Monogenetic diseases have already been successfully treated by gene therapy using retroviral vectors. By now, several diseases like X-linked severe combined immunodeficiency (X-SCID), adenosine deaminase (ADA)-deficient SCID, X-linked chronic granulomatous disease (X-CGD) and X-linked Wiskott Aldrich Syndrome (X-WAS) have been treated successfully. However, with the enhancement of retroviral gene transfer efficiency the occurrence of side effects increased. It was shown that the insertion of functional retrovirus LTRs can lead to insertional mutagenesis. By now, severe side effects have exclusively been observed in clinical trials using gamma-retroviral vectors as gene delivery vehicles. Individual patients enrolled in clinical gene therapy trials for X-SCID, X-CGD and X-WAS suffered from leukemia or myelodysplatic syndrome. For all reported cases of severe side effects, it was shown that vector-induced overexpression of nearby oncogenes (LMO2, CCND2, BMI1, MDS1-EVI1), was contributing as one step toward the malign transformation of individual cell clones. The occurrence of severe side effects highlighted the need for safer vectors systems and lead to the development HIV-1 based lentiviral vectors with self-inactivating (SIN) configuration. This vectors are a promising alternative to LTR-driven gamma-retroviral vectors, showing a reduced genotoxic risk in preclinical testing’s and so far no severe adverse events have been reported from clinical trials. We have performed an in depth integration site analysis of patient samples from the first clinical trial to treat a monogenetic cerebral disease using autologous hematopoietic stem cell transplantation with a HIV-1 based lentiviral SIN-vector. The cerebral disease has been stabilized in 2 out of 4 patients, the follow-up being too short in the 4th patient to draw any conclusion of clinical efficacy. So far, the correction of hematopoietic stem cell has not been accompanied by signs of clonal dominance or even premalignant disproportional distribution of cellular contributions in the 4 treated patients. Our large scale vector integration sites (IS) analysis performed on ex vivo transduced cells prior to reinfusion and on engrafted cells by LAM-PCR and subsequent 454 pyrosequencing showed a polyclonal hematopoietic reconstitution for samples from the first four patients treated by now. Downstream bioinformatics analysis of retrieved raw sequence data revealed the characteristic insertion profile reported for lentiviral vectors, showing gene coding regions as preferred targets for lentiviral vector integration (P1: 74%; P2: 74%; P3: 72%; P4: 71%), in line with a favored integration on chromosomes harboring gene dense regions. A successful ex vivo transduction of early hematopoietic progenitors is indicated by the presence of identical IS identified in myeloid and lymphoid lineages in P1, P2 and P3 (analysis is ongoing for P4). Interestingly, a preference of lentiviral vector insertion for particular genes has been observed by the identification of lentiviral vector integration sites as common integration sites in the same genes or genomic regions, observed in all four patients. Among others, the following genes were targeted by the vector: KDM2A, HLA and PACS1. High throughput distribution analysis of the IS repertoire indicates that lentivirus vectors offer great promise for safe and effective correction of human stem and progenitor cells.