Biomedical subjects

Boris Calmels

Publications and source records attributed to Boris Calmels.

8 recordsLinked to original sources

PubMed2005

Recurrent retroviral vector integration at the Mds1/Evi1 locus in nonhuman primate hematopoietic cells.

Recent reports linking insertional activation of LMO2 following gene therapy for X-linked severe combined immunodeficiency (X-SCID) have led to a re-evaluation of risks following gene therapy with retroviral vectors. In our analysis of 702 integration sites in rhesus macaques that underwent transplantation up to 7 years earlier with autologous CD34+ cells transduced with amphotropic murine leukemia virus (MLV)-derived retroviral vectors containing marker genes, we detected insertion into one locus, the Mds1/Evi1 region, a total of 14 times in 9 animals. Mds1/Evi1 integrations were observed stably long term, primarily in myeloid cells. We hypothesize that this over-representation likely results from an impact on the self-renewal and engraftment potential of CD34+ progenitor cells via insertional mutagenesis at this specific locus. There is no evidence of ongoing in vivo clonal expansion of the Mds1/Evi1 populations, and all animals are hematologically normal without evidence for leukemia. Characterization of integration sites in this relevant preclinical model provides critical information for gene therapy risk assessment as well as identification of genes controlling hematopoiesis.

Animals↗

PubMed2005

Clinical experience with the delivery of thawed and washed autologous blood cells, with an automated closed fluid management device: CytoMate.

BACKGROUND: Washing out of thawed autologous grafts, before reinfusion in poor-prognosis cancer patients who undergo high-dose chemotherapy, is desirable. The procedure allows for the reduction of infused dimethyl sulfoxide (DMSO) quantities and the performance of biologic controls on the infused cell product. STUDY DESIGN AND METHODS: Three-hundred four patients were treated with intensified chemotherapy and autologous transplantation at a single institution. Fifty-four of them received washed cell products, because three or more bags were to be reinfused. The recently available, closed, automated, and current good manufacturing practice-compliant device (CytoMate, Baxter Oncology) was used for this purpose. RESULTS: The performances of the device were similar to previously reported results, with greater than 75 percent CD34+ cell recovery. Neutrophil and platelet (PLT) recoveries were similar in the group of patients receiving washed cells and in the group of patients for whom cell products were extemporaneously thawed at the bedside. Adverse events that are typically reported after DMSO infusion were significantly less frequent and less severe in patients who received washed cells. Finally, the nurse staff on the transplant ward reported a decreased workload and more satisfactory procedure when infusing washed cell products. CONCLUSION: The CytoMate device allows for a significant reduction in DMSO infusion, with a diminished frequency and severity of immediate side effects and does not compromise neutrophil or PLT engraftment.

Adult↗

PubMed2004

Distinct genomic integration of MLV and SIV vectors in primate hematopoietic stem and progenitor cells.

Murine leukemia virus (MLV)-derived vectors are widely used for hematopoietic stem cell (HSC) gene transfer, but lentiviral vectors such as the simian immunodeficiency virus (SIV) may allow higher efficiency transfer and better expression. Recent studies in cell lines have challenged the notion that retroviruses and retroviral vectors integrate randomly into their host genome. Medical applications using these vectors are aimed at HSCs, and thus large-scale comprehensive analysis of MLV and SIV integration in long-term repopulating HSCs is crucial to help develop improved integrating vectors. We studied integration sites in HSCs of rhesus monkeys that had been transplanted 6 mo to 6 y prior with MLV- or SIV-transduced CD34(+)cells. Unique MLV (491) and SIV (501) insertions were compared to a set of in silico-generated random integration sites. While MLV integrants were located predominantly around transcription start sites, SIV integrants strongly favored transcription units and gene-dense regions of the genome. These integration patterns suggest different mechanisms for integration as well as distinct safety implications for MLV versus SIV vectors.

Animals↗

PubMed2004

Low-dose total body irradiation causes clonal fluctuation of primate hematopoietic stem and progenitor cells.

Due to high frequency of side effects caused by high-dose total body irradiation (TBI) the nonmyeloablative regimen together with cytotoxic agents is currently used especially for elderly patients. However, immediate and long-term effects of low-dose irradiation used in allogeneic transplantation on stem cells is less well known. We have studied the effect of low-dose 3 Gy TBI on the number of hematopoietic stem cell (HSC) clones contributing simultaneously to granulocyte production in rhesus macaque. The number of clones after 3 Gy TBI decreased markedly by 2 to 3 weeks after 3 Gy TBI, followed by a period of clonal instability, and recovery to almost pre-3 Gy TBI clonal diversity. The clones accounting for this recovery contributed before 3 Gy TBI, suggesting the profound initial impact of TBI was on a pool of progenitor cells, whereas most of the more primitive HSCs remained unaffected and were able to again contribute to hematopoiesis after recovery. Clonal fluctuation may indirectly suggest the presence of short-term/long-term HSC populations in rhesus macaque bone marrow as reported in a mouse model. The results indicate that even low-dose irradiation affects hematopoietic clonal dynamics and have implications for design of conditioning regimens for transplantation purposes.

