`over 1bc last dccude, post-lraosplant disease relapse bas
`emerged as tbtc major impediment to improving 1bc outcome
`of Ibis pa1icnt. popu la1ion [G-10), The inability of rnaxi.mally
`iateosive prepar;11ive regimens combintd with immunologic:
`graft-vtrsus-tumor reactivity to eradieale minim.ti residual
`disease is th.: mechanism of lrca1men1 failure in allogeneic
`transpluntalion while. in lhc aulologous selling, tumor con(cid:173)
`tamination o.f lbe stem cell graft can ~lso contribute lo
`posHransplant relapse [ 11). Targeting minimal residual dis(cid:173)
`ease early af11~r transplantation is one stralcgy to conso!ichile
`tile lumor cytorodut:tion acbievod witb rnyeloahl,ttive pro(cid:173)
`paralive regimens and pUige, in vivo, malignant cells trans(cid:173)
`ferred witb aulologous stem cell grafts. The utility of thera(cid:173)
`peutic modalitit::s for targeting minimal residual disease
`shortly following stem cell rescue is depeadent on both a
`limited i;pcctru01 of toxicity and the susceptibilit.y of residual
`tumor cells 110 the modality's antilumor effector mccha(cid:173)
`uism(s). The successful elimination of pcrsis1en1 minimal
`residual dise::a.sc .should not unly have a major impac1 on tbe
`uu1come of trnnspluntation for hematologic malignancy uti(cid:173)
`lizing cu1rca1 myeloabla1ive preparative regimens but may
`also providt opportunities to decreaSt: the iotcnsi1y of 1hes~
`regimens aod their attendant loxicities.
`[0004] TI1e prognosis for patients wi1h bcr-abl positive
`Acute Lymphioblaslic Leukemia (ALL) trcaled with chemo(cid:173)
`therapy is poor and allogeneic transpla111ation has offered a
`curative O[Jlio10 for m11oy paticots when an approprialc clooor
`was ttvailable. For ex!I.Olple al the City of !lope, 76 putients
`with bcr-abl positive ALL W!lrtl treated with :iUogcneic 130111!
`M,1rrow Tra.n:;planla.lion ( 13MT) Crom a I I LA matched donor.
`Of these palients, 26 were in first remission, 35 were
`traasvluoted ~1ftcr first remission. The two year probahiUty of
`disease free survival was 68% with a 10% relapse rate in
`those palien~, transplao1cd in lust remission whereas l'or
`those palients 1raosplanted after first remission, the., d.isc:asc(cid:173)
`free survival and relapse rate wt:rc 36% aod 38%. rcspcc(cid:173)
`Lively [ 12). Post-transplant Polymerase Chain Ren ct ion
`
`(PCR) screening of blood aod marrow for bcr-abl traoscript
`is under eva!trntion :1s a molecular screening tool for idcn(cid:173)
`Li.fyiog early tbose transplant recipients at bigb risk for late.r
`development of overt relapse [13,14). Patients for whom
`detectable pl 90 1ranscriJ.1t was detected following .BMT hau
`a 6.7 higher iocidcaco of oven relapsu lban PCH uugativc
`patieats. Tbe mediun lime from thi: developmenl of a
`po.-;itive s ignal Le, n1orphc1logic relapse was 80-90 d~1ys in
`these patients, 'l11c identification of patient.'; in the enrlicst
`phases of post-traosplan1 rdapsc affords the opportuni1y for
`making therapeutic iutervenlions when tumor burden is low
`and potenrially most arncn~blc to alvagc therapy,
`[0005] Recent advanc.es in the lh:ld of immunology have
`elucidaled many of the mo!ccular underpinnings of immune
`syJ;tem rei:,<ulatfoa and have provided novel opportunities Cm
`therapeutic immune syslem manipulation, including tumor
`immunolberapy. Evidence supporting 1he poteati;:u of
`immune-mediated eradication of residual tumor coll:s fol(cid:173)
`lowing al!ogencic transplantation can be inferred by com(cid:173)
`pariag the tlisparnlo relapse ratos between recipiCDts of
`syngenei..: and non-T cell depleted matched sibling lmns(cid:173)
`plants. l'aticnts with chronic myelogonous leukemia in
`chronic p base (CML-CP), acu1e myelogenous leukemia in
`Ctrst com_plete remission ( l'' CR), aod acute lympboblastic
`leukemia in I" CR wbo received a ma rrow traasplant from
`a syngeneic donor bad an actutlriaJ probability or relapse al
`3 yc~rs of 45%, 49%, and 41 %. respectively, wbcrcas lhc
`rates for recipients of a non-T dc.,pleted marrow traoi;plant
`From an HLA identical sibling for the same diseases were
`12%, 2D%, and 24%, rcspec.:tively [15-17]. The reclucti.oa of
`relapse rntts following allogeollic bone marrow 1ransplao(cid:173)
`tatioo has been most signiflt.:ant in patients who dcvelop
`acute and/o, chronic GVJ ID. C.'tirreolly, efforts are fo,cused
`on developing strateg-Ies lo i;ehxtively augmenl the graft(cid:173)
`vcrsus-lc11kemia (GVL) response in order to reduce post(cid:173)
`transplnnl relapse rates wilhoul lhc attendant toxicitiies of
`augmllntud GVHD.
