Vol. 4, 2741-2746, November 1998 Clinical Cancer Research 2741

3 The abbreviations used are: PDT, photodynamic therapy; NPe6. mono-

L-aspartyl chlorin e6; ECG, electrocardiogram.

Photodynamic Therapy Using Mono-L-aspartyl Chlorin e6 (Npe6)

for the Treatment of Cutaneous Disease: A Phase I

Clinical Study1

Scott W. Taber,2 Victor H. Fingar,

Carolyn T. Coots, and T. Jeffery Wieman

Department of Surgery, Division of Surgical Oncology, University of

Louisville, Kentucky 40292

ABSTRACTThe activity of a new photosensitizer, mono-L-aspartyl

chlorin e6 (Npe6), was assessed in an ascending dose Phase I

study for patients with superficial tumor. Eleven patients,

with a total of 14 tumor sites, were treated with photody-

namic therapy (PDT) using Npe6. Lesions included recur-

rent adenocarcinoma of the breast, basal cell carcinoma,

and squamous cell carcinoma. The phototherapy protocolconsisted of a single i.v. injection of 0.5-3.5 mg/kg Npe6,

followed 4 h later by 25-100 j/cm2 at 664 nm of light. PDTusing Npe6 caused no significant toxicity with the exception

of temporary generalized skin photosensitivity. In all cases,

light treatment caused immediate tissue blanching, followed

by a marked necrosis of the tumor mass. Regression of

tumor occurred over 24-48 h after the light treatment and

was followed by the formation of a heavy eschar over the

tumor site. Tumor regression was short-lived at Npe6 doses

of 1.65 mg/kg and below. In two of three patients, tumor

regression was either incomplete or tumors recurred within

the 12-week observation period. Increasing the Npe6 dose to

2.5 or 3.5 mg/kg combined with 100 J/cm2 of light energyresulted in better control of tumor regrowth with 66% (6/9)

of sites remaining tumor-free through 12 weeks observation.This increased tumor response came at the expense of the

tissue selectivity observed at Npe6 doses of 1.65 mg/kg and

below. There was no apparent selectivity for destruction of

tumor compared with normal skin at Npe6 doses of 2.5

mg/kg and above. These data demonstrate that Npe6 is bothan effective and safe photosensitizer for use in PDT and

provide the impetus for continued study in Phase II clinical

trials.

Received 6/2/98; revised 8/20/98; accepted 8/24/98.The costs of publication of this article were defrayed in part by thepayment of page charges. This article must therefore be hereby markedadvertisement in accordance with 18 U.S.C. Section 1734 solely to

indicate this fact.

C This investigation was sponsored by Nippon Petrochemical and Beck-

loff Associates and was supported by the Department of Surgery and theJames Graham Brown Cancer Center at the University of Louisville.2 To whom requests for reprints should be addressed, at Department ofSurgery, University of Louisville, James Graham Brown Cancer Center,

529 South Jackson Street, Louisville, KY 40292. Phone: (502) 629-6950; Fax: (502) 629-3183; E-mail: swtabe0l @pdt.bcc.louisville.edu.

INTRODUCTION

PDT3 is an evolving cancer treatment shown highly suc-

cessful for treatment malignant neoplasrns. The relative simplic-

ity and cost effectiveness of the administration of PDT make it

a very promising modality in both the curative and palliative

treatment of a variety of solid tumors. PDT has been used for

treatment of a wide range of malignancies including skin (I, 2),

breast (3), head and neck (4, 5), esophageal (6), lung (7, 8), and

gastric cancers (8, 9). Complete understanding of the mecha-

nisms by which PDT destroys tumor has not yet been attained.

