In this population-based case–control study, we examined the risk of childhood ALL associated with several genes involved in fetal growth and body size regulation, utilizing a haplotype-tagging approach to maximize capture of genetic variation. We identified haplotypes of several genes that were significantly associated with childhood ALL, including IGF1, IGF2, and IGF2R. To our knowledge, no previous studies have expressly examined the role of genes involved in the IGF axis or body size regulation in risk of childhood ALL. Our findings support the hypothesis that factors involved in fetal growth and body size regulation, specifically components of the IGF axis, play a role in mediating risk of childhood ALL.
In this study, we found two haplotypes in the same 6-htSNP window of IGF1
to be significantly associated with an increased risk of childhood ALL. The haplotype window of association here stretches across 25 kb, from the 5′ promoter region into the second intron. IGF1, also known as somatomedin C, is expressed by both the mother and the fetus during pregnancy and plays a critical role in modulating cellular proliferation, differentiation, and apoptosis [6
]. While IGF1 controls growth directly in utero, it becomes subject to regulation by growth hormone in the postnatal period [9
We identified a significant 3-htSNP haplotype of IGF2
associated with a markedly increased risk of childhood ALL. In contrast to IGF1
is thought to play a more critical role in growth and development in utero versus postnatally [6
]. Serological IGF2 levels are higher during the in utero versus the postnatal period, and studies of IGF2
knockout mice show stunted fetal growth but normal postnatal growth [20
]. Although IGF2 is an imprinted gene, meaning that only one allele is usually expressed, there are no consistent findings to support the hypothesis that loss of imprinting of IGF2
is associated with an increased risk of childhood ALL [6
Finally, we found significant associations of overlapping but distinct IGF2R
haplotype windows with risk of childhood ALL among Hispanics and non-Hispanics separately. Interestingly, the two overlapping htSNPs in each of these windows, rs9457799 and rs6455678, had significantly different single-htSNP effects in Hispanics versus non-Hispanics. The differential effect of these two htSNPs was present in the haplotype analysis as well: The risk haplotype among Hispanics included C-G at these two loci and was associated with a borderline significant increased risk, while among non-Hispanics the haplotype including C-G at these two loci showed no association. Conversely, the haplotype with A-A at these two loci was associated with a significantly increased risk among non-Hispanics and null risk among Hispanics. The IGF2R
gene product serves no signal transduction purpose; rather, by binding IGF2, it appears to serve mainly to regulate IGF2 levels in utero [21
The putative functional IGF1
SNP (rs35767) found to be significantly associated with childhood ALL risk is in the IGF1
gene promoter region. It is not functional itself but is in strong linkage disequilibrium with -841(CA)n
, a common microsatellite that has been linked to circulating IGF1 levels as well as birth weight and other measures of body size [10
]. This SNP is part of the 6-htSNP haplotype window we found to be significantly associated with childhood ALL risk.
We also examined body size regulation genes outside the IGF axis. These genes include LEP
, whose gene product leptin signals satiety and has been found to correlate with neonatal birth weight [23
] and size for gestational age [24
], and LEPR, whose gene product mediates the effects of leptin. Ghrelin, encoded by GHRL
and secreted primarily by the stomach, serves to stimulate appetite and promote adiposity and may have a compensatory effect on intra-uterine growth restriction, serving to boost growth in the postnatal period [7
]. Neuropeptide Y, encoded by NPY
, is a neurotransmitter that also leads to increased appetite and storage of energy as fat, as well as decreased physical activity. However, we found no significant haplotype associations between these genes and risk of childhood ALL.
