Photodynamic Therapy

Basic Principles and Clinical Applications

Barbara W. Henderson

• Publisher: CRC Press
• Year: 2020
• eBook ISBN: 9781000148015
• Topic: Art
• Subtopic: Photography

Covering all aspects of photodynamic therapy, 70 expert contributors from the fields of photochemistry, photobiology, photophysics, pharmacology, oncology and surgery, provide multidisciplinary discussions on photodynamic therapy - a rapidly-developing approach to the treatment of solid tumours.;Photodynamic Therapy: Basic Principles and Clinical Applications describes the molecular and cellular effects of photodynamic treatment; elucidates the complex events leading to photodynamics tissue destruction, particularly vascular and inflammatory responses; discusses the principles of light penetration through tissues and optical dosimetry; examines photosensitizer pharmacology and delivery systems; reviews in detail photosensitizer structure-activity relationships; illustrates novel devices that aid light dosimetry and fluorescence detection; and extensively delineates clinical applications, including early diagnosis and treatment.;A comprehensive and up-to-date reference, this book should be useful for oncologists, pharmacologists, surgeons, photobiologists, optical engineers, laser technicians, biologists, physicists, chemists and biochemists involved in cancer research, as well as graduate-level students in these disciplines.

1

Historical Perspective

Thomas J. Dougherty, Barbara W. Henderson

Roswell Park Memorial Institute, Buffalo, New York

Samuel Schwartz

Minneapolis Medical Research Foundation, Minneapolis, Minnesota

James W. Winkelman

Brigham and Women’s Hospital, Boston, Massachusetts

Richard L. Lipson *

University of Vermont College of Medicine, Burlington, Vermont

Although photodynamic therapy as a clinical modality began to take shape in the late 1970s, the potential of certain compounds, in particular porphyrins, to localize in neoplastic lesions and to destroy those lesions upon light exposure, had been recognized much earlier by Drs. Schwartz, Winkelman, and Lipson.^* It was our privilege to honor these three scientists at the third Biennial Meeting of the International Photodynamic Association. The following contributions represent their personal reminiscences of PDT’s early days.

The Editors

I. Samuel Schwartz

I would like to summarize briefly some historical factors that led to my studies of the interrelations of porphyrins, tumors, and radiations. In my case, these radiations have been primarily ionizing x-rays and γ-rays, and I would hope that their brief discussion here will help broaden perspectives in the kindred field of phototherapy in ways that might have decisive future benefits.

_________________

^* Editor’s note: We are saddened by the recent death of Dr. Lipson. He will be fondly remembered by the PDT community

We start with the early embryonic days (Table 1) when, in 1934, I began working in Dr. Cecil Watson’s laboratory, at the University of Minnesota, as a freshman student, dishwasher, and laboratory assistant. Here I learned about the fascinating world of porphyrin pigments; of their physical properties, such as solubility, fluorescence, and light absorption; and of their essential roles in human and plant biology. During this period, too, I learned of the amazingly diverse manifestations of the different disease forms of hereditary porphyria. I read about the famous patient, Petry, who suffered from so-called congenital (now termed “erythropoietic”) porphyria, with its severe and debilitating photosensitivity. I was also much impressed with reports by Blum [1] and others of numerous photosensitizing effects of porphyrins, effects that could lead to death of paramecia or of mice, to lysis of red blood cells, and to destruction of enzyme activity.

Table 1 The Development of a Porphyrin Scientist

Embryonic Days	The 1930s
C. J. Watson Laboratories, University of Minnesota, 1934
Porphyrins, porphyrias, poetry, photosensitivity
Fetal Days	The 1940s
Manhattan Project, University of Chicago
Ionizing Radiations	Effects
1. Define
2. Prevent	Porphyrin, protect, potentiate?
3. Take Advantage of (shared effects of light and X-rays)
Infancy	The 1950s
Hematoporphyrin (HP)
Variable commercial products
Variable tumor localization	Cancer—porphyrin uptake?
Fractionate or make “derivatives”
(Urethane, HOAc/H2SO4, etc.)
The 1950s	Therapy trials
HP plus X-irradiation
A. Subjects	B. Materials
1. Humans (1955, inoperable CA)	1. Free porphyrins (many sources)
2. Mice	2. Metal complexes
3. Dogs
4. Paramecia
5. Enzymes

