Photosensitization by 8-Methoxypsoralen1

PHOTOSENSITIZATION BY 8METHOXYPSORALEN* JANET HOWELL CLARK, Pn.D.

It has been known for sometime that certain psoralen compounds can act as photosensitizers

exposed to long ultraviolet and short visible light, using Sylvania blaeklight fluorescent lamps. The

2). In these experiments crystalline 8-

lamps give radiation from X310 to 440 m with maximum energy output at 350 mp. The energy by the Paul B. Elder Company. This is the delivered by two 1 5TSBL lamps at six inches is compound sometimes used to promote pigmenta- 2.35 X 1W ergs/sec/cm2. (Figure furnished by tion in vitiligo. It also is referred to as meth- the Engineering Department of Sylvania Electric oxsalen and sold under the trade name oxsoralen. Products, Inc.) The material is extremely insoluble, but if fed by Animals were exposed after local application of mouth in water suspension or mixed with food, it oxsoralen to the ears and backs and also after oral is absorbed in the gastrointestinal tract and administration of S-MOP. Some groups were produces photosensitivity in guinea-pigs and exposed at 12 inches under four lamps. Others mice an hour after feeding. A 1% solution of were exposed from the side with two lamps on "oxsoralen" in an acetone, alcohol, propylene each side at six inches. (1,

methoxypsoralen (8-MOP) was used, furnished

glycol base, furnished by the Paul B. Elder Company, produced local photosensitization when applied to the skin in humans, guinea-pigs and mice. EXPERIMENTAL

In preliminary experiments a quartz mercury arc with a glass filter was used. Far ultraviolet

RESULTS

External Application (1 % Oxsoralen Solution)

Mice: Twenty female mice (Swiss albino) were painted with oxsoralen on the ears and backs and exposed at 12 inches under four BL lamps. The mice scratched, spreading oxsoralen

radiation was therefore eliminated, and the to the facial areas, and probably ingested some effects observed were due to X> 310 m. These from their paws. As early experiments had experiments, in which albino guinea-pigs were established the fact that there is a long latent used, were concerned chiefly with the time be- period in animals radiated after sensitization

tween oral administration of 8-MOP and with psoralen, these animals were given one-hour radiation. It was found that animals fed one day and radiated the next showed no effect so that there is not storage of 8-MOP in the tissues. This agrees with previous observations (3). Comparison of animals radiated immediately with those radiated one to two hours after feeding showed that maximum sensitization occurs about one and one-half hours after feeding. After repeated

exposures at four- or five-day intervals. There were 23 exposures given from November 6 to June 4 (total dose = 9.66 X 10 ergs/cm2). The mice proved very sensitive. The ears were

red after one exposure, very red after two exposures, and after four radiations there was hair loss on the backs. When radiations were ter-

minated 3 had very sore ears, 14 sore backs and 4 had developed cataracts, probably due to some ears were ied, peeling and sore and later became ingestion of the oxsoralen solution. Three months crumpled with extensive tissue loss. There was later all surviving animals still had bare areas on hair loss around the eyes and animals that lived the backs. One animal had a large growth behind a year or more after radiation was discontinued the left ear which was sectioned and found to be developed corneal opacities. a malignant fibrosareoma, and another showed After these preliminary experiments groups of precancerous changes in the skin of the back. albino guinea-pigs and Swiss albino mice were Oral Administration (Crystalline * From the Department of Environmental 8-Methoxypsoralen) Medicine, School of Hygiene and Public Health, The Johns Hopkins University, Baltimore, MaryA. Guinea-pigs: Albino guinea-pigs were fed land. S-MOP in water suspension. The standard dose HeceiTed for publication November 28, 1960.

exposures, followed by severe reactions, the

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used was 20 mg/kilo. This is larger than the were given 30 radiations in five months (total dose used in therapy (4). There was no evidence dose 12.6 x 10 ergs/cm2 on each side). of illness as a result of feeding the standard dose. Group III (four animals) had the standard Those fed twice the standard dose failed to gain dose arid was given 25 half-hour radiations in weight hut showed normal growth when the five months from the side (total (lose 10.5 X 10 dose was reduced to the standard. Control animals fed 8-MOP without radiation

crgs/cm2 on each side).

or given radiation without 8-MOP showed no

When radiation was given from the side, the effect on the eyes was more pronounced and

effects.

corneal opacities developed in some cases by the

First, the effect of varying the dose of 8-MOP was tried and three groups were fed respectively

time radiation was stopped. Corneal opacities

standard dose, standard dose and twice the standard dose. After waiting one hour they were given a two-hour radiation at 12 inches from four BL lamps. This produced a definite effect in the

group with 3 standard dose, a severe effect in the group with the standard dose and a very

developed in the majority of the animals in Groups II and III and lentieular opacities in two. One animal in each group showed precancerous keratotie changes in the eyelids. B. Mice: No attempt was made to administer a measured dose of 8-MOP orally. Small pellets

of flour were made up containing crystalline 8-methoxypsoralen and given to the mice after other food had been withheld for a short time. fects that radiation had to be discontinued, and Animals were radiated one hour after feeding. in further experiments the dose was reduced to 3z Effects varied with the amount ingested. Control standard or standard and the radiation time was animals radiated without 8-MOP feeding showed reduced to hour. Three groups were then no effects. severe effect in the group with twice the standard dose. Repeated exposures produced such severe ef-

exposed as follows:

