Methylation‐associated DOK1 and DOK2 down‐regulation: Potential biomarkers for predicting adverse prognosis in acute myeloid leukemia

Abstract

DOK-1 and DOK-2 (DOK1/2) are closely related members of downstream of tyrosine kinase (DOK) family genes, which are found to be frequently rearranged in several hematopoietic cancers. However, the clinical implications of DOK1/2 in acute myeloid leukemia (AML) remain largely unknown. To investigate the clinical significance, real-time quantitative PCR (RQ-PCR) was carried out to detect DOK1/2 expressions in 125 de novo AML patients and 28 healthy controls. Real-time quantitative methylation-specific PCR (RQ-MSP) and bisulfite sequencing PCR (BSP) were applied to detect DOK1/2 methylation level and density. DOK1/2 expressions were significantly down-regulated in AML patients. The promoters of DOK1/2 were highly hypermethylated and negatively correlated with DOK1/2 expressions in AML patients. In addition, we also confirmed that DOK1/2 expressions could be restored by DOK1/2 demethylation using 5-aza-2'-deoxycytidine in leukemia cell line THP-1. Survival analyses showed that low-expressed DOK1/2 were associated with markedly shorter overall survival and leukemia free survival in both whole-cohort AML and non-M3 AML patients. Multivariate analyses further revealed that DOK1/2 were act as independent prognostic factors in AML patients. These findings indicate that decreased DOK1/2 expressions associated with their promoter hypermethylations predict adverse prognosis in AML.

Comprehensive analysis of DOK family genes expression, immune characteristics, and drug sensitivity in human tumors

“…DOK family genes disorders have been reported in many cancers [9] , [18] , [19] , [20] . Although DOK family genes expression is closely associated related with pathological progression and poorer prognosis in some tumors, there has been no comprehensive analysis of DOK family genes in different cancer.…”

“…Recent studies have shown that abnormal DOK gene expression is closely related to leukemia [9] , lung cancer [10] , gastric cancer [11] , colorectal cancer [11] , [12] , and breast cancer [13] , [14] . However, most of the studies are conducted in cell lines and/or animal models, and systematic studies have not been conducted in human tumors.…”

Reduced protocadherin17 expression in leukemia stem cells: the clinical and biological effect in acute myeloid leukemia

“…Total RNA was extracted using TRIzol reagent (Invitrogen, Carlsbad, CA, USA) and reverse transcribed into cDNA as described previously [26]. Real-time quantitative PCR (RT-qPCR) was performed using AceQ qPCR SYBR Green Master Mix (Vazyme Biotech Co., Piscataway, NJ, USA) on a 7500 Thermo cycler (Applied Biosystems, CA).…”

“…Real-time quantitative PCR (RT-qPCR) was performed using AceQ qPCR SYBR Green Master Mix (Vazyme Biotech Co., Piscataway, NJ, USA) on a 7500 Thermo cycler (Applied Biosystems, CA). Reaction conditions and PCR cycling were conducted as previously described [26], adjusting only for optimized primer annealing temperatures (60 °C). PCR primers were designed using Primer Premier 6 (Premier Biosoft, Palo Alto, CA, USA), and the primer sequences were listed in Additional file 1: Table S2.…”

“…To assess the changes in methylation level, real-time methylation-specific polymerase chain reaction (RQ-MSP) was employed using primers listed in Additional file 1: Table S2. PCR conditions were as described previously [26], except that the annealing temperature for methylation primers was 59 °C.…”