                         SEQUENCE LISTING

<110>  National Research Council of Canada
 
<120>  SINGLE DOMAIN ANTIBODIES TARGETING CA-IX AS WELL AS COMPOSITIONS 
       COMPRISING SAME

<130>  2020-115-02

<140>  PCT/IB2022/050554
<141>  2022-01-21

<150>  US 63/141,123
<151>  2021-01-25

<160>  39    

<170>  PatentIn version 3.5

<210>  1
<211>  130
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence for sdCA9-1- VHH

<400>  1

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


Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp Asp Trp 
            20                  25                  30          


Ala Ile Gly Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu Gly Val 
        35                  40                  45              


Ser Cys Ile Ser Lys Arg His Gly Thr Thr His Tyr Ala Asp Ser Val 
    50                  55                  60                  


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


Leu Arg Met Asn Gly Leu Lys Pro Glu Asp Thr Ala Val Tyr Tyr Cys 
                85                  90                  95      


Ala Ala Ser Ser Trp Gly Ser Cys Thr Val Ala Thr Met Arg Asp Val 
            100                 105                 110         


Asp Arg Tyr Asp Tyr Asp Tyr Trp Gly Gln Gly Thr Gln Val Thr Val 
        115                 120                 125             


Ser Ser 
    130 


<210>  2
<211>  8
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence of CDR1 of sdCA9-1- VHH

<400>  2

Gly Phe Thr Phe Asp Asp Trp Ala 
1               5               


<210>  3
<211>  8
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence of CDR2 of sdCA9-1- VHH

<400>  3

Ile Ser Lys Arg His Gly Thr Thr 
1               5               


<210>  4
<211>  23
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence of CDR3 of sdCA9-1- VHH

<400>  4

Ala Ala Ser Ser Trp Gly Ser Cys Thr Val Ala Thr Met Arg Asp Val 
1               5                   10                  15      


Asp Arg Tyr Asp Tyr Asp Tyr 
            20              


<210>  5
<211>  133
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence for sdCA9-2-VHH

<400>  5

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


Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Leu Asp Tyr Tyr 
            20                  25                  30          


Ala Ile Gly Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu Gly Val 
        35                  40                  45              


Ser Cys Phe Ser Ser Val Asp Gly Ser Thr Tyr Tyr Ala Asp Ser Val 
    50                  55                  60                  


Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Met Val Tyr 
65                  70                  75                  80  


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


Ala Ala Glu Leu Tyr Gly Glu Leu Gly Val Ala Thr Val Gln Ala Met 
            100                 105                 110         


Cys Arg Leu Thr Thr Pro Gly Gly Asp Tyr Trp Gly Gln Gly Thr Gln 
        115                 120                 125             


Val Thr Val Ser Ser 
    130             


<210>  6
<211>  8
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence of CDR1 of sdCA9-2-VHH

<400>  6

Gly Phe Thr Leu Asp Tyr Tyr Ala 
1               5               


<210>  7
<211>  8
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence of CDR2 of sdCA9-2-VHH

<400>  7

Phe Ser Ser Val Asp Gly Ser Thr 
1               5               


<210>  8
<211>  26
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence CDR3 of sdCA9-2-VHH

<400>  8

Ala Ala Glu Leu Tyr Gly Glu Leu Gly Val Ala Thr Val Gln Ala Met 
1               5                   10                  15      


Cys Arg Leu Thr Thr Pro Gly Gly Asp Tyr 
            20                  25      


<210>  9
<211>  118
<212>  PRT
<213>  Artificial sequence

<220>
<223>  A 118 amino acid sequence for CA9-3-VHH

<400>  9

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


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


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


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


Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Glu Asn Thr Val Tyr 
65                  70                  75                  80  


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


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


Gln Val Thr Val Ser Ser 
        115             


<210>  10
<211>  8
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence of CDR1 of sdCA9-3-VHH

<400>  10

Gly Gly Thr Phe Ser Arg Tyr Gly 
1               5               


<210>  11
<211>  8
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence of CDR2 of sdCA9-3-VHH

<400>  11

Ile Ser Gly Thr Gly Leu Asn Thr 
1               5               


<210>  12
<211>  11
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Amino acid sequence of CDR3 of sdCA9-3-VHH

<400>  12

Ala Arg Asp Arg Gly Gly Val Glu Tyr Asp Tyr 
1               5                   10      


<210>  13
<211>  23
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Leader amino acid sequence

<400>  13

Glu Phe Ala Thr Met Glu Phe Gly Leu Ser Trp Val Phe Leu Val Ala 
1               5                   10                  15      


