Polyclonal Hematopoietic Repopulation in Clinical Wiskott-Aldrich Syndrome (WAS) Gene Therapy Trial.

KERSTIN SCHWARZWAELDER; MANFRED SCHMIDT; KAAN BOZTUG; HANNO GLIMM; CHRISTINA LULAY; RICARDO A. DEWEY; INES AVEDILLO DÍEZ; JANA DIESTELHORST; SONJA NAUNDORF; KLAUS KUEHLKE; IRINA KONDRATENKO; LAZLO MARODI; KARL WELTE; CHRISTOPH KLEIN; CHRISTOPH VON KALLE

Boston, USA

Congreso; American Society of Gene Therapy 11th Annual Meeting; 2008

American Society of Gene Therapy

 The potential of gene therapy to treat monogenetic immunodeficiencies has been proven in clinical trials of the lymphoid and myeloid compartment. In the underlying clinical gene therapy trial two patients suffering from Wiskott-Aldrich Syndrome (WAS), a disease concerning the lymphoid as well as the myeloid lineage, were successfully treated. Both patients received autologous CD34+ cells which were transduced with a retroviral vector encoding the therapeutic Wiskott-Aldrich Syndrome protein (WASP) gene. We accomplished linear amplification mediated PCR (LAM-PCR) and Sanger sequencing to analyse the clonal inventory of the patients. The hematopoietic repopulation of patient 1 has been polyclonal until 401 days post transplantation and the repopulation of patient 2 has been polyclonal until 364 days after treatment which were the last time points analysed. So far we identified 128 and 207 unique integration sites (IS) from patient 1 and 2 by Sanger sequencing, respectively. Among these we detected common insertion sites (CIS) of 2. (2 IS within 30 kb), 3. (3 IS within 50 kb) or 4. (4 IS within 100 kb) order. 4 integration sites were located in the PRDM16 locus, 3 integration sites were located in the UMODL1-, CCND2 and ARNTL2 locus and 2 integration sites were located in a non coding region of chromosome 14, in the TAL1-, C1orf83-, TAGLN2-, SERPINB6-, BACH2-, ZYX-, MBP-, CALN1- as well as in the MDS1 locus. Furthermore we detected 1 integration in the LMO2 locus. None of the integration clones was detected at more than 3 time points analysed. Our results show that the hematopoietic repopulation of two successful treated WAS patients stays polyclonal more than 1 year after treatment without the apparent presence of dominant clones. Analysis of the 454 pyrosequencing results will give us deeper insights into the clonal inventory of the hematopoietic repopulation of these patients.