@ @\ CANCER RESEARCH VOLUME 18 SEPTEMBER 1958 NUMBER 8 Tissue Culture in Cancer Chemotherapy Screening* ERICH HIRSCHBERG (Institute of Cancer Research and Department of Biochemistry, Columbia University Collegeof Physicians and Surgeens, New York 8@,N.Y.) The sponsorship of this Symposium represents on a secure basis and the availability of trans the first formal joint venture of the Tissue Culture plantable tumors facilitated the first attempts Association and the American Association for Can to develop systematic approaches to cancer chemo cer Research. As these two groups meet together therapy. today to appraise each other's problems and The year 1907 was an important one in both accomplishments, it is appropriate to recall that fields of investigation. Paul Ehrlich (@6) laid the tissue culture and experimental cancer research foundations of modern chemotherapy in his Har had their beginnings at about the same time and ben Lectures in London; Ross Harrison (40) made have developed over the years in close chronologi tissue culture a practical tool by explanting ner cal parallel and with increasing mutual interaction. vous tissues in a simple assembly and observing The history of both of these fields dates back their development in isolation. By 1910 and 1911, a little more than two generations, to the 1880's.' Alexis Carrel and Montrose T. Burrows (16, 17) During that decade, many attempts were made had started to maintain chick embryo fibroblasts to grow tissues or cells outside the body, as continuously in tissue culture and were cultivating illustrated by the work of Wilhelm Roux, who tumors in vitro; cancer chemotherapy also entered in 1885 achieved the isolation and maintenance a period of intense activity following the reports of the medullary plate of the chick embryo in of August von Wassermann and his associates warm saline solution (cf. 67). Four years later, (77, 78) on the inhibition of animal tumors by Arthur Hanau (39) carried out the first successful the intravenous or subcutaneous administration homologous transplantation of an animal tumor, of selenium and eosin. an achievement which signaled the start of modern This is not the occasion for a detailed account experimental oncology. By 1903, Carl Jensen (49) of the growth and development of the two fields was able to report that a mouse tumor could be during the period between the two World Wars. transplanted for nineteen successive generations; However, it is of interest to recall a few of the this important laboratory technic was thus put events of the past dozen years. In 1946, a notable Conference on Tissue Culture was convened in * Most of this material was presented at the 49th Annual Meeting of the American Association for Cancer Research in Hershey, Pa., which brought together a group Philadelphia, April 11, 1958, as part of a Symposium on “Re of active workers to evaluate the past, assess cent Contributions of Tissue Culture to Cancer Research.― future needs and objectives, and establish the The experimental work of the author and his associates men Tissue Culture Association. In 1953, Margaret tioned in this paper has been supported by U.S. Public Health Service Research Grants C-1894 and C-@33@and by the R. Murray and Gertrude Kopech (58) completed Charles Ulrick and Josephine Bay Gift for Research in Brain the monumental task of bringing together the Tumor Chemotherapy. tissue culture literature up to 1950 in a classified ‘Auseful guide to the early history of tissue culture is found bibliography of some 15,000 references; a first in Parker (6@); the beginnings of experimental cancer research supplement covering the more recent literature have been fully detailed by Woglom (80), and the development is in preparation. In 1954, a conference on tissue of cancer chemotherapy has been traced by Dyer (@0). culture technics in pharmacology at the New Received for publication May 14, 1958. York Academy of Sciences (63) stimulated the 869 This on@ [email protected] _____ Downloaded from cancerres.aacrjournals.org on September 24, 2021. © 1958 American Association for Cancer Research. 870 Cancer Re8earch Vol. 18, September, 1958 interest of experimental chemotherapists in this inhibition, the comments of several other reviewers useful investigative tool. Two years ago, the De (1, 15, 18, 50, 53, 57, 61, 71, 7@) have been of cennial Review Conference on Tissue Culture at great assistance. Woodstock, Vt. (79), bore witness to the vastly increased scope and versatility of these technics SYSTEMATIC APPLICATION OF TISSUE in many diverse areas of investigation. CULTURE TO SCREENING In experimental cancer chemotherapy, there During the last decade, much information has has been a similar period of intense and ever been gathered on the effects of a variety of chemi expanding research effort and periodic critical cal compounds on tumor cells in tissue culture, assessment. In the summers of 1945 and 1946, with or without normal cell controls. Many of conferences