Ile Leu Lys Gly Val Gln Cys 
            20              


<210>  14
<211>  16
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Linker amino acid sequence

<400>  14

Ala Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro 
1               5                   10                  15      


<210>  15
<211>  216
<212>  PRT
<213>  Artificial sequence

<220>
<223>  hIgG1Fc

<400>  15

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


Pro Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val 
            20                  25                  30          


Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr 
        35                  40                  45              


Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu 
    50                  55                  60                  


Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His 
65                  70                  75                  80  


Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys 
                85                  90                  95      


Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln 
            100                 105                 110         


Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu 
        115                 120                 125             


Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro 
    130                 135                 140                 


Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn 
145                 150                 155                 160 


Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu 
                165                 170                 175     


Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val 
            180                 185                 190         


Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln 
        195                 200                 205             


Lys Ser Leu Ser Leu Ser Pro Gly 
    210                 215     


<210>  16
<211>  385
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Leader-sdCA9-1-VHH-Linker-hIgG1Fc

<400>  16

Glu Phe Ala Thr Met Glu Phe Gly Leu Ser Trp Val Phe Leu Val Ala 
1               5                   10                  15      


Ile Leu Lys Gly Val Gln Cys Gln Val Gln Leu Val Glu Ser Gly Gly 
            20                  25                  30          


Gly Leu Val Gln Ala Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser 
        35                  40                  45              


Gly Phe Thr Phe Asp Asp Trp Ala Ile Gly Trp Phe Arg Gln Ala Pro 
    50                  55                  60                  


Gly Lys Glu Arg Glu Gly Val Ser Cys Ile Ser Lys Arg His Gly Thr 
65                  70                  75                  80  


Thr His Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Ser Asp 
                85                  90                  95      


Asn Ala Lys Asn Thr Val Tyr Leu Arg Met Asn Gly Leu Lys Pro Glu 
            100                 105                 110         


Asp Thr Ala Val Tyr Tyr Cys Ala Ala Ser Ser Trp Gly Ser Cys Thr 
        115                 120                 125             


Val Ala Thr Met Arg Asp Val Asp Arg Tyr Asp Tyr Asp Tyr Trp Gly 
    130                 135                 140                 


Gln Gly Thr Gln Val Thr Val Ser Ser Ala Glu Pro Lys Ser Cys Asp 
145                 150                 155                 160 


Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly 
                165                 170                 175     


Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile 
            180                 185                 190         


Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu 
        195                 200                 205             


Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His 
    210                 215                 220                 


Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg 
225                 230                 235                 240 


Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys 
                245                 250                 255     


Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu 
            260                 265                 270         


Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr 
        275                 280                 285             


Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln Val Ser Leu 
    290                 295                 300                 


Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp 
305                 310                 315                 320 


Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val 
                325                 330                 335     


Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp 
            340                 345                 350         


Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His 
        355                 360                 365             


Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro 
    370                 375                 380                 


Gly 
385 


<210>  17
<211>  388
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Leader-sdCA9-2-VHH-Linker-hIgG1Fc

<400>  17

Glu Phe Ala Thr Met Glu Phe Gly Leu Ser Trp Val Phe Leu Val Ala 
1               5                   10                  15      


Ile Leu Lys Gly Val Gln Cys Gln Val Lys Leu Glu Glu Ser Gly Gly 
            20                  25                  30          


Asp Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser 
        35                  40                  45              


Gly Phe Thr Leu Asp Tyr Tyr Ala Ile Gly Trp Phe Arg Gln Ala Pro 
    50                  55                  60                  


Gly Lys Glu Arg Glu Gly Val Ser Cys Phe Ser Ser Val Asp Gly Ser 
65                  70                  75                  80  


Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp 
                85                  90                  95      


Asn Ala Lys Asn Met Val Tyr Leu Gln Met Asn Ser Leu Lys Pro Glu 
            100                 105                 110         


Asp Thr Ala Ile Tyr Tyr Cys Ala Ala Glu Leu Tyr Gly Glu Leu Gly 
        115                 120                 125             


Val Ala Thr Val Gln Ala Met Cys Arg Leu Thr Thr Pro Gly Gly Asp 
    130                 135                 140                 


Tyr Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser Ala Glu Pro Lys 
145                 150                 155                 160 


Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu 
                165                 170                 175     


Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr 
            180                 185                 190         


Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val 
        195                 200                 205             


Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val 
    210                 215                 220                 


Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser 
225                 230                 235                 240 


Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu 
                245                 250                 255     


Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala 
            260                 265                 270         


Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro 
        275                 280                 285             


Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn Gln 
    290                 295                 300                 


Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala 
305                 310                 315                 320 


Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr 
                325                 330                 335     


Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu 
            340                 345                 350         


Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser 
        355                 360                 365             


Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser 
    370                 375                 380                 


Leu Ser Pro Gly 
385             


<210>  18
<211>  373
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Leader-sdCA9-3-VHH-Linker-hIgG1Fc

<400>  18

Glu Phe Ala Thr Met Glu Phe Gly Leu Ser Trp Val Phe Leu Val Ala 
1               5                   10                  15      


Ile Leu Lys Gly Val Gln Cys Gln Val Gln Leu Val Glu Ser Gly Gly 
            20                  25                  30          


Gly Leu Val Gln Ala Gly Asp Ser Leu Arg Leu Ser Cys Ala Ala Ser 
        35                  40                  45              


Gly Gly Thr Phe Ser Arg Tyr Gly Met Gly Trp Phe Arg Gln Ala Pro 
    50                  55                  60                  


Gly Lys Glu Arg Glu Phe Val Ala Ala Ile Ser Gly Thr Gly Leu Asn 
65                  70                  75                  80  


Thr Tyr Tyr Met Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp 
                85                  90                  95      


Asn Ala Glu Asn Thr Val Tyr Leu Gln Met Asn Ser Leu Lys Pro Glu 
            100                 105                 110         


Asp Thr Ala Val Tyr Tyr Cys Ala Arg Asp Arg Gly Gly Val Glu Tyr 
        115                 120                 125             


Asp Tyr Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser Ala Glu Pro 
    130                 135                 140                 


Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu 
145                 150                 155                 160 


Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp 
                165                 170                 175     


Thr Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp 
            180                 185                 190         


Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly 
        195                 200                 205             


Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn 
    210                 215                 220                 


Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp 
225                 230                 235                 240 


Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro 
                245                 250                 255     


Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu 
            260                 265                 270         


Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu Leu Thr Lys Asn 
        275                 280                 285             


Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile 
    290                 295                 300                 


Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr 
305                 310                 315                 320 


Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys 
                325                 330                 335     


Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys 
            340                 345                 350         


Ser Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu 
        355                 360                 365             


Ser Leu Ser Pro Gly 
    370             


<210>  19
<211>  38
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Nucleic acid sequence of sense primer VHH Bbsl

<400>  19
tatgaagaca ccaggcccag gtgcagctgg tggagtct                               38


<210>  20
<211>  31
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Nucleic acid sequence of anti-sense primer VHH-BamHI

<400>  20
cgcgggatcc tgaggagacg gtgacctggg t                                      31


<210>  21
<211>  390
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Nucleic acid sequence of sdCA9-1-VHH

<400>  21
caggtacagc tggtggagtc tgggggaggc ttggtgcagg ctggggggtc tctgagactc       60

tcctgtgcag cctctggatt cactttcgat gattgggcca taggctggtt ccgccaggcc      120

ccagggaagg agcgtgaggg ggtctcatgt attagtaaga ggcatggtac tacacactat      180

gcagactccg tgaagggccg attcaccatc tccagtgaca acgccaagaa cacggtgtat      240

ctgcgaatga acggcctgaa acctgaggac acggccgttt attactgtgc agcatcctcc      300

tggggatcat gtactgtagc gactatgagg gacgttgata gatatgacta tgactattgg      360

ggccagggga cccaggtcac cgtctcctca                                       390


<210>  22
<211>  399
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Nucleic acid sequence of sdCA9-2-VHH

<400>  22
caggtaaagc tggaggagtc tgggggagac ttggtgcagc ctggggggtc tctgagactc       60

tcctgtgcag cctctggatt cactttggat tattatgcca taggctggtt ccgccaggcc      120

ccagggaagg agcgtgaggg ggtctcatgt tttagtagtg ttgatggtag cacatactat      180

gcagactccg tgaagggccg attcaccatc tccagagaca acgccaagaa catggtgtat      240

ctgcaaatga acagcctgaa acctgaggac acagccattt attactgtgc agcagaattg      300

tacggggagt tgggagtggc tactgttcag gctatgtgtc gcctgactac tccggggggt      360

gactactggg gccaggggac ccaggtcacc gtctcctca                             399


<210>  23
<211>  354
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Nucleic acid sequence of sdCA9-3-VHH