`[0006] Studies in animal models have established that
`donor MHC-rcstrictcd Cos ► and CD4•att3· Tcolls specific
`for minor bistocompalibility antigens encoded by p(1lymor(cid:173)
`phic genes that differ between the donor and recipient are tbe
`principle mediators of a.::ute GVIIO and GVL [IH-21].
`Reccolly, p~t.icnts with C'ML iu ch.rook phase wbo relapse
`:i.fter allogeoeic BMT bave been identiticd as a patieat
`populatioa fur wbom !be iofusioa of donor lymphocytes
`(DU) SIICccssf-ully promotes a GVl effect [22,23]. Com(cid:173)
`plete response rates of approxima1t:1y 75% are acbitvcd wi1b
`DLI cell doses in the range of ().25-1'2.3x 10" mononuclear
`cells/kg [24]. t\.llhough the antitumclr activity ol' donor
`lymphocyte infusion underscores the potential of ce;Jhil~r
`immunotberapy for C'ML, the clinical benefit of DU has ool
`been gencrali7.ablc to all forms of hematologic maligniancy.
`Relapsed ALL is much less responsive to DLI with a
`reported CR ra te of less ti.Jan 20%; wbea tumo r res)lons.L'S arc
`observed, they are typically assodalcd witb significant
`GVHD morbidity and mortality [25). ln order lo iocrea:sc tbe
`therapeutic rnlio of DU , genetic modification of doaor
`Jympbocytus lo express :i suicide geoc is being evalua1led as
`a $lrntegy to perm ir the in vivo ablation or dooor lympho(cid:173)
`cytes should toxicity l'rnm GVJID warra.at this ma neuver
`[2<1,27]. Alicrna1cly, efforts are underway to idcnlify gene:,
`encoding minor hislocomp&tibility antigens (mHA's) wirb
`rcs1ric11:d bcma1opoie1ic cxpressioo thal elicit donor ~nnigen(cid:173)
`specific T cell responses. The isolation, ex vivo expausioa,
`
`Miltenyi Ex. 1007 Page 11
`
`

`

`US 2004/0126363 Al
`
`Jul. l, 2004
`
`2
`
`aad rc-iofusi<Jn of dooor-derived doncs specific for tbese
`rnl lt\'s has t,be potential of selectively augrncating_ GVL
`foUowing allogcneit: bone marrow traosplanlatioo [28-30].
`
`ootoxio B43-PAP is currently undergoing investigation in
`pha.~e I/II clinical trials in pa Lien ts with high risk pre-fl: ALL.
`[52).
`
`[0007) Non-transformed B-ct:lls and malignant B-cells
`express an array of cell-surface mlllet:ulcs tbat define lbeir
`lineage comnniLment and stage of maturation. Tbese were
`ideotilicd initially by murinc monoclonal notibodies and
`more re<-'enlly by molecular genetic techniques. Expression
`of se11eral or these cell-surface molecules is highly restricted
`IO B-cells and their malignant counterparts. CD20 is a
`clinically us\:ful cell-surface rurgct for B-cell lymphoma
`immunotbera.py wilb :uiti-CD2C) rnonoclona.l antibodies.
