Patients with acute myeloid leukemia (AML) carrying high-risk genetic lesions or high residual disease levels after therapy are particularly exposed to the risk of relapse. Here, we identified the ...long non-coding RNA CDK6-AS1 able to cluster an AML subgroup with peculiar gene signatures linked to hematopoietic cell differentiation and mitochondrial dynamics. CDK6-AS1 silencing triggered hematopoietic commitment in healthy CD34+ cells, whereas in AML cells the pathological undifferentiated state was rescued. This latter phenomenon derived from RUNX1 transcriptional control, responsible for the stemness of hematopoietic precursors and for the block of differentiation in AML. By CDK6-AS1 silencing in vitro, AML mitochondrial mass decreased with augmented pharmacological sensitivity to mitochondria-targeting drugs. In vivo, the combination of tigecycline and cytarabine reduced leukemia progression in the AML-PDX model with high CDK6-AS1 levels, supporting the concept of a mitochondrial vulnerability. Together, these findings uncover CDK6-AS1 as crucial in myeloid differentiation and mitochondrial mass regulation.
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•CDK6-AS1 acts in concert with CDK6•High CDK6-AS1 levels trigger RUNX1 early differentiation arrest in myeloid cells•CDK6-AS1 controls mitochondrial mass of AML blasts•CDK6-AS1 levels impact on mitochondrial-targeted agents sensitivity
Molecular biology; Cell biology; Cancer
To overcome the lack of effective pharmacological treatments for high-risk neuroblastoma (HR-NB), the development of novel in vitro and in vivo models that better recapitulate the disease is ...required. Here, we used an in vitro multiclonal cell model encompassing NB cell differentiation stages, to identify potential novel pharmacological targets. This model allowed us to identify, by low-density RT-PCR arrays, two gene sets, one over-expressed during NB cell differentiation, and the other up-regulated in more malignant cells. Challenging two HR-NB gene expression datasets, we found that these two gene sets are related to high and low survival, respectively. Using mouse NB cisplatin-treated xenografts, we identified two genes within the list associated to the malignant stage (MCM2 and carbonic anhydrase 9), whose expression is positively correlated with tumor growth. Thus, we tested their pharmacological targeting as potential therapeutic strategy. We measured mice survival and tumor growth rate after xenografts of human NB treated with cisplatin in the presence of MCM2/carbonic anhydrase 9 inhibitors (ciprofloxacin and acetazolamide). MCM2 or carbonic anhydrase 9 inhibition significantly increased cisplatin activity, supporting their possible testing for NB therapy.
Neuroblastoma (NB) is the most common extra-cranial malignancy in preschool children. To portray the genetic landscape of an overly aggressive NB leading to a rapid clinical progression of the ...disease, tumor DNA collected pre- and post-treatment has been analyzed. Array comparative genomic hybridization (aCGH), whole-exome sequencing (WES), and pharmacogenetics approaches, respectively, have identified relevant copy number alterations (CNAs), single nucleotide variants (SNVs), and polymorphisms (SNPs) that were then combined into an integrated analysis. Spontaneously formed 3D tumoroids obtained from the recurrent mass have also been characterized. The results prove the power of combining CNAs, SNVs, and SNPs analyses to assess clonal evolution during the disease progression by evidencing multiple clones at disease onset and dynamic genomic alterations during therapy administration. The proposed molecular and cytogenetic integrated analysis empowers the disease follow-up and the prediction of tumor recurrence.
The members of the BCL-2 associated athanogene (BAG) family participate in the regulation of a variety of interrelated physiological processes, such as autophagy, apoptosis, and protein homeostasis. ...Under normal circumstances, the six BAG members described in mammals (BAG1-6) principally assist the 70 kDa heat-shock protein (HSP70) in protein folding; however, their role as oncogenes is becoming increasingly evident. Deregulation of the BAG multigene family has been associated with cell transformation, tumor recurrence, and drug resistance. In addition to BAG overexpression, BAG members are also involved in many oncogenic protein–protein interactions (PPIs). As such, either the inhibition of overloading BAGs or of specific BAG–client protein interactions could have paramount therapeutic value. In this review, we will examine the role of each BAG family member in different malignancies, focusing on their modular structure, which enables interaction with a variety of proteins to exert their pro-tumorigenic role. Lastly, critical remarks on the unmet needs for proposing effective BAG inhibitors will be pointed out.
Oncogenic activation of the PI3K signalling pathway plays a pivotal role in the development of glioblastoma multiforme (GBM). A central node in PI3K downstream signalling is controlled by the ...serine-threonine kinase AKT1. A somatic mutation affecting residue E17 of the AKT1 gene has recently been identified in breast and colon cancer. The E17K change results in constitutive AKT1 activation, induces leukaemia in mice, and accordingly, may be therapeutically exploited to target the PI3K pathway. Assessing whether AKT1 is activated by somatic mutations in GBM is relevant to establish its role in this aggressive disease.
We performed a systematic mutational analysis of the complete coding sequence of the AKT1 gene in a panel of 109 tumor GBM samples and nine high grade astrocytoma cell lines. However, no somatic mutations were detected in the coding region of AKT1.
Our data indicate that in GBM oncogenic deregulation of the PI3K pathway does not involve somatic mutations in the coding region of AKT1.
