PMC

Different subtypes of leukemia are coded by a specific colour. Yellow: ALL samples without methylation, Blue: ALL samples with a least one methylated gene.

DFS curves for all patients enrolled in this study (panel A), childhood ALL (B) and adult ALL (C) according to the methylation profile. OS curves for all patients enrolled in this study (D), childhood ALL (E) and adult ALL (F) according to the methylation profile. EFS curves for all patients enrolled in this study (G), childhood ALL (H) and adult ALL (I) according to the methylation profile.

The top bar beneath the dendrogram refers to the subtypes of ALL and specific controls from healthy donors, while the lower bar indicates ALL or control. Different subtypes of leukemia or controls are color coded (ALL: Acute Lymphoblastic Leukemia; PBMNC: Peripheral Blood mononuclear cells from healthy donors, CD19⁺ cells from PB and CD3⁺ cells from PB). Red: methylated; Green: non-methylated.

A. Hypermethylated genes in more that 10% of ALL samples. B. Hypomethylated genes in more than 10% of ALL samples. The top bar beneath the dendrogram refers to the subtypes of ALL and specific controls from healthy donors, while the lower bar indicates ALL or control. Subtypes of leukemia or controls are color coded (ALL: Acute Lymphoblastic Leukemia; PBMNC: Peripheral Blood mononuclear cells from healthy donors, CD19⁺ cells from PB and CD3⁺ cells from PB). Red: methylated; Green: non-methylated.

A. MSP analysis of the methylated and un-methylated sequences of genes implicated in TP53 pathway in ALL derived cell lines. UC: un-methylated control (DNA of peripheral blood from a healthy donor). MC: universally methylated control, M: 100 bp molecular marker. B. Graphic representation of the results of methylation of genes of TP53 pathway and its involvement in TP53 dependent apoptosis, cell cycle or regulation of TP53. Information regarding CpG composition and whether these genes are target of PCR2 in ESC is shown. Red: methylated, Green: unmethylated.

TOM-1 and NALM-20 cells were treated with 5-aza-2-deoxycitidine, Curcumin and Nutlin-3. A. Apoptosis induced by 5-aza-2-deoxycitidine treatment: Annexin V analysis by flow citometry (grey: early apoptotic; black: late apoptotic/death cells), Caspase-3 analysis by flow citometry and Western blot analysis of the 85-kDa fragment of PARP. B–D. Curcumin treatment; Western blot analysis and quantification of Caspase 8. PRO-C8: full length caspase-8 fragment; p43/p41: cleaved intermediate p43/p41 fragment of caspase-8 and p18: caspase-8 active fragment p18. Numbers at the bottom of the figure represent the quantification of these three fragments of caspase-8 (B); Activation of apoptosis measure by Annexin V (grey: early apoptotic; black: late apoptotic/death cells) and Caspase-3 (FACS) and by the detection of the 85-kDa fragment of PARP (C). Upregulation in the amount of acetylated histone 3 by Western blot is shown. Total histone 3 and β-actin are loading controls (C); qRT-MSP and pyrosequencing of hypermethylated genes implicated in TP53 pathway before and after treatment with Curcumin (D). E. Western blot analysis of p21 and p53, levels of CDKN1A and TP53 mRNA by Q-RT-PCR and detection of apoptosis by Annexin V (grey: early apoptotic; black: late apoptotic/death cells), Caspase-3 and detection of the 85-kDa fragment of PARP by western blot after treatment with Nutlin-3. β-actin was used as a loading control in all cases. A,C and E, the mean ± SD of at least 3 different experiments is shown. A-E, a representative example of at least 3 different experiments is shown.