<400>  23
caggtgcagc tggtggagtc tgggggagga ttggtgcagg ctggggactc tcttagactc       60

tcctgtgcag cctctggcgg caccttcagt aggtatggca tgggctggtt ccgccaggct      120

ccagggaagg agcgtgagtt tgtcgcagct attagcggga ctggtctgaa tacatactat      180

atggactccg tgaagggccg attcaccatc tccagagaca acgccgagaa cacggtgtat      240

ctgcaaatga acagcctgaa acctgaggac acggccgtgt attactgtgc cagagaccga      300

ggaggcgttg agtatgacta ctggggccag gggacccagg tcaccgtctc ctca            354


<210>  24
<211>  1095
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Nucleic acid sequence of sdCA9-1-linker- hIgG1Fc

<400>  24
caggtacagc tggtggagtc tgggggaggc ttggtgcagg ctggggggtc tctgagactc       60

tcctgtgcag cctctggatt cactttcgat gattgggcca taggctggtt ccgccaggcc      120

ccagggaagg agcgtgaggg ggtctcatgt attagtaaga ggcatggtac tacacactat      180

gcagactccg tgaagggccg attcaccatc tccagtgaca acgccaagaa cacggtgtat      240

ctgcgaatga acggcctgaa acctgaggac acggccgttt attactgtgc agcatcctcc      300

tggggatcat gtactgtagc gactatgagg gacgttgata gatatgacta tgactattgg      360

ggccagggga cccaggtcac cgtctcctca gctgaaccca agtcctgcga caagacccac      420

acctgtcccc cctgccctgc ccctgaactg ctgggcggac cttccgtgtt cctgttcccc      480

ccaaagccta aggacaccct gatgatctcc cggacccccg aagtgacctg cgtggtggtg      540

gacgtgtccc acgaggaccc tgaagtgaag ttcaattggt acgtggacgg cgtggaagtg      600

cacaacgcca agaccaagcc cagagaggaa cagtacaact ccacctaccg ggtggtgtcc      660

gtgctgaccg tgctgcacca ggactggctg aacggcaaag agtacaagtg caaggtctcc      720

aacaaggccc tgcctgcccc catcgaaaag accatcagca aggccaaggg ccagccccgc      780

gagccccagg tgtacaccct gccccctagc cgggacgagc tgaccaagaa tcaggtgtcc      840

ctgacctgtc tggtgaaagg cttctacccc tccgatatcg ccgtggaatg ggagtccaac      900

ggccagcccg agaacaacta caagaccacc ccccctgtgc tggactccga cggctcattc      960

ttcctgtact ccaagctgac cgtggacaag tcccggtggc agcagggcaa cgtgttctcc     1020

tgctccgtga tgcacgaggc cctgcacaac cactacaccc agaagtccct gtccctgagc     1080

cccggctgag gatcc                                                      1095


<210>  25
<211>  1104
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Nucleic acid sequence of sdCA9-2-linker- hIgG1Fc

<400>  25
caggtaaagc tggaggagtc tgggggagac ttggtgcagc ctggggggtc tctgagactc       60

tcctgtgcag cctctggatt cactttggat tattatgcca taggctggtt ccgccaggcc      120

ccagggaagg agcgtgaggg ggtctcatgt tttagtagtg ttgatggtag cacatactat      180

gcagactccg tgaagggccg attcaccatc tccagagaca acgccaagaa catggtgtat      240

ctgcaaatga acagcctgaa acctgaggac acagccattt attactgtgc agcagaattg      300

tacggggagt tgggagtggc tactgttcag gctatgtgtc gcctgactac tccggggggt      360

gactactggg gccaggggac ccaggtcacc gtctcctcag ctgaacccaa gtcctgcgac      420

aagacccaca cctgtccccc ctgccctgcc cctgaactgc tgggcggacc ttccgtgttc      480

ctgttccccc caaagcctaa ggacaccctg atgatctccc ggacccccga agtgacctgc      540

gtggtggtgg acgtgtccca cgaggaccct gaagtgaagt tcaattggta cgtggacggc      600

gtggaagtgc acaacgccaa gaccaagccc agagaggaac agtacaactc cacctaccgg      660

gtggtgtccg tgctgaccgt gctgcaccag gactggctga acggcaaaga gtacaagtgc      720

aaggtctcca acaaggccct gcctgccccc atcgaaaaga ccatcagcaa ggccaagggc      780

cagccccgcg agccccaggt gtacaccctg ccccctagcc gggacgagct gaccaagaat      840

caggtgtccc tgacctgtct ggtgaaaggc ttctacccct ccgatatcgc cgtggaatgg      900

gagtccaacg gccagcccga gaacaactac aagaccaccc cccctgtgct ggactccgac      960

ggctcattct tcctgtactc caagctgacc gtggacaagt cccggtggca gcagggcaac     1020

gtgttctcct gctccgtgat gcacgaggcc ctgcacaacc actacaccca gaagtccctg     1080

tccctgagcc ccggctgagg atcc                                            1104


<210>  26
<211>  1059
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Nucleic acid sequence of sdCA9-3-linker- hIgG1Fc