PDT is a two-step process. Light sensitive dyes or photo-

sensitizing agents are injected iv. and allowed to circulate and

be taken up by tissues including tumor (10). Tissues are then

illuminated with exposure to nonthermal light at a wavelength

specific to the photosensitizing agent. The activation of the

photosensitizer by light is an oxygen-dependent process that

results in the generation of highly cytotoxic species including

singlet oxygen ( 1 1 ). The release of these reactive molecules

results in damage to both tumor cells and to the tumor micro-

environment. There are several photosensitizers presently under

clinical investigation, and Photofrin (QLT Phototherapeutics,

Vancouver, BC), a first-generation agent, has been approved

recently for use against esophageal cancer ( 1 2) and early lung

cancer ( 1 3). Npe6 is a new photosensitizer that has been shown

to be effective in preclinical and clinical studies (14-16). Npe6

has a strong absorption peak at 664 nm and produces high yields

of triplet oxygen when excited. This absorption peak allows

activation at long wavelengths and provides deeper penetration

of the light into the target tissue, thereby increasing the volume

of tissue that can be ablated ( 17). Npe6 is cleared rapidly from

tissues and in preliminary studies appears to offer a reduced

duration of cutaneous photosensitivity (18, 19). This article

reviews an 18-month clinical study of PDT using Npe6 as the

photosensitizing agent. The design of this investigation was a

single ascending dose, open-label study of I 1 patients with a

variety of solid tumors.

MATERIALS AND METHODS

PDT therapy with Npe6 was used in the treatment of

superficial malignancies in five women and six men ranging in

age from 51 to 91 years. The objectives of this study were to

determine the safety and tolerance of iv. administration of

single ascending doses of Npe6 and the effectiveness of PDT

using Npe6. Tumor palliation and/or cure were secondary ob-

jectives.

Patients with superficial malignancies were asked to enroll

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2742 PDT Using Npe6

into the study if they had either failed or refused conventional

treatments for their disease and met specific criteria. Inclusion

criteria included superficial malignancies, either primary or

metastatic that were accessible to treatment, including primary

skin cancers, metastatic or recurrent cutaneous tumors, soft

tissue sarcomas, and primary head and neck cancers. Subjects

who met one of the above criteria were enrolled into the study

if they had: a life expectancy of at least 12 weeks, a Karnofsky

Performance Status Score of �40%, the ability to tolerate re-

gional anesthesia, adequate renal function with no prior history

of renal disease, adequate hepatic function with a serum total

bilirubin of � I .5 mg/dl, the ability to give informed consent,

and the willingness to be institutionalized with close monitoring

for 48 h after treatment and to return for regular follow-up visits.

Exclusion criteria included: the failure to meet any of the

above inclusion criteria; a history of porphyria; a patient history

or family history of seizure disorders; an abnormal coagulation

profile; a history of sun hypersensitivity or photosensitive der-

rnatitis; significant gastrointestinal, cardiovascular, hematolog-

ical, endocrine, neurological, respiratory, or psychiatric dis-

eases; a history of significant allergies; previous treatment with

doxorubicin or related products (due to production of skin

photosensitivity); concurrent administration of any other mcdi-

cations affecting skin sensitivity; systemic or localized cancer

treatment within 30 days prior to PDT; and a positive serology

for hepatitis or HIV. Subjects were also excluded if they had a

history of ongoing alcohol or narcotic abuse. All subjects signed

the informed consent in compliance with the Institutional Re-

view Board of the University of Louisville School of Medicine.

Doses of Npe6 and the timing between photosensitizer

injection and light treatment were based on data from preclinical

studies. The initial dose of the protocol (0.5 mg/kg) was 1/50th

the dosage that had no measurable effect in dogs. The Npe6

doses used were 0.5, 1.0, 1.65, 2.5, and 3.5 mg/kg with no

dose-limiting toxicity reported. Light treatment was adminis-

tered 4 h after the iv. injection of Npe6.

Of 15 patients enrolled in the study, all were eligible to

participate. Three patients withdrew prior to receiving Npe6.