`This 33-kOa protein has structural [eatures consistent with
`its ability TO function as a calcium ion chunuel and is
`ex11resscd on normal pre-8 and mature B cells, but not
`hemntopoieti<: stem cells □or plasma cells [31-33). CD20
`doe-s nol mod1ulntc nor does it shed from the cell surface [34].
`In vitro studies bave demonstrated that CD20 croAAlinking
`by anti-C'D20 monoclonal antibodies can trigger apoplosis
`uf lymphoma ct:ll;:; [35,36), Clinical trials evaluating the
`amitlllllor activity of chlmeric ami-CD20 amibody IDl2C(cid:173)
`C2J38 (Rituxi.mab) ia patients with (elapsed follicular lym(cid:173)
`phoma have documeoted tumor rnspooses in nearly haJr the
`patients treated, although tbe clinical effect is usually tran(cid:173)
`sient [37-40]. Despite the prolonged ablation of' nom1ul
`CD'.W+ 8 -cell.s, patients receiving Rjtu x.imab bavc not mani(cid:173)
`fe:sred compl.icatioos attributable to B-cell lymphc1penia
`[41). Radioimmunotherapy with m l-conjugated and ooy_
`conjugated anti-C020 antibodies also has shown prnmising
`clinical activity in patients with rchlpsed/r.ifraetory bigh(cid:173)
`grade Non-! l•odgkins Lymphoma but hcm,itopoietic tm.ici(cid:173)
`ties from raclliation have been significant, often requiring
`stem cell support [ 42).
`
`((H)0S) UnUkc CD20, CDl9 is expressed on a.I! human
`B-cclls bcginoing from 1bc initial comrnitmcot nfsrom cells
`to tbc Il lineage and persisting until terminal tlill'crllntiation
`iato plasma ,cells [43]. CDJ9 is a typo l transmcrnbraac
`protein Lhut associutcs witb tbe comph!ment 2 (CD21),
`TAPA-1, and 1.euJ3 antigeos forming a B-c.:11 signal 1raru;(cid:173)
`duc1io11 complex. This compkx participates io tbe regulation
`of R-ccU proliferation [44), Although CD19 does 001 shed
`frnm the cell surface, it docs inreroaJize [ 45]. Accordingly,
`targeting CDJl9 with monocloaal antibodies conjugated with
`toxin rnolecullcs is currently being investigated as a strategy
`lt) speciflc:11ly dcliv,;.r cytottlxic agents to 1be iotracdlufar
`tompartment of maljgnant B-c:clls [ 46-48]. /\nli-CD 19 anti(cid:173)
`bmfy c:onjug11ted 10 blocked ricin and poke-wt:ed antiviral
`protein (P/\.P) dramalically increase specJficity and potency
`ol' leukemia cell killing both in ex vivo bont: marrow purging
`procedures and when administered lo NOO-SC'ID animals
`inoculated with CD19+ leukemia cells (49). In. vitro leuke(cid:173)
`mia progenitor cell as.,;ays bave provided evidence tbal lb.c
`srnalJ percentage of l.:ukemic blasts wil!J tbc capacity for
`sdf-rcnt!wal ,~xprt!ss CD19 on lht:ir ct:ll surface. Th.is t:on(cid:173)
`clusion was .derived [rom the observations thlll lcukt!mic(cid:173)
`progcnitur activity is observed exclusively in frcsb marrow
`samples sorted for en J 9 positive cells und is not observed
`in ihe CD19 negative oeU population [SO]. Additionally
`843-PAP treatment of nilapscd leukemic marrow specimens
`ablates progEmitor ccU activi1y whih:: a PAf> conju.galecl
`antibody with an irrdcvaOL speciJicily had no sucb activity
`[51). Systemic administration of tht: C019-specific imrnu-
`
`[0009) Despite !he i1ntitumor activity or monoclonal a□ti­
`CD20 and anti-CD19 11ntibody lberapy observed in cliuiL'al
`trials. lbt: high rate of relapse in these palients uaclerscores
`the Limited capacity of current antibody-based immuno(cid:173)
