                         SEQUENCE LISTING

<110>  National University of Singapore
       Png, Yi Tian
       Vinanica, Natasha
       Kamiya, Takahiro
       Campana, Dario
 
<120>  BLOCKADE OF CD7 EXPRESSION AND CHIMERIC ANTIGEN RECEPTORS FOR 
       IMMUNOTHERAPY OF T-CELL MALIGNANCIES

<130>  119419-5002-WO

<140>  Filed Herewith
<141>  2017-11-22

<150>  US 62/425,398
<151>  2016-11-22

<150>  US 62/543,696
<151>  2017-08-10

<160>  40    

<170>  PatentIn version 3.5

<210>  1
<211>  117
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  VH domain of TH69 antibody (anti-CD7 scFv)

<400>  1

Glu Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Lys Pro Gly Gly 
1               5                   10                  15      


Ser Leu Lys Leu Ser Cys Ala Ala Ser Gly Leu Thr Phe Ser Ser Tyr 
            20                  25                  30          


Ala Met Ser Trp Val Arg Gln Thr Pro Glu Lys Arg Leu Glu Trp Val 
        35                  40                  45              


Ala Ser Ile Ser Ser Gly Gly Phe Thr Tyr Tyr Pro Asp Ser Val Lys 
    50                  55                  60                  


Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Arg Asn Ile Leu Tyr Leu 
65                  70                  75                  80  


Gln Met Ser Ser Leu Arg Ser Glu Asp Thr Ala Met Tyr Tyr Cys Ala 
                85                  90                  95      


Arg Asp Glu Val Arg Gly Tyr Leu Asp Val Trp Gly Ala Gly Thr Thr 
            100                 105                 110         


Val Thr Val Ser Ser 
        115         


<210>  2
<211>  112
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  VL domain of TH69 antibody (anti-CD7 scFv)

<400>  2

Ala Ala Tyr Lys Asp Ile Gln Met Thr Gln Thr Thr Ser Ser Leu Ser 
1               5                   10                  15      


Ala Ser Leu Gly Asp Arg Val Thr Ile Ser Cys Ser Ala Ser Gln Gly 
            20                  25                  30          


Ile Ser Asn Tyr Leu Asn Trp Tyr Gln Gln Lys Pro Asp Gly Thr Val 
        35                  40                  45              


Lys Leu Leu Ile Tyr Tyr Thr Ser Ser Leu His Ser Gly Val Pro Ser 
    50                  55                  60                  


Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Tyr Ser Leu Thr Ile Ser 
65                  70                  75                  80  


Asn Leu Glu Pro Glu Asp Ile Ala Thr Tyr Tyr Cys Gln Gln Tyr Ser 
                85                  90                  95      


Lys Leu Pro Tyr Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys Arg 
            100                 105                 110         


<210>  3
<211>  42
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  intracellular signaling domain of 4-1BB

<400>  3

Lys Arg Gly Arg Lys Lys Leu Leu Tyr Ile Phe Lys Gln Pro Phe Met 
1               5                   10                  15      


Arg Pro Val Gln Thr Thr Gln Glu Glu Asp Gly Cys Ser Cys Arg Phe 
            20                  25                  30          


Pro Glu Glu Glu Glu Gly Gly Cys Glu Leu 
        35                  40          


<210>  4
<211>  112
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  intracellular signaling domain of CD3zeta

<400>  4

Arg Val Lys Phe Ser Arg Ser Ala Asp Ala Pro Ala Tyr Gln Gln Gly 
1               5                   10                  15      


Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg Arg Glu Glu Tyr 
            20                  25                  30          


Asp Val Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu Met Gly Gly Lys 
        35                  40                  45              


Pro Arg Arg Lys Asn Pro Gln Glu Gly Leu Tyr Asn Glu Leu Gln Lys 
    50                  55                  60                  


Asp Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly Met Lys Gly Glu Arg 
65                  70                  75                  80  


Arg Arg Gly Lys Gly His Asp Gly Leu Tyr Gln Gly Leu Ser Thr Ala 
                85                  90                  95      


