PRIMA-1MET induces nucleolar translocation of Epstein-Barr virus-encoded EBNA-5 protein

The low molecular weight compound, PRIMA-1MET restores the transcriptional transactivation function of certain p53 mutants in tumor cells. We have previously shown that PRIMA-1MET induces nucleolar translocation of p53, PML, CBP and Hsp70. The Epstein-Barr virus encoded, latency associated antigen EBNA-5 (also known as EBNA-LP) is required for the efficient transformation of human B lymphocytes by EBV. EBNA-5 associates with p53-hMDM2-p14ARF complexes. EBNA-5 is a nuclear protein that translocates to the nucleolus upon heat shock or inhibition of proteasomes along with p53, hMDM2, Hsp70, PML and proteasome subunits. Here we show that PRIMA-1MET induces the nucleolar translocation of EBNA-5 in EBV transformed B lymphoblasts and in transfected tumor cells. The PRIMA-1MET induced translocation of EBNA-5 is not dependent on the presence of mutant p53. It also occurs in p53 null cells or in cells that express wild type p53. Both the native and the EGFP or DSRed conjugated EBNA-5 respond to PRIMA-1MET treatment in the same way. Image analysis of DSRed-EBNA-5 expressing cells, using confocal fluorescence time-lapse microscopy showed that the nucleolar translocation requires several hours to complete. FRAP (fluorescence recovery after photobleaching) and FLIP (fluorescence loss in photobleaching) measurements on live cells showed that the nucleolar translocation was accompanied by the formation of EBNA-5 aggregates. The process is reversible since the aggregates are dissolved upon removal of PRIMA-1MET. Our results suggest that mutant p53 is not the sole target of PRIMA-1MET. We propose that PRIMA-1MET may reversibly inhibit cellular chaperons that prevent the aggregation of misfolded proteins, and that EBNA-5 may serve as a surrogate drug target for elucidating the precise molecular action of PRIMA-1MET

Characterization of Human Malignancies Caused by Human Herpesvirus 8

Humán herpeszvírus-8 által okozott emberi malignitások jellemzése Ötvös Rita Témavezető: Dr. Kónya József, PhD Debreceni Egyetem Klinikai Központ Mikrobiológiai Intézet Gyógyszerészeti Tudományok Doktori Iskola, Mikrobiológiai Program ÖSSZEFOGLALÁS A humán herpesvírus-8 (HHV-8) a Kaposi sarcoma (KS) patogenezisében bizonyítottan etiológiai szerepet játszik, amelynek felfedezése óta 20 év telt. A vírust AIDS-hez társuló testüregi B-sejtes lymphomák esetén, valamint multicentrikus Castleman-betegségben is kimutatták. A malignus betegségben szenvedők kezelési eredményeinek javítása a jövőben a terápia agresszivitásásnak növelése helyett egyre inkább a személyre szabott terápiától várható. Az in vitro gyógyszerérzékenységi vizsgálatok segítségével meghatározott hatásos citosztatikumok alkalmazása javíthatja a betegség kimenetelét. A HHV-8 kutatás sokat köszönhet a testüregi B-sejtes lymphomának, ugyanis ez ebből eredő immortalizált sejtvonalak in vitro kultúrában könnyen fenntarthatóak és a vírust látens episzomális formában hordozzák. 11 BC sejtvonal in vitro gyógyszerérzékenységét vizsgáltuk kutatásaink során. Az élő és elpusztult sejtek számát automatizált lézer konfokális mikroszkóp segítségével határoztuk meg. A sejtvonalak eredetűktől függetlenül hasonló gyógyszerérzékenységi mintázatot mutattak a 27 tesztelt citosztatikummal szemben. Vizsgálataink szerint a BC sejtvonalak különösen érzékenyek a mikrotubulusokra ható tumorellenes szerekre és az anthracyclin származékokra, de ezek közül is az epirubicin, a daunorubicin, a paxlitacel és a vinorelbin bizonyult a leghatásosabbnak. Mivel a HHV-8 indukálta testüregi B-sejtes lymphoma viszonylag ritka betegség, nincs kialakult kezelési protokoll ezen betegség kezelésére, a B-sejtes lymphomák sémáját követik, mely a betegség stádiumától függően különböző polikemoterápiát javasol. Eredményeink alapján javasoljuk a fent említett gyógyszerek használatát és beépítését betegségek során használt protokollokba. A HHV-8 orf-K1 membránprotein szokatlanul magas szintű genetikai variabilitását mutat, melyen a filogenetikai kutatások alapulnak, ez alapján a HHV-8 hét fő szubtípusát írták le: A, B, C, D, E, F és Z. Kutatásaink során az orf-K1 génrégiót amplifikáltuk fel PCR segítségével és szekvenáltuk hazai Kaposi sarcomas betegek mintáin. Összesen 17 betegtől 36 formalin-fixált paraffinba ágyazott preparátumot vizsgáltunk. Az orf-K1 génrégió alapján az A szubtípust identifikáltuk, azonbelül az A1 és A2 szubtípust. Két beteg mintájában az orf-K1 két különböző variánsát mutattuk ki. Az általunk végzett munka fontos adatokat szolgáltat a HHV-8 magyarországi elterjedéséről és eredetéről, mely alapján megállapíthatjuk, hogy a vizsgálatainkban talált szubtípusok illeszkednek a régióban elterjedt törzsekhez. Kulcsszavak / Keywords: HHV-8 / HHV-8 K1 törzs / K1 train Testüregi B-sejtes lymphoma / Body-cavity based lymphoma Characterization of Human Malignancies Caused by Human Herpesvirus 8 by Rita Ötvös supervisor: Kónya József MD, PhD Department of Medical Microbiology, University of Debrecen Doctoral School of Pharmaceutical Sciences, Microbiology Program SUMMARY The human herpesvirus 8 (HHV-8) is the etiologic agent in Kaposi´s sarcoma which was discovered 20 years ago. It was found to be associated with some rare types of lymphoma in AIDS, namely body-cavity based lymphoma and the plasmablastic variant of multicentric Castleman´s disease and some cases was shown in bone marrow samples of myeloma multiplex and benign monoclonal gammopathy disease. The survival of patients with malignant disease can be increased in the future using a more individualized therapy. Using of effective drugs determined by in vitro drug sensitivity test might result in a better clinical outcome. As body cavity cell lines (BCBLs) are well established in vitro models for body-cavity based lymphoma we have assessed 11 BCBLs for cytotoxic drug sensitivity. The precise number of living and dead cells was determined using a custom made automated laser confocal fluorescent microscope. Independently from their origin, BCBLs showed very similar patterns against 27 frequently used cytostatic drugs. BCBLs were highly sensitive for epirubicin, daunorubicin, paclitaxel and vinorelbin. The prognosis of PEL is poor, as the median survival in the previously published series does not exceed 6 months. Despite the improvement in therapeutical strategies during the last few years, there is no evidence of a cure for BCBL patients with conventional systemic chemotherapy addressed to aggressive NHL. Our data suggest that inclusion of the above drug into BCBL chemotherapy protocols may be justified. Based on the sequence variation in the open reading frame (orf) K1, HHV-8 is now classified into subtypes A, B, C, D, E, F and Z. We attempted to develop a typing system based on amplification and sequencing of the K1 region. A total of 36 paraffin-embedded biopsies were tested in 17 patients. Based on the K1 region we identified subtype A within subtype A1 and A2. We determined different K1 strains in two specimens. The main outcome is that it provides useful data for molecular epidemiological studies in Hungary and the found genotype is considered to originate from Europe. Kulcsszavak / Keywords: HHV-8 / HHV-8 K1 strain / K1 törzs Body-cavity based lymphoma / Testüregi B-sejtes lymphomaN

