Novel therapies for the treatment of cystic fibrosis: new developments in gene and stem cell therapy

Cystic fibrosis (CF) was one of the first target diseases for lung gene therapy. Studies of lung gene transfer for CF have provided many insights into the necessary components of successful gene therapy for lung diseases. Many advancements have been achieved with promising results in vitro and in small animal models. However, studies in primate models and patients have been discouraging despite a large number of clinical trials. This reflects a number of obstacles to successful, sustained, and repeatable gene transfer in the lung. Cell-based therapy with embryonic stem cells and adult stem cells (bone marrow or cord blood), have been investigated recently and may provide a viable therapeutic approach in the future. In this article, the authors review CF pathophysiology with a focus on specific targets in the lung epithelium for gene transfer and summarize the current status and future directions of gene- and cell-based therapies.     

Gene therapy for lung disease: hype or hope?

Gene therapy, the treatment of any disorder or pathophysiologic state on the basis of the transfer of genetic information, was a high-priority goal in the 1990s. The lung is a major target of gene therapy for genetic disorders, such as cystic fibrosis and alpha1-antitrypsin deficiency, and for other diseases, including lung cancer, malignant mesothelioma, pulmonary inflammation, surfactant deficiency, and pulmonary hypertension. This paper examines general concepts in gene therapy, summarizes the results of published clinical trials, and highlights areas of research aimed at overcoming challenges in the field. Although progress has been slower than anticipated, gene transfer has been safely achieved in patients with lung diseases. Recent advancements in understanding of the molecular basis of lung disease and the development of improved vector systems make it likely that gene therapy will be an important tool for the 21st-century clinician.     

Flexible bronchoscopy in molecular biology.

Flexible fiberoptic bronchoscopy has allowed researchers to use the bench to bedside approach in the study and therapy of lung diseases. Through bronchoscopy, the lung is a relatively convenient source of samples for the direct evaluation of human gene expression and function. Sampling of respiratory epithelium is performed by brushing with a cytology brush, whereas the epithelial lining fluid and the inflammatory cells in the bronchoalveolar space are obtained by bronchoalveolar lavage. Furthermore, bronchoscopy has been a cornerstone essential to gene therapy trials for lung disease.     

Gene therapy for inherited lung disorders: an insight into pulmonary defence

This review summarizes the latest developments in viral and nonviral gene delivery systems to the lung, and the problems that have to be overcome. Gene delivery has the potential to offer effective treatment to patients with life-threatening lung diseases such as cystic fibrosis and alpha(1)-antitrypsin deficiency, and could modify gene-environment relationships in asthma and other respiratory diseases. Phase I clinical trials conducted in the early 1990s showed that in principle gene transfer to the lung was safe. Although the preliminary results gave encouraging laboratory data, gene expression from viral or nonviral gene delivery systems was too inefficient or transient to offer clinical benefit. Initial optimism gave way to the realization that gene therapy to the lung was unlikely to be straightforward. The host innate and acquired immune system, which protects against infection from inhaled bacteria and viruses, represents a major barrier to successful gene transfer to the lung. A better understanding of the immunological barriers which exist in the lung may allow the development of pharmacological and/or immunological agents that modulate the host immune system to allow for a more continuous and regulated level of gene expression following gene transfer.     

Gene therapy for lung neoplasms

Both advanced-stage lung cancer and malignant pleural mesothelioma are associated with a poor prognosis. Advances in treatment regimens for both diseases have had only a modest effect on their progressive course. Gene therapy for thoracic malignancies represents a novel therapeutic approach and has been evaluated in several clinical trials. Strategies have included induction of apoptosis, tumor suppressor gene replacement, suicide gene expression, cytokine-based therapy, various vaccination approaches, and adoptive transfer of modified immune cells. This review considers the clinical results, limitations, and future directions of gene therapy trials for thoracic malignancies.     

Prospects for gene therapy in acute lung injury.

