Low Tumorigenic Potential Research Articles

Multiple steps are required for the induction of tumors by Abelson murine leukemia virus

Helper virus-free Abelson murine leukemia virus (A-MuLV) was used to induce monoclonal pre-B-cell tumors in mice. The clonality, patterns of immunoglobulin heavy-chain gene rearrangement, tumorigenicity, and v-abl oncogene expression in individual preleukemic and leukemic colonies were compared. Our results indicate that A-MuLV preleukemic cells with low or undetectable tumorigenic potential give rise to leukemic cells with high tumorigenic potential by a process of subclone selection. The levels of v-abl oncogene product in preleukemic and leukemic cell populations were not significantly different. These results suggest that an additional event(s) unrelated to the level of the v-abl protein product is required for A-MuLV-transformed cells to become fully malignant.

Chemotactic factor and P15E-related chemotaxis inhibitor in human melanoma cell lines with different macrophage content and tumorigenicity in nude mice.

The present study was designed to characterize the production of chemoattractants by human melanoma lines with high (M4Be, M3Da, NTerDa) or low tumorigenic (Doc8, M1Do) potential when heterotransplanted in nude mice. Supernatants from the Doc8 and M1Do cell lines were strongly chemotactic in vitro for mononuclear phagocytes. Chemotactic activity was destroyed by proteolytic enzymes, and upon gel filtration on Sephadex G75, it eluted in the cytochrome c region corresponding to an apparent m.w. of 12,000. Upon chromatofocusing, the Sephadex-separated tumor-derived chemotactic factor (TDCF) showed an isoelectric point of 5.5 to 6. Cell lines with high tumorigenic potential contained low or no detectable chemotactic activity. When culture supernatants of cell lines with modest (M3Da) or no (M4Be) chemotactic activity were exposed to immobilized monoclonal antibodies directed against the retroviral transmembrane protein P15E, appreciable chemotactic activity was detectable (M4Be) or preexisting levels increased (M3Da). The material eluted from Sepharose-bound anti-P15E antibodies inhibited the migration of monocytes in response to chemoattractants. These findings demonstrate the coexistence in some human melanoma cell line supernatants of factors (TDCF and P15E-related inhibitor) with opposite influence on monocyte chemotaxis. That tumor cell products play a pivotal role in regulating the extravasation of monocytes into neoplastic tissues is suggested by the close correlation observed between macrophage levels in melanomas grown in nude mice and levels of chemotactic activity detectable in culture supernatants.

Karyotype instability of Chinese hamster cells during in vivo tumor progression.

The extent of karyotype instability in spontaneously transformed Chinese hamster cells was determined after tumor formation by cytogenetic analysis of karyotype heterogeneity. The degree of karyotype heterogeneity among tumors formed in nude mice correlated with tumor latent period. The karyotypes of tumors formed after a short latent period by cells of high tumorigenic potential were similar to each other and to the injected cells. The karyotypes of tumors from cells of low tumorigenic potential and long latents periods were diverse, however. No chromosome aberration was common to every tumor. These results suggest that preneoplastic cells whose phenotypes are not directly capable of tumor formation can progress in vivo and that karyotype instability plays an important role in providing cell variants for tumor progression.

Mlt Mutation in the polyomavirus genome impairing a function of the middle T protein.

The DNA from polyomavirus mlt mutant P155 transforms cells in culture as efficiently as wild-type DNA but has a much lower tumorigenic potential when injected into newborn rodents. The mutant has a 12-base-pair deletion between nucleotides 1347 and 1360, i.e., in a region which encodes parts of the middle and large T antigens (G elinas et al., J. Virol. 43:1072-1081, 1982). To determine which of the two viral gene functions was affected by the mutation, we transferred the latter into a modified polyomavirus genome encoding exclusively the middle T protein. Our results show that the P155 mutation alters a function of the polyomavirus middle T protein required for the induction of the tumorigenic process in vivo. Beside the 12-base-pair deletion at 96.3 map units, there is no other alteration in the coding sequence of P155 middle T with respect to that of P16, the wild-type parental strain. We conclude, therefore, that the deletion is the lesion affecting the tumorigenic potential of mutant P155 .

The low tumorigenic potential of PRCII, among viruses of the Fujinami sarcoma virus subgroup, corresponds to an internal (fps) deletion of the transforming gene.

