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iAMP21 xenografts
gression remains uncertain, as it was co-selected with other abnormalities, including an NRAS mutation and bi- allelic loss of CDKN2A/B. Among the four other cases transplanted, three were affected by concordant or discor- dant CDKN2A deletions, two bi-allelic and one mono-allel- ic, detected by MLPA only and without apparent involve- ment of CDKN2B. Further suggesting strong selective pres- sure for loss of CDKN2A/B in the xenografts, as evidenced by RNA sequencing data, the bi-allelic deletions were all highly clonal. As deletion of this locus only occurs in about 12% of iAMP21-ALL patients,21 these observations support previous reports that CDKN2A/B loss is associated with rapid disease manifestation27 and is selected for in B-ALL xenografts,4 and are also in keeping with a xenograft-spe- cific expression signature enriched for cell cycle genes.31 Alternative mutations of NRAS and KRAS were also strongly selected and both apparently cooperated with an NF1 mutation in xenografts. To our knowledge NF1 and RAS mutations have always been reported as mutually exclusive in B-ALL patients, although their co-occurrence in juvenile myelomoncytic leukemia has been associated with aggressive disease. In mouse models, a combination of NF1 deficiency and KRAS activating mutation reduced the latency of myeloid malignancy compared with either abnormality alone.13,32,33
Other chromosomal regions were strongly implicated in the progression of ALL, as targets of overlapping abnormal- ities segregated in different clones of xenografts. These included genes known to be involved in B-ALL; ETV6, SH2B3 and BLNK (SLP-65),34-36 as well as novel candidate tumor suppressor genes. Two distinct large deletions, selected in different xenografts, resulted in conversion to homozygosity of a micro-deletion involving PIK3AP1 and LCOR. PIK3AP1 encodes an adaptor protein linking the B- cell receptor and CD19 to activation of PI3K/Akt.37-39 A sim- ilar function in the transduction of pre-B-cell receptor sig- naling is likely and, although not previously implicated in childhood leukemia, focal deletions of PIK3AP1 have been
reported in three cases of adult B-ALL.40,41 Combined with our evidence of strong selective pressure for its conversion from mono- to bi-allelic deletion, PIK3AP1 is an interesting candidate, possibly worthy of addition to the growing list of pre-B-cell receptor-related genes disrupted in childhood B-ALL.42 However a role for LCOR, which functions as a co-repressor of several nuclear hormone receptors,43 and has been reported to interact with methyltransferase com- plexes including polycomb repressive complex 2,44 should not be discounted.
In conclusion, we present the first successful xenografts of iAMP21-ALL and demonstrate their potential as experi- mental models for functional investigation of this poorly understood genetic subtype. These xenografts could also serve as models in pre-clinical trials or for personalized medicine, with the caveat that systemic niche destruction occurred in some cases. Tracking of CNA, to investigate genomic evolution in xenografts, revealed a surprisingly high rate of instability and examples of marked divergence in CN status of known leukemia driver genes. Further genomic screening of iAMP21-ALL xenografts is likely to reveal many more clonal abnormalities undetected in patients’ samples, augmenting data generated from clinical trial cohorts, as well as potentially guiding treatment in individual cases.
Acknowledgments
The authors would like to thank the Department of Cellular Medicine, Royal Victoria Infirmary, Newcastle-upon-Tyne NHS Foundation Trust for fixing and staining histological samples. Tracey Davey and Kath White, Electron Microscopy Research Services, Newcastle University for EM preparations and images and Clare Orange and Lynn Stevenson, University of Glasgow, for the brain histology and imaging. The brain histology slides were scanned by Glasgow University slide scanning and image analysis service at the QEUH. We would also like to thank the Bloodwise Childhood Leukaemia Cell Bank for providing primary childhood leukemia samples.
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