Fate decreed that I join the Manhattan Project at the University of Chicago in 1943, and horizons expanded. Here, our primary goal was to define and test biochemical effects of ionizing radiations, especially those of x-rays and γ-rays. A growing interest for me, too, was the modification of these radiation effects; first, how to prevent or protect against their damage and, second, how to potentiate their effects for special purposes such as tumor therapy. I wrote a proposal in 1944, or perhaps in early 1945, asking permission from the National Medical Director to test what I predicted would be radioprotective compounds, including cysteine, glutathione, and methylene blue. Permission was denied, but others in the project later tested cysteine. It worked, and the modern age of ra-dioprotection was born. Other investigators tried glutathione, methylene blue, and others, and they also worked.

These successes were encouraging, but I was now more interested in other possibilities involving my beloved porphyrins. Most of all, I was impressed by the many similarities in chemical and biological effects shared by ionizing radiations and by light plus porphyrin sensitizers.

During the war years and later, I tabulated reasons why porphyrins might be radioprotective, why they might be radiosensitizers, why either effect might be modified by light, and why cancer might respond differently from normal tissues. Reported tissue differences in heme enzyme concentrations and in consequent respiratory versus glycolytic activity were of special interest. It was not until about 1950, however, that the opportunity came to investigate the all-too-many variables associated with these different radiation responses.

The 1950s began with studies on porphyrin uptake by mouse tumors. Like Figge [2] and others, we chose to work with so-called hematoporphyrin (HP), which was then the only porphyrin available commercially in large amounts. Our solubility and chromatography tests, however, quickly showed that all commercial preparations were very impure, and only about 30–65% of the porphyrin included was actually hematoporphyrin. With partial purification, surprisingly, the hematoporphyrin-rich fractions were among the poorest localizers in tested tumors. Interestingly, too, Mann Chemical Co. supplied us with evaporated residue from the HCl mother liquor remaining after removal of the crystalline “hematoporphyrin” hydrochloride. Porphyrins in this normally discarded residue localized in tumors very much better than did the crystallized HP! Accordingly, we focused on tests of non-HP fractions, and made many new preparations. These included complexes with numerous metals, with urethane, then used for chemotherapy, and others. Among these new porphyrin preparations was something we called our acetic-sulfuric acid porphyrin. This material later came to be known as hematoporphyrin derivative (HPD), and ultimately, as Photofrin.

The story of how I first came to make HPD is quite unknown, so this may be a good time to tell the tale briefly. As background, I would call attention to our then basic purification system: HP, dissolved in dilute NaOH, was extracted into a mixture of ethyl acetate/glacial acetic acid. After washing with H₂O, HP was extracted into 0.5–1% HCl, and then other remaining porphyrins were extracted into 5–10% HCl. Hemin, if present, stayed with the ethyl acetate (in these early days, some commercial preparations also contained significant amounts of bro-minated porphyrin, formed as intermediates in the preparation of HP from hemin). Porphyrins were precipitated by neutralizing the HCl extracts with an excess of sodium acetate. Porphyrins from the 5% HCl fraction localized in tumors much better than did the purified HP extracted by 1% HCl.

We wanted to improve purification by use of column chromatography of the methyl esters. For this, the initial ethyl acetate/acetic acid extract was vacuum distilled by water suction, and the residue was esterified in a 20:1 mixture of methanol/H₂SO₄, extracted into CHCl₃, and dried. Some of this dried residue was hydrolyzed in 25% HCl to again yield the free porphyrin. To our surprise this hydrolyzed material localized in tumors much better than did the starting HP!

To make more material, we esterified directly by adding HP to methanol/ H₂SO₄. But this did not work; the product still localized poorly in mouse tumor. I then remembered that, in the initial preparation, the distillate residue from several liters of ethyl acetate/acetic acid contained about 10–15 ml of glacial acetic acid, and I had added the methanol/H₂SO₄ to this acetic acid residue. Therefore, I dissolved some HP in glacial acetic acid (HOAc) and then added methanol/ H₂SO₄. This product localized well in tumor, so acetic acid seemed essential. Subsequently, we found that use of methanol (and esterification) was unnecessary. The HP, instead, was dissolved directly in various ratios of HOAc/H₂SO₄ either at room temperature or at 60°–100°C, and kept at these temperatures, for various times up to 36 hr before precipitation with approximately 10 volumes of 3% sodium acetate. These simplified procedures, as you know, yielded products that localized well in tumors. The chemical nature of the localizing porphyrin mixture was unknown, although we showed that acetylation in acetic anhydride was ineffective. Other chemical changes were clearly involved. In addition, the products obtained varied considerably depending on the temperature and heating duration used. This variable deserves further careful study. I would note, too, that our preparations were dissolved and kept in dilute alkali for at least a couple of hours before testing in biological systems.

