ABSTRACT
Epigenetic dysregulation is a hallmark of cancer executed by a number of complex processes the most important of which converge on DNA methylation and histone protein modifications. Epigenetic marks are potentially reversible and thus promising drug targets. In the setting of acute lymphoblastic leukemia (ALL) they have been associated with clinicopathological features including risk of relapse or molecular subgroups of the disease. Here, using immunocytochemistry of bone marrow smears from diagnosis, we studied global histone H4 acetylation, whose loss was previously linked to treatment failure in adults with ALL, in pediatric patients. We demonstrate that preserved global histone H4 acetylation is significantly associated with favorable outcome (RFS, EFS, OS) in children with B cell progenitor (BCP) ALL, recapitulating the findings from adult populations. Further, for the first time we demonstrate differential histone H4 acetylation in molecular subclasses of BCP-ALL including cases with ETV6-RUNX1 fusion gene or PAX5 deletion or deletions in genes linked to B cell development. We conclude global histone H4 acetylation is a prognostic marker and a potential therapeutic target in ALL.
Subject(s)
Core Binding Factor Alpha 2 Subunit/physiology , Epigenesis, Genetic , Histones/metabolism , Oncogene Proteins, Fusion/physiology , PAX5 Transcription Factor/deficiency , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/genetics , Acetylation , Adolescent , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Child , Child, Preschool , Chromosomes, Human, Pair 12/ultrastructure , Chromosomes, Human, Pair 21/ultrastructure , Disease-Free Survival , Female , Humans , Infant , Male , Multiplex Polymerase Chain Reaction , PAX5 Transcription Factor/genetics , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/mortality , Prognosis , Proportional Hazards Models , Protein Processing, Post-Translational , Remission Induction , Translocation, Genetic , Treatment OutcomeABSTRACT
Arginine deprivation, either by nutritional starvation or exposure to ADI-PEG20, induces adaptive transcriptional upregulation of ASS1 and ASL in glioblastoma multiforme ex vivo cultures and cell lines. This adaptive transcriptional upregulation is blocked by neoplasia-specific CpG island methylation in either gene, causing arginine auxotrophy and cell death. In cells with methylated ASS1 or ASL CpG islands, ADI-PEG20 initially induces a protective autophagic response, but abrogation of this by chloroquine accelerates and potentiates cytotoxicity. Concomitant methylation in the CpG islands of both ASS1 and ASL, observed in a subset of cases, confers hypersensitivity to ADI-PEG20. Cancer stem cells positive for CD133 and methylation in the ASL CpG island retain sensitivity to ADI-PEG20. Our results show for the first time that epigenetic changes occur in both of the two key genes of arginine biosynthesis in human cancer and confer sensitivity to therapeutic arginine deprivation. We demonstrate that methylation status of the CpG islands, rather than expression levels per se of the genes, predicts sensitivity to arginine deprivation. Our results suggest a novel therapeutic strategy for this invariably fatal central nervous system neoplasm for which we have identified robust biomarkers and which overcomes the limitations to conventional chemotherapy imposed by the blood/brain barrier.
Subject(s)
Apoptosis , Argininosuccinate Lyase/metabolism , Argininosuccinate Synthase/metabolism , Autophagy , Epigenomics , Arginine/metabolism , Argininosuccinate Lyase/genetics , Argininosuccinate Synthase/antagonists & inhibitors , Argininosuccinate Synthase/genetics , Azacitidine/analogs & derivatives , Azacitidine/pharmacology , Central Nervous System Neoplasms/metabolism , Central Nervous System Neoplasms/pathology , Chloroquine/toxicity , CpG Islands , DNA Methylation/drug effects , Decitabine , Glioblastoma/metabolism , Glioblastoma/pathology , Humans , Hydrolases/pharmacology , Polyethylene Glycols/pharmacology , RNA Interference , RNA, Small Interfering/metabolism , Stilbenes/pharmacology , Tumor Cells, Cultured , Up-Regulation/drug effectsABSTRACT
BACKGROUND: Prolyl hydroxylation is a post-translational modification that affects the structure, stability and function of proteins including collagen by catalysing hydroxylation of proline to hydroxyproline through action of collagen prolyl hydroxylases3 (C-P3H) and 4 (C-P4H). Three C-P3Hs (nomenclature was amended according to approval by the HGNC symbols and names at http://www.genenames.org/ and Entrez database at http://www.ncbi.nlm.nih.gov/gene) leucineproline-enriched proteoglycan (leprecan) 1 (Lepre1), leprecan-like 1 (Leprel1), leprecan-like 2 (Leprel2) and two paralogs Cartilage-Related Protein (CRTAP) and leprecan-like 4 (Leprel4) are found in humans. The C-P4Hs are tetrameric proteins comprising a variable α subunit, encoded by the P4HA1, P4HA2 and P4HA3 genes and a constant ß subunit encoded by P4HB. METHODS: We used RT-PCR, qPCR, pyrosequencing, methylation-specific PCR, western blotting and immunohistochemistry to investigate expression and regulation of the C-P3H and C-P4H genes in B lymphomas and normal bone marrow. RESULTS: C-P3H and C-P4H are downregulated in lymphoma. Down-regulation is associated with methylation in the CpG islands and is detected in almost all common types of B-cell lymphoma, but the CpG islands are unmethylated or methylated at lower levels in DNA isolated from normal bone marrow and lymphoblastoid cell lines. Methylation of multiple C-P3H and C-P4H genes is present in some lymphomas, particularly Burkitt's lymphoma. CONCLUSIONS: Methylation of C-P3H and C-P4H is common in B lymphomas and may have utility in differentiating disease subtypes.