Eleven of 12 patients completed 4 weeks on the study. The data

from these 1 1 patients were analyzed. Lesions treated included

recurrent adenocarcinoma of the breast, adenocarcinoma of the

colon, basal cell carcinoma of the chest wall, squamous cell

carcinoma of the hypopharynx, shoulder, and parotid, and epi-

dermoid cancer of the urethra. More than one lesion per patient

was treated if separation of the lesions was adequate to insure

proper shielding.

Npe6 was supplied as a lyophilized powder by Beckloff

Associates, Inc. (Overland Park, KS)fNippon Petrochemicals

Co., Ltd. (Tokyo, Japan), study monitor and study sponsor,

respectively. It was stored at a temperature of 25#{176}Cand was

protected from light. The Npe6 was reconstituted with sodium

chloride injection USP to a concentration of 25 mg/ml imme-

diately before use. A single iv. dose of Npe6 was given 4 h

before light treatment.

For photosensitizer activation, either a Spectra-Physics

(Mountain View, CA) Model 171 or a Laserlonics (Orlando,

FL) model 1400-l2A argon ion laser was used to pump a

Spectra-Physics model 375B dye laser. The wavelength of ac-

tivating light was adjusted to 664 nm for activation of Npe6 and

was verified by a monochrornator (model DMC1-02; Optomet-

rics, Inc., Ayer, MA). A fiberoptic delivery system was used and

was terminated with either a single-use microlens or a cylindri-

cal diffusing fiber [PDT Systems (now Mirivant), Santa Bar-

bara, CA]. The fiberoptic tip was examined for defects, func-

tion, and power output before treatment. Power output was

monitored before and after light treatment with a calibrated

meter [PDT Systems (Mirivant) model 2015]. A flat tumor or

raised tumor < 1.5 cm in thickness was irradiated superficially

with a microlens. A raised tumor > 1 .5 cm and �2.0 cm in

thickness was irradiated interstitially with a cylindrical diffuser.

A raised tumor >2.0 cm in thickness was not treated in this

study. The tumor and a perimeter of normal tissue (0.5 cm) were

irradiated at power densities ranging between 35 and 100 rnW/

cm2 for total light doses classified as low (25 J/cm2), interme-

diate (50 or 100 J/cm2), or high (200 J/cm2). All treatments

using cylindrical diffusing fibers were at a power density of 200

mW/linear cm for total doses of 100 Jllinear cm.

Cutaneous photosensitivity/phototoxicity testing was ac-

complished using a solar simulator (Oriel Corporation, Strat-

ford, CT). A small circular area was outlined on the patient’s

lower back and irradiated at a light intensity of 80 mW/cm2 for

5 mm (24 J/cm2). Phototoxicity was graded as: 0, no visible

reaction; 1, minimal erythema; 2, deep, clearly defined cry-

thema; or 3, intense erythema and edema. Skin photosensitivity/

phototoxicity testing was done prestudy and at frequent intervals

after dosing through week 1 and then at weeks 2, 3, 4, and 6 if

a moderate or greater skin reaction persisted.

Patients given Npe6 were instructed to wear protective

sunglasses for 6 weeks from the time of Npe6 injection (or

longer if photophobia was experienced). Protective clothing was

also to be worn until testing with the solar simulator showed no

reaction. Patients were still required to report any cutaneous

photosensitivity for the duration of the study.

Study patients were evaluated prestudy during treatment,

including 48 h in the hospital, and during a 4-week period

posttreatment. Additional assessments were made at 6 and 12

weeks after treatment. The prestudy evaluation included a corn-

prehensive medical history, physical examination with vital

signs, neurological examination, ECG, chest X-ray, clinical

laboratories, skin photosensitivity testing, and ophthalmological

examination. A positive biopsy was required within 3 months of

study enrollment. A posttreatment biopsy of the treated area

could be performed at the discretion of the investigator.

During the 48-h hospitalization, evaluation included car-

diac monitoring for 24 h, beginning just before NPe6 injection,

an ECG at 48 h, frequent clinical laboratories (complete blood

count, liver function tests, and serum electrolytes) and daily

photosensitivity testing. Physical and neurological examinations

were conducted immediately predosing and daily thereafter.

