A thiol-bound drug reservoir enhances APR-246-induced mutant p53 tumor cell death
Ontology highlight
ABSTRACT:
SUBMITTER: Ms. Sophia Ceder
PROVIDER: S-SCDT-EMM-2019-10852 | biostudies-other |
REPOSITORIES: biostudies-other
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Project description:The tumor suppressor gene TP53 is the most frequently mutated gene in cancer. The compound APR-246 (PRIMA-1Met/Eprenetapopt) is converted to methylene quinuclidinone (MQ) that targets mutant p53 protein and perturbs cellular antioxidant balance. APR-246 is currently tested in a phase III clinical trial in myelodysplastic syndrome (MDS). By in vitro, ex vivo, and in vivo models, we show that combined treatment with APR-246 and inhibitors of efflux pump MRP1/ABCC1 results in synergistic tumor cell death, which is more pronounced in TP53 mutant cells. This is associated with altered cellular thiol status and increased intracellular glutathione-conjugated MQ (GS-MQ). Due to the reversibility of MQ conjugation, GS-MQ forms an intracellular drug reservoir that increases availability of MQ for targeting mutant p53. Our study shows that redox homeostasis is a critical determinant of the response to mutant p53-targeted cancer therapy.
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Project description:The TP53 tumor suppressor gene is frequently inactivated in human tumors by missense mutations in the DNA binding domain. TP53 mutations lead to protein unfolding, decreased thermostability and loss of DNA binding and transcription factor function. Pharmacological targeting of mutant p53 to restore its tumor suppressor function is a promising strategy for cancer therapy. The mutant p53 reactivating compound APR-246 (PRIMA-1Met) has been successfully tested in a phase I/IIa clinical trial. APR-246 is converted to the reactive electrophile methylene quinuclidinone (MQ), which binds covalently to p53 core domain. We identified cysteine 277 as a prime binding target for MQ in p53. Cys277 is also essential for MQ-mediated thermostabilization of wild-type, R175H and R273H mutant p53, while both Cys124 and Cys277 are required for APR-246-mediated functional restoration of R175H mutant p53 in living tumor cells. These findings may open opportunities for rational design of novel mutant p53-targeting compounds.
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Project description:TP53 is the most frequently mutated gene in human cancer and thus an attractive target for novel cancer therapy. Several compounds that can reactive mutant p53 protein have been identified. APR-246 is currently being tested in a phase II clinical trial in high-grade serous ovarian cancer. We have used RNA-seq analysis to study the effects of APR-246 on gene expression in human breast cancer cell lines. Although the effect of APR-246 on gene expression was largely cell line dependent, six genes were upregulated across all three cell lines studied, i.e., TRIM16, SLC7A11, TXNRD1, SRXN1, LOC344887, and SLC7A11-AS1. We did not detect upregulation of canonical p53 target genes such as CDKN1A (p21), 14-3-3?, BBC3 (PUMA), and PMAIP1 (NOXA) by RNA-seq, but these genes were induced according to analysis by qPCR. Gene ontology analysis showed that APR-246 induced changes in pathways such as response to oxidative stress, gene expression, cell proliferation, response to nitrosative stress, and the glutathione biosynthesis process. Our results are consistent with the dual action of APR-246, i.e., reactivation of mutant p53 and modulation of redox activity. SLC7A11, TRIM16, TXNRD1, and SRXN1 are potential new pharmacodynamic biomarkers for assessing the response to APR-246 in both preclinical and clinical studies.
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Project description:TP53 mutations frequently occur in head and neck squamous cell carcinoma (HNSCC) patients without human papillomavirus infection. The recurrence rate for these patients is distinctly high. It has been actively explored to identify agents that target TP53 mutations and restore wild-type (WT) TP53 activities in HNSCC. PRIMA-1 (p53-reactivation and induction of massive apoptosis-1) and its methylated analogue PRIMA-1Met (also called APR-246) were found to be able to reestablish the DNA-binding activity of p53 mutants and reinstate the functions of WT p53. Herein we report that piperlongumine (PL), an alkaloid isolated from Piper longum L., synergizes with APR-246 to selectively induce apoptosis and autophagic cell death in HNSCC cells, whereas primary and immortalized mouse embryonic fibroblasts and spontaneously immortalized non-tumorigenic human skin keratinocytes (HaCat) are spared from the damage by the co-treatment. Interestingly, PL-sensitized HNSCC cells to APR-246 are TP53 mutation-independent. Instead, we demonstrated that glutathione S-transferase pi 1 (GSTP1), a GST family member that catalyzes the conjugation of GSH with electrophilic compounds to fulfill its detoxification function, is highly expressed in HNSCC tissues. Administration of PL and APR-246 significantly suppresses GSTP1 activity, resulting in the accumulation of ROS, depletion of GSH, elevation of GSSG, and DNA damage. Ectopic expression of GSTP1 or pre-treatment with antioxidant N-acetyl-L-cysteine (NAC) abrogates the ROS elevation and decreases DNA damage, apoptosis, and autophagic cell death prompted by PL/APR-246. In addition, administration of PL and APR-246 impedes UMSCC10A xenograft tumor growth in SCID mice. Taken together, our data suggest that HNSCC cells are selectively sensitive to the combination of PL and APR-246 due to a remarkably synergistic effect of the co-treatment in the induction of ROS by suppression of GSTP1.