dealing with the current status of the these experiments, however, have been concerned field were held at Gibson Island; much of this with one or two agents at a time; only a handful material was published in 1947 (56). Helen Dyer's of studies have attempted a systematic explora very useful “Index of Tumor Chemotherapy― tion of larger series, thus permitting a more direct (@0) appeared in 1949. A concerted national pro evaluation of in vitro technics as screening tools. gram in cancer chemotherapy was initiated by A brief outline of the various systems which have the U.S. Public Health Service in 1953, and the been employed in this way will provide an indica Cancer Chemotherapy National Service Center tion of the diversity of approaches while furnishing @ began operations years later (@8). A number some background information for the general corn of conferences at the New York Academy of rnents which will be made later. Sciences and elsewhere afforded an evaluation Investigatioiu of Biesele and associales.—An im of particular types of chemotherapeutic agents, pressive series of publications (@—14,47, 48, 74) e.g., folic acid antagonists and other antimetabo has provided data on the in vitro effects of about lites (65, 81), 6-mercaptopurine (64), and alkylat 300 compounds, particularly purines, purine nu ing agents (51), and of various tools and technics cleosides, pyrirnidines, benzirnidazoles, antifolics, used for the selection of potential agents of chemo and amino acid antagonists. In most of these therapeutic interest (41, 73). studies, short-term primary explants of a trans ofcourse,asbothoftheseinvestigatiyeareas, plantable mouse tumor, principally mouse Sar tisssue culture and cancer chemotherapy, have coma 180, T@41, or Ma887, were placed into roller expanded and flourished, more and more points tubes; in addition to several tumor fragments, of contact between them have become apparent. each tube also contained explants of mouse em The availability of compounds with a documented bryonic skin as the normal control. The medium inhibitory effect on tumor growth has been of consisted of balanced salt solution, chick embryo great value in studies of the growth and behavior extract, human placental serum, and horse serum. of cells in vitro; problems dealing with the chemis The agent to be tested was incorporated into try of mitosis and the mechanism of action of the medium at several concentrations, ranging mitotic poisons have concerned investigators in generally from 0.1 to 4.0 millimoles/mi, and al both fields; inter-comparisons of the direct in lowed to act on the tumor and embryonic cells vitro effects and the often indirect in vivo action for 24 hours. Morphological evidence of damage of chemical agents on cells have proved a fruitful was then sought, and counts of mitotic and pyknot area of research; the applicability of tissue culture ic nuclei were often carried out to obtain a more technics to the search for carcinost.atic and car quantitative result. cinolytic agents has become an important question The main objective of these experiments has in cancer chemotherapy screening methodology. been to find compounds with selective or differ Each of these topics merits review and thorough ential toxicity for tumor cells. Several agents, discussion, but only the last will be considered notably 2,6-diaminopurine, 6-mercaptopurine, and in detail in this survey. No systematic attempt other 2- and 6- substituted purines, caused signif has been made to deal with the multitudinous icant damage to tumor cells at concentrations investigations in which nontumor cells or tissues which were entirely nontoxic to the embryonic (rat fibrocytes, chick and mouse embryonic tissues, controls, but most of them exhibited no such organ cultures, normal cell lines, and so on) were selectivity (cf. 3). A second objective has been exposed to various chemical agents. The pages to study the mechanism of action of these com to follow will deal primarily with those recent pounds by appropriate reversal experiments with studies in which tissue cultures of human and presumed normal metabolites. Experiments with animal tumors have been used; in considering the purine antagonists, in particular, have contributed relevance of these in vitro results to in vivo tumor to our understanding of nucleic acid synthesis and Downloaded from cancerres.aacrjournals.org on September 24, 2021. © 1958 American Association for Cancer Research. HIRSCHBERG—Ti&@Ue Culture in Chemotherapy Screening 871 its blockade by compounds like 6-mercaptopurine Re8ulis obtained by Wright, Cobb,andassociates.— or 2,6-diaininopurine. An attempt to employ tissue culture in a more Some of the more recent studies (7—9)have direct approach to the problems of clinical chemo been extended to human cell strains of cancer therapy, by-passing animal tumors entirely, has and normal tissue origin grown in the semisyn been made (19, 82, 83) by using a biopsy sample thetic medium of Eagle (cf.