Supplementary files
PAX5 mutations occur frequently in adult B-Cell Progenitor Acute Lymphoblastic Leukemia
(BCP-ALL) and are associated with BCR-ABL1 and TCF3-PBX1 fusion genes. A GRAALL
study
Figure S1
FISH analysis of two patients (#119339 and #462837) with a der(19)t(1;19) translocation
(TCF3-PBX1 fusion) showing a secondary deletion of PAX5 RP11-243F8 probe. TCF3 split
signal probes labeled with Texas red (TCF3 5’) and FITC (TCF3 3’) visible with the FITCTexas red-DAPI filter and PAX5 RP11-243F8 probe labeled with Rhodamine visible both with
FITC-Texas red-DAPI filter and FITC-Rhodamine-DAPI filter.
Patient #119339. Top panel: metaphase with normal PAX5 signals; bottom panel: nucleus
with PAX5 deletion (only one RP11-243F8 probe signal). Left: FITC-Texas red-DAPI filter
allows to detect Texas red TCF3 5’ signals on chromosomes 19 and der(19); TCF3 3’ FITC
signal on normal chromosome 19. The der(19) has lost the 3’ signal as the consequence of
the TCF3-PBX1 fusion. The PAX5 Rhodamine signals are also detected with this filter. Right
panel. FITC-Rhodamine-DAPI filter allows to visualize specifically two (Top) or one (Bottom)
PAX5 Rhodamine signals and TCF3 3’ FITC signals on the normal chromosome 19. This
patient has 80% of the nucleus with a TCF3-PBX1 fusion and 20% of these nucleus show a
secondary deletion of PAX5.
Patient #462837. Top panel: nucleus with normal PAX5 signals; bottom panel: nucleus with
PAX5 deletion (only one RP11-243F8 probe signal). Left: FITC-Texas red-DAPI filter allows
to detect Texas red TCF3 5’ signals on chromosomes 19 and der(19); TCF3 3’ FITC signal
on normal chromosome 19. The der(19) has lost the 3’ signal as the consequence of the
TCF3-PBX1 fusion. The PAX5 Rhodamine signals are also detected with this filter. Right
panel. FITC-Rhodamine-DAPI filter allows to visualize specifically two (Top) or one (Bottom)
PAX5 Rhodamine signals and TCF3 3’ FITC signals on the normal chromosome 19. This
patient has 90% of the nucleus with a TCF3-PBX1 fusion and 60% of these nucleus show a
secondary deletion of PAX5.
Figure S2
HRM analysis for PAX5 exons 1, 4, 5, 6 and 10. PAX5 exon 3 analysis was used as
validation of the HRM screening.
Figure S3
Analysis of PAX5 point mutations
Sequence chromatographs of PAX5 mutants performed on genomic DNA at diagnosis (dg)
and follow up (fu) when available. Nucleotide sequence is shown on the top of each panel.
Patient number are given on top of each frame..
Table S1
Primer sequences
Table S2
Genomic PAX5 quantification and classification of the GRAALL-2003/GRAAPH-2003
patients in 4 groups regarding the PAX5 mutations.
This table shows the study identification number of patients and raw data of quantification of
each PAX5 exons. Four groups were defined as Normal, Deletion, Mutation and
Amplification.
Table S3
PAX5 FISH validation of normal cases (normal PAX5), deletion-only cases (deleted PAX5)
and mutant cases (mutant PAX5) using the PAX5 DAKO probe.
Table S4
Deletion 9p regarding PAX5 status. 0: no deletion; 1: deletion.
Table S5
Clinical and biological data
Clinical features of GRAALL-2003/GRAAPH-2003 patients included in this analysis. The first
column shows the study identification number of patients. The second column shows their
PAX5 status. The third column indicates their age in years. The fourth column indicates their
gender (F: female, M: male). The fifth column reports the corticosensitivity (S) or
corticoresistance (R) during the 7-day prephase of steroid therapy (NA when information is
not available). The sixth column reports the chemosensitivity (S) or resistance (R) during the
induction course. The seventh column provides the graft status (Yes when engrafted). The
eighth column indicates the achievement (Yes) or not (No) of complete remission (CR1) by
the patient. The ninth column indicates whether relapse occurred after the first complete
remission as relapse (Yes) or absence (No). The tenth column provides the patient status as
dead (Yes) or alive (No). The last column shows cytogenetic/molecular subgroups. Normal :
no cytogenetic/molecular abnormality; BCR-ABL1 : Ph chromosome/BCR-ABL1; MLL : MLL
rearrangement; TCF3-PBX1; hypo-triploid : hypodiploidy 30-40 chromosomes and/or near-
triploidy 60-80 chromosomes; 51-65 : high hyperdiploidy 51-65 chromosomes; 47-50 : low
hyperdiploidy 47-50 chromosomes; 41-45: hypodiploidy 41-45 chromosomes; complex : 5 or
more chromosomal abnormalities; other : chromosomal abnormality not otherwise
classified.PAX5 mutations occur frequently in adult B-Cell Progenitor Acute Lymphoblastic
Leukemia (BCP-ALL) and are associated with BCR-ABL1 and TCF3-PBX1 fusion genes. A
GRAALL study