Multidrug transporter proteins are best known for their contributions to chemoresistance through the efflux of anticancer drugs from cancer cells. However, a considerable body of evidence also points ...to their importance in cancer extending beyond drug transport to fundamental roles in tumour biology. Currently, much of the evidence for these additional roles is correlative and definitive studies are needed to confirm causality. We propose that delineating the precise roles of these transporters in tumorigenesis and treatment response will be important for the development of more effective targeted therapies.
Abstract The extrusion of anticancer drugs by members of the ATP-binding cassette (ABC) transporter family is one of the most widely recognized mechanisms of multidrug resistance, and can be ...considered a hijacking of their normal roles in the transport of xenobiotics, metabolites and signaling molecules across cell membranes. While roles in cancer multidrug resistance have been clearly demonstrated for P-glycoprotein (P-gp), Breast Cancer Resistance Protein (BCRP) and Multidrug Resistance Protein 1 (MRP1), direct evidence for a role in multidrug resistance in vivo is lacking for other family members. A less well understood but emerging theme is the drug efflux-independent contributions of ABC transporters to cancer biology, supported by a growing body of evidence that their loss or inhibition impacts on the malignant potential of cancer cells in vitro and in vivo . As with multidrug resistance, these contributions likely represent a hijacking of normal ABC transporter functions in the efflux of endogenous metabolites and signaling molecules, however they may expand the clinical relevance of ABC transporters beyond P-gp, BCRP and MRP1. This review summarizes established and emerging roles for ABC transporters in cancer, with a focus on neuroblastoma and ovarian cancer, and considers approaches to validate and better understand these roles.
Myc transcriptional activity is frequently deregulated in human cancers, but a Myc-driven gene signature with prognostic ability across multiple tumor types remains lacking. Here, we selected 18 ...Myc-regulated genes from published studies of Myc family targets in epithelial ovarian cancer (EOC) and neuroblastoma. A Myc family activity score derived from the 18 genes was correlated to
/
/
expression in a panel of 35 cancer cell lines. The prognostic ability of this signature was evaluated in neuroblastoma, medulloblastoma, diffuse large B-cell lymphoma (DLBCL), and EOC microarray gene expression datasets using Kaplan-Meier and multivariate Cox regression analyses and was further validated in 42 primary neuroblastomas using qPCR. Cell lines with high
, and/or
gene expression exhibited elevated expression of the signature genes. Survival analysis showed that the signature was associated with poor outcome independently of well-defined prognostic factors in neuroblastoma, breast cancer, DLBCL, and medulloblastoma. In EOC, the 18-gene Myc activity signature was capable of identifying a group of patients with poor prognosis in a "high-
" molecular subtype but not in the overall cohort. The predictive ability of this signature was reproduced using qPCR analysis of an independent cohort of neuroblastomas, including a subset of tumors without
amplification. These data reveal an 18-gene Myc activity signature that is highly predictive of poor prognosis in diverse Myc-associated malignancies and suggest its potential clinical application in the identification of Myc-driven tumors that might respond to Myc-targeted therapies.
.
The majority of patients with neuroblastoma due to MYCN oncogene amplification and consequent N-Myc oncoprotein over-expression die of the disease. Here our analyses of RNA sequencing data identify ...the long noncoding RNA lncNB1 as one of the transcripts most over-expressed in MYCN-amplified, compared with MYCN-non-amplified, human neuroblastoma cells and also the most over-expressed in neuroblastoma compared with all other cancers. lncNB1 binds to the ribosomal protein RPL35 to enhance E2F1 protein synthesis, leading to DEPDC1B gene transcription. The GTPase-activating protein DEPDC1B induces ERK protein phosphorylation and N-Myc protein stabilization. Importantly, lncNB1 knockdown abolishes neuroblastoma cell clonogenic capacity in vitro and leads to neuroblastoma tumor regression in mice, while high levels of lncNB1 and RPL35 in human neuroblastoma tissues predict poor patient prognosis. This study therefore identifies lncNB1 and its binding protein RPL35 as key factors for promoting E2F1 protein synthesis, N-Myc protein stability and N-Myc-driven oncogenesis, and as therapeutic targets.
Around 10% of acute leukemias harbor a rearrangement of the MLL/KMT2A gene, and the presence of this translocation results in a highly aggressive, therapy‐resistant leukemia subtype with survival ...rates below 50%. There is a high unmet need to identify safer and more potent therapies for MLL‐rearranged (MLL‐r) leukemia that can be combined with established chemotherapeutics to decrease treatment‐related toxicities. The curaxin, CBL0137, has demonstrated nongenotoxic anticancer and chemopotentiating effects in a number of preclinical cancer models and is currently in adult Phase I clinical trials for solid tumors and hematological malignancies. The aim of our study was to investigate whether CBL0137 has potential as a therapeutic and chemopotentiating compound in MLL‐r leukemia through a comprehensive analysis of its efficacy in preclinical models of the disease. CBL0137 decreased the viability of a panel of MLL‐r leukemia cell lines (n = 12) and xenograft cells derived from patients with MLL‐r acute lymphoblastic leukemia (ALL, n = 3) in vitro with submicromolar IC50s. The small molecule drug was well‐tolerated in vivo and significantly reduced leukemia burden in a subcutaneous MV4;11 MLL‐r acute myeloid leukemia model and in patient‐derived xenograft models of MLL‐r ALL (n = 5). The in vivo efficacy of standard of care drugs used in remission induction for pediatric ALL was also potentiated by CBL0137. CBL0137 exerted its anticancer effect by trapping Facilitator of Chromatin Transcription (FACT) into chromatin, activating the p53 pathway and inducing an Interferon response. Our findings support further preclinical evaluation of CBL0137 as a new approach for the treatment of MLL‐r leukemia.