<400>  26
caggtgcagc tggtggagtc tgggggagga ttggtgcagg ctggggactc tcttagactc       60

tcctgtgcag cctctggcgg caccttcagt aggtatggca tgggctggtt ccgccaggct      120

ccagggaagg agcgtgagtt tgtcgcagct attagcggga ctggtctgaa tacatactat      180

atggactccg tgaagggccg attcaccatc tccagagaca acgccgagaa cacggtgtat      240

ctgcaaatga acagcctgaa acctgaggac acggccgtgt attactgtgc cagagaccga      300

ggaggcgttg agtatgacta ctggggccag gggacccagg tcaccgtctc ctcagctgaa      360

cccaagtcct gcgacaagac ccacacctgt cccccctgcc ctgcccctga actgctgggc      420

ggaccttccg tgttcctgtt ccccccaaag cctaaggaca ccctgatgat ctcccggacc      480

cccgaagtga cctgcgtggt ggtggacgtg tcccacgagg accctgaagt gaagttcaat      540

tggtacgtgg acggcgtgga agtgcacaac gccaagacca agcccagaga ggaacagtac      600

aactccacct accgggtggt gtccgtgctg accgtgctgc accaggactg gctgaacggc      660

aaagagtaca agtgcaaggt ctccaacaag gccctgcctg cccccatcga aaagaccatc      720

agcaaggcca agggccagcc ccgcgagccc caggtgtaca ccctgccccc tagccgggac      780

gagctgacca agaatcaggt gtccctgacc tgtctggtga aaggcttcta cccctccgat      840

atcgccgtgg aatgggagtc caacggccag cccgagaaca actacaagac caccccccct      900

gtgctggact ccgacggctc attcttcctg tactccaagc tgaccgtgga caagtcccgg      960

tggcagcagg gcaacgtgtt ctcctgctcc gtgatgcacg aggccctgca caaccactac     1020

acccagaagt ccctgtccct gagccccggc tgaggatcc                            1059


<210>  27
<211>  657
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Nucleic acid sequence  encoding hIgG1Fc (SEQ ID NO: 15)

<400>  27
gcccctgaac tgctgggcgg accttccgtg ttcctgttcc ccccaaagcc taaggacacc       60

ctgatgatct cccggacccc cgaagtgacc tgcgtggtgg tggacgtgtc ccacgaggac      120

cctgaagtga agttcaattg gtacgtggac ggcgtggaag tgcacaacgc caagaccaag      180

cccagagagg aacagtacaa ctccacctac cgggtggtgt ccgtgctgac cgtgctgcac      240

caggactggc tgaacggcaa agagtacaag tgcaaggtct ccaacaaggc cctgcctgcc      300

cccatcgaaa agaccatcag caaggccaag ggccagcccc gcgagcccca ggtgtacacc      360

ctgcccccta gccgggacga gctgaccaag aatcaggtgt ccctgacctg tctggtgaaa      420

ggcttctacc cctccgatat cgccgtggaa tgggagtcca acggccagcc cgagaacaac      480

tacaagacca ccccccctgt gctggactcc gacggctcat tcttcctgta ctccaagctg      540

accgtggaca agtcccggtg gcagcagggc aacgtgttct cctgctccgt gatgcacgag      600

gccctgcaca accactacac ccagaagtcc ctgtccctga gccccggctg aggatcc         657


<210>  28
<211>  1675
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Nucleic acid sequence  encoding CAR modular construct (SEQ ID NO:
       28)


<220>
<221>  MISC_FEATURE
<222>  (62)..(63)
<223>  Insertion site for an antigen binding domain encoding sequence is
       between residues 62 and 63.

<220>
<221>  MISC_FEATURE
<222>  (62)..(63)
<223>  Insertion site for an antigen binding domain encoding sequence is
       between nucleotides 62 and 63.

