Skin Like New

Repair of aging skin could become more than merely a cosmetic concern if current research fulfills its promise. Recent efforts to eliminate wrinkles and regenerate skin damaged by the years--or by injury, acne, or infection--may have applications for improved wound repair in the elderly, and potentially for protection against serious aging-related skin conditions. Geron Corp of Menlo Park, Calif., perhaps best known for its research into inhibition of telomerase to treat cancer, is also investigating the benefits of activating the enzyme. Telomerase adds nucleotide repeats called telomeres to the ends of chromosomes, thus compensating for the loss of telomeric DNA that occurs with each cell division. In humans, telomeres act as the counter of how many times a cell can replicate. Geron chief scientific officer Calvin B. Harley explains that the loss of telomeres, and the subsequent end to a cell's replicative life, are characteristics of chronic skin ulcers in the elderly, a potentially fatal condition. "It's a serious unmet medical need, and one in which we think telomerase therapy might be first implemented." Geron's telomerase activation therapy research is currently focused on the skin and liver. Last August, Harley and collaborators published a paper showing that telomerase expression can cause old skin to assume the phenotype of young skin.1 In what Harley calls "an important proof of concept paper," researchers demonstrated that telomerase therapy can not only "immortalize" cells, causing them to continue replicating indefinitely but the therapy can replace the characteristic blistering or splitting near the dermal-epidermal junction of aged skin with a youthful, nonblistering phenotype. Courtesy of William K. Boss, Jr.BeforeAfterAbout 1,450 patients in the U.S. and Europe have undergone a procedure involving cultured fibroblasts to improve their skin. Senior author Warren K. Hoeffler, then an assistant professor of dermatology at Stanford University School of Medicine, developed a cell-sorting technology that enabled the researchers to obtain these results in the fibroblasts of a human "skin equivalent." Hoeffler is founder and president of the Burlingame, Calif.-based Xgene Corp., and has licensed the technology exclusively to Stanford University. The skin is cultured on the backs of mice in a cell slurry containing two human cell types: dermal fibroblasts, which form connective collagen fibers, and keratinocytes from the epidermis. The cell types then sort out, to reconstitute the dermis and stratified epidermis of human skin. Hoeffler's company also has a patent pending on a method of growing skin in a culture dish in which a biopsy taken from a patient is disrupted, thus breaking the tissue into separate cells. The cells then sort into the proper layers through Xgene's technology and the reconstituted skin is then grafted onto the patient. Addition of telomerase to the cultured cells would extend their in vivo replicative life indefinitely. "We think this fits in very well with being able to replace either damaged or aged skin," he declares. This single-step process also will be cheaper and quicker than current composite techniques that build skin in individual layers, he says. Hoeffler believes his method will avoid the blistering and failure of the graft to "take" that plague composite skins. A major cause of graft failure is that the skin is grown on an inert scaffold, such as a biodegradable polymer, which creates a barrier between the cultured tissue and nutrients derived from the wound bed. "What we've started to do is construct the skin upside down," he explains. The epidermis is on the bottom, next to the scaffold. When the culture is ready for grafting, the skin is flipped over. The dermis is placed directly on the vascular bed, and the scaffolding on top becomes part of the wound dressing. Courtesy of William K. Boss, Jr.BeforeAfter Human Cell Therapy Promising results in restoring aged skin in humans using a different technology have already been recorded. William K. Boss Jr., vice chairman of the plastic surgery department at Hackensack University Medical Center, developed a method that involves cultured fibroblasts. After culturing a specimen of the patient's skin (usually from a biopsy behind the ear), the expanded fibroblasts are reinjected into the treatment area of the face. Wrinkles, scars, acne irregularities, and laser burns are among the conditions that have been treated. About 1,450 patients in the United States and Europe have undergone the procedure, including 94 at Boss's clinic. Of the latter group, 92 percent reported satisfaction with their skin improvements, and 70 percent showed continued correction at follow-ups ranging from 36 to 48 months.2 Similar results were obtained at other centers, including a pilot trial of 10 patients that was conducted through the division of head and neck surgery at the University of California, Los Angeles.3 