Inhibition of G Protein-Coupled Receptor Kinase 2 Promotes Unbiased Downregulation of IGF1 Receptor and Restrains Malignant Cell Growth.

The ability of a receptor to preferentially activate only a subset of available downstream signal cascades is termed biased signaling. Although comprehensively recognized for the G protein-coupled receptors (GPCR), this process is scarcely explored downstream of receptor tyrosine kinases (RTK), including the cancer-relevant insulin-like growth factor-1 receptor (IGF1R). Successful IGF1R targeting requires receptor downregulation, yet therapy-mediated removal from the cell surface activates cancer-protective ß-arrestin-biased signaling (ß-arr-BS). As these overlapping processes are initiated by the ß-arr/IGF1R interaction and controlled by GPCR-kinases (GRK), we explored GRKs as potential anticancer therapeutic targets to disconnect IGF1R downregulation and ß-arr-BS. Transgenic modulation demonstrated that GRK2 inhibition or GRK6 overexpression enhanced degradation of IGF1R, but both scenarios sustained IGF1-induced ß-arr-BS. Pharmacologic inhibition of GRK2 by the clinically approved antidepressant, serotonin reuptake inhibitor paroxetine (PX), recapitulated the effects of GRK2 silencing with dose- and time-dependent IGF1R downregulation without associated ß-arr-BS. In vivo, PX treatment caused substantial downregulation of IGF1R, suppressing the growth of Ewing's sarcoma xenografts. Functional studies reveal that PX exploits the antagonism between ß-arrestin isoforms; in low ligand conditions, PX favored ß-arrestin1/Mdm2-mediated ubiquitination/degradation of IGF1R, a scenario usually exclusive to ligand abundancy, making PX more effective than antibody-mediated IGF1R downregulation. This study provides the rationale, molecular mechanism, and validation of a clinically feasible concept for "system bias" targeting of the IGF1R to uncouple downregulation from signaling. Demonstrating system bias as an effective anticancer approach, our study reveals a novel strategy for the rational design or repurposing of therapeutics to selectively cross-target the IGF1R or other RTK. SIGNIFICANCE: This work provides insight into the molecular and biological roles of biased signaling downstream RTK and provides a novel "system bias" strategy to increase the efficacy of anti-IGF1R-targeted therapy in cancer.

Cancer-associated rs6983267 SNP and its accompanying long noncoding RNA CCAT2 induce myeloid malignancies via unique SNP-specific RNA mutations.

The cancer-risk-associated rs6983267 single nucleotide polymorphism (SNP) and the accompanying long noncoding RNA CCAT2 in the highly amplified 8q24.21 region have been implicated in cancer predisposition, although causality has not been established. Here, using allele-specific CCAT2 transgenic mice, we demonstrate that CCAT2 overexpression leads to spontaneous myeloid malignancies. We further identified that CCAT2 is overexpressed in bone marrow and peripheral blood of myelodysplastic/myeloproliferative neoplasms (MDS/MPN) patients. CCAT2 induces global deregulation of gene expression by down-regulating EZH2 in vitro and in vivo in an allele-specific manner. We also identified a novel non-APOBEC, non-ADAR, RNA editing at the SNP locus in MDS/MPN patients and CCAT2-transgenic mice. The RNA transcribed from the SNP locus in malignant hematopoietic cells have different allelic composition from the corresponding genomic DNA, a phenomenon rarely observed in normal cells. Our findings provide fundamental insights into the functional role of rs6983267 SNP and CCAT2 in myeloid malignancies.

Retrospective study on osteosarcoma and ewing sarcoma - our experience.

