Three Faces of Mercaptopurine Cytotoxicity In Vitro: Methylation, Nucleotide Homeostasis, and Deoxythioguanosine in DNA

Coulthard, Sally A.; McGarrity, Sarah; Sahota, Kalvin; Berry, Philip; Redfern, Christopher P.F.

doi:10.1124/dmd.118.081844

Cited by 10 publications

(5 citation statements)

References 29 publications

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“…When 6-thioguanine is incorporated into DNA, DNA-protein crosslinks and single-strand breaks result. Thiopurine also causes the level and activity of the enzyme DNA (cytosine-5)-methyltransferase (DNMT1) 1 to decrease, which lowers DNA methylation [40]. In the present investigation, HepG2 treated with 30 µ g/mL 6-MP had greater percentages of cells in the S phase (21.8%) and G2/M (32.6%) than untreated HepG2 did: (17.3%) and (27.5%), respectively.…”

Section: Text Correctionmentioning

confidence: 52%

Preparation of 6-Mercaptopurine Loaded Liposomal Formulation for Enhanced Cytotoxic Response in Cancer Cells

et al. 2022

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6-Mercaptopurine (6-MP) is a well-known immunosuppressive medication with proven anti-proliferative activities. 6-MP possesses incomplete and highly variable oral absorption due to its poor water solubility, which might reduce its anti-cancer properties. To overcome these negative effects, we developed neutral and positively charged drug-loaded liposomal formulations utilizing the thin-film hydration technique. The prepared liposomal formulations were characterized for their size, polydispersity index (PDI), zeta potential, and entrapment efficiency. The average size of the prepared liposomes was between 574.67 ± 37.29 and 660.47 ± 44.32 nm. Positively charged liposomes (F1 and F3) exhibited a lower PDI than the corresponding neutrally charged ones (F2 and F4). Entrapment efficiency was higher in the neutral liposomes when compared to the charged formulation. F1 showed the lowest IC50 against HepG2, HCT116, and MCF-7 cancer cells. HepG2 cells treated with F1 showed the highest level of inhibition of cell proliferation with no evidence of apoptosis. Cell cycle analysis showed an increase in the G1/G0 and S phases, along with a decrease in the G2/M phases in the cell lines treated with drug loaded positively charged liposomes when compared to free positive liposomes, indicating arrest of cells in the S phase due to the stoppage of priming and DNA synthesis outside the mitotic phase. As a result, liposomes could be considered as an effective drug delivery system for treatment of a variety of cancers; they provide a chance that a nanoformulation of 6-MP will boost the cytotoxicity of the drug in a small pharmacological dose which provides a dosage advantage.

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Section: Text Correctionmentioning

confidence: 52%

Preparation of 6-Mercaptopurine Loaded Liposomal Formulation for Enhanced Cytotoxic Response in Cancer Cells

et al. 2022

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“…Generally, both prodrugs manifest cell death through the generation and incorporation of TGNs into DNA, but we validated a novel mechanism of resistance in RS4;11 by demonstrating no difference in DNA-TG levels between NT and NSD2 KD cells. Unlike 6-TG, 6-MP may impact cellular metabolism by inhibition of de novo purine synthesis and GTP homeostasis (45,46). Elucidating the exact mechanism(s) by which NSD2 EK mediates 6-MP resistance is critical, as 6-MP is a backbone of ALL maintenance therapy.…”

Section: Discussionmentioning

confidence: 99%

The NSD2 p.E1099K Mutation Is Enriched at Relapse and Confers Drug Resistance in a Cell Context–Dependent Manner in Pediatric Acute Lymphoblastic Leukemia

