Biomedical subjects

Gerard G M D'Souza

Publications and source records attributed to Gerard G M D'Souza.

8 recordsLinked to original sources

PubMed2006

Gene therapy of the other genome: the challenges of treating mitochondrial DNA defects.

Human mitochondrial DNA is a 16.5 kb circular DNA molecule located inside the mitochondrial matrix. Although accounting for only about 1% of total cellular DNA, defects in mitochondrial DNA have been found to have major effects on human health. A single mtDNA mutation may cause a bewildering variety of clinical symptoms mainly involving the neuromuscular system at any age of onset. Despite significant advances in the understanding of mitochondrial DNA defects at a molecular level, the clinical diagnosis of mtDNA diseases remains a significant challenge and effective therapies for such diseases are as yet unavailable. In contrast to gene therapy for chromosomal DNA defects, mitochondrial gene therapy is a field that is still in its infancy and attempts towards gene therapy of the mitochondrial genome are rare. In this review we outline what we believe are the unique challenges associated with the correction of mtDNA mutations and summarize current approaches to gene therapy for the "other genome".

Animals↗

PubMed2006

Liposomes and liposome-like vesicles for drug and DNA delivery to mitochondria.

Mitochondrial research is presently one of the fastest growing disciplines in biomedicine. Since the early 1990s, it has become increasingly evident that mitochondrial dysfunction contributes to a large variety of human disorders, ranging from neurodegenerative and neuromuscular diseases, obesity, and diabetes to ischemia-reperfusion injury and cancer. Most remarkably, mitochondria, the "power house" of the cell, have also become accepted as the "motor of cell death" reflecting their recognized key role during apoptosis. Based on these recent exciting developments in mitochondrial research, increasing pharmacological efforts have been made leading to the emergence of "Mitochondrial Medicine" as a whole new field of biomedical research. The identification of molecular mitochondrial drug targets in combination with the development of methods for selectively delivering biologically active molecules to the site of mitochondria will eventually launch a multitude of new therapies for the treatment of mitochondria-related diseases, which are based either on the selective protection, repair, or eradication of cells. Yet, while tremendous efforts are being undertaken to identify new mitochondrial drugs and drug targets, the development of mitochondria-specific drug carrier systems is lagging behind. To ensure a high efficiency of current and future mitochondrial therapeutics, colloidal vectors, i.e., delivery systems, need to be developed able to selectively transport biologically active molecules to and into mitochondria within living human cells. Here we review ongoing efforts in our laboratory directed toward the development of different phospholipid- and non-phospholipid-based mitochondriotropic drug carrier systems.

DNA↗

PubMed2005

Mitochondrial leader sequence--plasmid DNA conjugates delivered into mammalian cells by DQAsomes co-localize with mitochondria.

In the last decade the increase in therapeutic strategies aimed at mitochondrial targets has resulted in the need for novel delivery systems for the selective delivery of drugs and DNA into mitochondria. In this study, we have continued our efforts towards the development of the first mitochondriotropic drug and DNA delivery system (DQAsomes). Prepared from derivatives of the self-assembling mitochondriotropic bola-amphiphile dequalinium chloride, these vesicles bind and transport DNA to mitochondria in living mammalian cells where upon they have been shown to release the DNA on contact with mitochondrial membranes. We present data to demonstrate that oligonucleotides as well as plasmid DNA conjugated to a mitochondrial leader sequence (MLS) co-localize with mitochondria when delivered into mammalian cells by DQAsomes. In contrast to a commercially available DNA delivery vector, our vesicles appear to have a pronounced specificity for mitochondria. Further, the data strongly suggest that linear conjugates might be better suited to delivery into mitochondria and that in the absence of a mitochondria specific vector, the presence of a MLS-peptide conjugated to the DNA is alone not sufficient to direct the accumulation of DNA at mitochondria.

Cell Line, Tumor↗

PubMed2005

Mitochondriotropic liposomes.

Mitochondrial dysfunction contributes to a large variety of human disorders, ranging from neurodegenerative and neuromuscular diseases, obesity, and diabetes to ischemia-reperfusion injury and cancer. Increasing pharmacological efforts toward therapeutic interventions have been made leading to the emergence of "Mitochondrial Medicine" as a new field of biomedical research. The identification of molecular mitochondrial drug targets in combination with the development of methods for selectively delivering biologically active molecules to the site of mitochondria will eventually launch a multitude of new therapies for the treatment of mitochondria-related diseases, which are based either on the selective protection, repair, or eradication of cells. Yet, while tremendous efforts are being undertaken to identify new mitochondrial drugs and drug targets, the development of mitochondria-specific drug carrier systems is lagging behind. To ensure a high efficiency of current and future mitochondrial therapeutics, delivery systems need to be developed, which are able to selectively transport biologically active molecules to and into mitochondria within living human cells. In this study we present the first data demonstrating that conventional liposomes can be rendered mitochondria-specific via the attachment of known mitochondriotropic residues to the liposomal surface.