Gene transfection is an extremely useful tool with which to explore mechanisms of gene regulation, to examine the molecular physiology of proteins in cultured cells, and to produce transgenic organisms. Several transfection techniques that employ viral and nonviral vectors have been used successfully to transfect functional genes into lung cells in vivo. This report reviews some of the gene transfection techniques that have been applied to the intact lung, an organ that offers unique challenges and opportunities. Results indicate that somatic cell gene therapy is feasible and that gene-based therapies can be developed for acute and chronic lung diseases.     

hnRNP B1小干扰RNA在肺癌诊治中的研究进展

核内不均一核糖核蛋白B1(hnRNP B1)在肺癌各种组织类型中均有表达,是诊断早期肺癌灵敏度很高的指标.小干扰RNA在细胞内能介导RNA干扰效应,识别特异性mRNA,沉默同源基因表达,成功阻断RNA的表达,由mRNA编码的与疾病有关的任何蛋白都可以被小干扰RNA选择性清除.hnRNP B1特异性的小干扰RNA在体内可以稳定的表达,并且能特异、高效地抑制肺癌细胞hnRNP B1基因的表达,有望成为肺癌基因治疗的合理策略之一.     

腺相关病毒载体在肺疾病基因治疗中的应用

腺相关病毒（adeno-associated virus,AAV）载体具有安全性好、免疫原性低、能感染分裂细胞和非分裂细胞、能介导基因的长期稳定表达等优点。因此,作为一种基因导入系统,重组AAV载体在基因治疗的研究和开发中受到越来越多的关注和重视。本文着重就重组AAV载体在肺部疾病基因治疗中的应用以及改进重组AAV载体的策略作简要综述。     

Gene therapy for lung neoplasms

The field of gene therapy is still in its infancy, but significant accomplishments have been achieved. The ability to transfer genes safely and successfully into animals and patients clearly has been established. It is highly likely that in the near future, gene therapy will be shown to have clear therapeutic efficacy in diseases such as the treatment of hemophilia (using adeno-associated virus vectors) and the stimulation of angiogenesis in peripheral vascular disease and myocardial ischemia. Although only Phase 1 cancer gene therapy trials for thoracic malignancy have been conducted (usually in patients with large tumor burdens and at submaximal doses), there are some hints of efficacy at higher doses of vector in trials for localized malignancy. The studies reviewed in this article demonstrate the first attempts to use gene therapy vectors for lung cancer and mesothelioma. Although none of the diseases studied was "cured," valuable lessons have been learned from these trials, especially in defining the challenges of relatively inefficient and transient delivery of transgene in vivo. Using this knowledge, the second phase of gene therapy research has begun, with a strong focus on developing improved vector technology. Given the progress so far, there is little doubt that gene therapy will become a key approach for the treatment of thoracic malignancies in the near future.     

Aerosol gene delivery in vivo.

The ability to express transgenes selectively within the lung will greatly facilitate the development of gene therapy for a variety of human diseases. We have demonstrated that aerosol administration of a chloramphenicol acetyltransferase (CAT) expression plasmid complexed to cationic liposomes produces high-level, lung-specific CAT gene expression in mice in vivo. Significant levels of CAT activity are seen in the lungs for at least 21 days following aerosolization. In situ immunostaining for intracellular CAT protein reveals that the majority of airway epithelial and alveolar lining cells are transfected in vivo. Histological analyses show no apparent treatment-related damage. These results have important implications for the development of human gene therapy.     

Adeno-associated virus and lentivirus pseudotypes for lung-directed gene therapy

The enthusiasm for cystic fibrosis gene therapy that attended the initial cloning of the gene and in vitro correction of the genetic defect eventually diminished as we learned more about the limitations of vector technologies that were available in the 1980s and 1990s. Substantial progress has been made, however, over the last 5 years in developing second- and third-generation vector constructs that should be more useful in achieving gene transfer to the lung for the treatment of pulmonary diseases such as cystic fibrosis.     

