Biomedical subjects

Mahmoud Abd El-Latif

Publications and source records attributed to Mahmoud Abd El-Latif.

4 recordsLinked to original sources

PubMed2005

Liver-targeted gene therapy by SV40-based vectors using the hydrodynamic injection method.

Efficient reconstitution of defective genes in hepatocytes could be used to treat various liver and systemic diseases through gene therapy. To explore the potential of SV40-based vectors in liver gene therapy, we constructed SV/luc, an SV40 T-antigen replacement transduction vector, that was propagated on COS and COT cells, which supply the SV40 T-antigen in trans. For liver targeting, BALB/C mice were injected via the tail vein with SV/luc stocks containing 3 x 10(6) to 10(8) transducing units in a volume of 1-2 ml. Luciferase activity was monitored with a light-detection cooled charged-coupled device (CCCD) camera, which enables continuous in vivo measurement of luc expression. The SV40 vector proved to be efficient in gene delivery to the liver, leading to long-term (> or =107 days) transgene expression in hepatocytes. Optimal results were obtained with 3 x 10(6) to 3 x 10(7) transducing units. The hydrodynamic vector delivery method caused transient liver inflammatory changes, with full recovery within days. Low levels of SV40-neutralizing antibodies were detected in the sera of treated mice; however, there was no indication of vector or transgene-specific cellular immune responses. Vectors packaged in vitro, using recombinant capsid proteins and plasmid DNA, were also effective in liver transduction. These results suggest that SV40 vectors may be useful for liver gene therapy.

Alanine Transaminase↗

PubMed2003

The abundant nuclear enzyme PARP participates in the life cycle of simian virus 40 and is stimulated by minor capsid protein VP3.

The abundant nuclear enzyme poly(ADP-ribose) polymerase (PARP) functions in DNA damage surveillance and repair and at the decision between apoptosis and necrosis. Here we show that PARP binds to simian virus 40 (SV40) capsid proteins VP1 and VP3. Furthermore, its enzymatic activity is stimulated by VP3 but not by VP1. Experiments with purified mutant proteins demonstrated that the PARP binding domain in VP3 is localized to the 35 carboxy-terminal amino acids, while a larger peptide of 49 amino acids was required for full stimulation of its activity. The addition of 3-aminobenzamide (3-AB), a known competitive inhibitor of PARP, demonstrated that PARP participates in the SV40 life cycle. The titer of SV40 propagated on CV-1 cells was reduced by 3-AB in a dose-dependent manner. Additional experiments showed that 3-AB did not affect viral DNA replication or capsid protein production. PARP did not modify the viral capsid proteins in in vitro poly(ADP-ribosylation) assays, implying that it does not affect SV40 infectivity. On the other hand, it greatly reduced the magnitude of the host cytopathic effects, a hallmark of SV40 infection. Additional experiments suggested that the stimulation of PARP activity by VP3 leads the infected cell to a necrotic pathway, characterized by the loss of membrane integrity, thus facilitating the release of mature SV40 virions from the cells. Our studies identified a novel function of the minor capsid protein VP3 in the recruitment of PARP for the SV40 lytic process.

Animals↗

PubMed2002

A new packaging cell line for SV40 vectors that eliminates the generation of T-antigen-positive, replication-competent recombinants.

Simian virus 40 (SV40) vectors are efficient vehicles for gene delivery to hematopoietic and hepatic cells. To ensure their replication incompetence and because of safety considerations, it is critical that the vectors do not contain T-antigen sequences. Available packaging cell lines for T-antigen replacement vectors, COS and CMT4, contain considerable sequence identity with the vectors, leading to homologous recombination and reacquisition of the T-antigen gene. We constructed a packaging cell line, COT18, with minimal sequence identity to the vector. Vector stocks produced by passaging on COT18 had high transducing activity and undetectable levels of T-antigen-positive, replication-competent contaminants. This cell line provides a means for the preparation of safe SV40 vector stocks.

Animals↗

PubMed2002

The beta+-IVS-I-6 (T-->C) mutation accounts for half of the thalassemia chromosomes in the Palestinian populations of the mountain regions.

A study of the spectrum of beta-thalassemia mutations in the southern part of the West Bank of the Palestinian Authority revealed the presence of 10 different beta-globin mutations. The study included 41 patients and 54 carriers of beta-thalassemia and sickle cell anemia. The spectrum of mutations observed was typically Mediterranean. However, their relative frequencies was unique. The predominant allele was IVS-I-6 (T-->C), with an exceptionally high frequency of 48.5% for this mutation. The homozygous IVS-I-6 patients had widely variable clinical presentations, from typical transfusion-dependent thalassemia major to non-transfusion-dependent thalassemia intermedia phenotype. Since it is so widespread in these West Bank populations, the IVS-I-6 mutation may date back to ancient times. The nonsense mutation at codon 37 (G-->A) was found at a relatively high frequency of 11.3%, supporting the hypothesis that it originated in this region. The other mutations, at decreasing frequencies ranging from 9.5-1.5%, were: IVS-I-110 (G-->A), frameshift codon 5 (- CT), IVS-I-1 (G-->A), IVS-II-1 (G-->A), Hb S [beta6(A3)Glu-->Val], frameshift codons 8/9 (+G), codon 39 (C-->T), and -30 (T-->A). Our findings will improve health care for the Palestinian population, and also has implications for the study of the origin and spread of thalassemia in the Middle East.

Adolescent↗