Tumor assessments, which graded erythema, edema, firmness,

and the reaction of adjacent skin, were recorded, and photo-

graphs were taken at designated intervals.

The close follow-up of each study patient over a 4-week

period after PDT treatment with NPe6 consisted of the same

examinations, labs, testing, and assessments conducted during

hospitalization with the exception of the 24-h cardiac monitor-

ing. Ophthalmological examination was repeated at 12 weeks

after NPe6 injection.

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Patientno. Age Sex Diagnosis

No. of

treatedlesions Location

Lesion

size, cm(L/W/H)”

UL1-l 91 M Squamous cell 1 Parotid 8.7/3.9/0.8

ULI-2 63 M Basal cell I Chest 7.4/5.6/1.0UL1-3 73 M Malignant skin neoplasm I Shoulder 5.6/3.5/flatUL1-5 62 M Adenocarcinoma of colon 1 Flank 4.5/2.2/1.3

UL1-l4 74 M Squamous cell 1 Shoulder 3.0/2.1/0.8UL1-6 76 F Ductal carcinoma of breast 2 Chest

Chest1.1/0.5/flat0.8/0.5/flat

UL1-9 55 F Ductal adenocarcinoma of breast 1 Chest 2.8/2.8/flatUL1-lO 71 F Adenocarcinoma of breast 3 Back

ShoulderChest

3.3/3.3/flat

I .2/0.8/flat3.2/3.2/flat

UL1-Il 76 F Adenocarcinoma of breast I Chest 1.2/0.8/flatUL1-12 56 F Epidermoid carcinoma of urethra I Perineum 1.5/1.5/2.0

UL1-13 65 M Squamous cell 1 Hard and soft

palate2.0/2.0/flat

Lightdose

(J/cm2)

100100100100100

5050

100100100100100

NPe6 dose(mg/kg)

0.51.01.651.651.652.5

2.5

2.5

3.5

3.5

3.5

Lesion response”

Day Day Day28 42 84

F F FNE CR CRNE CR CR

F F F

NE NE FNE F FNE CR CR

NE NE CRNE NE CR

NE NE CR

NE NE CR

NE NE F

a L, length; W, width; H, height.b Lesion response was classified as failure (F), complete response (CR), or not evaluable (NE), as described in “Materials and Methods.”

Lens: 50 Diffuser: F100 J/cm

Lens: 25 Diffuser: CR25 J/cm

F

CR CR

Clinical Cancer Research 2743

Table 1 Clinical parameters and tumor response to PDT

Tumor response was evaluated at designated study inter-

vals with the final evaluation at 12 weeks after PDT treatment.

A complete response was defined as the absence of visual or

histopathological evidence of tumor within the area treated. A

>50% decrease in the size of the lesion treated was considered

a partial response. Failure was defined as a <50% decrease in

the size of the lesion treated.

Adverse experiences observed by the investigator or elic-

ited from the patient were recorded throughout the study. A

WHO scale was used to grade toxicity.

RESULTS

Tumor Response. Of the 15 tumors in 12 patients treated

with PDT, 14 were evaluated for response (Table 1). Patient

UL1-l5 died of metastatic disease at 2.5 weeks posttreatment.

Tumor response could not be adequately assessed in this patient,

and the patient was thus excluded from analysis. A dosage of 0.5

mg/kg NPe6 combined with 100 J/cm2 light was considered too

low to produce an effectual tumor response. The one patient

treated with this dose combination showed only minimal regres-

sion of the tumor mass. NPe6 doses of 1 .0-3.5 mg/kg combined

with 100 J/cm2 light were effective in producing tumor regres-

sion in the patient groups studied. In all cases, there was marked

blanching of the tissue by the completion of light treatment,

followed by edema and tissue discoloration. Regression of tu-

mor occurred over 24-48 h with gradual formation of a heavy

eschar over the tumor site. After eschar formation, the tissue

began to reepithelialize under the eschar, and the tissue healed

by secondary intention. The time required for complete healing

was variable but usually occurred by 8-12 weeks after photo-

therapy.

