MicroRNA Gene Networks in Oncogenesis

MicroRNAs are small non-coding RNAs that regulate gene expression at the transcriptional or posttranscriptional level. They are involved in cellular development, differentiation, proliferation and apoptosis and play a significant role in cancer. Examination of tumor-specific microRNA expression profiles has revealed widespread deregulation of these molecules in diverse cancers. Several studies have shown that microRNAs function either as tumor suppressor genes or oncogenes, whose loss or overexpression respectively has diagnostic and prognostic significance. It seems that microRNAs act as major regulators of gene expression. In this review, we discuss microRNAs’ role in cancer and how microRNAs exert their functions through regulation of their gene targets. Bioinformatic analysis of putative miRNA binding sites has indicated several novel potential gene targets involved in apoptosis, angiogenesis and metastatic mechanisms. Matching computational prediction analysis together with microarray data seems the best method for microRNA gene target identification. MicroRNAs together with transcription factors generate a complex combinatorial code regulating gene expression. Thus, manipulation of microRNA-transcription factor gene networks may be provides a novel approach for developing cancer therapies

Durvalumab: an investigational anti-PD-L1 monoclonal antibody for the treatment of urothelial carcinoma.

Our expanding knowledge of immunotherapy for solid tumors has led to an explosion of clinical trials aimed at urothelial carcinoma. The primary strategy is centered on unleashing the immune system by releasing the inhibitory signals propagated by programmed cell death-1 (PD-1) and its ligand programmed cell death ligand-1 (PD-L1). Many antibody constructs have been developed to block these interactions and are used in clinical trials. The Food and Drug Administration has already approved a number of checkpoint inhibitors such as anti-cytotoxic T-lymphocyte-associated protein 4 (CTLA4) monoclonal antibodies including ipilimumab; anti-PD-1 monoclonal antibodies including nivolumab and pembrolizumab; anti-PD-L1 antibodies including atezolizumab, avelumab, and durvalumab. One of the latest inhibitors is durvalumab, which is a high-affinity human immunoglobulin G1 kappa monoclonal antibody and blocks the interaction of PD-L1 with PD-1 and CD80. Currently, there are a number of ongoing trials in advanced urothelial carcinoma both using durvalumab monotherapy and in combination with other targeted therapies. In addition, durvalumab is being investigated in the non-muscle-invasive urothelial carcinoma, which is centered around intravenous formulations. These exciting developments have added a significant number of therapies in a previously limited treatment landscape

MiR-27b targets PPARγ to inhibit growth, tumor progression, and the inflammatory response in neuroblastoma cells

The PPARγ nuclear receptor pathway is involved in cancer, but it appears to have both tumor suppressor and oncogenic functions. In neuroblastoma cells, miR-27b targets the 3′UTR of PPARγ and inhibits its mRNA and protein expression. miR-27b overexpression or PPARγ inhibition blocks cell growth in vitro and tumor growth in mouse xenografts. PPARγ activates expression of the pH regulator NHE1, which is associated with tumor progression. Lastly, miR-27b through PPARγ regulates NF-κB activity and transcription of inflammatory target genes. Thus, in neuroblastoma, miR-27b functions as a tumor suppressor by inhibiting the tumor-promoting function of PPARγ, which triggers an increased inflammatory response. In contrast, in breast cancer cells, PPARγ inhibits NHE1 expression and the inflammatory response, and it functions as a tumor suppressor. We suggest that the ability of PPARγ to promote or suppress tumor formation is linked to cell-type specific differences in regulation of NHE1 and other target genes

Durvalumab: an investigational anti-PD-L1 monoclonal antibody for the treatment of urothelial carcinoma

Our expanding knowledge of immunotherapy for solid tumors has led to an explosion of clinical trials aimed at urothelial carcinoma. The primary strategy is centered on unleashing the immune system by releasing the inhibitory signals propagated by programmed cell death-1 (PD-1) and its ligand programmed cell death ligand-1 (PD-L1). Many antibody constructs have been developed to block these interactions and are used in clinical trials. The Food and Drug Administration has already approved a number of checkpoint inhibitors such as anti-cytotoxic T-lymphocyte-associated protein 4 (CTLA4) monoclonal antibodies including ipilimumab; anti-PD-1 monoclonal antibodies including nivolumab and pembrolizumab; anti-PD-L1 antibodies including atezolizumab, avelumab, and durvalumab. One of the latest inhibitors is durvalumab, which is a high-affinity human immunoglobulin G1 kappa monoclonal antibody and blocks the interaction of PD-L1 with PD-1 and CD80. Currently, there are a number of ongoing trials in advanced urothelial carcinoma both using durvalumab monotherapy and in combination with other targeted therapies. In addition, durvalumab is being investigated in the non-muscle-invasive urothelial carcinoma, which is centered around intravenous formulations. These exciting developments have added a significant number of therapies in a previously limited treatment landscape