Syndecan-1 in Cancer: Implications for Cell Signaling, Differentiation, and Prognostication

Syndecan-1, a cell surface heparan sulfate proteoglycan, is critically involved in the differentiation and prognosis of various tumors. In this review, we highlight the synthesis, cellular interactions, and the signalling pathways regulated by syndecan-1. The basal syndecan-1 level is also crucial for understanding the sequential changes involving malignant transformation, tumor progression, and advanced or disseminated cancer stages. Moreover, we focus on the cellular localization of this proteoglycan as cell membrane anchored and/or shed, soluble syndecan-1 with stromal or nuclear accumulation and how this may carry different, highly tissue specific prognostic information for individual tumor types

The D ~ Sense ex-vivo viability assay application in a patient with stage IV lung adenocarcinoma: a case report

Abstract Background The treatment resistance is a problem for lung cancer. In this study, we used a vitro tissue culturing system to select a new therapy strategy for a patient with tyrosine kinase inhibitors (TKIs) resistance. Case presentation A 42-year-old male Asian patient was diagnosed with advanced lung adenocarcinoma harboring an exon 19 deletion in the epidermal growth factor receptor (EGFR) gene. The patient was treated with Gefitinib, resulting in an almost complete remission for over a year. The patient relapsed after 13 months treatment, and received four cycles of chemotherapy. At 20 months, the patient had developed multiple lung metastases and a solitary cerebellar metastasis. An EGFR T790M mutation was identified in the peripheral blood sample. Subsequent treatment with Osimertinib resulted in a complete response of the intracranial metastasis. By 33 months, the patient had developed a mediastinal tumor mass that responded well to local radiotherapy. By 39 months, an EGFR C797S cis-mutation had been identified and the patient was treated with Brigatinib and Cetuximab. By 44 months, the tumor cells from the pleural effusion had been tested for sensitivity against 30 targeted and cytostatic drugs using the D ~ Sense ex-vivo viability assay. The assay identified 8 drugs with moderate to high sensitivity. Combination therapy of Gemcitabin and Lobaplatin had resulted in disease stabilization. Conclusions The case showed that individualized treatment aided by D ~ Sense ex-vivo viability assay can be a viable option for patients with advanced lung adenocarcinoma with pleural effusions

L'Auto-vélo : automobilisme, cyclisme, athlétisme, yachting, aérostation, escrime, hippisme / dir. Henri Desgranges

12 juillet 19291929/07/12 (A30,N10436)