`Lherapy to eliminate aU tumor cells (53]. lo conlrMl, tbc
`adoptive transfer of 1umor-spcdfic T cells can result in
`complete tumor er~dlca1ioo in animal models and a limited
`number of dinic<1lscniags [54,55). The ability uftraosferred
`T ccUs to directly recognize and lysc tumor 1argc1s, produce
`cy1okim:s tbal recrnit and activate aotigco non-specilk aoti(cid:173)
`turuor effector cells. migrate into tumor masses, and prolif(cid:173)
`erate l'ollowing tumor recognition all contribute lo !he
`immunologic clearance of tumor by T cells [56). fo;-prcs(cid:173)
`sion-cloning
`technologies have recently ptlrmillecl !he
`geac1ic idontificatioo of a growing number of genes
`expressed hy human tumors In which T cell responses have
`been isolated [57,58). 'l b dale leukemia and lymphoma(cid:173)
`s_peci1ic amigen., have not been ideatilied that arc both
`broadly .:-xprtissed by ma.ligonnt B-cells aod elicit "JI' cell
`responses. Consequently, prcdinica.1 and clinicnl investiga(cid:173)
`tion hiL, focused oo combining antibody targeting ofmmors
`witb T cell ctTcctor mcchunisms hy constructing bispccific
`antib(ldies consisting or CD20 or CDl9 binding sites and n
`binding site for a cell-surface CD3 oomplex cpitope. Such
`bispecitlc anribOdies cnn cu-localiLt: leukemia and lym(cid:173)
`phoma tnrg,cls with activated T cells resulting in targt:t cell
`lysis in vitro [59-6 l). The in vivo a111iturm>c activity of such
`bispecitk antibodies bas been limitt:d, however, bc,tb io
`anima.l models as well as in clinical practice [62). The
`discrepancy between in vitro activity and in vivo effect
`likely reflects the inherent limitations in antibody immuno(cid:173)
`lborapy compounclucl by the ohstacltlS associated with
`engaging T cells and tumor cells via a soluble linkcir in a
`manner tbat yields a persistent ttnd functional c~,llul~r
`immune response [63).
`
`[0010] The safety of adoptively lransfening antigen-spe(cid:173)
`cilk CTL clone~ in humans was originnJly examined in bone
`marrow transplant patients who received donor-dt,rived
`CMV-spccific T cell!\ [56). Previ()ltS studies have d~-O'JOll·
`stratctl lhat the recoostilution of endogenous CMV-~-pecific
`T cell responses following allogeneic hone ma rrow 1rans(cid:173)
`plnntn1ion (BM1) correlates witb. protect ioo from the clcvcl(cid:173)
`opmenr of severe CMV cliseuSc [64]. In :i.n effort to recon(cid:173)
`stitute delkient CMV immunity following RMT, COS ...
`CMV-spccific CTL clones wen:: generntcd from CMV s;erop(cid:173)
`ositive 1-n. A-matched sibling donors, expanded, and infused
`inlCJ sibling BMT recipients al risk for developing CMV
`disease. Fourteen pHlients were treated with four weekly
`escalating doses of these CMV-spccitlc CTL clones 10 a
`maxtmum cell dosu or .!09 ceUs/rn: wi1hou1 aay atlc-adaat
`toxicity [65]. Peripbera.l blood samples obtained [wm r,ecipi(cid:173)
`ents of adoptively lram;ferrotl T cell clones were evaJuated
`for in vivo persistence of transferred cells. Tbe recov,:rnble
`CMV-specillc C f'L activity increased after each succc:ssive
`infusion of CTL clone.~, and persisted at least 12 week::; after
`lh.: last infusion. 1 lowevcr, long term persistence of CDS+
`clones without a concurteot CD4+ helper response was aot
`observed. No palit:nts dewloped CMV viremia or disease.
