By Sean Henahan, Access Excellence

"This is encouraging news for people born with CGD," says NIAID Director Anthony S. Fauci, M.D. "While other therapeutic advances have improved the prognosis for CGD patients in recent years, the development of effective gene therapy would represent a big step forward."

The disease is caused by a defect in an enzyme called phagocyte NADPH oxidase, or phox. White blood cells use this enzyme to generate hydrogen peroxide, which the cells need to kill bacteria and fungi. Mutations in one of four different genes can cause this defect, which leads to frequent and often life-threatening infections of the skin, lungs and bones with localized, swollen collections of inflamed tissue called granulomas.

The researchers found that a new method of introducing genes resulted in prolonged production of genetically corrected cells. They removed stem cells, the  immune cells that form the basis of the immune system, from five patients with CGD. The researchers inserted the correct form of  the phox gene  into the stem cells and then transfused the corrected cells back into each patient. The researchers then sampled the patientsí blood periodically to see if the stem cells were producing white blood cells with functional phox genes.

"We detected phox activity in white blood cells from each patient for an average of three months after the gene-corrected stem cells were transfused. In one patient, phox activity was still present six months after transfusion. On average, the corrected phox gene was present in one out of every 5,000 cells. While the numbers of gene-corrected cells were small, the study demonstrates unequivocally that gene therapy of stem cells can produce functionally normal blood cells in patients for a prolonged period," said  Harry L. Malech, M.D., deputy chief of NIAIDís Laboratory of Host Defenses.

The primary benefit from the research could be an improvement in the treatment of  patients with CGD. "Since life-threatening infections caused by CGD may require many weeks or months of therapy and relapses are frequent, use of gene therapy to provide even short- to medium-term production of phox-positive cells may be clinically beneficial," says Dr. Malech.

Studies indicate that people having 3 to 5 percent phox-positive cells in their blood might be protected from infections associated with CGD, he notes. Although those levels are at least 150-fold higher than levels attained in the current study, Dr. Malech predicts that they might be achieved within the next five to 10 years.

"Until the tools are developed to achieve higher levels of permanent gene transfer to stem cells, our studies suggest that an achievable intermediate goal of gene therapy for CGD might be to augment white blood cell function in the treatment of severe infections," he added.

The techniques used in this study could also benefit gene therapy research in other areas. Designed to enhance the safety of gene therapy procedures involving stem cells, these techniques included the use of cell culture media containing no non-human proteins and a closed system of gas-permeable flexible plastic containers for culture and gene transfer.

The use of human-based culture medium is a key development. Animal proteins are widely used in most cell culture media. However, animal proteins taken up by human cells during prolonged culture can stimulate an immune response when the cells are transfused back into a patient undergoing gene therapy. Similarly, the closed system of  flexible plastic containers, similar to those used in blood banks, reduces the contamination risk associated with procedures where cells and culture media are transferred among flasks.

"To our knowledge, this is the first human gene therapy trial targeting stem cells in which animal proteins were eliminated and stem cells were grown in sealed gas-permeable flexible plastic containers," says Dr. Malech. "We showed that it is possible to incorporate these safety features without compromising stem cell viability or gene transfer efficiency."

CGD is one of more than 70 different inherited disorders known collectively as primary immune deficiencies. It is a rare disorder, affecting four to five of every million people. About 1,000 people in the United States have the disease.

The finding is reported in the Oct. 28, 1997 issue of the Proceedings of the National Academy of Sciences.