Thr Lys Asp Thr Tyr Asp Ala Leu His Met Gln Ala Leu Pro Pro Arg 
            100                 105                 110         


<210>  5
<211>  45
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Hinge region of anti-CD7 CAR

<400>  5

Thr Thr Thr Pro Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile Ala 
1               5                   10                  15      


Ser Gln Pro Leu Ser Leu Arg Pro Glu Ala Cys Arg Pro Ala Ala Gly 
            20                  25                  30          


Gly Ala Val His Thr Arg Gly Leu Asp Phe Ala Cys Asp 
        35                  40                  45  


<210>  6
<211>  24
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  transmembrane region of anti-CD7 CAR

<400>  6

Ile Tyr Ile Trp Ala Pro Leu Ala Gly Thr Cys Gly Val Leu Leu Leu 
1               5                   10                  15      


Ser Leu Val Ile Thr Leu Tyr Cys 
            20                  


<210>  7
<211>  21
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  CD8a signal peptide of anti-CD7 CAR

<400>  7

Met Ala Leu Pro Val Thr Ala Leu Leu Leu Pro Leu Ala Leu Leu Leu 
1               5                   10                  15      


His Ala Ala Arg Pro 
            20      


<210>  8
<211>  14
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Localization domain

<400>  8

Glu Gln Lys Leu Ile Ser Glu Glu Asp Leu Lys Asp Glu Leu 
1               5                   10                  


<210>  9
<211>  11
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Localizing domain

<400>  9

Gly Gly Gly Gly Ser Ala Glu Lys Asp Glu Leu 
1               5                   10      


<210>  10
<211>  68
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Localizing domain

<400>  10

Thr Thr Thr Pro Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile Ala 
1               5                   10                  15      


Ser Gln Pro Leu Ser Leu Arg Pro Glu Ala Cys Arg Pro Ala Ala Gly 
            20                  25                  30          


Gly Ala Val His Thr Arg Gly Leu Asp Phe Ala Cys Asp Ile Tyr Ile 
        35                  40                  45              


Trp Ala Pro Leu Ala Gly Thr Cys Gly Val Leu Leu Leu Ser Leu Val 
    50                  55                  60                  


Ile Thr Leu Tyr 
65              


<210>  11
<211>  15
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Localizing domain

<400>  11

Lys Tyr Lys Ser Arg Arg Ser Phe Ile Asp Glu Lys Lys Met Pro 
1               5                   10                  15  


<210>  12
<211>  20
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Linker

<400>  12

Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly 
1               5                   10                  15      


Gly Gly Gly Ser 
            20  


<210>  13
<211>  83
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Localizing domain

<400>  13

Thr Thr Thr Pro Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile Ala 
1               5                   10                  15      


Ser Gln Pro Leu Ser Leu Arg Pro Glu Ala Cys Arg Pro Ala Ala Gly 
            20                  25                  30          


Gly Ala Val His Thr Arg Gly Leu Asp Phe Ala Cys Asp Ile Tyr Ile 
        35                  40                  45              


Trp Ala Pro Leu Ala Gly Thr Cys Gly Val Leu Leu Leu Ser Leu Val 
    50                  55                  60                  


Ile Thr Leu Tyr Lys Tyr Lys Ser Arg Arg Ser Phe Ile Asp Glu Lys 
65                  70                  75                  80  


Lys Met Pro 
            


<210>  14
<211>  123
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  VH domain of 3alf antibody (anti-CD7 scFv)

<400>  14

Gln Val Gln Leu Gln Glu Ser Gly Ala Glu Leu Val Lys Pro Gly Ala 
1               5                   10                  15      


Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr 
            20                  25                  30          


Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile 
        35                  40                  45              


Gly Lys Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn Glu Lys Phe 
    50                  55                  60                  


Lys Ser Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr 
65                  70                  75                  80  


Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Cys 
                85                  90                  95      


Ala Arg Gly Gly Val Tyr Tyr Asp Leu Tyr Tyr Tyr Ala Leu Asp Tyr 
            100                 105                 110         