Tumor regression was often transient when NPe6 doses of

1 .65 mg/kg or below were used. Exceptions were the successful

treatment of a large basal cell carcinoma (7.4 cm X 5.6 cm) in

one patient (UL1-2) and an unclassified malignant skin neo-

plasm in a second patient (UL1-3). The response of tumor to

PDT was relatively selective with only minor damage to normal

tissues exposed to light during treatment. Increasing the NPe6

dose to 2.5 or 3.5 mg/kg combined with 100 J/cm2 light resulted

in better control of the tumor with 66% (six of nine) of the sites

remaining free of tumor throughout the completion of the study.

This increased response came at the expense of the tissue

selectivity observed at lower NPe6 doses. There was no appar-

ent selectivity of destruction to tumor compared with normal

skin at doses of 2.5 mg/kg or higher; both the tumor and the

surrounding normal tissues responded to light treatment with

similar degrees of tissue damage and necrosis. Phototherapy at

doses of 3.5 mg/kg showed no apparent increase in long-term

tumor cure compared with NPe6 doses of 2.5 mg/kg (five out of

six with complete response at 2.5 mg/kg versus one out of three

with complete response at 3.5 mg/kg). One of the failures at 3.5

mg/kg NPe6 and 100 J/cm2 light was a breast adenocarcinoma

that was refractory to previous PDT treatment at 2.5 mg/kg

NPe6 and 100 J/cm2 light (UL1 - 1 1 ). In both treatment failures,

the tumor and surrounding tissue did not show the characteristic

heavy necrosis and eschar formation seen in other patients. A

second patient with an invasive epidermal carcinoma that cx-

tended to a depth of 2 cm was treated with a surface illumination

of 50 J/cm2 light combined with 100 J/cm linear light from a

single cylindrical diffuser that was implanted along the major

axis of the tumor growth. The absence of complete tumor

regression in this patient was believed to result from incomplete

light delivery to the tumor. Subjects were not treated with the

high (200 J/cm2) light dosage.

Pain. Pain was a frequent reaction to light treatment of

photosensitized tissues both within and adjacent to the area or

areas treated. Pain was related to PDT treatment rather than to

NPe6 alone. The levels of pain experienced during and/or im-

mediately after PDT with NPe6 varied from no pain to moderate

pain, with no definitive correlation between the dosages of NPe6

and light and the degree of pain experienced.

Pain experienced during treatment was managed with mor-

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2744 PDT Using Npe6

Table 2 Photosensitivity testing”

Patient no. Age Sex

NPe6

dose(mg/kg) Baseline

Time after NPe6 dosing

24 h 48 h 96 h 1 week 2 weeks 3 weeks 4 weeks 6 weeks

ULI-l 91 M 0.5 0 0 0 <1

UL1-2 63 M 1.0 0 1 1 1 0

UL1-3 73 M 1.65 0 1 1 1 0

ULI-5 62 M 1.65 0 1 1 1 1 0

ULI-l4 74 M 1.65 0 1 1 2 2 1 1 1 0

ULI-6 76 F 2.5 0 1 1 1 1 1 0

ULI-9 55 F 2.5 0 1 1 1 1 0ULI-lO 71 F 2.5 0 1 1 1 1 1 0

UL1-Il 76 F 3.5 0 3 2 2 1 1 1 1 1

ULI-l2 56 F 3.5 0 1 1 1 1 1 1 1 0

ULI-13 65 M 3.5 0 0 0

ULI-l5 57 M 3.5 0 1 1 1 1 0

a A rating scale of 0-3 wa s utilized to assess cutan eous pho tosensitivity as described in “Materials and Methods.”

phine sulfate administered iv. by patient-controlled anesthesia

pump as a continuous infusion and/or a bolus infusion. Post-

treatment, during the 48-h hospitalization, any pain experienced

at the treatment site was treated with oral or iv. analgesics. If

pain persisted after hospitalization, it was effectively managed

with oral analgesics.