Pierro

Saliba

Narang

et al. 2020

Molecular Cancer Research

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The NSD2 p.E1099K (EK) mutation is observed in 10% of acute lymphoblastic leukemia (ALL) samples with enrichment at relapse indicating a role in clonal evolution and drug resistance. To discover mechanisms that mediate clonal expansion, we engineered B-ALL cell lines (Reh, 697) to overexpress wildtype (WT) and EK NSD2, but observed no differences in proliferation, clonal growth, or chemosensitivity. To address whether NSD2 EK acts collaboratively with other pathways, we used shRNAs to knockdown expression of NSD2 in B-ALL cell lines heterozygous for NSD2 EK (RS4;11, RCH-ACV, SEM). Knockdown resulted in decreased proliferation in all lines, decreased clonal growth in RCH-ACV, and increased sensitivity to cytotoxic chemotherapeutic agents, although the pattern of drug sensitivity varied among cell lines implying that the oncogenic properties of NSD2 mutations are likely cell context specific and rely on cooperative pathways. Knockdown of both Type II and REIIBP EK isoforms had a greater impact than knockdown of Type II alone, suggesting that both SET containing EK isoforms contribute to phenotypic changes driving relapse. Furthermore, in vivo models using both cell lines and patient samples revealed dramatically enhanced proliferation of NSD2 EK compared to WT and reduced sensitivity to 6-mercaptopurine in the relapse sample relative to diagnosis. Finally, EK-mediated changes in chromatin state and transcriptional output differed dramatically among cell lines further supporting a cell context specific role of NSD2 EK. These results demonstrate a unique role of NSD2 EK in mediating clonal fitness through pleiotropic mechanisms dependent on the genetic and epigenetic landscape. IMPLICATIONSNSD2 p.E1099K mutation leads to drug resistance and a clonal advantage in childhood B-ALL.

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“…A combination of many different factors has been proposed for their cytotoxicities, such as the inhibition of de novo purine synthesis (DNPS), alterations in DNA methylation state, disruption of guanosine-triphosphate (GTP) signaling, and the incorporation of thioguanine nucleotides (TGN) as bases into DNA. The incorporation of thioguanine nucleotides into DNA causes the induction of apoptosis 4 .…”

Section: Introductionmentioning

confidence: 99%

A bioinformatics approach to the identification of novel deleterious mutations of human TPMT through validated screening and molecular dynamics

Saxena¹,

Murthy²,

Chandrashekhar³

et al. 2022

Sci Rep

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Polymorphisms of Thiopurine S-methyltransferase (TPMT) are known to be associated with leukemia, inflammatory bowel diseases, and more. The objective of the present study was to identify novel deleterious missense SNPs of TPMT through a comprehensive in silico protocol. The initial SNP screening protocol used to identify deleterious SNPs from the pool of all TPMT SNPs in the dbSNP database yielded an accuracy of 83.33% in identifying extremely dangerous variants. Five novel deleterious missense SNPs (W33G, W78R, V89E, W150G, and L182P) of TPMT were identified through the aforementioned screening protocol. These 5 SNPs were then subjected to conservation analysis, interaction analysis, oncogenic and phenotypic analysis, structural analysis, PTM analysis, and molecular dynamics simulations (MDS) analysis to further assess and analyze their deleterious nature. Oncogenic analysis revealed that all five SNPs are oncogenic. MDS analysis revealed that all SNPs are deleterious due to the alterations they cause in the binding energy of the wild-type protein. Plasticity-induced instability caused by most of the mutations as indicated by the MDS results has been hypothesized to be the reason for this alteration. While in vivo or in vitro protocols are more conclusive, they are often more challenging and expensive. Hence, future research endeavors targeted at TPMT polymorphisms and/or their consequences in relevant disease progressions or treatments, through in vitro or in vivo means can give a higher priority to these SNPs rather than considering the massive pool of all SNPs of TPMT.

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Three Faces of Mercaptopurine Cytotoxicity In Vitro: Methylation, Nucleotide Homeostasis, and Deoxythioguanosine in DNA

Cited by 10 publications

References 29 publications

Preparation of 6-Mercaptopurine Loaded Liposomal Formulation for Enhanced Cytotoxic Response in Cancer Cells

Preparation of 6-Mercaptopurine Loaded Liposomal Formulation for Enhanced Cytotoxic Response in Cancer Cells

The NSD2 p.E1099K Mutation Is Enriched at Relapse and Confers Drug Resistance in a Cell Context–Dependent Manner in Pediatric Acute Lymphoblastic Leukemia

A bioinformatics approach to the identification of novel deleterious mutations of human TPMT through validated screening and molecular dynamics

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