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Current status of CF gene therapy

Gene therapy for the treatment of the pulmonary manifestations of cystic fibrosis (CF) has been at the forefront of gene therapy research over the last decade. During this time, however, despite immense efforts, controlled clinical trials with CF patients have failed to demonstrate significant and reproducible “correction” of the CF bioelectrical functional defect. The target tissue requiring “correction” in CF lung disease is the respiratory epithelium that lines the airways of the lung, and evidence is now emerging that the epithelium has evolved to elude the uptake of potential pathogens, including viruses, bacteria, and gene transfer vectors. The majority of studies aimed at gene transfer to the airway epithelium have used the adenovirus (Ad) as the gene delivery vector, since high-efficiency gene transfer to airway epithelial cells grown in culture can be demonstrated. However, when these vectors are tested in the airways of animals and humans in vivo, the efficiency of gene transfer is low. The factors responsible for the discrepancies between in vitro and in vivo gene transfer efficiency with Ad are now being elucidated and for the lung it is becoming apparent that several barriers to gene transfer exist. Identification of extracellular barriers to gene transfer (mucus, glycocalyx, absence of viral receptors) has indicated these barriers as rate-limiting for successful gene transfer to the airway epithelium and has subsequently led to the development of strategies that may be able to test the hypothesis that “corrective gene transfer” to the lungs of CF patients may one day be a valid approach to the treatment of this devastating disease.

The functional repertoire of the human genome is amplified by the differential assortment of exons. Spliceosome-mediated RNA trans-splicing can mobilize these packets of genetic information to reprogram mRNAs. In principle, this process could repair defective transcripts in loss-of-function genetic disorders in humans. We developed a tractable lacZ repair system to serve as a model for these genetic disorders. Targeted pre-trans-splicing RNA molecules efficiently and specifically repaired mutated lacZ transcripts and restored enzymatic activity in human cells. The development of this model confirms the potential for spliceosome-mediated RNA trans-splicing in genetic repairs and provides a powerful tool for rational design and in vitro evolution of pre-trans-splicing molecules.

Soluble factors normally produced by cells of the human body are of increasing importance as potential therapeutic agents. Although considerable progress has been made in understanding the etiology and pathogenesis of disease, in developing animal models and newer experimental therapeutics, few discoveries have been translated into clinically effective ways of delivering the multiple therapeutic agents obtained from living mammalian cells. This review examines the use of transplanted cells as alternatives to conventional delivery systems to deliver a variety of protein based therapeutic agents. The chapter begins with a set of questions to establish the complexity and challenges of this form of drug delivery. The following section focuses the discussion on our understanding of genetic engineering, tissue engineering, and some areas of developmental biology as they relate to the development of this nascent field. Much of the discussion has a neuro/endocrine emphasis. The chapter ends by listing the basic ingredients needed to push the use of transplanted cells toward medical practice and some general comments about future developments.

Gene therapy involves the introduction of normal, healthy genes into cells to correct the underlying cause of a wide variety of inherited and acquired diseases. Future progress in developing effective clinical protocols involving gene therapy for the treatment of cellular dysfunction associated with disease may incorporate metabolic engineering. Metabolic engineering can be applied to gene therapy for the successful identification of disease genes; elucidation of disease pathways; development of safe and efficient gene-delivery systems; and regulation and control of gene expression. Cystic fibrosis, cancer, and diabetes are reviewed as examples of diseases where gene therapy approaches are being studied.

The goal of this article was to review the status of gene therapy involving cystic fibrosis (CF), particularly the progress that has been made over the past decade in adenovirus (Ad)-mediated CF lung disease.

CF is theoretically an ideal disease to use in the study of gene therapy. Over the past decade, considerable progress has been made, with the path from CF gene discovery to clinical application being cleared. Clinical trials, mainly using Ad vectors, have been conducted, involving>200 patients with CF.

The results of the past decade of study have been both encouraging and frustrating. Good evidence exists that there are low levels of CF transmembrane conductance regulator (CFTR) gene transfer and limited evidence of small changes in ion transport. Although Ad vectors are the most advanced method for CF gene transfer to human airways, the current vectors are not yet suitable for in vivo application. The main problem involves the inefficiency of the Ad vectors when they are applied to the apical surface of the airway epithelia. Therefore, further applications of current Ad vectors in gene therapy in vivo will require improvements in efficiency and minimization of the toxicity/inflammatory responses in patients with CF. Although the efficiency of gene transfer is not sufficient to correct the functional defect of CF airways, many strategies to improve this gene transfer are being explored.

Adenovirus infection requires that the virus attach to cells and be internalized. Interaction between the viral fiber protein and specific cell surface receptors, such as the 46-kDa coxsackievirus and adenovirus receptor (CAR), is responsible for attachment; a second interaction between the viral penton base and cell surface integrins facilitates virus internalization. Expression of receptors may determine whether tissues are susceptible to adenovirus infection and adenovirus-mediated gene delivery.