The avian sarcoma viruses FSV, PRCII, PRCIIp, and PRCIV share a related class of hybrid onc genes (delta gag-fps) defined by a specific nucleotide sequence fps and by delta gag-fps proteins of different sizes. Among these viruses, PRCII appears to have a lower tumorigenic potential than the others. Here we have compared fibroblast-transforming function and onc gene structure of these viruses. The fibroblast transforming ability of PRCII was lower than those of FSV, PRCIIp, and PRCIV. By gel electrophoresis the genomic RNA of PRCII measured 3.5 kb and those of FSV, PRCIIp, and PRCIV 4.5 kb; the delta gag-fps protein of PRCII measured 105 kilodaltons (kd), that of FSV 140 kd, and those of PRCIIp and PRCIV about 150 kd. By fingerprinting viral RNAs hybridized with molecularly cloned viral DNA the delta gag regions of PRCII and PRCIIp were defined to be 1.45 kb and that of FSV to be 1.3 kb. Fingerprint analysis of viral RNA-proto fps DNA hybrids showed the fps regions (approximately 2.8 kb) of FSV and PRCIIp to be isogenic. Compared to FSV and PRCIIp, the fps sequence of PRCII lacked a 1-kb region which maps between 0.3 and 1.3 kb from the 5' end of fps in FSV and PRCIIp. Based on oligonucleotide analysis, the shared fps complements of PRCII and PRCIIp were indistinguishable while that of FSV differed from those of the PRC viruses in scattered point mutations amounting to 1-2% of the RNA. Since all other regions of PRCII are isogenic with those of the highly tumorigenic variants PRCIIp, PRCIV, and FSV, it is concluded that the low fibroblast-transforming and oncogenic potential of PRCII reflects the internal fps deletion. Since the fps deletion reduces but does not eliminate transforming function, we suggest that the complete onc genes of viruses in the FSV subgroup include either several functional, or a regulatory and a functional fibroblast transforming domain. It has been reported that the 3' domains of the onc genes of viruses in the Fujinami subgroup and the onc genes of certain feline sarcoma viruses are distantly related. Since full transforming potential of the avian viruses depends on the 5' fps region not shared with the feline sarcoma viruses, we suggest that despite their structural homology, the avian and feline onc genes must have functionally different domains.

Polyoma virus mutant with normal transforming ability but impaired tumorigenic potential

Cloned DNA from the P155 mutant of polyoma virus transforms cells in culture as efficiently as wild-type DNA, but has a much lower tumorigenic potential when injected into newborn rats. Like cells transformed by wild-type DNA, cells transformed by the mutant DNA grow in low serum concentrations, form colonies in agar suspension, and grow to high saturation densities compared with untransformed cells. They are, however, much less tumorigenic since they transplant 100- to 2,000-fold less efficiently than cells transformed by wild-type DNA. Substitution of the region between 89.7 and 1.8 map units by the corresponding region of P155 DNA decreased the tumorigenicity of wild-type DNA. When this region was isolated from wild-type DNA and substituted in P155 DNA, the tumorigenicity of the latter increased to values comparable to those of wild-type DNA. This showed that the lesion affecting tumorigenicity occurred between 89.7 and 1.8 map units on the polyoma virus genome. Sequence analysis in this region revealed a 12-base-pair deletion between nucleotides 1,347 and 1,360. This identified P155 as an mlt mutant, i.e., a mutant with a deletion from a region which encodes parts of the large and middle T antigens.

Hypolipidemics Carcinogenicity and Extrapolation of Experimental Results for Human Safety Assessments

Dyslipoproteinemias represent a group of disorders closely related to alterations of cholesterol and triglycerides. The alterations of these lipids are considered important risk factors in coronary heart disease and indicate the need for clinically effective and safe drugs. Hypolipidemic agent therapy, however, does not appear without risk since the administration of these agents is by necessity, on a long-term basis. In the conduct of animal safety studies with some hypolipidemics, hyperplastic nodules or tumors developed in the liver of rodents. Data from the literature seem to indicate that the tumor response in rodents varies with the type of hypolipidemic drug administered. This paper summarizes the studies with the new lipid-regulating agent gemfibrozil. Aside from conventional long-term studies in rodents, the ultrastructural aspects of the liver were analyzed in several species and genotoxicity assays and short-term tests for hepatocarcinogenicity were conducted. Thus, it was possible to obtain an overview of these biological phenomena in order to allow for safety extrapolations. The biological behavior of these liver nodules showed that gemfibrozil and clofibrate-induced hepatocytes had not undergone malignant transformation. Further, the phenomenon of peroxisome proliferation, a characteristic event that follows hypolipidemic administration in rodents, was not confirmed in primate or human liver. Peroxisome proliferation has been linked to the process of hepatocarcinogenesis in rodents, although genotoxicity assays were negative and initiation/promotion tests failed to elicit tumors or nodules in a system where hepatocarcinogens manifest their activity. Thus, hypolipidemics such as gemfibrozil or Clofibrate may possess low tumorigenic potential with low risk due to the lack of correlation between these tests. Nevertheless, these agents are indicated for specific lipoprotein phenotype alteration with the resulting clinical benefits.