INTRODUCTION: Primary bone tumors are relatively rare types of cancer. Their relative frequency is not yet well established and still there is more information needed regarding the evolution and prognosis of those patients. OBJECTIVES: We analyzed several factors (site of lesion, tumor stage, tumor volume, disease related complications, therapy related complications) that influenced the evolution of bone tumor in a lot of patients diagnosed with osteosarcoma or Ewing sarcome. MATERIAL AND METHODS: A retrospective review was conducted on hospital-based registry from the Emergency Hospital for Children "Louis Turcanu" Timisoara. Patients with newly diagnosed osteosarcoma and Ewing sarcoma, hospitalised in our clinic during a period of 10 years (1996-2006) were included. Records were analyzed for patient demographics, site of lesion, treatment and outcomes. The study group was composed of 36 patients with bone tumors, with ages betwen 3-23 years, who came from Timis and several counties around it. RESULTS: We found Ewing Sarcoma (ES) in 52.94% of cases and osteosarcoma (OS) in 47.06% of cases analyzed. We found diseases in advanced stages in 33.3% of cases in stage III and in 27.7% in stage IV. Tumoral volume had more than 200 cm3 in 53.3% of OS patients and in 21% of cases of ES. Treatment was accomplished according to the European protocols, COSS 96 in 66.6% of OS cases, EWING 99 in 73.6% of ES cases. Disease related complications were found in 26.6% of OS cases and in 51% of ES patients. CONCLUSION: In this study, the patients survival rate at 5 years after diagnosis was lower than in other studies. A possible explaination for such a high rate of mortality could be the delayed diagnosis and the advanced stage of the neoplasia, especially for Ewing sarcoma where only 16.66% of the patients were stage I or II. For the short time survival it was found a corelation with the period of time between the simptoms appearance and the moment of diagnosis, tumor stage, metastasis and severity of the complications.

Novel mechanisms of regulation of IGF-1R action: functional and therapeutic implications.

The IGF-1R pathway is essential for the initiation and progression of many cancers. In contrast to other receptor tyrosine kinases involved in cancer, it is not frequently mutated or amplified. The classical model of signaling through the IGF-1R centers on ligand initiated kinase activation, allowing binding of adaptor molecules and downstream activation of the MAPK and PI3K pathways. The signaling is terminated through receptor ubiquitination and subsequent degradation. To date, therapies targeting IGF-1R have been designed solely aiming to block phosphorylation mediated signaling by preventing receptor-ligand interaction or by limiting kinase activation. Yet, the classical model is insufficient to explain receptor behavior induced by some IGF-1R inhibitors. This review advocates an updated model of IGF-1R signaling, accommodating the "classical" kinase signaling and the IGF-1R-kinase independent signaling thus providing the theoretical background for receptor downregulation induced by IGF-1R inhibitors. This model should be considered for future design of effective therapies targeting the IGF-1R pathway.

Cytotoxic antibodies--valuable prognostic factor for long term kidney allograft survival.

BACKGROUND: Since the first attempts of kidney transplant, the inflammation mediated by T lymphocytes was considered one of the most important processes implicated in graft rejection but, multiple acute and chronic graft rejects revealed that the inflammation process is not singular and humoral mechanisms may play a role in the development of chronic vascular rejection. MATERIAL AND METHODS: We evaluated 500 Romanian patients registered on the kidney transplant waiting list. We performed anti-HLA class I and class II antibodies screening and identification. Laboratory tests were performed at Centre for Immunogenetics and Virology, Fundeni Clinical Institute, Bucharest, Romania. The methods used are represented by ELISA (GTI Diagnosis, USA) and Luminex (Tepnel, USA) RESULTS: pretransplant evaluation of the subjects illustrates that 145 patients (29%) have been sensitized and 355 patients (71%) have not been sensitized. The most frequent types of anti-HLA antibodies were A2 (13%), B42 (10%), DR7 and DR11 (13%). Post transplant, the most cases with de novo antibodies were observed in the first 6 months post transplantation. High serum levels of Il-2 Receptor, TNF-alpha and neopterin in post transplant sensitized patients were observed following de novo cytotoxic antibodies occurrence. CONCLUSION: post renal transplantation, patients present high risk in developing de novo cytotoxic antibodies, especially those who had HLA mismatch with the donor. These antibodies are predictors for acute graft rejection and for graft failure.