The story would probably have ended here, except for a happy circumstance. An old adage tells us to “Publish or Perish,” but I was not ready to publish. Instead, the good Lord sent an alternative; I found a friend to spread the word. Dick Lipson visited me from Mayo Clinic to discuss use of HP. I urged him, instead, to try the HOAc/H₂SO₄ material. He did, and he liked it. After completing some elegant studies, pressures were put on him to publish his results and his master’s degree thesis [3]. I insisted that he do so and not wait for me. He did, with very kind and complete acknowledgments. The rest is history [4] and I, and all of us, are grateful to him and others who pursued the story farther.

During the 1950s (and later), we did extensive studies on the relations of porphyrins and ionizing radiations. Because I believe these studies may have profound implications for phototherapy as well, and because too little attention has been paid to them (perhaps because I have submitted too little for publication), I would like to emphasize them briefly. About 55 human cancer patients, more than 20,000 mice, many dogs, rabbits, and paramecia, and some enzymes were included in our studies. Both local and total-body irradiation were used in the mouse and dog studies. More than 100 different porphyrin preparations were tested. A general review and preliminary report of our human studies was published in 1955 [5]. A national “Conference on Some Relationships of Porphyrins, Tumors, and Radiosensitivity” (undoubtedly the first of its kind) was held at the University of Minnesota in 1956, sponsored in part by Merck & Co. Numerous clinical and biological findings were reviewed, and tumor and normal tissue uptake of our acetic/sulfuric acid porphyrin (HPD) was compared with that of other porphyrin fractions [unpublished].

Response to ionizing radiation depends heavily on three factors: porphyrin dose, porphyrin type, and tissue type. The porphyrin dose is particularly critical. Small doses of some porphyrins markedly and consistently enhanced radio-sensitivity of tumors, but not of normal tissues. On the other hand, large doses of the same and other porphyrins and metalloporphyrins were radioprotective for these same tumors. Normal tissues, or total-body-irradiated normal mice were protected by both small and large doses of porphyrin. This opposite response of tumors and normal tissues to small porphyrin doses appears to be of critical importance, since it leads to a marked increase in the therapeutic ratio (response of tumor/normal tissue). Ehrlich ascites tumor cells irradiated in vitro were protected, even when the porphyrin was added up to 30 min after irradiation. Similar time studies were not done in animals. As noted in the following, paramecia behaved like cancer cells in their variable porphyrin dose response. Two brief illustrative examples will be given, and comments on our human studies are also included:

1. The dose dependency of radiation response in transplanted rhabdomyosarcoma tumors in mice was shown most strikingly in studies with Cohen [6]. About 2300 R was delivered to tumors transplanted to thighs of 153 mice. Three hours earlier the mice had been injected intraperitoneally with bicarbonate vehicle alone or with 10, 50, 250, or 1250 μg of a crude hematoporphyrin or its copper complex (the copper complex does not fluoresce, and is not photosensitizing). Results with both materials were similar and have been pooled. One hundred percent cures (27 of 27 mice) followed injection of 50 μg of either porphyrin preparation; only three cures were found in the remaining 126 mice during the 84- to 86-day study. The lowest dose, 10 μg/mouse, significantly improved tumor regression and mouse survival, but not enough to yield cures. I might also note that the optimum dose f...

Table of contents

• Cover
• Half Title
• Title Page
• Copyright Page
• Preface
• Introduction
• Contents
• Contributor
• 1 Historical Perspective
• I Molecular and Cellular Effects of Photodynamic Therapy
• II Tissue Effects of Photodynamic Therapy
• III Clinical Applications of Photodynamic Therapy
• IV Photophysics and Photodynamic Therapy Technology
• Index
• About the Editors