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The Role of RING Ubiquitin Ligases in Biologic and Oncologic Processes in Tissues of Mesenchymal Origin (RING UB LIGASE)

22. februar 2015 oppdatert av: Rambam Health Care Campus

Stage 1:

Bone marrow will be collected from otherwise healthy patients undergoing orthopedic surgery - arthroplasty for the treatment of degenerative joint disease or traumatised patients. The bone marrow will be collected from disposable tissue that is removed during the normal sequence of the surgery. These cells will be used for:

Establishment of the role of RING ubiquitin ligases in osteogenic progenitors proliferation and differentiation in culture.

I. To determine the levels of RING ubiquitin ligases mRNA and protein in differentiating human mesenchymal precursor cells: To test the correlation between levels of RING ubiquitin ligases and degree of osteogenic differentiation, we will extract mesenchymal precursor cells from the collected tissue and test the mRNA protein level of RING ubiquitin ligases upon induction of differentiation. Specifically, we will compare the levels of RING ubiquitin ligases before and after the initiation of differentiational stimulus in time dependent manner by western blot and real time PCR analysis.

II. To test the impact of specific RING ubiquitin ligases on differentiation of human mesenchymal precursor cells: to test the impact of RING ubiquitin ligases on the differentiation of mesenchymal precursor cell from the collected tissue by testing the expression of classical differentiational bone markers by flow cytometry (fibronectin, CD105) and by Alkaline phosphatase (ALP) activity assay. For this aim we developed both a constitutive and Dox-regulated conditional overexpression and shRNA lenti-viral systems that enables efficient modulation of these RING ubiquitin ligases level.

III. Determine the role of RING ubiquitin ligases in proliferation and survival of human mesenchymal precursor cells: Via inhibition or overexpression of ligases in mesenchymal progenitor cells we will test the role of these ligases in proliferation and survival of mesenchymal precursors by using MTT assay, Propidion-Iodid (PI) and tunnel assays in flow cytometry analysis.

Stage 2:

Collection of connective tissue from patients with malignancies of musculoskeletal origin. The tissue that will be used is part of the resected tumor specimens. The tissue will be used for:

The establishment of the role/s of RING ligases in musculoskeletal cancers using cell culture and in vivo activation. Test if the expression of selected positive candidates from stage 1 correlates with cancer development and progression in human-derived samples Independently of our mechanistic experiment we aim to determine the relevance of these ligases to human musculoskeletal cancers. As the first step we will screen primary tumor biopsies at the protein level correlates with cancer grade and prognosis. Toward this aim we recently generated a highly specific several anti-monoclonal antibodies in our laboratory as well use comercial available antibodies .

Studieoversikt

Status

Ukjent

Forhold

Detaljert beskrivelse

Stage 1:

Bone marrow will be collected from otherwise healthy patients undergoing orthopedic surgery - arthroplasty for the treatment of degenerative joint disease or traumatised patients. The bone marrow will be collected from disposable tissue that is removed during the normal sequence of the surgery. These cells will be used for:

Establishment of the role of RING ubiquitin ligases in osteogenic progenitors proliferation and differentiation in culture.

I. To determine the levels of RING ubiquitin ligases mRNA and protein in differentiating human mesenchymal precursor cells: To test the correlation between levels of RING ubiquitin ligases and degree of osteogenic differentiation, we will extract mesenchymal precursor cells from the collected tissue and test the mRNA protein level of RING ubiquitin ligases upon induction of differentiation. Specifically, we will compare the levels of RING ubiquitin ligases RING ubiquitin ligases before and after the initiation of differentiational stimulus in time dependent manner by western blot and real time PCR analysis.

II. To test the impact of RING ubiquitin ligases on differentiation of human mesenchymal precursor cells: to test the impact of RING ubiquitin ligases on the differentiation of mesenchymal precursor cell from the collected tissue by testing the expression of classical differentiational bone markers by flow cytometry (fibronectin, CD105) and by Alkaline phosphatase (ALP) activity assay. For this aim we developed both a constitutive and Dox-regulated conditional RING ubiquitin ligases overexpression and shRNA lenti-viral systems that enables efficient modulation of RING ubiquitin ligases level.

III. Determine the role of RING ubiquitin ligases in proliferation and survival of human mesenchymal precursor cells: We will test the role of their in proliferation and survival of mesenchymal precursors by using MTT assay, Propidion-Iodid (PI) and tunnel assays in flow cytometry analysis.

Stage 2:

Collection of connective tissue from patients with malignancies of musculoskeletal origin. The tissue that will be used is part of the resected tumor specimens. The tissue will be used for:

The establishment of the role/s of RING ubiquitin ligases in musculoskeletal cancers using cell culture and in vivo.

I. Test the role of our in proliferation and survival in musculoskeletal cancers: Using our lentiviral systems we will test the biological impact of inhibition and overexpression in musculoskeletal neoplastic cells.

II. Test whether inhibition or overexpression of these ligases impacts the biology of musculoskeletal cancers.We will check if inhibition of these ligases in malignant cells of musculoskeletal origin by lenti-viral infections induces differentiation. Infected cells will be analyzed for expression of differentiational markers in flow cytometry and immunohistochemistry.

