Heterogeneity in Phenotype, Functional Capacity, and Drug Sensitivity for Pediatric Acute Leukemia.
Authors: Wiedl Christina M. , Horton Terzah M. , Rossi Randall M , Neering Sarah J , Grose Valerie , Jordan Craig T. 
Published: October 2019

Abstract 2654

Poster Board II-630

Introduction:

With the institution of multidrug, multiphase chemotherapy regimens, major improvements in clinical outcomes have been made in pediatric acute leukemia patients in the last thirty years. However, there remains a substantial percentage of pediatric patients who relapse and die of their disease, particularly with high risk ALL, T cell ALL and AML. It is possible that these patients' disease initiates from a leukemic stem cell such as those found in adult myeloid disease, or at the very least, harbor a chemo-resistant population. Our research has two main aims: first to evaluate the functional and phenotypic heterogeneity within standard risk (SR), high risk (HR) and relapsed (RD) pediatric leukemia. Second, to evaluate current treatment regimens for the selection of a chemo-resistant or LSC populations and then attempt to target this population with novel treatments.

Methods:

In vitro studies for functional heterogeneity include colony-forming assays (CFU) using methylcellulose and limiting-dilution suspension culture studies. Phenotypic heterogeneity is evaluated with multi-color flow cytometry and detection of alterations in aldehyde dehydrogenase activity. Xenograft studies in immune deficient mice are used to evaluate self-renewal capability, serial engraftment kinetics, and alterations in phenotype. Drug studies are performed by evaluating the differences in phenotype and CFU over time when treating with conventional induction chemotherapy or novel agents.

Results:

We have evaluated several SR and HR ALL samples in addition to some RD samples, which are paired with HR diagnostic samples. In vitro studies revealed the SR samples had little to no colony forming ability (0-1%) while the HR samples had approximately 3-5% and the RD samples 8-10% colony-forming ability. Likewise, the SR samples failed to engraft NOD-SCID mice while the HR samples, from patients with infantile ALL and the MLL translocation or T cell ALL, had robust engraftment in primary and secondary recipients. The engraftment kinetics were uniformly faster in secondary recipients. These findings suggest that HR leukemia may be the result of a leukemia-initiating cell with stem cell-like characteristics while SR ALL may arise from a more committed lymphoid progenitor. Interestingly, in the RD samples, several of the phenotypic markers are similar to that of the primary sample after treatment with induction therapy, particularly with regards to percentages of CD 34, 133-1, 133-2 and aldehyde dehydrogenase levels. Several HR samples have been exposed to induction chemotherapy (Decadron, Cytarabine, Doxorubicin and Vincristine), and the CFU potential and phenotype evaluated over a two-week time course. Notably, the majority of bulk disease is effectively killed, the CFU content actually increases two to three-fold, when an equivalent number of viable cells are analyzed. Furthermore, the phenotype reveals brighter staining with several proposed stem cell markers (CD34, 117, 133-1, 133-2, 123, and measurement of aldehyde dehyrogenase). These data indicate the selection of a chemo-resistant or LSC population.

Conclusions:

Our results to this point suggest important differences both functionally and phenotypically, between SR, HR and RD pediatric leukemia. These findings are consistent with what would be expected given clinical differences in each of these disease states and begins to establish a means of identifying a LSC or chemo-resistant population, which can be targeted with novel treatment regimens. Likewise, these techniques may also provide a means of evaluating for minimal residual disease (MRD) in a LSC or chemo-resistant population by identifying that population's phenotype by passaging the initial sample through serial murine engraftments or in vitro drug studies.

Disclosures:

No relevant conflicts of interest to declare.