Ramucirumab plus docetaxel versus placebo plus docetaxel in patients with locally advanced or metastatic urothelial carcinoma after platinum-based therapy (RANGE): a randomised, double-blind, phase 3 trial

Few treatments with a distinct mechanism of action are available for patients with platinum-refractory advanced or metastatic urothelial carcinoma. We assessed the efficacy and safety of treatment with docetaxel plus either ramucirumab-a human IgG1 VEGFR-2 antagonist-or placebo in this patient population

Identification of novel microRNAs involved in hepatocellular carcinogenesis by high throughput screening

Identification of Novel MicroRNAs Involved in Hepatocellular Carcinogenesis by High Throughput screening. Abstract-background: Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death in men and the sixth in women. Those numbers reflect the aggressiveness of this disease while at the same time mirror the absence of effective therapeutic regimens. There have been conducted hundreds of clinical trials in which either combination chemotherapies or small molecule inhibitors and recently immune checkpoint inhibitors have been studied. However the only FDA approved drug for unresectable or metastatic HCC is sorafenib which is a tyrosine kinase inhibitor. Our interest was to identify genes that play a role in liver oncogenesis and so we directed our focus to study a novel class of small non coding RNAs, the microRNAs.MicroRNAs have been involved in the pathogenesis of different types of cancers, including liver cancer. Specific microRNA signatures have been identified to be deregulated in HCC patient tissues and also to correlate with different clinicopathological parameters including survival. Having all this in mind our mission was to identify microRNAs that have both functional and clinical relevance in HCC and examine their downstream signaling effectors. Methods: In this study, we have screened the human microRNAome, aiming to identify microRNAs that are potent regulators of HCC invasiveness. Initially we received 24 tissues from patients with liver cancer along with 14 from adjacent non tumor specimens. RNA extraction was performed and microRNA and gene expression levels were measured by quantitative real-time RT-PCR. We determined the expression levels of miR-9, miR-21 and miR-224 in those samples. Simultaneously a novel microRNA library screen was done after plating SNU-449 liver cancer cells in 96-well plates. Those were transfected with a microRNA library consisting of 316 microRNA mimics and 2 negative controls. 48hrs post transfection we evaluated the SNU-449 cell invasiveness. We also performed invasion assays in SNU-449 cells 24hrs after transfection specifically with miR-9 and anti-miR-9 and their respective controls. A TargetScan algorithm was used to identify miR-9 downstream direct targets based on bioinformatics. Those analyses were validated by performing 3’UTR luciferase assay. Precisely, we transfected SNU-449 cells with the reporter vectors carrying the 3’UTR of CDH1 or PPARA. The constructs harbored the seed sequence of miR-9 (wildtype) or had a deletion of this sequence (miR-9 mutant). At 24 hours, they were transfected with miR-9 or miR-control and at 48 hours luciferase activity was measured. We have also performed transfection with microRNA mimic for mir-9 overexpression as well as microRNA inihibitor to suppress the miR-9 activity and observed the expression levels of the downstream targets CDH1, PPARA, PDK4 and vimentin. Furthermore we performed cell proliferation as well as assessed sphere formation and colony assay in SNU-449 and HepG2 liver cancer cell lines, both of which were transfected with miR-9 and anti-miR-9 along with their respective controls. Results: First we performed a high-throughput microRNA screen in SNU-449 liver cancer cells and assessed their invasiveness while secondly we evaluated in tissue the expression levels of the microRNAs derived from the above screen. Overexpression of miR-9 was found to be the top inducer of SNU-449 cell invasiveness, cell growth and their ability to form colonies in soft agar. Furthermore miR-9 levels were found in tissue to increase during HCC progression. Bioinformatics and 3’UTR luciferase analyses identified E-cadherin (CDH1) and peroxisome proliferator-activated