Trp Gly Gln Gly Thr Thr Val Thr Val Ser Ser 
        115                 120             


<210>  15
<211>  108
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  VL domain of 3alf antibody (anti-CD7 scFv)

<400>  15

Asp Ile Glu Leu Thr Gln Ser Pro Ala Thr Leu Ser Val Thr Pro Gly 
1               5                   10                  15      


Asp Ser Val Ser Leu Ser Cys Arg Ala Ser Gln Ser Ile Ser Asn Asn 
            20                  25                  30          


Leu His Trp Tyr Gln Gln Lys Ser His Glu Ser Pro Arg Leu Leu Ile 
        35                  40                  45              


Lys Ser Ala Ser Gln Ser Ile Ser Gly Ile Pro Ser Arg Phe Ser Gly 
    50                  55                  60                  


Ser Gly Ser Gly Thr Asp Phe Thr Leu Ser Ile Asn Ser Val Glu Thr 
65                  70                  75                  80  


Glu Asp Phe Gly Met Tyr Phe Cys Gln Gln Ser Asn Ser Trp Pro Tyr 
                85                  90                  95      


Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys Arg 
            100                 105             


<210>  16
<211>  120
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  VH domain of T3-3A1 antibody (anti-CD7 scFv)

<400>  16

Asp Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 
1               5                   10                  15      


Ser Arg Lys Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Phe 
            20                  25                  30          


Gly Met His Trp Val Arg Gln Ala Pro Glu Lys Gly Leu Glu Trp Val 
        35                  40                  45              


Ala Tyr Ile Ser Ser Gly Ser Ser Thr Leu His Tyr Ala Asp Thr Val 
    50                  55                  60                  


Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Pro Lys Asn Thr Leu Phe 
65                  70                  75                  80  


Leu Gln Met Thr Ser Leu Arg Ser Glu Asp Thr Ala Met Tyr Tyr Cys 
                85                  90                  95      


Ala Arg Trp Gly Asn Tyr Pro His Tyr Ala Met Asp Tyr Trp Gly Gln 
            100                 105                 110         


Gly Thr Ser Val Thr Val Ser Ser 
        115                 120 


<210>  17
<211>  111
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  VL domain of T3-3A1 antibody (anti-CD7 scFv)

<400>  17

Asp Ile Val Met Thr Gln Ser Pro Ala Ser Leu Ala Val Ser Leu Gly 
1               5                   10                  15      


Gln Arg Ala Thr Ile Ser Cys Arg Ala Ser Lys Ser Val Ser Ala Ser 
            20                  25                  30          


Gly Tyr Ser Tyr Met His Trp Tyr Gln Gln Lys Pro Gly Gln Pro Pro 
        35                  40                  45              


Lys Leu Leu Ile Tyr Leu Ala Ser Asn Leu Glu Ser Gly Val Pro Ala 
    50                  55                  60                  


Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Asn Ile His 
65                  70                  75                  80  


Pro Val Glu Glu Glu Asp Ala Val Thr Tyr Tyr Cys Gln His Ser Arg 
                85                  90                  95      


Glu Leu Pro Tyr Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys 
            100                 105                 110     


<210>  18
<211>  4
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Localizing sequence

<400>  18

Lys Asp Glu Leu 
1               


<210>  19
<211>  4
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Localizing sequence


<220>
<221>  misc_feature
<222>  (3)..(4)
<223>  Xaa can be any naturally occurring amino acid

<400>  19

Lys Lys Xaa Xaa 
1               


<210>  20
<211>  3
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Localizing sequence


<220>
<221>  misc_feature
<222>  (2)..(2)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  MISC_FEATURE
<222>  (3)..(3)
<223>  Xaa can be Asp or Glu

<400>  20

Lys Xaa Xaa 
1           


<210>  21
<211>  4
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Localizing sequence

<400>  21

Tyr Gln Arg Leu 
1               


<210>  22
<211>  4
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Proteasome localizing sequence