Group I (four animals) was fed 3 standard dose (10 mg/kilo) and radiated under four BL lamps at 12 inches, one hour after feeding, for j hour twice a week. This produced a chronic condition of red peeling cars with some scab formation and hair loss around the eyes. The animals were given 50 radiations (total dose

Group I (20 mice) was radiated from above for one hour (12 inches from four BL lamps) twice a month (16 radiations in eight months). The total radiation received was 6.7 x 10 ergs/em2. After two radiations the eyes, ears and

tails were red and sore and the ears partially

destroyed. After three radiations there was hair 10.5 X 108 ergs/cm2), and when sacrificed a year loss on the face and back. After ten radiations later showed slightly thickened and crumpled (4.2 X 10 ergs/cm2) the faces were bare and shrivelled, the ears destroyed, and the animals cars and some cloudiness of the corneas. Group II (eight animals) had 3 standard dose began to develop cataracts. At the end of the and was radiated j hour from the side with two experiment 94% of the surviving animals had BL lamps on each side at 6 inches. The animals cataracts, and two animals had developed lumps TABLE I Female albino mice Radiation s-MOP

Results Condition

External appli-

20

cation

Total dose

12 in. from 4 BL lamps 9.66 X 108 e.rgs/cm2

from top

Oral administration

20

Oral

14

5% fibrosarcoma, 5% precancerous condition, 20% cataracts

12 in. from4BLlamps 6.7 X 10 crgs/cm from top

94% cataracts, 10% precan-

6in.from4BLlamps 5.0 1< 108 crgs/em2

70%cataracts,7%papilloma

from side

cerous changes

PHOTOSENSITIZATION BY 8-METHOXYPSORALEN

behind the ears which showed precancerous changes in the tissue.

173

There is a growing belief that skin cancer in

man is related to the degree of exposure to

Group II (14 mice) was given only six radi- ultraviolet radiation in sunlight (9). Bungeler (10) many years ago called attention to the fact six inches) over a 3k month period. The total that, since everyone does not get skin cancer on radiation received was 5 x 10 ergs/cm2. Cata- exposure to sunlight, some skins must contain racts began to develop after two radiations (1.7 sensitizers which accelerate the development of ations (one hour from the side, two DL lamps at

X 108 ergs/cm2), and svhen radiation was stopped,

70% had cataracts and one had a papilloma on the conjunctiva. All had badly burned faces and ears.

The results are summarized in Table I. DISCUSSION

It is evident from the foregoing experiments that 8-methoxypsoralen applied externally or administered orally sensitizes animals (guineapigs, mice) to radiation >X310 m so that extreme radiation injury is produced by exposure to near ultraviolet and short visible radiation. The most pronounced effect was the formation

of corneal and lenticular opacities when the sensitizer was given orally. Several observers have noted that sensitization with methoxsalen may increase the effectiveness

of cancer production by ultraviolet radiation (3, 5, 6). Recently Griffin (7) has reported cataract and tumor production in mice given

cancer. He was able to produce skin tumors after subcutaneous injections of sensitizers followed by

exposure to sunlight more rapidly than by exposure to sunlight alone. It is now evident that in animals, psoralen will accelerate the production

of skin cancer after exposure to ultraviolet radiation and will cause cataracts and skin cancer after exposure to near ultraviolet radiation and short visible light (6, 7). An exhaustive study on sensitizers (1) in which they were evaluated by a number of tests showed

that they fall into different groups. The group containing hematoporphyrin and chlorophyll will photooxidize and cause hemolysis but is relatively ineffectual in producing erythema. The psoralen compounds do not cause hemolysis but produce

dermatitis after either local application or injection. Sensitizers, such as psoralen, which produce photodermatitis effectively might be expected to accelerate skin cancer formation more effectively than those falling in the first

8-MOP intraperitoneally or orally after exposure group.

to a Woods type long-wave ultraviolet lamp. The incidence of cataract was similar to that

CONCLUSION

reported here, but the tumor incidence was much

It has been shown that 8-methoxypsoralen, higher, probably due to a larger total dose of applied externally or given orally, will sensitize radiation. The results of this investigation add animals to radiation >X310 m and extreme nothing to Griffin's results except to emphasize

radiation injury is produced by exposure to near

the fact that, after sensitization with meth- ultraviolet and short visible light. The most oxsalen, exposure to near ultraviolet radiation

pronounced effect was the production of 94% cataracts in albino mice when the sensitizer was with a dose of radiation too small for tumor given orally and the total dose of radiation was production. When radiation is given from the side as little as 1.7 x 108 ergs/cm2 may produce too low to produce tumors. It was found that a dose as low as 1.7 x 10 cataract. ergs/cm2 in the near ultraviolet will produce As the animals used in these experiments were all albinos, it is not possible to draw general cataract in sensitized mice. Although the dose conclusions about the effect in pigmented animals of methoxsalen was much larger than that used and in human beings. However, a recent paper in therapy and although no similar experiments by Cloud, Hakim and Griffin (8) brings out the have been made with pigmented animals, it has fact that although black animals show little or been shown (9) that the effect on the lens is no damage in the pigmented area (ears, lids), similar in pigmented and nonpigmented groups. when they are sensitized with methoxsalen and The extreme sensitivity of the sensitized animals exposed to long ultraviolet radiation, the effect to cataract emphasizes the possible danger to the will produce a very large percentage of cataracts

on the lens is similar to that produced in albinos.

eyes when this compound is used.

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