In this study, statistical adjustment for birth weight did not affect the associations observed for the significant risk haplotypes in IGF1
, and IGF2R
, or the single nominally significant (p
< 0.05) putative functional SNP in IGF1
. The independence of the observed associations from birth weight, coupled with the absence of direct associations of the haplotypes and htSNP in question with birth weight (data not shown), suggests that the effects of the risk haplotypes and alleles on childhood ALL risk are not directly mediated by birth weight. Indeed, data from a recent meta-analysis of genome-wide studies did not identify birth weight–associated loci in IGF axis genes [25
]. Because we did not begin collecting data on maternal height or pre-pregnancy weight until partway through recruitment of the study population, these data were unavailable for 49 % of study subjects, and therefore, we were unable to estimate percent optimal birth weight, a measure of appropriateness of fetal growth that takes into account maternal body size, gestational age, and other factors and may be more relevant to ALL risk than birth weight alone [4
Maternal components of the IGF axis are unable to cross the placenta [26
]; accordingly, fetal levels of these factors are likely to be determined by fetal tissues, lending support to the notion that fetal genotypes are particularly relevant. However, maternal genotypes influencing maternal metabolism and growth may have an impact on fetal growth; accordingly, investigation of maternal genotypes is warranted.
One of the key strengths of our study is the inclusion of U.S. Hispanics, an understudied population with the highest reported rates of childhood leukemia in California [27
]. Our htSNP selection strategy included elements designed to maximize capture of genetic variation in Hispanics. We examined Hispanics separately from non-Hispanics where there was significant heterogeneity in between-group effects of individual htSNPs in each gene. Although this approach may have limited our ability to detect associations in the population as a whole, we felt it was necessary given that genetic susceptibility may be different in Hispanics versus non-Hispanics due to the Hispanic population’s relatively recent genetic admixture [15
]. Results that differ between Hispanics and non-Hispanics may be due to differences in allele frequency and/or haplotype structure or may reflect underlying differences in exposures that modulate the effects of genes. Regardless, if the results are not spurious, they represent potential risk loci, and we present them in either or both ethnic groups for replication and further follow-up. The limited size of racial/ethnic sub-populations within the non-Hispanic group precluded further stratification of this group; as such, genetic heterogeneity within this group might have obscured results. However, adjustment for calculated genetic ancestry did not markedly change the race- and ethnicity-adjusted results we present here, indicating that the potential impact of population stratification is minimal.
Two large genome-wide association studies on childhood ALL have been published to date (with 907 cases and 2,398 adult and child controls, and 317 cases and 17,958 adult controls, respectively) [28
]. These studies have identified a number of novel loci; however, no significant associations were observed for genes in IGF axis and body size regulation genes. This may be due to stringent multiple testing adjustment (at the p
≤ 1 × 10−7
level) to account for the large number of individual variants tested in genome-wide studies. In contrast to the agnostic approach to discovery used in such studies, our study focused on relatively few genes representing key elements of the IGF axis and body size regulation pathways. It commenced prior to the recent publication of results linking birth weight to loci in two genes (ADCY5
]; as such, we were unable to investigate these genes. We concede that our study results may be due to chance and therefore must be replicated. However, the haplotype-tagging approach we adopted maximizes capture of total variation within each candidate gene, and the haplotype analysis increases statistical power to detect associations over analyses of individual variants. Finally, although the haplotype-tagging approach does not pinpoint potential causal variants, it does localize risk-associated regions for additional study, including fine-mapping.
Recent studies showing correlations of umbilical cord serum IGF1 levels with both cord stem cell levels and birth weight [30
] support the notion that increased fetal growth may increase cancer risk by either increasing pre-leukemic stem cell populations targeted for initiation, or promoting growth or survival of an initiated leukemic stem cell. Together with this, our results are consistent with the notion that the IGF axis influences the initiation or promotion of childhood ALL early in life. Further studies are needed to determine the specific roles played.
In summary, we set out to investigate the role of genes in the IGF axis and body size regulation pathways in risk of childhood ALL. Our results indicate that elements of the IGF axis are associated with childhood ALL risk. These findings are consistent with the evidence that childhood ALL initiation and/or promotion may begin in utero, when lymphoid progenitor cells are not yet fully differentiated and therefore more susceptible to malignant transformation. The associations and interactions identified here should be considered targets for replication in additional studies with larger sample size and finer coverage of variants in the identified associated gene regions.