What's new?
Rearrangement of the MLL/KMT2A gene results in a highly aggressive, therapy‐resistant leukemia subtype, with high incidence in infants and survival rates below 50%. In this preclinical study, the authors found that a new, small‐molecule curaxin drug called CBL0137 significantly reduced leukemia burden in xenograft models, and also enhanced the response to standard chemotherapy drugs. CBL0137 mediated this effect through activation of the p53 and IFN pathways. These findings support further preclinical evaluation of CBL0137 as a new approach for the treatment of MLL‐r leukemia.
Background
Although the prognostic value of the ATP-binding cassette, subfamily C (ABCC) transporters in childhood neuroblastoma is usually attributed to their role in cytotoxic drug efflux, certain ...observations have suggested that these multidrug transporters might contribute to the malignant phenotype independent of cytotoxic drug efflux.
Methods
A v-myc myelocytomatosis viral related oncogene, neuroblastoma derived (MYCN)-driven transgenic mouse neuroblastoma model was crossed with an Abcc1-deficient mouse strain (658 hMYCN
1/−, 205 hMYCN
+/1 mice) or, alternatively, treated with the ABCC1 inhibitor, Reversan (n = 20). ABCC genes were suppressed using short interfering RNA or overexpressed by stable transfection in neuroblastoma cell lines BE(2)-C, SH-EP, and SH-SY5Y, which were then assessed for wound closure ability, clonogenic capacity, morphological differentiation, and cell growth. Real-time quantitative polymerase chain reaction was used to examine the clinical significance of ABCC family gene expression in a large prospectively accrued cohort of patients (n = 209) with primary neuroblastomas. Kaplan-Meier survival analysis and Cox regression were used to test for associations with event-free and overall survival. Except where noted, all statistical tests were two-sided.
Results
Inhibition of ABCC1 statistically significantly inhibited neuroblastoma development in hMYCN transgenic mice (mean age for palpable tumor: treated mice, 47.2 days; control mice, 41.9 days; hazard ratio HR = 9.3, 95% confidence interval CI = 2.65 to 32; P < .001). Suppression of ABCC1 in vitro inhibited wound closure (P < .001) and clonogenicity (P = .006); suppression of ABCC4 enhanced morphological differentiation (P < .001) and inhibited cell growth (P < .001). Analysis of 209 neuroblastoma patient tumors revealed that, in contrast with ABCC1 and ABCC4, low rather than high ABCC3 expression was associated with reduced event-free survival (HR of recurrence or death = 2.4, 95% CI = 1.4 to 4.2; P = .001), with 23 of 53 patients with low ABCC3 expression experiencing recurrence or death compared with 31 of 155 patients with high ABCC3. Moreover, overexpression of ABCC3 in vitro inhibited neuroblastoma cell migration (P < .001) and clonogenicity (P = .03). The combined expression of ABCC1, ABCC3, and ABCC4 was associated with patients having an adverse event, such that of the 12 patients with the "poor prognosis" expression pattern, 10 experienced recurrence or death (HR of recurrence or death = 12.3, 95% CI = 6 to 27; P < .001).
Conclusion
ABCC transporters can affect neuroblastoma biology independently of their role in chemotherapeutic drug efflux, enhancing their potential as targets for therapeutic intervention.
Approximately 25% of human neuroblastoma is caused by amplification of the MYCN oncogene, which leads to overexpression of N‐Myc oncoprotein. The survival rate for this patient subtype is <50%. Here, ...we show that N‐Myc protein bound to the DEAD‐box RNA helicase DDX21 gene promoter and upregulated DDX21 mRNA and protein expression. Genome‐wide differential gene expression studies identified centrosomal protein CEP55 as one of the genes most dramatically downregulated after DDX21 knockdown in MYCN‐amplified neuroblastoma cells. Knocking down DDX21 or CEP55 reduced neuroblastoma cell cytoskeleton stability and cell proliferation and all but abolished clonogenic capacity. Importantly, DDX21 knockdown initially induced tumor regression in neuroblastoma‐bearing mice and suppressed tumor progression. In human neuroblastoma tissues, a high level of DDX21 expression correlated with a high level of N‐Myc expression and with CEP55 expression, and independently predicted poor patient prognosis. Taken together, our data show that DDX21 induces CEP55 expression, MYCN‐amplified neuroblastoma cell proliferation, and tumorigenesis, and that DDX21 and CEP55 are valid therapeutic targets for the treatment of MYCN‐amplified neuroblastoma.
