DNA Repair Enzyme Decreases Skin Cancer Incidence in Animals

BUENOS AIRES-Ultraviolet (UV) radiation contributes to skin cancerinduction not only by transforming normal cells to cancer cellsbut also by impairing the host immune response to skin cancer,said Daniel Yarosh, PhD, president of Applied Genetics, Inc.,Freeport, NY.

Speaking at the 6th World Congress on Cancers of the Skin, Dr.Yarosh described a series of experiments showing that a liposome-encapsulatedenzyme, T4N5, under development by Applied Genetics, may be ableto inhibit UV-induced DNA damage, thus reducing immune systemsuppression and ultimately lowering the risk of skin cancer.

Dr. Yarosh stated that UV radiation produces two major types ofDNA damage: pyrimidine dimer (PD) formation and 6-4 photoproducts.Mutations activating the ras oncogenes and inactivating p53 tumorsuppressor genes have been sequenced from human skin cancer DNA,and these mutations occur most frequently at cytosine-pyrimidinesequences, implicating UV-induced PD formation.

Dr. Yarosh's group, in collaboration with Dr. Margaret Kripke,University of Texas M.D. Anderson Cancer Center, tested the hypothesisthat UV-induced DNA damage is the triggering event for UV-inducedimmune suppression. Their approach involved the use of a DNA repairenzyme (T4 endonuclease V) that initiates the removal of UV-inducedPDs from DNA, activating DNA repair prior to cell division. Theenzyme is encapsulated in liposomes, called T4N5 liposomes, thatare used to deliver the enzyme into cells of the skin.

Initial Experiments

In the initial experiments, Dr. Yarosh said, purified T4 endonucleaseV, encapsulated in liposomes and delivered to UV-irradiated cellsin culture, increased PD removal and DNA repair synthesis, andenhanced survival of repair-deficient cells.

Subsequent animal studies involved UV irradiation of the SKH-1hairless mouse. Unirradiated mice do not produce spontaneous skintumors, but mice irradiated three times a week with 1 minimalerythemal dose of UV-B radiation develop skin squamous cell carcinomas.

The animals were treated immediately after irradiation (threetimes a week for 30 weeks) with either active T4N5 liposomes oran inactive lotion, and the induction of skin tumors was examined.

The incidence of tumors greater than 1 mm was reduced, survivalwas increased, and the mean time to development of the first tumorwas significantly increased in the group treated with active liposomes,compared with controls (see figures). The reduction in skin cancerwas T4N5 dose dependent.

Furthermore, Dr. Yarosh said, studies suggest that UV-inducedDNA damage triggers the production of cytokines by epidermal cells,and these cytokines play a key role in UV-induced immune suppression,via the formation of suppressor T-cell populations. In mice, applicationof T4N5 liposomes prevented the formation of these T-cell populations.

Murine skin cells exposed to ultraviolet radiation in vitro werefound to release immunosuppressive cytokines, including interleukin-10(IL-10), which caused immune suppression in vitro, he said. Introductionof T4N5 liposomes into the cultures reduced the production ofIL-10 and eliminated the in vivo suppressive activity.

"Thus," Dr. Yarosh commented, "the topical applicationof T4N5 liposomes can prevent the DNA damage, which in turn preventsthe cutaneous immune
suppression."

Human Trials Under Way

Initial human trials have begun with the topical application ofT4N5 liposomes in patients with the genetic disorder xerodermapigmentosum, Dr. Yarosh noted. These patients have a biochemicaldefect in the DNA repair of PD and develop skin cancers on sun-exposedsites at greatly increased frequency. In phase I trials, the liposomesproved safe, and there were no allergic reactions.

Phase II trials in patients with xeroderma pigmentosum who hada very small portions of their skin UV-irradiated showed thatDNA damage was reduced in 9 of 11 patients with the applicationof the liposome-encapsulated DNA repair enzyme, Dr. Yarosh said.

He noted that phase III clinical trials are planned in the largerpopulation of patients who have a history of skin cancer removal,with measurement of intermediate endpoints (the rate of actinickeratoses formation).