<400>  22

Pro Glu Ser Thr 
1               


<210>  23
<211>  351
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  VH domain of TH69 antibody (anti-CD7 scFv)

<400>  23
gaggtgcagc tggtcgaatc tggaggagga ctggtgaagc caggaggatc tctgaaactg       60

agttgtgccg cttcaggcct gaccttctca agctacgcca tgagctgggt gcgacagaca      120

cctgagaagc ggctggaatg ggtcgctagc atctcctctg gcgggttcac atactatcca      180

gactccgtga aaggcagatt tactatctct cgggataacg caagaaatat tctgtacctg      240

cagatgagtt cactgaggag cgaggacacc gcaatgtact attgtgccag ggacgaagtg      300

cgcggctatc tggatgtctg gggagctggc actaccgtca ccgtctccag c               351


<210>  24
<211>  351
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  VL domain of TH69 antibody (anti-CD7 scFv)

<400>  24
gaggtgcagc tggtcgaatc tggaggagga ctggtgaagc caggaggatc tctgaaactg       60

agttgtgccg cttcaggcct gaccttctca agctacgcca tgagctgggt gcgacagaca      120

cctgagaagc ggctggaatg ggtcgctagc atctcctctg gcgggttcac atactatcca      180

gactccgtga aaggcagatt tactatctct cgggataacg caagaaatat tctgtacctg      240

cagatgagtt cactgaggag cgaggacacc gcaatgtact attgtgccag ggacgaagtg      300

cgcggctatc tggatgtctg gggagctggc actaccgtca ccgtctccag c               351


<210>  25
<211>  369
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  VH domain of 3alf antibody (anti-CD7 scFv)

<400>  25
caggtccagc tgcaggagtc aggagctgag ctggtgaagc caggggcaag cgtcaaactg       60

tcctgcaagg cctctggata tacattcact agctactgga tgcactgggt gaaacagaga      120

cccggacagg gcctggagtg gatcggaaag attaacccta gcaatggcag gaccaactac      180

aacgaaaagt ttaaatccaa ggcaaccctg acagtggaca agagctcctc tacagcctac      240

atgcagctga gttcactgac ttcagaggat agcgcagtgt actattgcgc cagaggcggg      300

gtctactatg acctgtacta ttacgccctg gattattggg ggcagggaac cacagtgact      360

gtcagctcc                                                              369


<210>  26
<211>  324
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  VL domain of 3alf antibody (anti-CD7 scFv)

<400>  26
gacatcgagc tgacccagag tcctgctaca ctgagcgtga ctccaggcga ttctgtcagt       60

ctgtcatgtc gggcaagcca gtccatctct aacaatctgc actggtacca gcagaaatcc      120

catgaatctc cacgactgct gattaagagt gcctcacaga gcatctccgg cattccctcc      180

cggttctctg gcagtgggtc aggaactgac tttaccctga gtattaactc agtggagaca      240

gaagatttcg gcatgtattt ttgccagcag agcaattcct ggccctacac tttcggaggc      300

gggaccaaac tggagatcaa gcgg                                             324


<210>  27
<211>  360
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  VH domain of T3-4A1 antibody (anti-CD7 scFv)

<400>  27
gatgtgcagc tggtggagtc tgggggaggc ttagtgcagc ctggagggtc ccggaaactc       60

tcctgtgcag cctctggatt cactttcagt agctttggaa tgcactgggt tcgtcaggct      120

ccagagaagg ggctggagtg ggtcgcatac attagtagtg gcagtagtac cctccactat      180

gcagacacag tgaagggccg attcaccatc tccagagaca atcccaagaa caccctgttc      240

ctgcaaatga ccagtctaag gtctgaggac acggccatgt attactgtgc aagatggggt      300

aactaccctc actatgctat ggactactgg ggtcaaggaa cctcagtcac cgtctcctca      360


<210>  28
<211>  333
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  VL domain of T3-4A1 antibody (anti-CD7 scFv)

