Investigating the Biomarker Role of Long Non-Coding RNAs in Pediatric Precursor B-Cell Acute Lymphoblastic Leukemia

Introduction:

The diagnosis of childhood acute lymphoblastic leukemia (ALL) typically relies on cell detection from bone marrow (BM) and cerebrospinal fluid (CSF). Existing BM-based methods, while sensitive, are invasive, time-consuming, and laborious.

Central nervous system (CNS) involvement in ALL negatively impacts prognosis. However, routine CNS diagnostic methods yield a high percentage of false negative results. While blasts primarily adhere to the meninges in CNS involvement, molecules secreted by blasts might escape and diffuse more easily into the CSF. Long non-coding RNAs (lncRNAs) are non-coding polynucleotides with specific expression patterns for different cell types, maturation stages, and diseases.

Methods:

De novo precursor-B cell (pre-B) ALL patients under 18 years of age, diagnosed between 2016 and 2023 and treated as per the ALL IC-BFM 2009 protocol were enrolled. Samples from bone marrow aspirate (BM), peripheral blood (PB), and cerebrospinal fluid (CSF) were collected, centrifuged, and separate cell-containing and cell-free aliquots were stored.

An 89-target screening QPCR plate was built after a comprehensive literature search to find lncRNAs significant in ALL and B-cell development. Multiple RNA isolation methods were tested to suit the different body fluids. The lncRNA expression was measured in two experimental setups: 1) The QPCR screening was executed on 11 patients' day 0 and day 33 cell-free BM samples. 2) Five lncRNAs significantly overexpressed on day 0 were selected for droplet digital PCR (ddPCR) measurements on cell-free BM, PB, and CSF samples.

Results:

In the screening, five lncRNA targets had significantly higher expression in day 0 cell-free BM samples compared to day 33 samples, namely LINC01013 (p=0.007), AC002464.1 (p=0.013), AC002454.1 (p=0.035), BX293535.1 (p=0.006), LINC00958 (p=0.036). The 5-lncRNAs were well detected by ddPCR in cell-free BM and PB samples, but no significant signals were gained from the 300-700 μl volume CSF samples.

Conclusions:

The transcriptomic exploration in cell-free BM identified five lncRNAs as potential pre-B ALL biomarkers for further research. Low CSF lncRNA concentrations question their suitability as biomarkers in this compartment.

Erdelyi:Servier: Honoraria; Amgen: Honoraria; Sanofi: Research Funding; Pfizer: Research Funding; Novartis: Research Funding.