That study showed microscopic evidence of increased thickness and density of dermal-layer collagen, and skin improvements of 60-100 percent in nine of the 10 patients, using both subjective and objective measures. Deborah Watson, now an assistant professor of surgery at University of California, San Diego, performed the injections for that trial while a chief resident at UCLA. She thinks that the process "definitely offers another option for clinicians." She recruited 40 patients for an expanded trial before leaving UCLA, but she says "data acquisition" difficulties apparently ensued, and the results of that study have not been published. Boss notes that the Food and Drug Administration has determined that the proprietary treatment, known as Isolagen, should be classified as a drug. He has made an investigational new drug submission, but he says the process is now on "clinical hold" until funding for a Phase III trial can be obtained. Isolagen has been used as an adjunct to temporary treatments such as botulinum toxin injections and bovine collagen injections. However, its mechanism of action is unknown. "Exactly what turns these cells on when they're injected and are getting involved in their new environment, other than that they're reacting to conditions there, is hard to say," Boss acknowledges. He adds that the patient's age limit at which the treatment loses effectiveness is about 65. Harley points out that tissue cultures can isolate the most vigorous cells, those with relatively long telomeres and good replicative capacity, but there tend to be fewer of them as the individual ages. He notes, however, that whole-cell therapy such as the Isolagen technique offers the advantage of banking the cells for later use. Boss and colleagues do store autologous cultured fibroblasts in liquid nitrogen for later use. Boss notes that, "If people are to have a choice, they're going to want to use their own cells." Courtesy of William K. Boss, Jr.BeforeAfter Immortalization Dangers Kathleen J. Smith, codirector of the dermatopathology section at the University of Alabama, notes that another option used in Europe and America is to periodically harvest stem cells--specifically, stromal progenitor cells--that are known to exist in fat. With regard to telomerase therapy, she points out that upregulation of the enzyme has potential to benefit wound healing, especially in the elderly. But she cautions against permanently immortalizing skin cells, on the grounds that some cell populations that are going through various phases of oncogenic change might inadvertently be immortalized. "I think that's a real issue, and a real problem, because there are a lot of oncogenic changes in the skin that can occur. You need to be very careful." Harley agrees, explaining that Geron's approach is to transiently activate telomerase, to increase the life span of cells without immortalizing them. He adds that drug screens also might locate transient activators of endogenous telomerase, similar to the effects of adenoviral vectors. Smith offers yet another possible approach to skin rejuvenation. She cites work by Barbara A. Gilchrest, who chairs the dermatology department at Boston University. Gilchrest and colleagues have shown that DNA damage generated by ultraviolet radiation is at least one signal that stimulates the photoprotective response of tanning, and that this response also can be induced by treating skin with thymidine dinucleotides, small DNA fragments. Gilchrest postulates that topical application of these fragments can provide a photoprotective tan to human skin without the harmful effects of ultraviolet radiation.4 Smith's notion, which she acknowledges hasn't been tested, is that it might be possible to simultaneously regulate these skin repair enzymes and telomerase, for combined protection against carcinogenesis and aging effects. The cosmetic possibilities of such techniques, and the huge profits they could reap, are obvious. Yet, Geron's Harley observes that the appearance of aging skin also heralds numerous potential health problems. "The mission of our company is to treat serious conditions in individuals," he affirms, "but we won't ignore other opportunities." Steve Bunk (sbunk@uswest.net) is a contributing editor for The Scientist. References 1. W.D. Funk et al., "Telomerase expression restores dermal integrity to in vitro-aged fibroblasts in a reconstituted skin model," Experimental Cell Research, 258:270-8, 2000. 2. W.K. Boss, Jr., et al., "Autologous cultured fibroblasts: a protein repair system," Annals of Plastic Surgery, 44:536-42, 2000. 3. D. Watson et al., "Autologous fibroblasts for treatment of facial rhytids and dermal depressions. A pilot study," Archives of Facial Plastic Surgery, 1:165-70, 1999. 4. M.S. Eller, B.A. Gilchrest, "Tanning as part of the eukaryotic SOS response," Pigment Cell Research, Suppl. 8:94-7, 2000.

Skin Like New

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