| S-EPMC6014869 | biostudies-literature
Project description:The tumor suppressor p53 is commonly inactivated in human tumors, allowing evasion of p53-dependent apoptosis and tumor progression. The small molecule APR-246 (PRIMA-1Met) can reactive mutant p53 in tumor cells and trigger cell death by apoptosis. The thioredoxin (Trx) and glutaredoxin (Grx) systems are important as antioxidants for maintaining cellular redox balance and providing electrons for thiol-dependent reactions like those catalyzed by ribonucleotide reductase and peroxiredoxins (Prxs). We show here that the Michael acceptor methylene quinuclidinone (MQ), the active form of APR-246, is a potent direct inhibitor of Trx1 and Grx1 by reacting with sulfhydryl groups in the enzymes. The inhibition of Trx1 and Grx1 by APR-246/MQ is reversible and the inhibitory efficiency is dependent on the presence of glutathione. APR-246/MQ also inhibits Trxs in mutant p53-expressing Saos-2 His-273 cells, showing modification of Trx1 and mitochondrial Trx2. Inhibition of the Trx and Grx systems leads to insufficient reducing power to deoxyribonucleotide production for DNA replication and repair and peroxiredoxin for removal of ROS. We also demonstrate that APR-246 and MQ inhibit ribonucleotide reductase (RNR) in vitro and in living cells. Our results suggest that APR-246 induces tumor cell death through both reactivations of mutant p53 and inhibition of cellular thiol-dependent redox systems, providing a novel strategy for cancer therapy.
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Project description:Asparaginase depletes extracellular asparagine in the blood and is an important treatment for acute lymphoblastic leukemia (ALL) due to asparagine auxotrophy of ALL blasts. Unfortunately, resistance occurs and has been linked to expression of the enzyme asparagine synthetase (ASNS), which generates asparagine from intracellular sources. Although TP53 is the most frequently mutated gene in cancer overall, TP53 mutations are rare in ALL. However, TP53 mutation is associated with poor therapy response and occurs at higher frequency in relapsed ALL. The mutant p53-reactivating compound APR-246 (Eprenetapopt/PRIMA-1Met) is currently being tested in phase II and III clinical trials in several hematological malignancies with mutant TP53. Here we present CEllular Thermal Shift Assay (CETSA) data indicating that ASNS is a direct or indirect target of APR-246 via the active product methylene quinuclidinone (MQ). Furthermore, combination treatment with asparaginase and APR-246 resulted in synergistic growth suppression in ALL cell lines. Our results thus suggest a potential novel treatment strategy for ALL.
| S-EPMC8282662 | biostudies-literature
Project description:TP53 is the most frequently mutated gene in cancer. The p53 protein activates transcription of genes that promote cell cycle arrest or apoptosis, or regulate cell metabolism, and other processes. Missense mutations in TP53 abolish specific DNA binding of p53 and allow evasion of apoptosis and accelerated tumor progression. Mutant p53 often accumulates at high levels in tumor cells. Pharmacological reactivation of mutant p53 has emerged as a promising strategy for improved cancer therapy. Small molecules that restore wild type activity of mutant p53 have been identified using various approaches. One of these molecules, APR-246, is a prodrug that is converted to the Michael acceptor methylene quinuclidinone (MQ) that binds covalently to cysteines in p53, leading to refolding and restoration of wild type p53 function. MQ also targets the cellular redox balance by inhibiting thioredoxin reductase (TrxR1) and depleting glutathione. This dual mechanism of action may account for the striking synergy between APR-246 and platinum compounds. APR-246 is the only mutant p53-targeting compound in clinical development. A phase I/IIa clinical trial in hematological malignancies and prostate cancer showed good safety profile and clinical effects in some patients. APR-246 is currently tested in a phase Ib/II trial in patients with high-grade serous ovarian cancer.
