S is in line with the results of the present study wherein upregulated BMPRII gene expression andhypomethylation of BMPRII gene was noticed among unmutated subgroup of CLL patients. Alterations in methylation status and associated gene expression levels of another gene CRY1 have also been reported in prognostically distinct subsets of CLL [50] as well as in CML [51]. Our study confirms the possible influence of hypomethylation and upregulated expression of CRY1 in prognostically poor IGHV unmutated CLL and further emphasises its role as potential biomarker for relative risk of treatment initiation and TTFT in early stage CLL. In addition to CRY1, three other circadian rhythm genes NPAS2, BHLHE40, and ARNTL were also observed to be hypomethylated in the unmutated subgroup [52]. PAX9 is one of the nine members of "paired box" (PAX)-containing transcription factor family and its inhibition has been shown to induce apoptosis with increased PRIMA-1 cleavage of caspase-3 and PARP, increased expression of BAX and decreased expression of BCL-2 in oral squamous cell carcinoma [53]. In the recent years, it has emerged as one of the biomarkers of cell proliferation in lung cancer [54]. A significant association of PAX9 expression with stage, IPI score, relative risk of treatment initiation, TTFT and OS in the present studyRani et al. Clinical Epigenetics (2017) 9:Page 12 ofFig. 6 Kaplan-Meier survival curves representing overall survival in early stage CLL patients with a low (n = 47) and high (n = 46) mRNA expression of CRY1 b low (n = 46) and high (n = 46) mRNA expression of PAX9 and; c unmutated (n = 39) and mutated IGHV genes (n = 54)strengthens its role as an important marker of prognosis in CLL as well. Since levels of expression of either PAX9 or CRY1 did not show significant difference in CLL patients when compared to healthy controls but rather between patients subgrouped on the basis of the IGHV mutational status, it is plausible that these two genes may be involved in progression of CLL rather than development of the disease. This explanation is further supported by progressively increasing gene expression levels of PAX9 PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/18111632 and CRY1 in coherence with advanced Rai stage and higher IPI scores. The mechanism(s) underlying such an influence of these two genes in CLL pathogenesis are not known but might involve apoptotic [53, 55?7], or analogous pathways involved in cancer. Besides, several aberrantly methylated genes were also identified in IGHV mutational status based subgroups which could serve as potential markers in CLL. The major PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/22316373 limitation of the present study was that a limited number of genes were evaluated in a small cohort of early stage CLL patients. Further studies on large cohorts of early stage CLL patients for expression patterns of additional set of genes are required that may help in characterizing the functional role of the genes identified in the present study. Identification of relevant epigenetically influenced genes that have an impact on gene expression as well as clinical outcome may paveway for identification and development of therapeutically relevant drug targets.Conclusions The present study confirms the prognostic role of CRY1 in CLL as its aberrant methylation and expression is associated with high risk of treatment initiation and shorter time to first treatment. In addition, this study highlights PAX9 as a novel marker of prognostication in CLL as its expression was significantly associated with high risk of treatment in.