Localized Cutaneous Reactions. Erythema and edema

were common findings within or adjacent to sites of photody-

namic therapy. At doses of 0.5 mg/kg of NPe6 (one patient) and

1.0 mg/kg (one patient), mild erythema was seen immediately

posttreatment and resolved within 2-3 weeks. Increased cry-

therna and edema was observed after treatment in one patient

who received 1 .65 mg/kg NPe6 but was not observed in other

patients given the same NPe6 dose.

Of the three patients injected with 2.5 mg/kg of NPe6,

moderate to severe eiythema was noted in one patient. Mild to

moderate erythema was observed in the two patients. And mild

to moderate edema was observed in all patients.

One of the four patients injected with 3.5 mg/kg of NPe6

developed moderate to severe erythema, whereas the other three

patients showed only mild to moderate erythema. Moderate

edema was observed in this patient group.

Photosensitivity/Phototoxicity. Reactions to skin photo-

sensitivity testing were scored immediately after the completion

of the 24 J/cm2 light exposure and at 24- and 48-h intervals

thereafter. Table 2 shows the maximum skin photosensitivity

score at each test site over the study period. A reaction at the test

site was usually apparent immediately after the completion of

the light exposure and was sustained at that site through 24 h. In

one patient, a transient wheal and flare reaction was noted that

extended into the untreated skin surrounding the test area. Test

sites that initially showed minimal erythema (category 1) or

erythema (category 2) subsided within 48 h; test sites that

produced intense erythema (category 3) and edema in the first

24 h showed only minimal erythema at 48 h.

Test sites for patients given NPe6 dosages of 0.5 and 1.0

mg/kg produced only minimal erythema when testing was done

in the first week after NPe6 injection. Subsequent photosensi-

tivity testing in these patients showed no reaction. The treatment

groups that received Npe6 at doses of 1.65 and 5.5 mg/kg

exhibited phototoxicity scores that returned to baseline values

with 7-14 days.

One patient given NPe6 at the 3.5 mg/kg dose exhibited

intense erythema and edema at the test site when measured 24 h

after NPe6 injection. The patient developed erythema at test

sites through 96 h after injection and minimal erythema at test

sites to week 6 after injection. A second patient showed the

development of only minimal erythema at the test site, but that

minimal reaction persisted through week 4 of observation. A

third patient showed no apparent cutaneous photosensitivity at

any time before or after NPe6 injection.

Adverse Experiences. The physical and neurological cx-

aminations of patients given PDT using NPe6 were essentially

unchanged from the pretreatment evaluation through posttreat-

ment week 4. Clinical laboratory testing of blood revealed no

changes related to NPe6 alone or PDT with NPe6 treatment. An

elevation in results from liver function tests was noted in four

patients. This was considered to be unrelated to PDT with NPe6

but rather to the general diminishing health of these patients.

One patient (UL1- 12) presented with elevated bilirubin and

alkaline phosphatase levels during the study, but this was felt to

be secondary to the onset of metastatic disease in the liver.

Results from ECG and Holter monitoring after NPe6 injection

and PDT treatment revealed no changes from pretreatment eval-

uation. Ophthalmological examinations of the patients were

performed before NPe6 injection and 12 weeks after PDT treat-

ment. No clinically significant changes were observed. Three

patients did not receive an ophthalmological examination 12

weeks after phototherapy because they died of metastatic dis-

ease (UL1-2 and UL1-12) or were lost to follow-up (UL1-14).

The only adverse event among the study patients that was

considered “definitely related” to PDT with NPe6 was the

discharge of serosanguineous fluid at the treatment site. No side

effects were considered “probably” related to PDT with NPe6.

Other isolated complaints of nausea, pruritis, vomiting, head-

ache, diarrhea, dizziness, and heartburn were reported and clas-

sified as possibly related to the combination of NPe6 adminis-

tration with light treatment.

