In utero cytomegalovirus infection and development of childhood acute lymphoblastic leukemia

Abstract

It is widely suspected, yet controversial, that infection plays an etiologic role in the development of acute lymphoblastic leukemia (ALL), the most common childhood cancer and a disease with a confirmed prenatal origin in most cases. We investigated infections at diagnosis and then assessed the timing of infection at birth in children with ALL and age, gender, and ethnicity matched controls to identify potential causal initiating infections. Comprehensive untargeted virome and bacterial analyses of pretreatment bone marrow specimens (n = 127 ALL in comparison with 38 acute myeloid leukemia cases in a comparison group) revealed prevalent cytomegalovirus (CMV) infection at diagnosis in childhood ALL, demonstrating active viral transcription in leukemia blasts as well as intact virions in serum. Screening of newborn blood samples revealed a significantly higher prevalence of in utero CMV infection in ALL cases (n = 268) than healthy controls (n = 270) (odds ratio [OR], 3.71, confidence interval [CI], 1.56-7.92, P = .0016). Risk was more pronounced in Hispanics (OR=5.90, CI=1.89-25.96) than in non-Hispanic whites (OR=2.10 CI= 0.69-7.13). This is the first study to suggest that congenital CMV infection is a risk factor for childhood ALL and is more prominent in Hispanic children. Further investigation of CMV as an etiologic agent for ALL is warranted.

Figure 1.

Flowchart of approach. cDNA, complementary DNA.

Figure 2.

Nonhuman sequence search identification (“blast hits”) from pretreatment bone marrow. (A) Results of deep-sequencing RNA pools (73 ALLs in 4 pools and 28 AMLs in 2 pools) after ribosomal RNA depletion (RiboZERO). Raw Illumina reads were aggressively quality filtered, then aligned to hg19 using BowTie2; nonaligning reads were categorized using an E value less than or equal to 1 × E⁻¹⁰ with blastn and the nt database. Cytomegalovirus hits were normalized by sequencing effort per pool (blast hits/quality reads × 1.0 × E). Odds ratios were calculated by comparing presence/absence of virus in ALL versus AML pools; P values were computed by using Fisher’s exact test (for additional details, see the supplemental Methods section). (B) Results of particle isolation and 454 sequencing from an independent study of 36 ALL and 10 AML patients. Virus was isolated using centrifugation and Millipore filter, and then viral nucleic acids were extracted and sequenced using Roche 454 (for additional details, see the supplemental Methods section). Recovered sequences were assembled into contigs in which alignments to hg19 were removed using Bowtie2. Contigs were then categorized with a cutoff of E value less than or equal to 1.0 × E⁻¹⁰ using blastn and the refseq viral database. Cytomegalovirus contigs were normalized by recovered contigs per patient (blast hits/contigs assembled × 1 × E); odds ratios were calculated comparing presence/absence of virus in ALL versus AML patients; P values were computed using Fisher’s exact test (for additional details, see the supplemental Methods section). CMV was found to be the only virus that showed statistically significant variation between ALL and AML in the Illumina and 454 groups. CI, confidence interval; NA, not applicable; OR, odds ratio.