<400>  28
atgctcaggc tgctcttggc tctcaactta ttcccttcaa ttcaagtaac aggagggtct       60

tcgaagactt cctttgcgag acgacggtgg cgggggatca ggtggtggag gtagcggggg      120

agggggctca ggcggtacaa ctacgcctgc acctcgccca ccgaccccag caccaaccat      180

cgcttcacag cctttgagcc tgcgaccaga ggcatgtcgc cctgctgcgg gcggtgccgt      240

tcatactcgc ggacttgatt ttgcgtgtga cgtcgtctcg ccttctaagc ccttttgggt      300

gctggtggtg gttggtggag tcctggcttg ctatagcttg ctagtaacag tggcctttat      360

tattttctgg gtgaggaaac ggggcagaaa gaaactcctg tatatattca aacaaccatt      420

tatgcgacca gtacaaacta ctcaagagga agatggctgt agctgccgat ttccagaaga      480

agaagaagga ggatgtgaac tgctgagagt gaagttcagc aggagcgcag acgcccccgc      540

gtaccagcag ggccagaacc agctctataa cgagctcaat ctaggacgaa gagaggagta      600

cgatgttttg gacaagcgac gtggccggga ccctgagatg gggggaaagc cgcagagaag      660

gaagaaccct caggaaggcc tgtacaatga actgcagaaa gataagatgg cggaggccta      720

cagtgagatt gggatgaaag gcgagcgccg gaggggcaag gggcacgatg gcctttacca      780

gggactcagt acagccacca aggacaccta cgacgccctt cacatgcagg ccctgccccc      840

tcgcgctagc gccacgaact tctctctgtt aaagcaagca ggcgacgtgg aagaaaaccc      900

cggtcccatg gtgagcaagg gcgaggagga caacatggcc agcctgcccg ccacccacga      960

gctgcacatc ttcggcagca tcaacggcgt ggacttcgac atggtgggcc agggcaccgg     1020

caaccccaac gacggctacg aggagctgaa cctgaagagc accaagggcg acctgcagtt     1080

cagcccctgg attctggtgc cccacatcgg ctacggcttc caccagtacc tgccctaccc     1140

cgacggcatg agccccttcc aggccgccat ggtggacggc agcggctacc aggtgcacag     1200

gaccatgcag ttcgaggacg gcgccagcct gaccgtgaac tacaggtaca cctacgaggg     1260

cagccacatc aagggcgagg cccaggtgaa gggcaccggc ttccccgccg acggccccgt     1320

gatgaccaac agcctgaccg ccgccgactg gtgcaggagc aaaaagacct accccaacga     1380

caagaccatc atcagcacct tcaagtggag ctacaccacc ggcaacggca agaggtacag     1440

gagcaccgcc aggaccacct acaccttcgc caagcccatg gccgccaact acctgaagaa     1500

ccagcccatg tacgtgttca gaaagaccga gctgaagcac agcaagaccg agctgaactt     1560

caaggagtgg cagaaggcct tcaccgacgt gatgggcatg gacgagctgt acaagcccaa     1620

gaagaagagg aaggtggagg acccccccgc cgccaagagg gtgaagctgg actaa          1675


<210>  29
<211>  18
<212>  PRT
<213>  Homo sapiens

<400>  29

Met Leu Arg Leu Leu Leu Ala Leu Asn Leu Phe Pro Ser Ile Gln Val 
1               5                   10                  15      


Thr Gly 
        


<210>  30
<211>  17
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Synthetic Flexible Linker Domain (exemplary)

<400>  30

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


Gly 
    


<210>  31
<211>  45
<212>  PRT
<213>  Homo sapiens

<400>  31

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>  32
<211>  32
<212>  PRT
<213>  Homo sapiens

<400>  32

Pro Ser Lys Pro Phe Trp Val Leu Val Val Val Gly Gly Val Leu Ala 
1               5                   10                  15      


Cys Tyr Ser Leu Leu Val Thr Val Ala Phe Ile Ile Phe Trp Val Arg 
            20                  25                  30          


<210>  33
<211>  42
<212>  PRT
<213>  Homo sapiens

<400>  33

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>  34
<211>  114
<212>  PRT
<213>  Homo sapiens

<400>  34

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


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


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


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


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


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


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


Pro Arg 
        


<210>  35
<211>  5
<212>  PRT
<213>  Artificial sequence

<220>
<223>  Short flexible linker amino acid sequence

<400>  35

Gly Gly Gly Gly Ser 
1               5   


<210>  36
<211>  147
<212>  DNA
<213>  Artificial sequence

<220>
<223>  Bispecific T cell engager modular construct DNA sequence


<220>
<221>  MISC_FEATURE
<222>  (72)..(73)
<223>  Insertion site for an antigen binding domain encoding sequence is
       between nucleotides 72 and 73

<220>
<221>  MISC_FEATURE
<222>  (99)..(100)
<223>  Insertion site for an anti-CD3 variable heavy chain encoding 
       sequence is between nucleotides 99 and 100

