Nongenetic variability in susceptibility to oncogenesis.

Genetically homogeneous mice varied in susceptibility to tumor induction by 3-methylcholanthrene. The early appearance of an induced tumor identified an animal of relatively great susceptibility to tumor induction as compared to other animals of the same genotype.

The immune reaction as a stimulator of tumor growth.

Various numbers of spleen cells from specifically immunized mice were mixed with constant numbers of target tumor cells, and were inoculated subcutaneously into thymectomized, x-irradiated recipients. Small numbers of admixed immune spleen cells produced a statistically significant, and reproducible, acceleration of tumor growth in the inoculum as compared with controls of either nonimmune spleen cells or spleen cells from animals immune to a different, non-cross-reacting, tumor. Larger. numbers of specifically immune spleen cells, however, produced inhibition of tumor growth. These data imply that the normal immune reaction may have a dual function in relation to neoplasia: (i) stimulation of tumor growth, early in the course of the disease, or whenever the immune reaction is minimal; (ii) inhibition of tumor growth at other times.

Analysis of antigenic heterogeneity within individual 3-methylcholanthrene-induced mouse sarcomas.

A method of establishing sublines by trocar transplantation in vivo was used to analyze possible cellular heterogeneity with regard to tumor-specific antigens in individual methylcholanthrene-induced sarcomas. Differences in antigenic specificity were found in at least 1 of the 9 pairs of sublines obtained, respectively, from opposite poles of 9 different primary tumors. No evidence suggested such differences among 7 pairs of sublines similarly derived from later transplant generations. The lack of variation in antigenic specificity between members of subline pairs obtained from later transplant generation tumors suggested that antigenic specificity was a stable characteristic. Even when derived from later transplant generations, the sublines of a particular tumor sometimes differed in growth potential, and/or immunizing capacity, and/or responsiveness to immunity. There was little or no correlation between variations in the two parameters: immunizing capacity and responsiveness to immunity. These findings led to the interpretation that immunizing capacity and responsiveness to immunity behaved like partially independent variables, and that variations in these parameters probably depended on factors other than, or in addition to, cellular antigen content per se. The marked, random, spacial heterogeneity revealed within the tumors in cellular growth rate and antigenic properties suggests that changes seen during serial tumor transplantation are probably due to random clonal variation and subsequent selection.

Relationship of tumor immunogenicity to concentration of the oncogen.

Tumors were induced sc in (BALB/c x DBA/2)F1 female mice by various concentrations of 3-methylcholanthrene (MCA) in paraffin pellets. There was an inverse relationship between MCA concentration and tumor latency (interval between MCA implantation and detection of gross tumor). The tumors were transplanted into syngeneic recipients, and material from this first transplant generation was used to immunize a series of syngeneic mice; any resulting growth was excised. Nonimmunized mice were controls. Immunized and control mice were irradiated and given an sc inoculation of a near-threshold number of tumor cells. Tumor growth from that inoculation was measured weekly in both groups and the antigenicity ratio (mean tumor size in controls/mean tumor size in immunized mice) was calculated. In a series of tumors with similar latencies, the only ones with high antigenicity ratios were those resulting from the high MCA concentration. The results suggested that tumors induced by low levels of oncogen may be good models of spontaneous neoplasia, strengthened the hypothesis that "spontaneous' tumors may actually result from low levels of oncogen, and indicated that neoplastic transformation and the development of immunogenicity are, at least in chemically induced tumors, independent changes that may be produced in the same cell when the concentration of oncogen is sufficient.

Stimulatory effect of immunization on tumor induction by Moloney murine sarcoma virus.

Adult mice were immunized with varying doses of inactivated Moloney murine leukemia virus (M-MuLV). Eight weeks after immunization, mice were challenged with a dose of Moloney murine sarcoma virus (M-MuSV) that could induce tumors in approximately 50% of normal animals. Mice immunized with high doses of M-MuLV (10(10) particles) had significantly decreased tumor incidences, whereas mice immunized with low doses of M-MuLV (10(2) particles) had significnatly increased tumor incidences compared to those in nonimmunized controls. The stimulatory effect could be abrogated by the irradiation of mice with 450 rads 24 hours prior to M-MuSV challenge, whereas the inhibitory effect was resistant to this irradiation procedure. The results suggested that immunization with virus can either stimulate or inhibit virus-induced tumorigenesis, depending on the dose of virus used for immunization.

Influence of regenerative capacity and innervation on oncogenesis in the adult frog (Rana pipiens).

Twenty-two sarcomas were induced in 19 adult frogs (Rana pipiens) treated with 3-methylcholanthrene pellets. Thirteen of these tumors arose first in a denervated forelimb, and only 2 arose first in normal or nerve-supplemented control forelimbs (P = 0.004). The remaining tumors developed either as a second tumor in a tumor-bearing frog or in hindlimbs. The critical role of innervation in regenerative capacity suggests that the predilection to tumor formation in the denervated limbs may have resulted from their lessened regenerative capacity.