`Tbcse results demonstra1<: Lhul ex-vivo expanded CMV(cid:173)
`specific C11.. clones can be safely lransferr~d to BMT
`
`Miltenyi Ex. 1007 Page 12
`
`

`

`US 2004/0126363 Al
`
`Jul. 1, 2004
`
`3
`
`rccipieo1s aocl cao persist in vivo as functional elfocwr ccUs
`lhal may provide prolcclion from the development of CMV
`disl~d.sc.
`
`[(M)ll] A complication of bone marrow transplanta1ion.
`particularly when marrow is depleted of T cells, is the
`development of 1:.H V-associated lyo;pboproliforative dis(cid:173)
`eas~ [66]. TI1is rapidly progre$ive proliferation or EBV(cid:173)
`transformed 18-cells mimics immunobl,L<;tic lympli,1ma and
`is a consequence of delicient EBV-spe..·itic T cell immunily
`in iadividu:iLs harboring latent vims or imrnuoologically
`naive individuals receiving a vims inoculum with their
`rniirrow graft. Clioic11I trials by Rooney Ill nJ. bHw dornoo(cid:173)
`straled that adoptively transferred ex-vivo expaoded donor(cid:173)
`derived EB V-specilic T cell lines can protect palieots al b.igb
`risk fordevel,opmcnl of this complication ns well ns mediate
`the eradicati(Ja of clinically evident EBY-lraasformt:d 8
`cells [54]. No signitkanl toxicities were observed in the
`forty-one children trcnted with cell doses in U1e range of
`4xl0 · 1o l.2x JO" cclls/ni.
`
`[0012] Gcueti<.: rnodilkatioa of T l:t!lls used ia diaical
`trials b.is been utilized to mark cells for in vivo tracking aad
`tu endow T ct:11s with novel l"unctionaJ properties. Retroviral
`vectors have het:n used most extensively for this purpose
`due to their ri:lativc ly high transduction cfliciency nnd low
`ia vitro toxiciity IO T cells [67]. These vectors, however, are
`tirue consuming n.nd expensive 10 prepare as clinical grade
`material aad 1must be meticulousJv screened for lhe absence
`of rcpllcationt competent viral mutants [68), Roool!y C( aL
`1ransduc<1cl Ell3 V-reactive T ccll liacs with lbe Nt:0R gene 10
`facilitate assessment of cdl p<:rsistencc in vivo hy PCR
`specific for this marker gene [69]. Riddell el al. have
`conducted a Phase I trial lo augment lllV-spccific immunity
`in HfV seroposilive individuals by adoptive trnnsfer using
`HIV-specific cos+ C'TL clones [70). These clones were
`transduced with the rclroviral vector tgl .s+HyTK which
`directs tile synthesis of a bifunctioaal fusioa protein incor(cid:173)
`porating bygrnmycin phosphotraosforasc ~nd herpes virns
`tbymidimi kimasc (IISV-TK) permitting in vitro sulectioo
`wilb byg,rornycin and potuntial io vivo ablatioa of 1rnns(cid:173)
`ferred cells with ganoyclovir. Six I LIV infected patients wert
`trea.ted with a series of four escalating cell do!.e infw,ions
`without toxicities, with a maximum cell dose of Sx109
`ccl!S/m2 [70].
`
`[0013) As an alternate to viral gene therapy vectors, Nahel
`ct al. used plasmid DNA encoding an expression cassctlu for
`an anti,IIIY gene in a Phase I clinical trial. Plasmid ONA
`was introduc1id into T cells by parLide bombnrdruent with o
`geae gun [71]. (;eneticaJly modified T cells were expanded
`am.I infused back iato I IIV-iafecled study subjects. Although
`this study dernonslratcd the foasibil ity of w;ing a oon-viral
`genetic modificalion strategy for primary human Teel ls, one
`Limitation of t his approacb is the episomal propagation of the
`plasmid vtctqr in T Ct:lls. Unlike cbromusomaUy integrsted
`traosfcm:d 0,NA, episomal propagation of plasmid ONA
`carries the rii;k of lo&> or transferred genetic material with
`cell replicatic,n and o[ repetitive random cbromosomal inte(cid:173)
`gration event.s.
`
`clooal antibody's specificity [72]. The design of scFvFc:~
`receptors with target specificities for tumor cell-s