<220>
<221>  MISC_FEATURE
<222>  (144)..(145)
<223>  Insertion site for an anti-CD3 variable light chain encoding 
       sequence is between nucleotides 144 and 145

<400>  36
atggagtttg ggctgagctg ggttttcctc gttgctcttt ttagaggtgt ccagtgtaca       60

ggagggtctt cggaagactt ccttggagga ggcggaagtg gcggtggtgg gtcaggcggc      120

ggtgggagcg gaggaggtgg aagctag                                          147


<210>  37
<211>  389
<212>  PRT
<213>  Artificial sequence

<220>
<223>  sdCA9-1-VHH CAR amino acid sequence, exemplary sequence for a 
       single binder CAR construct

<400>  37

Met Leu Arg Leu Leu Leu Ala Leu Asn Leu Phe Pro Ser Ile Gln Val 
1               5                   10                  15      


Thr Gly Gln Val Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Ala 
            20                  25                  30          


Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Asp 
        35                  40                  45              


Asp Trp Ala Ile Gly Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu 
    50                  55                  60                  


Gly Val Ser Cys Ile Ser Lys Arg His Gly Thr Thr His Tyr Ala Asp 
65                  70                  75                  80  


Ser Val Lys Gly Arg Phe Thr Ile Ser Ser Asp Asn Ala Lys Asn Thr 
                85                  90                  95      


Val Tyr Leu Arg Met Asn Gly Leu Lys Pro Glu Asp Thr Ala Val Tyr 
            100                 105                 110         


Tyr Cys Ala Ala Ser Ser Trp Gly Ser Cys Thr Val Ala Thr Met Arg 
        115                 120                 125             


Asp Val Asp Arg Tyr Asp Tyr Asp Tyr Trp Gly Gln Gly Thr Gln Val 
    130                 135                 140                 


Thr Val Ser Ser Pro Ser Gly Gly Gly Gly Pro Ser Thr Thr Thr Pro 
145                 150                 155                 160 


Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile Ala Ser Gln Pro Leu 
                165                 170                 175     


Ser Leu Arg Pro Glu Ala Cys Arg Pro Ala Ala Gly Gly Ala Val His 
            180                 185                 190         


Thr Arg Gly Leu Asp Phe Ala Cys Asp Pro Ser Lys Pro Phe Trp Val 
        195                 200                 205             


Leu Val Val Val Gly Gly Val Leu Ala Cys Tyr Ser Leu Leu Val Thr 
    210                 215                 220                 


Val Ala Phe Ile Ile Phe Trp Val Arg Lys Arg Gly Arg Lys Lys Leu 
225                 230                 235                 240 


Leu Tyr Ile Phe Lys Gln Pro Phe Met Arg Pro Val Gln Thr Thr Gln 
                245                 250                 255     


Glu Glu Asp Gly Cys Ser Cys Arg Phe Pro Glu Glu Glu Glu Gly Gly 
            260                 265                 270         


Cys Glu Leu Leu Arg Val Lys Phe Ser Arg Ser Ala Asp Ala Pro Ala 
        275                 280                 285             


Tyr Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn Leu Gly Arg 
    290                 295                 300                 


Arg Glu Glu Tyr Asp Val Leu Asp Lys Arg Arg Gly Arg Asp Pro Glu 
305                 310                 315                 320 


Met Gly Gly Lys Pro Gln Arg Arg Lys Asn Pro Gln Glu Gly Leu Tyr 
                325                 330                 335     


Asn Glu Leu Gln Lys Asp Lys Met Ala Glu Ala Tyr Ser Glu Ile Gly 
            340                 345                 350         


Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His Asp Gly Leu Tyr Gln 
        355                 360                 365             


Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu His Met Gln 
    370                 375                 380                 


Ala Leu Pro Pro Arg 
385                 


<210>  38
<211>  392
<212>  PRT
<213>  Artificial sequence

<220>
<223>  sdCA9-2-VHH CAR amino acid sequence

<400>  38

Met Leu Arg Leu Leu Leu Ala Leu Asn Leu Phe Pro Ser Ile Gln Val 
1               5                   10                  15      


Thr Gly Gln Val Lys Leu Glu Glu Ser Gly Gly Asp Leu Val Gln Pro 
            20                  25                  30          


Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Leu Asp 
        35                  40                  45              


Tyr Tyr Ala Ile Gly Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu 
    50                  55                  60                  


Gly Val Ser Cys Phe Ser Ser Val Asp Gly Ser Thr Tyr Tyr Ala Asp 
65                  70                  75                  80  


Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala Lys Asn Met 
                85                  90                  95      


Val Tyr Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Ala Ile Tyr 
            100                 105                 110         


