PTPN11 and RAS Gene Mutation Pattern Identifies an Unique Feature of Upregulated RAS Function in Infant ALL.
Authors: Cazzaniga Giovanni , Martinelli Simone , den Boer Monique L. , Corral Lilia , Spinelli Monica , Basso Giuseppe , De Rossi Giulio , Pieters Rob , Tartaglia Marco , Biondi Andrea 
Published: October 2019

Abstract
Point mutations affecting RAS GTPase activity promote upregulation of RAS signaling, and are among the most common genetic alterations in human cancers. Oncogenic mutations in NRAS and KRAS2 genes are found with variable prevalence in hematological malignancies, including ALL. Upregulation of RAS signaling can also results from molecular lesions in genes coding transducers controlling RAS function. We recently provided evidence that PTPN11 is mutated in approximately one-third of pediatric ALL cases. Significantly, while PTPN11 mutations appeared to be preferentially associated with the common immunophenotype, mutations in NRAS or KRAS2 were uniformly distributed among the B-cell precursor ALL subtypes. Very few are the data on RAS pathway status in pediatric ALL patients aged <1 year (Infant). Infant ALL is a rare subgroup of leukemia peculiar for several characteristics, mainly related to a possible origin from a not-fully committed stem cell. In order to explore whether alterations of RAS signaling may identify specific subgroups within this heterogeneous patient population, we analyzed 87 Infant cases prospectively enrolled in the Interfant-99 International ALL protocol. This is the largest series of Infant ALL analyzed for alteration of the RAS pathway. PTPN11 (exon 3 and 13), NRAS (exon 1 and 2), and KRAS2 (exon 1 and 2) mutation analysis was performed by DHPLC analysis and direct sequencing. None of the 87 Infant cases carried a PTPN11 somatic mutation. By contrast, a high prevalence of mutations affecting NRAS and KRAS2 was observed (approximately 30%). Thirteen cases (14.9%) were positive for a mutation in exon 1 (codons 12 and 13, 10 cases, 11.5%) or exon 2 (codon 61, 3 cases, 3.4%) of the NRAS gene. Seventeen cases (19.5%) carried a variety of exon 1 KRAS2 mutations: codons 12 and 13, n=13 (14.9% of total); insAGC(30–32), n=1; G(40)C, n=1; C(53)A, n=1; G(57)C, n=1. Interestingly, in 9/27 cases (33.3% of RAS mutated cases, 10.3% of total) DHPLC and sequencing consistently showed the presence of the RAS mutation only in a fraction (10 to 40%) of the blast cells population. In one additional case a KRAS2 mutation was observed in a subset of cells along with a full NRAS mutation. In two more cases mutations affecting NRAS and KRAS2 coexisted on the same patient; in both cases the mutations were representative of a subgroup of the total cell population. In conclusion: differently from older children, PTPN11 gene mutations are a rare event in infant ALL. This could be related to the low incidence of the “common” immunophenotype in Infants. By contrast, the overall incidence of RAS gene mutations is high, accounting for about 30% of cases. Interestingly, in 10% of cases a mutation (or even more than one) was found in a fraction of the blast cell population. This could reflect the fact that RAS mutations were secondary to other genetic events, and in any case related to the oligoclonal characteristic of this age-specific ALL subgroup. The prognostic effect of these genetic events must be evaluated in a longer follow up time of prospective series of patient, as in the Interfant Consortium.