receptor alpha (PPARA) as direct downstream effectors of miR-9 activity. Inhibition of PPARA suppressed CDH1 mRNA levels, suggesting that miR-9 regulates CDH1 expression directly through binding in its 3’UTR and indirectly through PPARA. On the other hand, miR-9 inhibition of overexpression suppressed HCC tumorigenicity and invasiveness. PPARA and CDH1 mRNA levels were decreased in HCC relative to controls and were inversely correlated with miR-9 levels.Conclusions: Our study identified a novel microRNA signaling pathway, consisting of miR-9, PPARA and CDH1 that is deregulated in HCC patients affecting liver cancer cellular invasiveness and metastatic potential.Η διερεύνηση νέων MicroRNA που παίζουν ρόλο στην δημιουργία καρκίνου ήπατος μέσα από υψηλής τεχνολογίας ανάλυση. Περίληψη-ιστορικό: Το ηπατοκυταρρικό καρκίνωμα ή ηπάτωμα είναι ο συχνότερος πρωτοπαθής κακοήθης όγκος του οργάνου αυτού (90-95%) και η συχνότερη αναλογικά θανατηφόρος κακοήθης νεοπλασία. Συγκεκριμένα ο καρκίνος του ήπατος είναι η δεύτερη αιτία θανάτου λόγω καρκίνου για τους άντρες και η έκτη αιτία για τις γυναίκες. Αυτές οι στατιστικές αναλύσεις αντικατοπτρίζουν την επιθετικότητα αυτού του όγκου καθώς και την εως τώρα έλλειψη αποτελεσματικών θεραπειών στο συγκεκριμένο πεδίο. Έχουν διεξαχθεί εκατοντάδες κλινικές δοκιμές είτε με συνδυασμό χημικοθεραπευτικών σχημάτων είτε με μικρομοριακούς αναστολείς και πρόσφατα αναστολείς του ανοσοποιητικού συστήματος. Παρ’όλα αυτά το μόνο φάρμακο εγκεκριμένο από τον ΕΟΦ για ανεγχείρητους ή μεταστατικούς όγκους είναι το sorafenib που είναι αναστολέας της τυροσινικής κινάσης.Ο σκοπός μας ήταν να βρούμε γονίδια που παίζουν ρόλο στην δημιουργία του ηπατοκυτταρικού καρκίνου και έτσι κατευθύναμε τον στόχο μας στο να μελετήσουμε και να καταλάβουμε κάποια νέα μικρά γονίδια που δεν μεταφράζονται σε πρωτεΐνες και λέγονται microRNAs.Τα microRNAs έχουν εμπλακεί στην παθογένεια διαφορετικών καρκίνων συμπεριλαμβανομένου αυτού του ήπατος. Συγκεκριμένες microRNA υπογραφές έχουν βρεθεί να είναι απορρυθμισμένες στους ασθενείς με ηπατοκυτταρικό καρκίνωμα και να συσχετίζονται με την επιβίωση.ΜΕΘΟΔΟΙΣε αυτήν την μελέτη εξετάσαμε το ανθρώπινο microRNA γονιδίωμα. Επίπεδα έκφρασης των microRNA και γονιδίων μετρήθηκαν με ποσοτική πραγματικού χρόνου PCR σε ιστούς με ηπατοκυτταρικό καρκίνωμα και φυσιολογικούς ιστούς. Ο αλγόριθμος TargetScan χρησιμοποιήθηκε για να βρεθούν οι άμεσοι στόχοι καθοδικά του microRNA-9. Αποτελέσματα: Μέσα από υψηλής τεχνολογίας ανάλυση του ανθρώπινου microRNA γονιδιώματος βρήκαμε 28 microRNAs που είναι ρυθμιστές και προαγωγείς της επιθετικότητας των ηπατικών κυτταρικών σειρών. MiR-9, miR-21 και miR-224 ήταν οι 3 πρώτοι επαγωγείς της επιθετικότητας και η έκφραση τους ήταν πιο αυξημένη στους ιστούς με καρκίνο από ότι τους φυσιολογικούς. Ο συνδυασμός των κλινικών και μοριακών δεδομένων έδειξε ότι το mir-9 ήταν το πρώτο με κλινική και λειτουργική σπουδαιότητα. Τα επίπεδα του συσχετίζονταν με την σταδιοποίηση των ασθενών με ηπατοκυτταρικό καρκίνο. Είναι σημαντικό να σημειωθεί ότι η υπερέκφραση του miR-9 in vitro , επάγει στις SNU-449 και HepG2 κυτταρικές σειρές την κυτταρική ανάπτυξη, επιθετικότητα και ικανότητα τους να σχηματίζουν αποικίες σε μαλακό άγαρ. Μέσα από βιοπληροφορική και ανάλυση λουσιφεράσης βρήκαμε ότι η e-cadherin και το peroxisome proliferator-activated receptor alpha (PPARA) είναι οι άμεσοι στόχοι καθοδικά του microRNA-9. Αναστολή του PPARA καταστέλλει τα επίπεδα του mRNA της e-cadherin γεγονός που υποδηλώνει ότι το miR-9 ρυθμίζει την έκφραση της e-cadherin άμεσα μέσω σύνδεσης στο 3’UTR του γονιδίου και έμμεσα μέσω PPARA. Επιπρόσθετα η αναστολή της υπερέκφρασης του miR-9 μειώνει την ογκογονικότητα καθώς και την επιθετικότητα του όγκου. Τα επίπεδα έκφρασης του mRNA του PPARA και της e-cadherin ήταν μειωμένα στους ιστούς με ηπατοκυτταρικό καρκίνωμα σε σχέση με τους φυσιολογικούς και αντιστρόφως ανάλογα με τα επίπεδα του miR-9. Συμπέρασμα: Εν κατακλείδι, αυτή η μελέτη περιγράφει για πρώτη φορά τον σημαντικό ρόλο του μονοπατιού σηματοδότησης miR-9/PPARA/e-cadherin στην δημιουργία καρκίνου του ήπατος. Με βάση αυτήν την εργασία ο επιστημονικός στόχος μας είναι να χρησιμοποιήσουμε μη αναστρέψιμους αναστολείς του miR-9 σε κλινικές δοκιμές πρώτης φάσεως με ελπίδα να χρησιμοποιηθούν ως πιθανή θεραπεία του ηπατοκυταρρικού καρκίνου