Bromides↗

PubMed2004

Mitochondrial pharmaceutics.

Since the end of the 1980s, key discoveries have been made which have significantly revived the scientific interest in a cell organelle, which has been studied continuously and with steady success for the last 100 years. It has become increasingly evident that mitochondrial dysfunction contributes to a variety of human disorders, ranging from neurodegenerative and neuromuscular diseases, obesity, and diabetes to ischemia-reperfusion injury and cancer. Moreover, since the middle of the 1990s, mitochondria, the 'power house' of the cell, have also become accepted as the cell's 'arsenals' reflecting their increasingly acknowledged key role during apoptosis. Based on these recent developments in mitochondrial research, increased pharmacological and pharmaceutical efforts have lead to the emergence of 'Mitochondrial Medicine' as a whole new field of biomedical research. Targeting of biologically active molecules to mitochondria in living cells will open up avenues for manipulating mitochondrial functions, which may result in the selective protection, repair or eradication of cells. This review gives a brief synopsis over current strategies of mitochondrial targeting and their possible therapeutic applications.

Journal Article↗

PubMed2004

Approaches to mitochondrial gene therapy.

Since their discovery during the end of the 80's the number of diseases found to be associated with defects in the mitochondrial genome has grown significantly. Organs affected by mutations in mitochondrial DNA (mtDNA) include in decreasing order of vulnerability the brain, skeletal muscle, heart, kidney and liver. Hence neuromuscular and neurodegenerative diseases represent the two largest groups of mtDNA diseases. Despite major advances in understanding mtDNA defects at the genetic and biochemical level, there is however no satisfactory treatment available to the vast majority of patients. This is largely due to the fact that most of these patients have respiratory chain defects, i.e. defects that involve the final common pathway of oxidative metabolism, making it impossible to bypass the defect by administering alternative metabolic carriers of energy. Conventional biochemical treatment having reached an impasse, the exploration of gene therapeutic approaches for patients with mtDNA defects is warranted. For now mitochondrial gene therapy appears to be only theoretical and speculative. Any possibility for gene replacement is dependent on the development of an efficient mitochondrial transfection vector. In this review we describe the current state of the development of mitochondria-specific DNA delivery systems. We summarize our own efforts in exploring the properties of dequalinium and other similar cationic bolaamphiphiles with delocalized charge centers, for the design of a vector suited for the transport of DNA to mitochondria in living cells. Further, we outline some unique hurdles that need to be overcome if the development of such delivery systems is to progress.

DNA, Mitochondrial↗

PubMed2003

DQAsome-mediated delivery of plasmid DNA toward mitochondria in living cells.

DQAsomes are mitochondriotropic cationic 'bola-lipid'-based vesicles, which have been developed by us for the transport of drugs and DNA to mitochondria in living cells. This has made direct mitochondrial gene therapy feasible for the very first time. Our strategy for the delivery of DNA into the matrix of mitochondria is based upon the DQAsomal transport of a DNA-signal peptide conjugate to mitochondria, the selective liberation of this conjugate from DQAsomes at the mitochondrial membrane followed by DNA uptake via the mitochondrial protein import machinery. Using membrane-mimicking liposomes and isolated rat liver mitochondria we have shown earlier that DQAsome-DNA complexes (DQAplexes) selectively release pDNA when in contact with mitochondria-like membranes. Employing a newly developed protocol for selectively staining free pDNA in the cytosol of living cells and based on confocal fluorescence microscopic imaging we demonstrate here that DQAplexes appear to be able to escape from endosomes without loosing their pDNA load and transport the pDNA to the site of mitochondria at which at least a portion of the pDNA is released from its DQAsomal carrier. Free pDNA could not be detected anywhere else inside the cytosol of transfected cells demonstrating the target-selectivity of DQAsome-mediated DNA delivery to mitochondria.

Animals↗

PubMed2003

Cell transfection in vitro and in vivo with nontoxic TAT peptide-liposome-DNA complexes.

Liposomes modified with TAT peptide (TATp-liposomes) showed fast and efficient translocation into the cell cytoplasm with subsequent migration into the perinuclear zone. TATp-liposomes containing a small quantity (<or=10 mol %) of a cationic lipid formed firm noncovalent complexes with DNA. Here, we present results demonstrating both in vitro and in vivo transfection with TATp-liposome-DNA complexes. Mouse NIH/3T3 fibroblasts and rat H9C2 cardiomyocytes were transfected with such complexes in vitro. The transfection with the TATp-liposome-associated pEGFP-N1 plasmid encoding for the green fluorescent protein (GFP) was high, whereas the cytotoxicity was lower than that of commonly used cationic lipid-based gene-delivery systems. Intratumoral injection of TATp-liposome-DNA complexes into the Lewis lung carcinoma tumor of mice also resulted in an expression of GFP in tumor cells. This transfection system should be useful for various protocols of cell treatment in vitro or ex vivo as well as for localized in vivo gene therapy.

3T3 Cells↗