The status of gene therapy for cystic fibrosis

Cystic fibrosis (CF) has been a primary focus for gene therapy of lung diseases because the genetic cause is known and the airway epithelium is accessible for direct deoxyribonucleic acid (DNA) delivery. Soon after the mutated gene was identified in 1989, investigators demonstrated that transfer of a normal copy of the CF gene corrected ion transport abnormalities, thus validating the potential for use of gene therapy for this autosomal recessive disease. However, subsequent studies in a variety of in vitro and animal models, and more limited human studies, have revealed several obstacles to gene therapy for CF: (1) The incomplete understanding of CF lung disease pathogenesis, particularly the relative importance of ion transport and other cellular abnormalities (including glycoconjugate processing, pH regulation of intracellular organelles, and membrane trafficking), and of surface epithelial versus submucosal gland CF transmembrane regulator (CFTR) expression, generates uncertainty as to the necessary target cells for gene transfer and the optimum end point(s) for short-term human studies. (2) The airway epithelium has protective barriers against viral infection that impair gene transfer with several vectors, including recombinant viruses and DNA conjugates. Improvement in DNA transfer technology will be necessary for successful gene therapy. (3) Immune responses to recombinant viruses and inflammatory effects of bacterial DNA are only partially understood and appear to limit efficacy, particularly with repeated administration. Identification of these obstacles is prerequisite for progress, and recent studies with novel DNA delivery methods appear promising.     

Application of molecular biology to pulmonary disease

The techniques of molecular biology now make it possible to clone specific genes, determine the nature of their molecular message, produce their protein product, and study their function in health and disease. DNA probes, particularly those for ribosomal RNA, offer a new way for the diagnosis of infectious diseases affecting the lung, particularly TB. In addition, recombinant DNA libraries of mycobacteria can be used to isolate mycobacterial antigens recognized by patients with TB. This may allow development of better immunologic tests and vaccines. A specific chromosomal abnormality of human chromosome 3 has been found in small cell lung cancer. It is hypothesized that loss of genes from this region may play a role in the pathogenesis of lung cancer. Another important factor in development of the disease is the expression of cancer-associated oncogenes. In addition to insights into the biology of lung cancer, these oncogenes might provide a method to classify various types of lung cancer and predict response to therapy. Specialized DNA markers known as RFLPs have now been linked with CF. This has resulted in localization of the CF gene to human chromosome 7 and the detection of the gene in most of its carriers who have been studied. Knowing where the gene resides and use of techniques of genetic engineering will eventually allow isolation of the CF gene (or genes) on chromosome 11 and determination of the biochemical defect for which it codes. Similarly, the gene for human alpha 1-antitrypsin has also been cloned. A practical benefit is the production of normal and mutant enzyme for replacement therapy in patients.     

Lung transplantation in interstitial lung disease

Interstitial lung disease is a heterogeneous group of illnesses, some of which may progress to a fibrosing stage and cause respiratory failure. For selected candidates, lung transplantation is the ultimate therapeutic option. We review data on lung transplantation for various interstitial lung diseases. We address indications, procedures, and outcomes for patients undergoing transplantation. Unique issues affecting morbidity, mortality, and recurrence of disease are discussed. We review the literature of transplantation for specific interstitial lung diseases and the outcomes of transplantation for interstitial lung diseases. Candidates with idiopathic pulmonary fibrosis experience high mortality on the waiting list, but derive significant survival benefit from lung transplantation. Recurrence is reported for several interstitial lung diseases after lung transplantation. Survival with lung transplantation for interstitial lung diseases is comparable with that attained in recipients with other indications. Lung transplantation is a well-tolerated, effective therapy for respiratory failure in interstitial lung disease.     

肺癌的自杀基因放射导向治疗实验研究

目的：利用放射敏感性调控序列诱导单纯疱疹病毒胸苷激酶（HSV－TK）基因的肿瘤靶向表达，以提高肺癌自杀基因治疗的选择性和有效性。方法：利用基因重组构建放射可调控的HSV－TK表达载体；转染肺癌A549细胞系，观察γ线照射后细胞HSV－TK表达，以及丙氧鸟苷（GCV）作用下细胞相对存活率的变化。利用裸鼠肺癌模型观察不同处理后的抑瘤效应。结果：γ线可诱导HSV－TK在被转染肺癌细胞表达显增强，呈剂量依赖性；经放射诱导后，转染细胞对GCV的敏感性明显升高（P＜0．05），细胞生工活性显受抑；放射联合GCV可明显抑制感染tgEgr-HyTK的裸鼠移植瘤，并可使50％的肿瘤完全消退。结论：由辐射敏感性启动子诱导的自杀基因肿瘤靶向性表达，是一种高产、安全的肺癌基因治疗新策略。     

