Our study had three primary goals, to: 1) determine if germline TL was associated with risk of osteosarcoma, 2) identify associations between SNPs in telomere biology genes and osteosarcoma risk, and 3) determine if those SNPs were associated with TL. We hypothesized that since osteosarcoma somatic cells typically have significant chromosomal abnormalities and often use the alternative lengthening of telomeres pathway for telomere maintenance aberrations in telomere biology could contribute to osteosarcoma risk. Overall, we found that short TL was associated with osteosarcoma risk in females, SNPs in TERF1 were associated with decreased osteosarcoma risk, and that telomere biology gene SNPs were not strongly associated with TL.
TL in surrogate tissues (e.g.
, blood or buccal cells) has been postulated to be a biomarker of cancer risk. Several case-control studies have found statistically significant associations between shorter telomeres and risk of cancers such as bladder [14
], esophageal [17
], gastric [19
], head and neck [16
], lung [16
], ovarian [35
], and renal [16
]. A few studies have also suggested that longer telomeres are associated with risk of melanoma [37
], non-Hodgkin lymphoma [38
], and breast cancer [39
], although the breast cancer TL association studies have been inconsistent [39,41-43
]. Null associations with TL were reported in prospective studies of prostate [21
] and colorectal cancers [22
]. Overall, significant differences in TL between osteosarcoma cases and controls were not identified.
Our study and others suggest that healthy females have longer telomeres than males [44
]. We also found a statistically significant association between shorter TL and risk of osteosarcoma in females. This association was not noted in males or in the combined male-female dataset. This gender difference might reflect the effects of estrogen on telomere dynamics, possibly through the activation of the hTERT
gene promoter [48
], posttranslational regulation of hTERT
], or through its antioxidative capacity [50
]. It is also possible that this finding is a false positive due to small sample size. Alternatively, one could theorize that females with telomeres that are shorter than expected for their gender might be at even higher risk of cancer related to telomere shortening than males, as others have observed for other cancers [16
]. It is also possible that females could have different osteosarcoma risk factors than males. A recent study of the Pro72Arg TP53
polymorphism in osteosarcoma found that the variant allele was associated with osteosarcoma only in females [9
This pilot study was the first to explore the association between SNPs in telomere biology genes and osteosarcoma risk. We were able to augment our statistical power through the addition of controls from the PLCO study. With the addition of these controls, there was 80% power to detect an OR of 1.82 for SNPs with MAFs of at least 0.1. We chose to interpret the SNP data conservatively, by using the Bonferroni correction based on the number of SNPs per gene, because of the study's small sample size, and we used global gene- and pathway-level analyses to comprehensively evaluate our data.
This approach identified seven statistically significant SNPs in TERF1
after Bonferroni correction for the number of SNPs per gene. However, no associations remained significant if corrected for all 713 SNPs in the study. The SNPs in TERF1
were all inversely associated with osteosarcoma risk and were strongly correlated with each other. At the gene-level, TERF1
was also significantly associated with osteosarcoma after correction for multiple tests. TERF1
encodes TRF1, a member of the shelterin telomere protection complex which protects telomeres from degradation and inappropriate DNA repair [51
]. The role of TERF1
in osteosarcoma pathogenesis is not known. One small study did not find TERF1
mutations in osteosarcoma cell lines [52
We also evaluated the association between TL and SNPs in telomere biology genes in the BDISO participants, to better understand the role of common SNPs in TL regulation. A total of 20 SNPs in 13 genes were statistically significantly associated with TL before Bonferroni correction, but none remained significant (P
< 0.05) after this conservative statistical correction (). A recent genome-wide association study (GWAS) identified a SNP in the TERC
locus, rs 12696304, that was inversely associated with TL [53
]. This SNP was also associated with a reduction in TL in our dataset which was significant before correction (Beta -0.105, SE 0.05, P
= 0.034). Two other SNPs in our data-set in this region were also significant before Bonferroni correction. These three TERC
SNPs were all highly correlated in our dataset (r2
= 0.8-0.98). Recent genome-wide association studies have found variants in the TERT-CLPTM1L
locus associated with cancer risk [54
]. We evaluated 16 SNPs in the TERT
locus and did not find associations with osteosarcoma orTL.
Other studies have mapped loci influencing TL to chromosome 14q23.2 [56
], and to variants in the BICD1
], and VPS34/PIKC3C
] genes. Of these genes, only DDX11
was in our data set and its SNPs were not associated with TL. Another candidate gene study of TL and SNPs in 43 telomere biology genes found that SNPs in MEN1
were associated with TL [59
]. A SNP in MEN1
that was in both studies, rs670358, was significantly associated with TL before Bonferroni correction (P
= 0.008) in our study. In the current study this SNP was associated with an increase in TL, but the converse was true in the other study. This discrepancy may be due in part to differences in the age of the study populations (median age of 19 years in this study compared with 62 years in the other).
In summary, this pilot-study explored the potential role of telomere biology in osteosarcoma etiology. The results were very conservatively interpreted using Bonferroni correction which reduces the potential for false positive findings, but may be too stringent. The role of SNPs in TL regulation is an area of active investigation. This study confirms some of those associations, including an association between TL and SNPs in MEN1 and TERC. Common variants in TERF1 were inversely associated with risk of osteosarcoma. Additional studies of the role of TERF1 and other components of shelterin in osteosarcoma are warranted. Lastly, we found that females with shorter teiomeres had higher risks of osteosarcoma than males. The sample size was small and larger studies are required to better understand this gender difference.