Tyr Cys Ala Ala Glu Leu Tyr Gly Glu Leu Gly Val Ala Thr Val Gln 
        115                 120                 125             


Ala Met Cys Arg Leu Thr Thr Pro Gly Gly Asp Tyr Trp Gly Gln Gly 
    130                 135                 140                 


Thr Gln Val Thr Val Ser Ser Pro Ser Gly Gly Gly Gly Pro Ser Thr 
145                 150                 155                 160 


Thr Thr Pro Ala Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile Ala Ser 
                165                 170                 175     


Gln Pro Leu Ser Leu Arg Pro Glu Ala Cys Arg Pro Ala Ala Gly Gly 
            180                 185                 190         


Ala Val His Thr Arg Gly Leu Asp Phe Ala Cys Asp Pro Ser Lys Pro 
        195                 200                 205             


Phe Trp Val Leu Val Val Val Gly Gly Val Leu Ala Cys Tyr Ser Leu 
    210                 215                 220                 


Leu Val Thr Val Ala Phe Ile Ile Phe Trp Val Arg Lys Arg Gly Arg 
225                 230                 235                 240 


Lys Lys Leu Leu Tyr Ile Phe Lys Gln Pro Phe Met Arg Pro Val Gln 
                245                 250                 255     


Thr Thr Gln Glu Glu Asp Gly Cys Ser Cys Arg Phe Pro Glu Glu Glu 
            260                 265                 270         


Glu Gly Gly Cys Glu Leu Leu Arg Val Lys Phe Ser Arg Ser Ala Asp 
        275                 280                 285             


Ala Pro Ala Tyr Gln Gln Gly Gln Asn Gln Leu Tyr Asn Glu Leu Asn 
    290                 295                 300                 


Leu Gly Arg Arg Glu Glu Tyr Asp Val Leu Asp Lys Arg Arg Gly Arg 
305                 310                 315                 320 


Asp Pro Glu Met Gly Gly Lys Pro Gln Arg Arg Lys Asn Pro Gln Glu 
                325                 330                 335     


Gly Leu Tyr Asn Glu Leu Gln Lys Asp Lys Met Ala Glu Ala Tyr Ser 
            340                 345                 350         


Glu Ile Gly Met Lys Gly Glu Arg Arg Arg Gly Lys Gly His Asp Gly 
        355                 360                 365             


Leu Tyr Gln Gly Leu Ser Thr Ala Thr Lys Asp Thr Tyr Asp Ala Leu 
    370                 375                 380                 


His Met Gln Ala Leu Pro Pro Arg 
385                 390         


<210>  39
<211>  222
<212>  PRT
<213>  Artificial sequence

<220>
<223>  sdCAIX2-CD3scFv amino acid sequence, exemplary sequence for a CAIX
       bispecific immune engager construct


<220>
<221>  SITE
<222>  (216)..(217)
<223>  Insertion site for a CD3-specific scFv sequence is between 
       residues 216 and 217

<400>  39

Met Glu Phe Gly Leu Ser Trp Val Phe Leu Val Ala Leu Phe Arg Gly 
1               5                   10                  15      


Val Gln Cys Thr Gly Gln Val Lys Leu Glu Glu Ser Gly Gly Asp Leu 
            20                  25                  30          


Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe 
        35                  40                  45              


Thr Leu Asp Tyr Tyr Ala Ile Gly Trp Phe Arg Gln Ala Pro Gly Lys 
    50                  55                  60                  


Glu Arg Glu Gly Val Ser Cys Phe Ser Ser Val Asp Gly Ser Thr Tyr 
65                  70                  75                  80  


Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ala 
                85                  90                  95      


Lys Asn Met Val Tyr Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr 
            100                 105                 110         


Ala Ile Tyr Tyr Cys Ala Ala Glu Leu Tyr Gly Glu Leu Gly Val Ala 
        115                 120                 125             


Thr Val Gln Ala Met Cys Arg Leu Thr Thr Pro Gly Gly Asp Tyr Trp 
    130                 135                 140                 


Gly Gln Gly Thr Gln Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly 
145                 150                 155                 160 


Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Thr Thr Thr Pro Ala 
                165                 170                 175     


Pro Arg Pro Pro Thr Pro Ala Pro Thr Ile Ala Ser Gln Pro Leu Ser 
            180                 185                 190         


Leu Arg Pro Glu Ala Cys Arg Pro Ala Ala Gly Gly Ala Val His Thr 
        195                 200                 205             


Arg Gly Leu Asp Phe Ala Cys Asp His His His His His His 
    210                 215                 220         