Chromosome instability has a pivotal role among the hallmarks of cancer, but its transcriptional counterpart is rarely considered a relevant factor in cell destabilization. To examine transcription ...instability (TIN), we first devised a metric we named TIN index and used it to evaluate TIN on a dataset containing more than 500 neuroblastoma samples. We found that metastatic tumors from high‐risk (HR) patients are characterized by significantly different TIN index values compared to low/intermediate‐risk patients. Our results indicate that the TIN index is a good predictor of neuroblastoma patient's outcome, and a related TIN index gene signature (TIN‐signature) is also able to predict the neuroblastoma patient's outcome with high confidence. Interestingly, we find that TIN‐signature genes have a strong positional association with superenhancers in neuroblastoma tumors. Finally, we show that TIN is linked to chromatin structural domains and interferes with their integrity in HR neuroblastoma patients. This novel approach to gene expression analysis broadens the perspective of genome instability investigations to include functional aspects.
This study involved the systematic investigation of transcription instability (TIN) as a genome‐wide phenomenon related to cancer. In neuroblastoma, pediatric cancer characterized by a low mutational load, TIN‐related genes has a strong positional association with superenhancers and with chromatin structural domains whose integrity is disrupted in high‐risk compared to low/intermediate‐risk patients.
Therapeutic management of Locally Advanced Rectal Cancer (LARC) involves pre-operative chemoradiotherapy (pCRT) followed by surgery. However, after pCRT the complete pathological response is ...approximately 20%, whereas in 20 to 40% of patients the response is poor or absent.
Cancer biopsy specimens (n= 38) and serum samples (n= 34) obtained before pCRT from 38 LARC patients were included in the study. Patients were classified in responders (R, tumor regression grade TRG 1-2; n= 16) and non-responders (NR, TRG 3-5; n= 22) according to the pathological response observed upon surgery. We performed miRNA microarrays analysis on biopsy specimens, and validated the selected candidates both by qRT-PCR (tissue and serum) and by in situ hybridization (tissue, miR-125b) analyses.
Eleven miRNAs were significantly different between R and NR (miR-154, miR-409-3p, miR-127-3p, miR-214*, miR-299-5p and miR-125b overexpressed in NR; miR-33a, miR-30e, miR-338-3p, miR-200a and miR-378 decreased). In particular, miR-125b resulted to be the best candidate to discriminate the two groups (AUC of 0.9026; 95% CI, 0.7618-1.043). Additionally, miR-125b serum levels were significantly overexpressed in NR patients compared to R (p-value=0.0087), with an excellent discriminating power (AUC of 0.782; 95% CI, 0.6123-0.9518).
The obtained results further support the clinical impact of miRNA analysis. High miR-125b expression in tissue and serum were associated with a poor treatment response in LARC patients, therefore miR-125b could be considered as a possible novel non-invasive biomarker of response in LARC treatment.
Neuroblastoma (NB) is a threatening childhood malignancy. Its prognosis is affected by several morphological, and biological characteristics, including the constitutive expression of ALK tyrosine ...kinase. In this study we examined the therapeutic potential of a novel ALK inhibitor, entrectinib, in obliterating NB tumor cells. Entrectinib showed the growth-inhibitory effects on NB cells with a 50% inhibitory concentration range of 0.03-5 μM. In the ALK-dependent cells, entrectinib mediated G1-arrest, which was associated with modified expression of multiple cell-cycle regulators. Down-regulation of Ki-67, and attenuated phosphorylation of ERK1/2, and STAT3, correlated with observed antiproliferative capacity of entrectinib. Initial cytostatic activity of entrectinib was followed by concentration-dependent apoptotic cell death, and Caspase-3 activation. However, we delineated a reduced sensitivity of ALK mutated NB cells to entrectinib, and demonstrated strong activation of autophagy in SH-SY5YF1174L NB cell line. Abrogation of autophagy by chloroquine increased significantly the toxicity of entrectinib, as confirmed by enhanced death rate, and PARP protein cleavage in SH-SY5YF1174L cells. In aggregate, our data show that entrectinib inhibits proliferation, and induces G1-arrest, and apoptosis in NB cells. We propose entrectinib for further consideration in treatment of NB, and recommend pharmacological inhibition of autophagy to be explored for a combined therapeutic approach in NB patients that might develop resistance to entrectinib.
Neuroblastoma (NB) is an embryonal tumor with low cure rate for patients classified as high-risk. This class of NB tumors shows a very complex genomic background and requires aggressive treatment ...strategies. In this work we evaluated the efficacy of the novel multi-kinase inhibitor TP-0903 in impairing NB cells’ growth, proliferation and motility. In vitro studies were performed using cell lines with different molecular background, and in vivo studies were done using the zebrafish experimental model.
Our results confirmed a strong cytotoxicity of TP-0903 already at the sub-micro molar concentrations. The observed cytotoxicity of TP-0903 was irreversible and the resulting apoptosis was caspase dependent. In addition, TP-0903 impaired colony formation and neurosphere creation. Depending on the molecular background of the selected NB cell lines, TP-0903 influenced either their capacity to migrate, to complete their cell cycle or both. Likewise, TP-0903 reduced NB cells intravasation in vitro and in vivo. Importantly, TP-0903 showed remarkable pharmacological efficacy not only as a mono-treatment, but also in combination with conventional chemotherapy drugs (ATRA, cisplatin, and VP16) in different types of NB cells.
In conclusion, the multi-kinase activity of TP-0903 allowed the impairment of several biological processes required for expansion of NB cells, making them more vulnerable to the conventional chemotherapeutics. Altogether, our results support the eligibility of TP-0903 for further (pre)clinical assessments in NB.
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