Animals↗

PubMed2004

Long-term clinical and molecular follow-up of large animals receiving retrovirally transduced stem and progenitor cells: no progression to clonal hematopoiesis or leukemia.

There has been significant progress toward clinically relevant levels of retroviral gene transfer into hematopoietic stem cells (HSC), and the therapeutic potential of HSC-based gene transfer has been convincingly demonstrated in children with severe combined immunodeficiency syndrome (SCID). However, the subsequent development of leukemia in two children with X-linked SCID who were apparently cured after transplantation of retrovirally corrected CD34+ cells has raised concerns regarding the safety of gene therapy approaches utilizing integrating vectors. Nonhuman primates and dogs represent the best available models for gene transfer safety and efficacy and are particularly valuable for evaluation of long-term effects. We have followed 42 rhesus macaques, 23 baboons, and 17 dogs with significant levels of gene transfer for a median of 3.5 years (range 1-7) after infusion of CD34+ cells transduced with retroviral vectors expressing marker or drug-resistance genes. None developed abnormal hematopoiesis or leukemia. Integration site analysis confirmed stable, polyclonal retrovirally marked hematopoiesis, without progression toward mono- or oligoclonality over time. These results suggest that retroviral integrations using replication-incompetent vectors, at copy numbers achieved using standard protocols, are unlikely to result in leukemogenesis and that patient- or transgene-specific factors most likely contributed to the occurrence of leukemia in the X-SCID gene therapy trial.

Animals↗

PubMed2002

The cleavage/polyadenylation activity triggered by a U-rich motif sequence is differently required depending on the poly(A) site location at either the first or last 3'-terminal exon of the 2'-5' oligo(A) synthetase gene.

Production of the two mRNAs encoding distinct forms of 2'-5'-oligoadenylate synthetase depends on processing that involves the recognition of alternative poly(A) sites and an internal 5'-splice site located within the first 3'-terminal exon. The resulting 1.6- and 1.8-kb mRNAs are expressed in fibroblast cell lines, whereas lymphoblastoid B cells, such as Daudi, produce only the 1.8-kb mRNA. In the present study, we have shown that the 3'-end processing at the last 3'-terminal exon occurs independently of the core poly(A) site sequence or the presence of regulatory elements. In contrast, in Daudi cells, the recognition of the poly(A) site at the first 3'-terminal exon is impaired because of an unfavorable sequence context. The 3'-end processing at this particular location requires a strong stabilization of the cleavage/polyadenylation factors, which can be achieved by the insertion of a 25-nucleotide long U-rich motif identified upstream of the last poly(A) site. Consequently, we speculate that in cells expressing the 1.6-kb mRNA, such as fibroblasts, direct or indirect participation of a specific mechanism or cell type-specific factors are required for an efficient polyadenylation at the first 3'-terminal exon.

2',5'-Oligoadenylate Synthetase↗

PubMed2002

Ikaros gene expression and leukemia.

The Ikaros (Ik) protein, or LyF1, was initially described as a protein binding to regulatory sequences of a number of genes expressed in murine lymphoid cells. Ikaros is a critical regulator of normal hematopoietic stem cell differentiation, as evidenced by dramatic defects in the lymphoid compartments, in homozygous animals with gene inactivation. Because differential splicing produces multiple isoforms with potentially different functions, Ikaros provides a unique model to study how post-transcriptional mechanisms may be involved in neoplastic processes. Indeed, several groups including ours have underlined evidences that expression of different Ikaros isoforms vary among different types of leukemias. The predominance of short isoforms in certain subsets is intriguing. Here, additional observations reinforced the hypothesis that Ikaros expression may be deregulated in human leukemias. Whether this is a cause or a consequence of the leukemic process remains speculative. Other human diseases however, provide examples of abnormal post-transcriptional regulations that have been further characterized.

Alternative Splicing↗

PubMedDate not supplied

[Immunotherapy of acute myeloid leukemias: development of vaccines and cell therapy approaches].

Prognosis of acute myeloblastic leukemias (AML) remains with relapse, at a time when progress in chemotherapy regimens have increased the rate of complete remission, and improved biological tools have helped in defining diagnostic and prognostic criteria for this heterogeneous group of diseases. Allogeneic stem cell transplantation offers an example of a situation in which various immune effectors can contribute to the eradication of residual leukemic cells. However, use of allogeneic transplantation is restricted to a minority of patients. Thus, the development of immunotherapy strategies that could be used for all patients, appears highly desirable. Progress in fundamental immunology, and a better understanding of the role of the immune system in the physiopathology of leukemias now open new avenues to design innovative therapies. We here review recently published observations in this field, and propose new vaccine programs that use autologous AML blasts differentiated into dendritic cells.

Acute Disease↗