Hypolipidemics Carcinogenicity and Extrapolation of Experimental Results for Human Safety Assessments

Dyslipoproteinemias represent a group of disorders closely related to alterations of cholesterol and triglycerides. The alterations of these lipids are considered important risk factors in coronary heart disease and indicate the need for clinically effective and safe drugs. Hypolipidemic agent therapy, however, does not appear without risk since the administration of these agents is by necessity, on a long-term basis. In the conduct of animal safety studies with some hypolipidemics, hyperplastic nodules or tumors developed in the liver of rodents. Data from the literature seem to indicate that the tumor response in rodents varies with the type of hypolipidemic drug administered. This paper summarizes the studies with the new lipid-regulating agent gemfibrozil. Aside from conventional long-term studies in rodents, the ultrastructural aspects of the liver were analyzed in several species and genotoxicity assays and short-term tests for hepatocarcinogenicity were conducted. Thus, it was possible to obtain an overview of these biological phenomena in order to allow for safety extrapolations. The biological behavior of these liver nodules showed that gemfibrozil and clofibrate-induced hepatocytes had not undergone malignant transformation. Further, the phenomenon of peroxisome proliferation, a characteristic event that follows hypolipidemic administration in rodents, was not confirmed in primate or human liver. Peroxisome proliferation has been linked to the process of hepatocarcinogenesis in rodents, although genotoxicity assays were negative and initiation/promotion tests failed to elicit tumors or nodules in a system where hepatocarcinogens manifest their activity. Thus, hypolipidemics such as gemfibrozil or clofibrate may possess low tumorigenic potential with low risk due to the lack of correlation between these tests. Nevertheless, these agents are indicated for specific lipoprotein phenotype alteration with the resulting clinical benefits.

Adhesive Characteristics of Tumor Cell Variants of High and Low Tumorigenic Potential<xref ref-type="fn" rid="FN2">2</xref>

A variant subpopulation of C57BL/6 mouse fibrosarcoma cells that had very low tumorigenic potential was isolated from a highly tumorigenic parent fibrosarcoma cell culture. The adhesive characteristics of parent cells and variant cells were compared. The low-tumorigenic variant cells were released from the surfaces of plastic dishes, from protein-coated dishes, or from monolayers of fibroblasts or endothelial cells by protease treatment much more readily than were the parent cells. There was no difference between the variant cells and the parent cells in EDTA sensitivity or sensitivity to mechanical agitation under the conditions used. Also, no difference existed between the variant cells and the parent cells in rates of attachment to the surfaces of plastic dishes or to monolayers of endothelial cells. The variant cells were characterized by high levels of chymotrypsin-like esterase activity (two to three times increased over parent cell levels), but there was only a slight difference between the variant cells and the parent cells in caseinolytic or fibrinolytic activity.

Comparative studies of two types of “spontaneous” malignant alteration of ST/A mouse lung fibroblasts propagated in vitro

Two types of apparently spontaneous malignant alterations of fibroblastlike ST/a mouse lung cells (ST-L cells) grown in vitro are described. One type is characterized by a high tumorigenic potential of the altered cells in nonconditioned syngeneic recipients, a fibroblastlike morphology with cell surface showing very few microvilli by scanning electron-microscopy (SEM), and a growth pattern typical of nontransformed cells. These cells were described as R- cells. The other type is characterized bya low tumorigenic potential in non-conditioned, immunocompetent syngeneic recipients, rounding up of the cells which by SEM showed numerous microvilli on the surface, and a growth pattern typical of transformed cells. These cells were described as round cells or R+ cells. In immunoincompetent mice, R+ cells readily produced sarcomas, which grew faster than those produced by R- cells. Both types of ST-L cells expressed murine leukemia virus (MuLV) when tested in a peroxidase anti-p30 plaque test. The concentration of murine leukemia virus envelope glycoprotein (gp70) has previously (5) been shown to be threefold higher in R+ cells compared to R- cells. Furthermore, round-cell transformation was accompanied by the development of crossreacting rejection antigens protective against a secondary shallenge with Ehrlich ascites tumor and with syngeneic dimethylbenzanthracene induced ST/a mouse leukemia (STABAL). A similar protection was obtained by preimmunization with a cloned embryonic feral mouse cell line (SC-1) infected with ST-L virus as well as with virus-free SC-1 cells, suggesting the presence of rejection antigens both of viral (gp70) and nonviral origin.

The distribution of the mouse mammary tumor agent and its antigenicity in milk.

AbstractInbred mice with a propensity for mammary tumor development contain the mammary tumor agent and a specific soluble antigen in their milk. These were found in approximately equal proportions in both the supernatant and the pellet after ultracentrifugation. Over 90% of the agent and the highest antigen titer were located in a density range of 1.06–1.075 in ficoll density gradients. The low tumorigenic potential and the antigenicity of the B particle band (p = 1.12) remained with the particles after removal of proteinaceous contaminant by gel filtration. The immunogen from the upper portion (p = 1.06–1.075) of ficoll gradients appears to aggregate when concentrated and eluted from gel filtration columns. The tumorigenic material isolated from ECTEOLA and carboxymethylcellulose chromatographic columns does not show a high positive correlation with the distribution of the virus‐like B particles, which was consistent with the finding that the agent was concentrated in a narrow region with an average density of 1.28 in cesium chloride density gradients. The antigen associated with B particles and the lower molecular weight active component are immunologically identical.