Expression profiling in granulomatous lung disease

Granulomatous lung diseases, such as sarcoidosis, hypersensitivity pneumonitis, Wegener's granulomatosis, and chronic beryllium disease, along with granulomatous diseases of known infectious etiologies, such as tuberculosis, are major causes of morbidity and mortality throughout the world. Clinical manifestations of these diseases are highly heterogeneous, and the determinants of disease susceptibility and clinical course (e.g., resolution vs. chronic, progressive fibrosis) are largely unknown. The underlying pathogenic mechanisms of these diseases also remain poorly understood. Within this context, these diseases have been approached using genomic and proteomic technologies to allow us to identify patterns of gene/protein expression that track with clinical disease or to identify new pathways involved in disease pathogenesis. The results from these initial studies highlight the potential for these "-omics" approaches to reveal novel insights into the pathogenesis of granulomatous lung disease and provide new tools to improve diagnosis, clinical classification, course prediction, and response to therapy. Realizing this potential will require collaboration among multidisciplinary groups with expertise in the respective technologies, bioinformatics, and clinical medicine for these complex diseases.     

精准医学概念在肺癌靶向治疗中的应用

精准医学是临床转化医学的组成部分,是对医学的进一步理解,其服务于患者,有助于健康医疗,基于肺癌驱动基因指导下的肺癌靶向治疗是精准治疗的重要部分.文中就精准医学概念在肺癌靶向治疗中的应用作一综述,以肺癌靶向治疗研究进展为例,阐述对精准医学的理解,并展望肺癌靶向治疗在临床上的前景.     

Cell-based therapies for lung vascular diseases: lessons for the future

Pulmonary arterial hypertension or acute respiratory distress syndrome (ARDS) represent diseases caused in large part by lung endothelial injury and inflammation. Pulmonary arterial hypertension is a progressive and debilitating lung disorder, which is characterized by hallmark pathological features in the small peripheral pulmonary arteries, including intimal hyperplasia and fibrosis, medial hypertrophy, increased deposition of extracellular matrix, infiltration of inflammatory mediators, and in many instances of advanced disease, plexiform lesions. ARDS is a clinically important complication of severe acute lung injury and a significant cause of morbidity and mortality in critically ill patients. Acute lung injury/ARDS is characterized by breakdown of the air-blood barrier, with the accumulation of proteinaceous fluid and influx of inflammatory cells into the alveolar airspace, all of which are likely mediated by endothelial dysfunction and disruption of the pulmonary endothelial barrier. Preclinical studies suggest that cellular therapies may offer unique and effective alternatives for respiratory diseases that currently have limited (or no) treatment options. Encouraging results from studies using progenitor or mesenchymal stem cells suggest that these cells may exert their therapeutic benefits by promoting vascular repair and regeneration and/or by modulating the pathological immune responses. Furthermore, by combining genetic engineering with cell therapy, it may be possible to enhance the regenerative function of these cells, particularly in the context of adult autologous cell therapy, and therefore provide additional benefits that may overcome many of the limitations of cell or gene therapy alone.     

New perspectives for gene therapy in endocrinology

Gene therapy for endocrine diseases represents an exciting new type of molecular intervention that may be a curative one. Endocrine disorders that might be treated by gene therapy include monogenic diseases, such as GH deficiency and hypothalamic diabetes insipidus, and multifactorial diseases, such as diabetes mellitus, obesity and cancer. Premises seem promising for endocrine tumours, but many combined approaches of cell and gene therapy are foreseeable also for other endocrine disorders. This review outlines the principles of gene therapy, describes the endocrine disorders that might take advantage of gene transfer approaches, as well as the gene therapy interventions that have already been attempted, their major limitations and the problems that remain to be solved.