Multiple Treatment with NPe6. A patient not listed

above (UL1- 16) was selected for treatment in a special protocol

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Clinical Cancer Research 2745

designed to assess the effect of multiple NPe6 injections and

PDT treatments over a short time span. This patient had exten-

sive breast carcinoma that encompassed 60% of her chest wall

and extended around the left side to the back. The total treatment

area was divided into four sites that would be treated at desig-

nated intervals based on the previous treatment results. A total

of four NPe6 treatments was given over a period of 12 weeks,

each involving injection of photosensitizer and light treatment

4 h later. The first NPe6 dose given was 3.5 mg/kg. Subsequent

NPe6 doses were given at I .65 mg/kg. The plasma levels of

NPe6 were carefully recorded from the time of initial NPe6

injection through the completion of the fourth treatment. Light

treatment was 50 or 100 i/cm2, 4 h after NPe6 injection. PDT

using NPe6 produced tumor regression at all sites treated over

the four courses of therapy. Tissues containing tumor showed

necrosis followed by the formation of a heavy eschar. The

treated areas showed evidence of reepithelization and healing

2-4 weeks after treatment. The response of all areas, irrespec-

tive of the NPe6 dose administered, was classified as complete

at 12 weeks observation. Additional areas of tumor growth were

noted outside the light treatment fields during this study. We did

not observe any cumulative effects of the multiple NPe6 admin-

istration for skin photosensitivity. Cutaneous photosensitivity in

this patient was similar to that observed in other patients given

the same photosensitizer doses. Plasma levels of NPe6 showed

the expected decay profile observed when patients received a

single photosensitizer dose (data not shown). Repeated injec-

tions of NPe6 at 4-week intervals did not alter the baseline for

photosensitizer retention.

DISCUSSION

This Phase I clinical study demonstrates that PDT using

NPe6 is an effective and safe treatment for recurrent breast

cancer and specific skin cancers. No side effects from iv.

administration of NPe6 were observed at doses up to and in-

cluding 3.5 mg/kg with the exception of a generalized cutaneous

photosensitivity.

Light treatment was delivered 4 h after injection of NPe6

and was based on preclinical studies (15). The tumor response

and cure in these studies was found to be greatest when light

treatment was given when the plasma level of NPe6 was high

(19) and decreased rapidly as the time between NPe6 injection

and light treatment progressed from 2-12 h. Phototherapy under

conditions when plasma levels of photosensitizer are high im-

plies a vascular mechanism of tissue destruction. Several recent

investigations have concluded that tumor destruction after PDT

using NPe6 is dependent on vascular damage and blood flow

stasis (20, 21).

Tissue blanching and discoloration were the first signs of

response of photosensitized tissues treated with 664 nm of light

in the clinical study. Shrinkage of tumor and formation of eschar

followed this. Phototherapy with NPe6 doses of 1 .65 mg/kg or

less produced a highly selective response. Tumor shrinkage and

necrosis were observed without significant damage to the sur-

rounding skin within the light treatment field. Slight edema and

erythema were observed on the normal skin within the treatment

field, but this did not lead to necrosis. This subsided after light

application ended. In our small study treatment, NPe6 doses of

1 .65 mg/kg and 100 i/cm2 light produced tumor regression

through 12 weeks of observation in one of three patients. The

recurrent growth of tumor appeared to originate from the edges

of the original tumor site, suggesting that there was incomplete

destruction of tumor cells at the interface between tumor and

normal tissue. These observations are similar to findings of

failure in tumor response and cure noted in preclinical experi-

ments using Photofrin (22). Long-term tumor regression and

cure were found only if damage occurred to both the tumor and

the perimeter of normal skin surrounding the tumor (23, 24).

Cells at the periphery of the tumor mass appear to survive PDT

treatment due to either maintained tissue oxygenation or reoxy-

genation of the cells by the microvasculature of the surrounding

skin.