<400>  28
gacattgtga tgacccagtc tcctgcttcc ttagctgtat ctctggggca gagggccacc       60

atctcatgca gggccagcaa aagtgtcagt gcatctggct atagttatat gcactggtac      120

caacagaaac caggacagcc acccaaactc ctcatctatc ttgcatccaa cctagaatct      180

ggggtccctg ccaggttcag tggcagtggg tctgggacag acttcaccct caacatccat      240

cctgtggagg aggaggatgc tgtaacctat tactgtcagc acagtaggga gcttccgtac      300

acgttcggag gggggaccaa gctggaaata aaa                                   333


<210>  29
<211>  5
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Polypeptide linker

<400>  29

Gly Gly Gly Gly Ser 
1               5   


<210>  30
<211>  63
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  CD8alpha signal peptide

<400>  30
atggctctgc ctgtgaccgc actgctgctg cccctggctc tgctgctgca cgccgcaaga       60

cct                                                                     63


<210>  31
<211>  60
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  VH-VL Linker

<400>  31
ggaggaggag gaagcggagg aggaggatcc ggaggcgggg gatctggagg aggaggaagt       60


<210>  32
<211>  42
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  ER localization domain, KDEL tethered to scFv with myc ("myc 
       KDEL")

<400>  32
gagcagaaac tgattagcga agaggacctg aaagatgaac tg                          42


<210>  33
<211>  78
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  localization domain, "link.(20)AEKDEL"

<400>  33
ggtggtggcg gcagtggtgg cggtggctca ggcggtggtg gctccggtgg cggtggctct       60

gcagaaaaag atgagttg                                                     78


<210>  34
<211>  249
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  localization domain, "mb DEKKMP"

<400>  34
accactacac ctgcaccaag gcctcccaca cccgctccca ctatcgcttc ccagccactg       60

tccctgaggc ccgaggcctg caggccagca gctggcggag ccgtgcatac tagggggctg      120

gacttcgctt gcgacatcta catctgggcc ccactggcag ggacatgcgg agtcctgctg      180

ctgtccctgg tcatcacact ttacaaatac aaaagcagac gcagttttat tgatgaaaag      240

aaaatgcct                                                              249


<210>  35
<211>  126
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Nucleic acid sequence of intracellular signaling domain of 4-1BB

<400>  35
aagcgggggc gcaaaaaact gctgtatatc tttaagcagc ctttcatgag accagtgcag       60

acaacccagg aggaagatgg gtgctcatgc cggtttcccg aggaggagga aggcggctgc      120

gagctg                                                                 126


<210>  36
<211>  336
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  intracellular signaling domain of CD3zeta

<400>  36
agggtgaagt tttcccgctc agcagatgct cctgcctacc agcagggcca gaaccagctg       60

tataatgagc tgaacctggg cagacgcgaa gagtatgatg tgctggacaa aaggcgggga      120

agagaccccg aaatgggagg gaagccaagg cggaaaaacc cccaggaggg cctgtacaat      180

gagctgcaga aggacaaaat ggcagaggct tacagtgaga ttgggatgaa gggagagaga      240

cggaggggaa aagggcacga tggcctgtac caggggctga gcacagcaac caaagatact      300

tatgacgcac tgcacatgca ggcactgcca cccaga                                336


<210>  37
<211>  204
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Hinge and transmembrane domain of CD8alpha

<400>  37
accactacac ctgcaccaag gcctcccaca cccgctccca ctatcgcttc ccagccactg       60

tccctgaggc ccgaggcctg caggccagca gctggcggag ccgtgcatac tagggggctg      120

gacttcgctt gcgacatcta catctgggcc ccactggcag ggacatgcgg agtcctgctg      180

ctgtccctgg tcatcacact gtac                                             204


<210>  38
<211>  15
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  synthetic oligonucleotide primer

<400>  38
atggccgggc ctccg                                                        15


<210>  39
<211>  18
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  synthetic oligonucleotide primer

<400>  39
tcactggtac tggttggg                                                     18


<210>  40
<211>  10
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Localizing domain

<400>  40

Glu Gln Lys Leu Ile Ser Glu Glu Asp Leu 
1               5                   10  