III. Test if expression of RING ubiquitin ligases correlates with cancer development and progression in human-derived samples Independently of our mechanistic experiment we aim to determine the relevance of RING ubiquitin ligases to human musculoskeletal cancers.

Patients consent will be signed after properly informed at time of pre-surgical preparation visit. In cases when pre-surgical visit is not held consent will be signed prior to the day of surgery.

Studietype

Observasjonsmessig

Registrering (Forventet)

90

Kontakter og plasseringer

Denne delen inneholder kontaktinformasjon for de som utfører studien, og informasjon om hvor denne studien blir utført.

Studiesteder

  • Israel
      • Haifa, Israel
        • Rekruttering
        • Rambam MC
        • Ta kontakt med:
        • Ta kontakt med:
        • Hovedetterforsker:
          • David Nikomarov, MD
        • Underetterforsker:
          • Rostislav Novak, MD/PhD
        • Underetterforsker:
          • Amir Oryan (Orian), MD/PhD

Deltakelseskriterier

Forskere ser etter personer som passer til en bestemt beskrivelse, kalt kvalifikasjonskriterier. Noen eksempler på disse kriteriene er en persons generelle helsetilstand eller tidligere behandlinger.

Kvalifikasjonskriterier

Alder som er kvalifisert for studier

18 år og eldre (Voksen, Eldre voksen)

Tar imot friske frivillige

Nei

Kjønn som er kvalifisert for studier

Alle

Prøvetakingsmetode

Ikke-sannsynlighetsprøve

Studiepopulasjon

  1. Healthy patients undergoing orthopedic surgery - arthroplasty for the treatment of degenerative joint disease or traumatised patients.
  2. Patients with malignancies of musculoskeletal origin.

Beskrivelse

Group 1:

  • Inclusion Criteria:
  • Healthy patients undergoing orthopedic surgery - arthroplasty for the treatment of degenerative joint disease or traumatised patients.
  • Exclusion Criteria:
  • Any malignancy in the past or active infection.

Group 2:

  • Inclusion Criteria:
  • patients with malignancies of musculoskeletal origin.
  • Exclusion Criteria:
  • Any malignancy of unknown origin or malignancy of multiple origins or active infection.

Studieplan

Denne delen gir detaljer om studieplanen, inkludert hvordan studien er utformet og hva studien måler.

Hvordan er studiet utformet?

Designdetaljer

Kohorter og intervensjoner

Gruppe / Kohort
The role of RING ubiquitin ligases in osteogenic progenitors
Bone marrow (BM) will be collected from healthy patients undergoing orthopedic surgery. BM will be collected from disposable tissue that is removed during the normal sequence of the surgery.
The role of RING ubiquitin ligases in musculoskeletal cancer
Collection of connective tissue from patients with tumors of musculoskeletal origin - a part of the resected tumor specimens.

Hva måler studien?

Primære resultatmål

Resultatmål
Tiltaksbeskrivelse
Tidsramme
Protein and mRNA levels of RING ubiquitin ligases during osteogenesis
Tidsramme: 5 years
To determine the levels of RING ubiquitin ligases mRNA and protein in differentiating human mesenchymal precursor cells. We will extract mesenchymal precursor cells from the collected tissue and test the mRNA and protein levels of RING ubiquitin ligases upon induction of differentiation.
5 years
Protein and mRNA levels of RING ubiquitin ligases in musculoskeletal tumors and its correlation to staging and grading
Tidsramme: Time Frame: * 5 years
It is not clear if RING ubiquitin ligases protein levels are elevated in soft tissue-derived cancers (primary and metastatic tumors) and whether there is a correlation between RING ubiquitin ligases levels and staging and grading of soft tissue-derived cancers. For this reason, we will check whether RING ubiquitin ligases expression correlates with cancer development and progression in human-derived samples. We will determine the relevance of RING ubiquitin ligases to human musculoskeletal cancers. To do this we will screen primary tumor biopsies for RNA and protein levels of RING ubiquitin ligases .
Time Frame: * 5 years

Samarbeidspartnere og etterforskere

Det er her du vil finne personer og organisasjoner som er involvert i denne studien.

Sponsor

Rambam Health Care Campus

Samarbeidspartnere

Technion, Israel Institute of Technology

Studierekorddatoer

Disse datoene sporer fremdriften for innsending av studieposter og sammendragsresultater til ClinicalTrials.gov. Studieposter og rapporterte resultater gjennomgås av National Library of Medicine (NLM) for å sikre at de oppfyller spesifikke kvalitetskontrollstandarder før de legges ut på det offentlige nettstedet.

Studer hoveddatoer

Studiestart

1. oktober 2014

Primær fullføring (Forventet)

1. juli 2019

Studiet fullført (Forventet)

1. juli 2019

Datoer for studieregistrering

Først innsendt

29. juli 2014

Først innsendt som oppfylte QC-kriteriene

30. september 2014

Først lagt ut (Anslag)

3. oktober 2014

Oppdateringer av studieposter

Sist oppdatering lagt ut (Anslag)

24. februar 2015

Siste oppdatering sendt inn som oppfylte QC-kriteriene

22. februar 2015

Sist bekreftet

1. februar 2015

Mer informasjon

Begreper knyttet til denne studien

Andre studie-ID-numre

  • 0180-14-RMB

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