Drug Response Profiling of Childhood T-Cell Acute Lymphoblastic Leukemia to Inform Functional Precision Medicine

Background/Aims: Despite the advancement in risk-directed, chemotherapy-based protocols, the prognosis of children with T-cell acute lymphoblastic leukemia (T-ALL) especially for those who were refractory to existing treatments or relapsed from remissions remains poor. The complexity of mutational landscape and lack of actionable genomic lesions collectively pose significant challenges for implementing targeted therapies based solely on genomics. To address this issue, this study seeks to utilize drug sensitivity and resistance test (DSRT) to directly assess the ex vivo response of primary T-lymphoblasts to a portfolio of targeted agents, aiming to establish its drug sensitivity profile and identify drug candidates of immediate clinical relevance to tailor functional precision medicine.

Methods: A serum-free, mesenchymal stem cell (MSC)-based culture system was adopted to perform ex vivo DSRT on primary or patient-derived xenograft (PDX) samples of pediatric T-ALL. The latter was generated by transplanting NSG mice with paucicellular specimens with splenocytes collected at leukemia engraftment as the sources of patient-derived T-lymphoblasts. The 42-drug panel consisted of 35 targeted agents (1 in Phase I trials, 6 in Phase II trials, 6 in Phase III trials, 22 FDA-approved) and 7 standard chemotherapeutics. The DSRT tested six serial drug concentrations ranging from 0.1 to 10,000 nM. After 96 hours of co-culturing with MSCs, T lymphoblasts were identified by CD7 staining, while apoptotic cells were characterized by Annexin V/7-AAD. The half maximal inhibitory concentration (IC50) of individual drugs were curated from the dose-response curves. Unsupervised clustering was performed to map the overall drug response pattern. A case series of relapsed or refractory T-ALL were managed with targeted therapies guided by drug profiling results and evaluated for clinical responses.

Results: DSRT was conducted on 19 primary and 14 PDX pediatric T-ALL samples from 20 patients. For primary samples, two had been tested for 42 drugs, ten were tested for 16 drugs and the rest underwent testing with varying numbers of drugs depending on cellularity and sample quality. All PDX samples were subjected to testing for 42 drugs. PDX samples retained the original pattern of drug response in matched patient materials (R ² = 0.77-0.94, 8 paired samples). Unsupervised clustering identified 5 distinct groups of drug response: (i) highly active compounds with median IC50 <10 nM (4 drugs including the proteasome inhibitors bortezomib and carfizomib, BCL2 inhibitor navitoclax and HDAC inhibitor panobinostat); (ii) generally active compounds (median IC50 <230 nM, 10 drugs), including BCL2 inhibitors, MCL1 inhibitors, JAK2/3 inhibitors and MDM2 inhibitors; (iii) compounds with bimodal activities (wide IC50 ranges, 4 drugs), including dasatinib (tyrosine kinase inhibitor), sirolimus and everolimus (mTOR inhibitors) and standard chemotherapeutics; (iv) generally inactive drugs with sporadic exceptions (12 drugs); and (v) completely inactive compounds (12 drugs). Four children with T-ALL received treatment according to drug profiling results. Two patients were in first relapse, one in third relapse and one in a non-remission state. Three of these patients received bortezomib treatment, while the fourth patient received venetoclax. All patients responded to the treatments with complete or partial remission. One patient was bridged to allogeneic stem cell transplantation, and two were bridged to CD7-directed CAR therapies.

Conclusions: Our study unveiled a pediatric-specific drug response profile for T-ALL and highlighted opportunistic drug candidates with clinical relevance. The successful implementation of functional precision medicine in our clinic elucidated its practicality and potential to enable personalized treatment for high-risk pediatric T-ALL.