The MYCN oncogene encodes an ‘undruggable’ protein N‐Myc. Here, we show that N‐Myc protein binds to the DDX21 gene promoter, leading to increased CEP55 expression, neuroblastoma cell cytoskeletal stability, and cell proliferation. Knocking down DDX21 suppressed tumor progression in neuroblastoma‐bearing mice. Our data demonstrate that DDX21 and CEP55 are valid therapeutic targets for the treatment of MYCN‐amplified neuroblastoma.
The prognosis for high-risk childhood acute leukaemias remains dismal and established treatment protocols often cause long-term side effects in survivors. This study aims to identify more effective ...and safer therapeutics for these patients.
A high-throughput phenotypic screen of a library of 3707 approved drugs and pharmacologically active compounds was performed to identify compounds with selective cytotoxicity against leukaemia cells followed by further preclinical evaluation in patient-derived xenograft models.
Auranofin, an FDA-approved agent for the treatment of rheumatoid arthritis, was identified as exerting selective anti-cancer activity against leukaemia cells, including patient-derived xenograft cells from children with high-risk ALL, versus solid tumour and non-cancerous cells. It induced apoptosis in leukaemia cells by increasing reactive oxygen species (ROS) and potentiated the activity of the chemotherapeutic cytarabine against highly aggressive models of infant MLL-rearranged ALL by enhancing DNA damage accumulation. The enhanced sensitivity of leukaemia cells towards auranofin was associated with lower basal levels of the antioxidant glutathione and higher baseline ROS levels compared to solid tumour cells.
Our study highlights auranofin as a well-tolerated drug candidate for high-risk paediatric leukaemias that warrants further preclinical investigation for application in high-risk paediatric and adult acute leukaemias.
Epithelial ovarian cancer (EOC) is the most lethal gynaecological malignancy with over 80% of cases already disseminated at diagnosis and facing a dismal five-year survival rate of 35%. EOC cells ...often spread to the greater omentum where they take-up cholesterol. Excessive amounts of cholesterol can be cytocidal, suggesting that cholesterol efflux through transporters may be important to maintain homeostasis, and this may explain the observation that high expression of the ATP-binding cassette A1 (ABCA1) cholesterol transporter has been associated with poor outcome in EOC patients.
ABCA1 expression was silenced in EOC cells to investigate the effect of inhibiting cholesterol efflux on EOC biology through growth and migration assays, three-dimensional spheroid culture and cholesterol quantification.
ABCA1 suppression significantly reduced the growth, motility and colony formation of EOC cell lines as well as the size of EOC spheroids, whilst stimulating expression of ABCA1 reversed these effects. In serous EOC cells, ABCA1 suppression induced accumulation of cholesterol. Lowering cholesterol levels using methyl-B-cyclodextrin rescued the effect of ABCA1 suppression, restoring EOC growth. Furthermore, we identified FDA-approved agents that induced cholesterol accumulation and elicited cytocidal effects in EOC cells.
Our data demonstrate the importance of ABCA1 in maintaining cholesterol balance and malignant properties in EOC cells, highlighting its potential as a therapeutic target for this disease.
ATP-binding cassette (ABC) transporters play various roles in cancer biology and drug resistance, but their association with outcomes in serous epithelial ovarian cancer (EOC) is unknown.
The ...relationship between clinical outcomes and ABC transporter gene expression in two independent cohorts of high-grade serous EOC tumors was assessed with real-time quantitative polymerase chain reaction, analysis of expression microarray data, and immunohistochemistry. Associations between clinical outcomes and ABCA transporter gene single nucleotide polymorphisms were tested in a genome-wide association study. Impact of short interfering RNA-mediated gene suppression was determined by colony forming and migration assays. Association with survival was assessed with Kaplan-Meier analysis and log-rank tests. All statistical tests were two-sided.
Associations with outcome were observed with ABC transporters of the "A" subfamily, but not with multidrug transporters. High-level expression of ABCA1, ABCA6, ABCA8, and ABCA9 in primary tumors was statistically significantly associated with reduced survival in serous ovarian cancer patients. Low levels of ABCA5 and the C-allele of rs536009 were associated with shorter overall survival (hazard ratio for death = 1.50; 95% confidence interval CI =1.26 to 1.79; P = 6.5e-6). The combined expression pattern of ABCA1, ABCA5, and either ABCA8 or ABCA9 was associated with particularly poor outcome (mean overall survival in group with adverse ABCA1, ABCA5 and ABCA9 gene expression = 33.2 months, 95% CI = 26.4 to 40.1; vs 55.3 months in the group with favorable ABCA gene expression, 95% CI = 49.8 to 60.8; P = .001), independently of tumor stage or surgical debulking status. Suppression of cholesterol transporter ABCA1 inhibited ovarian cancer cell growth and migration in vitro, and statin treatment reduced ovarian cancer cell migration.
Expression of ABCA transporters was associated with poor outcome in serous ovarian cancer, implicating lipid trafficking as a potentially important process in EOC.
PDF
