For nearly all successful gene therapy studies worldwide that have the aim of curing immunodeficiencies, the group surveyed the clonal composition of the hematopoietic system after transplantation. These studies also provided precious insight into the biology of stem cells, physiology of hematopoietic (and other tissue) regeneration and development of malignancies. Moreover, the integration site analysis, combined with next-generation sequencing and bioinformatic data management, serves as a highly valuable platform for more than 40 national and international collaborative projects that investigate molecular mechanisms underlying gene and molecular therapeutical studies.

As a partner of the Helmholtz-Alliance for Immunotherapy, the group is involved in a complex project that aims to generate tumor reactive cells by transfer of tumor-specific T cell receptor (TCR) genes. To assess the potential risks associated with TCR gene therapy, the clonal repertoire of T cell receptors was analyzed in preclinical models. In a next step, this strategy will be transferred to the clinical setting to analyze samples from patients undergoing T cell therapy.

• performed clonality and biosafety analyses in multiple preclinical gene therapy studies using low- and high-frequency integrating vectors (oncoretrovirus, lentivirus, adeno-associated virus) to monitor the clonal composition of post-transplant hematopoiesis (Blood 2007, 110:1779-87).
• monitored repopulation kinetics and clonal behavior of individual transduced cells in clinical gene therapy studies (Nat Med 2006, 12:401-9; J Clin Invest 2007, 117:2225-31; J Clin Invest 2007, 117:2241-9).
• analyzed and modeled the genome-wide distribution of common restriction enzyme(s) used for digestion of genomic DNA and devised theoretical (mathematical/computational) and practical (non-restrictive LAM-PCR) approaches to enable the comprehensive retrieval of vector insertion sites with maximum genomic access (Nat Med 2009, 15:1431-6).
• made its methodology for the identification of individual integration sites of viral gene transfer vectors in highly polyclonal samples applicable to clinical routine (Nat Meth 2007, 4:1051-7; Meth Mol Biol 2009, 506:363-72; Nat Protoc 2010, 5:1379-95).
• showed in a first application of lentiviral-mediated gene therapy in humans that hematopoietic stem cells can provide clinical benefit in patients with X-linked adrenoleukodystrophy (ALD) (Science 2009, 326:818-23).
• identified, in collaboration with other partners, molecular mechanisms in the pathogenesis of human myelodysplastic syndrom (MDS) following MDS1/EVI1 gene activation (Nat Med 2010, 16:198-204).
  

Schmidt M, Schwarzwaelder K, Bartholomae C, Zaoui K, Ball C, Pilz I, Braun S, Glimm H, von Kalle C: High-resolution insertion-site analysis by linear amplification-mediated PCR (LAM-PCR). Nat Methods 2007, 4:1051-7

Gabriel R, Eckenberg R, Paruzynski A, Bartholomae C, Nowrouzi A, Arens A, Howe SJ, Recchia A, Cattoglio C, Wang W, Faber K, Schwarzwaelder K, Kirsten R, Deichmann A, Ball CR, Balaggan KS, Yáñez-Muñoz RJ, Ali RR, Gaspar HB, Biasco L, Aiuti A, Cesana D, Montini E, Naldini L, Cohen-Haguenauer O, Mavilio F, Thrasher AJ, Glimm H, von Kalle C, Saurin W, Schmidt M: Comprehensive genomic access to vector integration in clinical gene therapy. Nat Med 2009, 15:1431-6

Boztug K, Schmidt M, Schwarzer A, Banerjee PP, Avedillo Díez I, Dewey R, Böhm M, Ball CR, Nowrouzi A, Naundorf S, Kühlcke K, Blasczyk R, Kondratenko I, Maródi L, Orange JS, von Kalle C, Klein C: Stem-cell gene therapy for the Wiskott-Aldrich syndrome. N Engl J Med 2010, 363:1918-27

Paruzynski A, Arens A, Gabriel R, Bartholomae CC, Scholz S, Wang W, Wolf S, Glimm H, Schmidt M, von Kalle C: Genome-wide high-throughput integrome analyses by nrLAM-PCR and next-generation sequencing. Nat Protoc 2010, 5:1379-95

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Email: manfred.schmidt@nct-heidelberg.de