Invasion and metastasis of a xenogeneic tumor in nude mice.

When aliquots of a malignant hamster tumor were transplanted to nude mice and to hamsters, the tumor showed 1) more rapid growth in the hamster, 2) local invasion and metastasis in both species, and 3) occasional spontaneous regression in the nude mouse.

Tumor-specific antigens as altered growth factor receptors.

Most induced, as opposed to spontaneous, tumors possess tumor-specific transplantation antigens. Data suggest that the prevalence of these antigens, at least among tumors induced chemically in diffusion chambers, is dependent upon the density of the culture at the time of carcinogen application. Other data show that density inhibition of cell growth is mediated by the interaction of various "growth factors" with their respective cell surface receptors. The juxtaposition of these two observations leads me to hypothesize that the tumor-specific transplantation antigen is an altered growth factor receptor. Development of the hypothesis leads to rational explanations of some paradoxical features of tumor growth in nude mice, to a possible understanding of why spontaneous tumors are nonimmunogenic, and to an explanation of the phenomenon of facilitation of tumor growth by a weak immune reaction.

Many growth factors may not be growth factors.

Both organ growth and tumor growth are dependent upon a biased ratio of cell births to cell deaths; this ratio is independent of the frequency of mitosis. Thus, so-called growth factors that affect only the frequency of mitosis are not really growth factors. I shall advance two hypotheses: that the normal adult ratio of cell births to cell deaths is maintained by the activity of a factor or factors produced by the stem cells; and that the differentiating cells, as well as the stem cells, produce a different factor that limits the frequency of mitosis.

Stimulatory effects of immune reactions upon the growths of untransplanted tumors.

The accumulated literature suggests that altering the immune capacities of animals or humans seldom has detectable effects on tumor incidence nor, it seems, does the growth of an untransplanted tumor, either spontaneous or induced, immunize the host against the growth of a subsequent implant of that tumor. These observations suggest that even clearly immunogenic transplanted tumors may not have had their growths modulated by an immune reaction when they were as yet untransplanted in their primary hosts. Also, there are many data that have been interpreted to show that spontaneous tumors of rodents are, even when transplanted, nonimmunogenic. A reconsideration of the available studies, especially those in which either host immune capacity or tumor immunogenicity was titrated, has led me to different conclusions. I believe that the data suggest that probably all tumors, including spontaneous ones, are immunogenic but that the weak immune response in the primary host to the antigens of even the more immunogenic induced tumors usually produces stimulation of tumor growth rather than growth inhibition. The prevalence of immunogenicity suggests that vaccination or other forms of immunotherapy will eventually succeed; however, the analysis also suggests that, in the case of weakly immunogenic tumors, increasing the immune reaction, unless the increase is massive, may have little effect or may actually stimulate rather than inhibit the growths of these tumors in their primary hosts. Immunosuppression might actually be therapeutic in these cases; this may be an unrecognized benefit of the chemotherapy of many human tumors.

The inhibition of tumor growth by tumor mass.

Evidence suggests that a tumor behaves, in its pattern of growth, like an integrated organ rather than a collection of independently growing cells. Tumor growth tends to slow progressively as size increases and to undergo compensatory growth after partial resection. Consequently, therapies that reduce tumor mass may tend to accelerate the growth of the remaining tumor and tumor metastases. An approach to therapy based upon a simulated increase in tumor mass may be worthy of consideration.

The paradoxical association of regression with a poor prognosis in melanoma contrasted with a good prognosis in keratoacanthoma.

Partial regression in cutaneous malignant melanoma has been reported by a number of observers, albeit not all, to be associated with a relatively poor prognosis; in contrast, a keratoacanthoma, which eventually regresses, does not metastasize. The Hammond effect could explain the possibly poor prognosis of the thin regressing melanoma. Hammond(W.G. Hammond et al., Cancer J., 8: 130-138, 1995) showed that the speed of biological progression to less differentiated phenotypes is directly related to the immunocompetences of the tumor hosts. If partial regression is a sign of an unusually strong immune reaction, then the melanoma that partially regresses might have a relatively poor prognosis because of the greater risk of biological progression among the surviving tumor clones. A Hammond effect is not associated with regression of a keratoacanthoma. I postulate that the growth of this tumor is accelerated, rather than restrained, by the immune reaction and that the ultimate regression of the tumor is the result, not of immune cytotoxicity, but of a rapid terminal differentiation (a reverse Hammond effect); alternatively, very rapid growth might lead to an exhaustion of growth potential before progression to clonal immortality could occur.

Two competing influences that may explain concomitant tumor resistance.

