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The gradual depletion of CD4+T lymphocytes during the development of AIDS may be due, at least in part, to a process referred to as apoptosis. This process involves a Ca2+dependent nuclear endonuclease that cleaves the chromatin at internucleosomal junctions. In addition, we have recently provided evidence that apoptosis may be responsible not only for the progressive loss of CD4+T lymphocytes but may be operative in CD8+T lymphocytes as well. Here, we describe mechanisms which by direct and indirect pathways may induce apoptosis during HIV infection and thus leading to elimination of T cells.

Le lichen plan érosif (LPE) est une maladie chronique dysimmunitaire rare et invalidante caractérisée par des lésions muqueuses inflammatoires érosives, avec une infiltration de lymphocytes T cytotoxiques CD8+. La spécificité de ces lymphocytes n’a jamais été analysée. Dans le but d’identifier les mécanismes moléculaires sous-jacents, nous avons étudié la diversité et la spécificité du récepteur à l’antigène (TCR) exprimé par les lymphocytes T CD8+par analyse en PCR quantitative, immunoscope, clonage et séquençage et marquage par dextramères, chez 10 patients atteints de LPE et traités par photochimiothérapie extracorporelle (PEC). Une expansion des lymphocytes T CD8+utilisant Vβ3 pour leur TCR a été identifiée dans le sang et dans les lésions muqueuses. Les études de séquences ont permis d’identifier des clonotypes spécifiques chez chaque patient. Ces expansions clonales étaient enrichies en lymphocytes T CD8+spécifiques du Human Papilloma Virus (HPV) 16 comme le démontrait la reconnaissance de l’épitope immunodominant E711–20 chez les patients HLA-A*0201+.Sous traitement par PEC, les expansions clonotypiques des lymphocytes T CD8+diminuaient quantitativement parallèlement à l’obtention de la rémission clinique. Ces résultats permettent d’établir un lien entre l’infection par HPV et le LPE en mettant en évidence une expansion clonale de lymphocytes T CD8+spécifiques de HPV16, évoluant parallèlement à l’évolution clinique de la maladie, et ouvrent de nouvelles perspectives thérapeutiques dans le LPE.

We have previously reported that ex vivo retroviraly-mediated gc gene transfer into CD34 (+) bone marrow precursor cells led to the correction of the immunodeficiency in 9 out of 10 patients with X-linked severe combined immunodeficiency. Follow-up now reaches more that 6 years for the first 2 treated patients. Patients’immune function has been restored. The distribution of both TCR Vb family usage and TCR Vb CDR3 length still reveals a broadly diversified T cell repertoire. Moreover 6 years after treatment the thymus is still seeded by transduced progenitor cells as attested by the presence of TRECS in peripheral blood RTE. Among these patients, three (P4, P5 and P10) developed at 30 to 34 months after gene therapy a monoclonal T cell proliferation requiring a chemotherapy. P4 received also an allogenic HSCT from a MUD but died 26 months after the occurence of the lymphoproliferation. For P5 and P10, chemotherapy has led to an overall control of the clonal proliferation. These two patients are doing well and P5 is off treatment with a good immunological recovery. Genetic analysis of the blastic cells showed that in the two first cases the vector had integrated within or upstream of the LMO2 locus causing an insertional activation of LMO2 transcription. The last case revealed the involvement of several targeted sites, but their exact contribution to the lymphoproliferation is still under investigation. The repeated involvment of LMO2 as a site of vector integration in the proliferating T-cells points to an insertional activation of this gene as at least one of the causes of the oncogenic process. However, the long latency observed in all cases (> 30 months) suggests that additional “hits” have been required for overt desease. Synergy with gc expression and thereby induced proliferative signals (explaining occurrence in SCID-X1 patients only) is the most obvious hypothesis which we are trying to analyse in a mouse model. A deep analysis of retroviral integration patterns has been performed on patients’PBMCS by LAM-PCR to estimate the frequency of potentially harmful integration events and to assess the risk factors associated with the LTR’s strong enhancer effect of the MLV-based retroviral vector. 708 unique integration sites (IS) have been obtained from all analysed patients post-gene therapy and among them, 577 could be mapped unequivocally to the human genome. *Most of these insertions (63%) are located in the vicinity of 10kb or within the coding sequence of a known gene*. A significant peak of insertion frequency is related closely to the transcription strart site *among the 577 IS, 43 are common integration sites. Among the latter, we found out a high selection of genes involved in human oncogenic process.

We have previously reported that ex vivo retroviraly-mediated gc gene transfer into CD34 (+) bone marrow precursor cells led to the correction of the immunodeficiency in 9 out of 10 patients with X-linked severe combined immunodeficiency. Follow-up now reaches more that 6 years for the first 2 treated patients. Patients'immune function has been restored. The distribution of both TCR Vb family usage and TCR Vb CDR3 length still reveals a broadly diversified T cell repertoire. Moreover 6 years after treatment the thymus is still seeded by transduced progenitor cells as attested by the presence of TRECS in peripheral blood RTE. Among these patients, three (P4, P5 and P10) developed at 30 to 34 months after gene therapy a monoclonal T cell proliferation requiring a chemotherapy. P4 received also an allogenic HSCT from a MUD but died 26 months after the occurence of the lymphoproliferation. For P5 and P10, chemotherapy has led to an overall control of the clonal proliferation. These two patients are doing well and P5 is off treatment with a good immunological recovery. Genetic analysis of the blastic cells showed that in the two first cases the vector had integrated within or upstream of the LMO2 locus causing an insertional activation of LMO2 transcription. The last case revealed the involvement of several targeted sites, but their exact contribution to the lymphoproliferation is still under investigation. The repeated involvment of LMO2 as a site of vector integration in the proliferating T-cells points to an insertional activation of this gene as at least one of the causes of the oncogenic process. However, the long latency observed in all cases (> 30 months) suggests that additional “hits” have been required for overt desease. Synergy with gc expression and thereby induced proliferative signals (explaining occurrence in SCID-X1 patients only) is the most obvious hypothesis which we are trying to analyse in a mouse model. A deep analysis of retroviral integration patterns has been performed on patients'PBMCS by LAM-PCR to estimate the frequency of potentially harmful integration events and to assess the risk factors associated with the LTR's strong enhancer effect of the MLV-based retroviral vector. 708 unique integration sites (IS) have been obtained from all analysed patients post-gene therapy and among them, 577 could be mapped unequivocally to the human genome. *Most of these insertions (63%) are located in the vicinity of 10kb or within the coding sequence of a known gene*. A significant peak of insertion frequency is related closely to the transcription strart site *among the 577 IS, 43 are common integration sites. Among the latter, we found out a high selection of genes involved in human oncogenic process.