| S-EPMC4737887 | biostudies-literature
Project description:p53 mutation is common and highly related to radiotherapy resistance in rectal cancer. APR-246, as a small molecule, can restore the tumor-suppressor function to mutant p53. There is no study on combining APR-246 with radiation in rectal cancer; therefore, we examined whether APR-246 sensitized colorectal cancer cells with different p53 status to radiation. The combination treatment had synergistic effects on HCT116p53-R248W/-(p53Mut) cells, followed by HCT116p53+/+(p53WT) cells, and exhibited an additive effect on HCT116p53-/-(p53Null) cells through inhibiting proliferation, enhancing reactive oxygen species, and apoptosis. The results were confirmed in zebrafish xenografts. Comparison of the transcriptome in colorectal cancer cells with different p53 status 72hrs post-treatment with APR-246, irradiation, or the combination of APR-246 with irradiation was carried out. p53Mut and p53WT cells shared more activated pathways and differentially expressed genes following the combination treatment, compared to p53Null cells, although the combination treatment regulated individual pathways in the different cell lines. APR-246 mediated radio-sensitization effects through p53-dependent and -independent ways.
2023-05-12 | GSE232305 | GEO
Project description:Though p53 mutations are rare in ES, there is a strong indication that p53 mutant tumours form a particularly bad prognostic group. As such, novel treatment strategies are warranted that would specifically target and eradicate tumour cells containing mutant p53 in this subset of ES patients.PRIMA-1(Met), also known as APR-246, is a small organic molecule that has been shown to restore tumour-suppressor function primarily to mutant p53 and also to induce cell death in various cancer types. In this study, we interrogated the ability of APR-246 to induce apoptosis and inhibit tumour growth in ES cells with different p53 mutations.APR-246 variably induced apoptosis, associated with Noxa, Puma or p21(WAF1) upregulation, in both mutant and wild-type p53 harbouring cells. The apoptosis-inducing capability of APR-246 was markedly reduced in ES cell lines transfected with p53 siRNA. Three ES cell lines established from the same patient at different stages of the disease and two cell lines of different patients with identical p53 mutations all exhibited different sensitivities to APR-246, indicating cellular context dependency. Comparative transcriptome analysis on the three cell lines established from the same patient identified differential expression levels of several TP53 and apoptosis-associated genes such as APOL6, PENK, PCDH7 and MST4 in the APR-246-sensitive cell line relative to the less APR-246-sensitive cell lines.This is the first study reporting the biological response of Ewing sarcoma cells to APR-246 exposure and shows gross variability in responses. Our study also proposes candidate genes whose expression might be associated with ES cells' sensitivity to APR-246. With APR-246 currently in early-phase clinical trials, our findings call for caution in considering it as a potential adjuvant to conventional ES-specific chemotherapeutics.
| S-EPMC3833220 | biostudies-literature
Project description:The prevalence and dire implications of mutations in the tumour suppressor, p53, highlight its appeal as a chemotherapeutic target. We recently showed that impairing cellular antioxidant systems via inhibition of SLC7A11, a component of the system xc- cystine-glutamate antiporter, enhances sensitivity to mutant-p53 targeted therapy, APR-246. We investigated whether this synergy extends to other genes, such as those encoding enzymes of the pentose phosphate pathway (PPP). TKT, one of the major enzymes of the PPP, is allegedly regulated by NRF2, which is in turn impaired by accumulated mutant-p53 protein. Therefore, we investigated the relationship between mutant-p53, TKT and sensitivity to APR-246. We found that mutant-p53 does not alter expression of TKT, nor is TKT modulated directly by NRF2, suggesting a more complex mechanism at play. Furthermore, we found that in p53null cells, knockdown of TKT increased sensitivity to APR-246, whilst TKT overexpression conferred resistance to the drug. However, neither permutation elicited any effect on cells overexpressing mutant-p53 protein, despite mediating oxidative stress levels in a similar fashion to that in p53-null cells. In sum, this study has unveiled TKT expression as a determinant for sensitivity to APR-246 in p53-null cells.
| S-EPMC7904805 | biostudies-literature