The first Japanese biobank of patient-derived pediatric acute lymphoblastic leukemia xenograft models

Abstract

A lack of practical resources in Japan has limited preclinical discovery and testing of therapies for pediatric relapsed and refractory acute lymphoblastic leukemia (ALL), which has poor outcomes. Here, we established 57 patient-derived xenografts (PDXs) in NOD.Cg-Prkdc^scid ll2rg^tm1Sug /ShiJic (NOG) mice and created a biobank by preserving PDX cells including three extramedullary relapsed ALL PDXs. We demonstrated that our PDX mice and PDX cells mimicked the biological features of relapsed ALL and that PDX models reproduced treatment-mediated clonal selection. Our PDX biobank is a useful scientific resource for capturing drug sensitivity features of pediatric patients with ALL, providing an essential tool for the development of targeted therapies.

Keywords: biobank; leukemia; pediatric; preclinical; xenograft.

FIGURE 1

Establishment of a biobank of pediatric ALL PDX models. (A) Engraftment of leukemia cells in PDX mice over time. Primary leukemic cells were transplanted into recipient mice, and the proportions of human CD19‐ or CD3‐positive ALL cells were detected in BM by flow cytometry. Each line represents data from a single mouse. (B) In vivo expansion rate of leukemic cells. Numbers above are mean expansion rates with standard deviation. BM (B‐ALL, n = 14), CSF (B‐ALL, n = 3; T‐ALL, n = 2), and testis (B‐ALL, n = 1).

FIGURE 2

PDX mice preserve the characteristics of primary samples. (A–D) Immunohistochemical analysis of leukemic cell proliferation in (A) spleen, (B) liver, (C) brain, and (D) testis from a xenograft mouse generated from a sample from patient ALL#25 relapsed in testis. (E) Unsupervised hierarchical clustering of expression of patterns using DNA microarray data among four primary ALL samples and the corresponding PDX cells derived from BM and spleen.

FIGURE 3

PDX cells retain gene‐expression features corresponding to their cytogenetic abnormalities. Unsupervised hierarchical clustering of expression of patterns of 500 genes among 16 PDX cells with cytogenetic abnormalities.

FIGURE 4

PDX cells are useful resources for in vitro drug testing. (A, B) In vitro drug treatment of leukemic cells from the same B‐ALL (A) and T‐ALL (B) xenograft mice. Single agent VCR, DEX, l‐Asp, Ara‐C, or Ara‐G therapies were assessed and sensitivity to each drug is shown as the ratio of that to VCR. (C) Heatmap of in vitro responses of 80 drugs tested against ALL PDX cells. Responses are shown for each drug against each PDX tested as scored according to the following formula: drug effective score = [log₁₂₅ × {1 − (survival rate at less than ×1/125 concentration)} + log₂₅ × {1 − (survival rate at less than ×1/25 concentration)} + log₅ × {1 − (survival rate at less than ×1/5 concentration)} + {1 − (survival rate at less than ×1 concentration)}] × 100/(log₁₂₅ + log₂₅+ log₅ + 1).

FIGURE 5

Treatment‐induced clonal selection reproduced in a patient‐derived mouse model. (A) Multiplexed‐FISH analysis showing the BCR‐ABL1 fusion signal in patient samples collected at diagnosis (left) and at relapse (right). Original magnification ×100. Arrowheads, yellow fusion signal; arrows, probe signals. Probe combinations were as follows: BCR, green; ABL1, red. (B) Human leukemic chimerism levels in the BM of PDX mice over time in untreated (left, n = 5) and dasatinib treated (right, n = 3) mice. Data are presented as means with standard deviations. (C) Patterns of cytogenetic subclonal populations. Clonal stability, with no change in the dominant clone between transplantation without treatment, is observed in upper right and lower left charts. Clonal selection with a minor population becoming dominant at relapse in the clinical setting was reproduced in PDX mice (lower right chart).