Escalation of the NPe6 dose to 2.5 and 3.5 mg/kg and light

treatment with 50-100 J/crn2 light caused necrosis of both

tumor and normal tissue within the treatment area. No apparent

selectivity for damage to either tumor or normal skin was

observed. The increase in relative damage to both of these

tissues did result in complete tumor regression in six of nine

patients through 12 weeks of observation after PDT (i.e., Npe6

and light). It remains unclear whether this response is a result of

increased damage within the tumor or as a result of the destruc-

tion of the normal skin surrounding the tumor and presumed

destruction of the vasculature at the periphery of the tumor

mass.

It remains unknown whether the destruction of the normal

tissue surrounding the tumor is a necessary condition for tumor

cure. The use of different NPe6 or light dose conditions that may

have maximized the destruction of tumor compared with normal

tissue was not investigated in this Phase I clinical study. Studies

under Phase II should investigate the effect of increasing the

light dose above 100 J/cm2 at NPe6 doses where tissue selec-

tivity was seen in this Phase I trial ( 1 .65 mg/kg NPe6).

A significant advantage of PDT using NPe6 is the reduced

duration of cutaneous photosensitivity as compared with other

photosensitizers. Photosensitivity testing of patients given NPe6

doses of up to 3.5 mg/kg generally showed only minimal or mild

erytherna within the first 96 h after NPe6 injection and no

significant skin photosensitivity after that time. Two patients

were exceptions to this finding and exhibited mild cutaneous

photosensitivity at 4 weeks after NPe6 injection. Because of the

wide range in results from photosensitivity testing, no direct

correlation of NPe6 dosage to the degree of cutaneous photo-

sensitivity was established in the present study. None of the

groups given a dose of NPe6 <3.5 mg/kg showed prolonged

photosensitivity. This, however, represents a small number of

patients and further testing for photosensitivity would need to be

performed.

The short duration of photosensitivity corresponds to ani-

mal studies by Roberts et a!. (18), who found no skin photo-

toxicity 24 h after NPe6 injection in mice and guinea pigs. These

findings coincide with the rapid clearance of NPe6 from plasma

and tissues seen by other groups (15, 19). The effect of NPe6 on

skin photosensitivity is different from that observed with Pho-

tofrin, which can be associated with some degree of photosen-

sitivity for periods of 6 weeks or longer after injection (24, 25).

The rapid clearance of NPe6 from tissues permits addi-

tional PDT treatments to be given over a relatively short time

Research. on May 10, 2018. © 1998 American Association for Cancerclincancerres.aacrjournals.org Downloaded from

2746 PDT Using Npe6

period. Our experience with one patient given four sequential

PDT treatments over 12 weeks indicates that this therapy can be

delivered safely and effectively with no accumulation of resid-

ual photosensitizer. Multiple treatments of single tumors may

allow smaller and less damaging NPe6 doses and light doses to

be used. This could be useful to preserve the surrounding normal

tissues or vital structures and to treat large areas or invasive

tumors that cannot be satisfactorily managed in a single treat-

ment.

NPe6 appears to offer a practical and safe alternative to

other presently available photosensitizers used in PDT. A lim-

ited duration of photosensitivity is the only expected and sig-

nificant side effect. The addition of NPe6 as a photosensitizer

for use in PDT provides the clinician with an additional tool for

the delivery of phototherapy and treatment of solid tumors. The

potential of this photosensitizer for use in PDT provides an

important motivation for continued investigation of NPe6 in

Phase II studies.

ACKNOWLEDGMENTS

We thank Dr. Malcolm N. Johnston, Kimberly McMahon, Pamela

Moore, and Cooper Buschemeyer for expert assistance with the clinical

studies and Susan Kemper for assistance in preparing the manuscript.

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1998;4:2741-2746. Clin Cancer Res   S W Taber, V H Fingar, C T Coots, et al.   for the treatment of cutaneous disease: a Phase I clinical study.Photodynamic therapy using mono-L-aspartyl chlorin e6 (Npe6)

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