A second tumor inoculum is often inhibited in its growth by the presence in the recipient animal of an earlier implanted, growing tumor. The tumor resulting from the first inoculum may, paradoxically, continue to grow despite the simultaneous inhibition of the growth of the second inoculum, a phenomenon usually called "concomitant immunity." Evidence now suggests that the phenomenon can be observed in the absence of any recognizable type of immune reaction and might often be named more appropriately "concomitant tumor resistance." Consideration of a variety of probably related observations suggests that concomitant tumor resistance can best be explained by the competitive interaction of two opposing influences: a local diffusible, tumor-facilitating environment, produced by both tumor and normal tissues, that is counteracted by circulating inhibitors that are also produced by both tumor and by normal tissues. In an implanted small tumor, because of geometric considerations and diffusion, the action of the local facilitating environment is weak; in a larger tumor the local facilitating environment has a relatively greater influence and thus the larger tumor can continue to grow despite levels of circulating tumor inhibitors capable of inhibiting the smaller growth.

Cancers beget mutations versus mutations beget cancers.

Despite the plethora of "oncogenes" and "tumor suppressor genes," the hypothesis that cancer is usually the result of genomic mutations may be wrong. We should at least examine the alternative hypothesis, for which there is considerable evidence, that mutations do not commonly beget cancer, but rather that cancer phenotypes result from confused or aberrant patterns of normal-gene expression; the abnormal patterns are postulated to result from epigenetic mechanisms rather than from mutations. The epigenetic hypothesis that I am proposing suggests that cancers may exhibit mutations primarily because replicative errors at inactive sites in the cancer genome may be repaired slowly or not at all, but the mutations so produced, occurring at already inactivated sites in the genome, may have limited biological significance. Thus, it may be more correct to say that cancers beget mutations than it is to say that mutations beget cancers.

On the prevention and therapy of prostate cancer by androgen administration.

It has been widely suggested that elevated androgen levels may be critically involved in the genesis of prostate cancer. Despite the dependency of the normal prostate and of most prostatic cancers upon androgens and the fact that tumors can be produced in some rodent models by androgen administration, I will argue that, contrary to prevalent opinion, declining rather than high levels of androgens probably contribute more to human prostate carcinogenesis and that androgen supplementation would probably lower the incidence of the disease. I will also consider the possibility that the growth of androgen-independent prostate cancers might be reduced by the administration of androgens.

The autoimmune nature of cancer.

Four different kinds of data from the 3-methylcholanthrene-induced sarcoma system of the mouse show that the immune system stimulates oncogenesis; i.e., the presence of a tumor-specific immune reaction is a positive aid to tumor development. It seems proper, therefore, to consider cancer, at least in part, an autoimmune disease.

Clinical and immunological significance of human melanoma cytotoxic antibody.

The activity of a complement-dependent cytotoxic antibody in the sera of 21 melanoma patients was investigated using a microcytotoxicity assay. Heat-inactivated sera were caused to react against mechanically dispersed fresh tumor cells in the presence of exogenous blood group AB complement. Cytotoxicity was evaluated relative to pooled normal sera as a control. Sera were cytotoxic against autochthonous tumor cells in 9 of 10 patients with localized or regional melanoma and in 1 of 11 patients with disseminated metastases. Cytotoxicity of sera was unrelated to size of tumor burden. Six of 7 antibody-positive sera (autochthonous system) were noncytotoxic to between 2 and 7 different allogeneic melanoma tumor cell preparations. Immunological reactivity of the cytotoxic antibody-positive and -negative groups was similar with respect to their capacity to be sensitized to dinitrochlorobenzene, produce positive skin tests to microbial antigens, and produce antibodies to typhoid vaccination; serum immunoglobulins were comparable. These results support the reported findings of the presence of cytotoxic antibody in the sera of melanoma patients without disseminated metastases.

Are tumor-associated transplantation antigens of chemically induced sarcomas related to alien histocompatibility antigens?

The hypothesis tested was that tumor-specific transplantation antigens of chemically induced tumors cross-react with allogeneic histocompatibility antigens. This hypothesis makes several predictions that can be tested experimentally. First, tumors should grow better and be less immunogenic in certain F1 hybrids than in their syngeneic parents, owing to the hypothecated cross-reactivity of the tumor-specific transplantation antigens with F1 antigens. This is in contrast to the more common observation that parental strain tumors grow worse in the F1 hybrids than they do in the parent. Also certain allogeneic skin grafts might immunize the parental strain mice against their syngeneic tumors, and, finally, immunizing parental mice with syngeneic tumor might cause accelerated rejection of certain skin allografts. The results show that certain tumors grew better in the F1 mice than they did in the parents but that the tumors were not less immunogenic in the F1 hybrids. Mice immunized against alloantigens showed a dose-dependent enhancement of syngeneic tumor growth. Finally, mice immunized with syngeneic tumors demonstrated an apparent prolongation of certain skin allografts. The discussion considers possible alternatives explaining these results.

The flip side of tumor immunity.

A large amount of data suggest that tumors are, to some degree, dependent for their growth on a positive level of immune reaction, a level that is unique for each tumor. Each tumor gradually adjusts its immunogenicity to the level that will, in the immunologic context of its own particular host, maximize its growth. Thus, it follows that immunosuppression may be as likely as immunoaugmentation to have a therapeutic effect.