MicroRNA-gene signaling pathways in pancreatic cancer

Pancreatic cancer is the fourth most frequent cause of cancer-related deaths and is characterized by early metastasis and pronounced resistance to chemotherapy and radiation therapy. Despite extensive esearch efforts, there is not any substantial progress regarding the identification of novel drugs against pancreatic cancer. Although the introduction of the chemotherapeutic agent gemcitabine improved clinical response, the prognosis of these patients remained extremely poor with a 5-year survival rate of 3-5%. Thus, the identification of the novel molecular pathways involved in pancreatic oncogenesis and the development of new and potent therapeutic options are highly desirable. Here, we describe how microRNAs control signaling pathways that are frequently deregulated during pancreatic oncogenesis. In addition, we provide evidence that microRNAs could be potentially used as novel pancreatic cancer therapeutics through reversal of chemotherapy and radiotherapy resistance or regulation of essential molecular pathways. Further studies should integrate the deregulated genes and microRNAs into molecular networks in order to identify the central regulators of pancreatic oncogenesis. Targeting these central regulators could lead to the development of novel targeted therapeutic approaches for pancreatic cancer patients

Anti-PD-L1 therapy and the onset of diabetes mellitus with positive pancreatic autoantibodies.

An 84-year-old woman with metastatic squamous cell carcinoma of the nasopharynx and no history of diabetes was started on the antiprogrammed cell death ligand-1 (anti-PD-L1) antibody durvalumab. Four months later, she presented in diabetic ketoacidosis with glucose 488 mg/dL, anion gap 16, positive serum ketones and A1C9.1%. Antiglutamic acid decarboxylase 65 (GAD) antibody was 13 U/mL (normal, <0.5 U/mL), c-peptide 0.4 ng/dL (normal, 1.1-4.3 ng/mL) and glucose 142 mg/dL. A man with metastatic papillary urothelial carcinoma was treated with the PD-L1 inhibitor atezolizumab. He had no history of diabetes. Nine weeks after initiation, he developed fatigue and polyuria with blood glucose 336 mg/dL, c-peptide 0.6 ng/mL, A1C8.2% and GAD antibodies 28.4 U/mL (normal, <1 U/mL). Due to the diagnosis of autoimmune diabetes, both patients were treated with insulin. Autoimmune diabetes is a rare immune-related adverse effect of PD-L1 inhibitors. We present the first two cases with documented positive pancreatic autoantibodies

Rapidly progressive neurologic decline and morbilliform rash presenting in a patient with lymphoma

A 67-year-old male with past medical history of mantle cell lymphoma and atrial fibrillation presented with a truncal rash, bilateral lower extremity weakness, and confusion. Within three days of presentation, his condition rapidly deteriorated with the onset of diffuse flaccid paralysis, aphasia, and severe alteration in mental status. Initial results from serum studies, lumbar puncture, magnetic resonance imaging, and electroencephalogram were not diagnostic. However, on the ninth day after initial presentation, the West Nile Virus (WNV) immunoglobulin M antibody returned positive from the cerebrospinal fluid. West Nile Virus encephalitis is endemic worldwide, and is the most common viral encephalitis in the United States. WNV presents in a variety of ways, and the recognition by physicians is crucial due to the estimated 2- 12% mortality rate and significant longterm morbidity of neuroinvasive disease. The initial management and long term prognosis are points of ongoing research. This case represents a particularly profound example of neuroinvasive WNV. Our patient made a significant recovery after his initial presentation with aggressive supportive care, however still suffers from bilateral lower extremity weakness more than a year later