Pulmonary Tumors In Mice Research Articles

Non-canonical genomic driver mutations of urethane carcinogenesis.

The carcinogen urethane induces pulmonary tumors in mice initiated by an incredibly specific Q61L/R oncogenic mutation in the proto-oncogene Kras. Previous Whole-Exome Sequencing of urethane-induced tumors revealed a bias towards A➙T/G and G➙A substitutions. Subsequent ultra-sensitive Maximum-Depth Sequencing of Kras shortly after urethane exposure suggest a further refinement to CA➙CT/G substitutions. As C182AA➙C182T/GA substitutions in Kras result in Q61L/R mutations, the extreme bias of urethane towards these genomic driver mutations can be ascribed to the specificity of the carcinogen for CA➙CT/G substitutions. However, we previously found that changing rare codons to common in the Kras gene to increase protein expression shifted mutations in urethane-induced tumors away from Kras, or when detected in Kras, to G12D mutations that are usually rarely detected in such tumors. Moreover, the loss of p53 partially reversed this effect, generating tumors with either Q61L/R or G12D oncogenic Kras mutations, or no Kras mutations, presumably due to other genomic driver mutations. Determining the origin of these G12D and other unknown non-canonical genomic driver mutations would provide critical insight into the extreme bias of carcinogens for specific genomic driver mutations. We thus compared the types of Single Nucleotide Variations detected by previously performed Maximum-Depth Sequencing immediately after urethane exposure to the mutation signatures derived from Whole Exome Sequencing of urethane-induced tumors. This identified two types of non-canonical mutations. First, a V637E oncogenic mutation in the proto-oncogene Braf that conforms to the mutation signature of urethane, suggesting that the mutational bias of the carcinogen may account for this non-canonical mutation, similar to that for canonical Q61L/R mutations in Kras. Second, G12D and Q61H mutations in Kras that did not fit this mutation signature, and instead shared similarity with Single Nucleotide Variations detected by Maximum-Depth Sequencing from normal cells, suggesting that perhaps these mutations were pre-existing. We thus posit that when canonical Kras mutations are selected against that the carcinogen may instead promote the expansion of pre-existing genomic driver mutations, although admittedly we cannot rule out other mechanisms. Interrogating the mutation signatures of human lung cancers similarly identified KRAS genomic driver mutations that failed to match the mutation signature of the tumor. Thus, we also speculate that the selection for non-canonical genomic driver mutations during urethane carcinogenesis may reflect the process by which discordance between genomic driver mutations and mutational signatures arises in human cancers.

Abstract 4782: Inhibition of FGFR in breast cancer metastasis

Abstract Metastatic breast cancer is the most advanced stage of breast cancer. This type of cancer can spread other organs and cause 90% of cancer-related death. However, little is known about the molecular mechanisms which drive metastasis in breast cancer. Epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) promote tumor invasion and metastasis. Previously, it has been reported that fibroblast growth receptor 1 plays a key role during EMT: MET cycle. Therefore, optimizing FGFR inhibitors is crucial for therapeutic targeting of late-stage breast cancer. Here, we examined the efficacies of three FGFR kinase inhibitors, AZD4547, JNJ-42756493 and FIIN4, in the 4TO7 cell line both in vitro and in vivo. The 4TO7 cell line is a syngeneic model of systemic dormancy. Mammary fat pad engraftment of the 4T07 cells onto immune competent Balb/C mice results in systemic dissemination, but no macroscopic metastasis. In contrast, the 4T07 cells can escape cellular dormancy in 3D culture and form robust pulmonary tumors in mice upon tail vein inoculation. Furthermore, the 4TO7 cell line robustly responses to exogenous stimulation with FGF2. The phosphorylation of Erk1/2 downstream of FGFR was stimulated after a five-minute treatment with FGF2 and was effectively blocked with all the inhibitors tested. Furthermore, we developed a 3D culture approach that combines tumor spheroid formation in a non-adherent round bottom dish followed by placement on a bed of matrix. Our in vitro results demonstrate that both JNJ-42756493 and FIIN4 have more potent anti-proliferative activities in 3D culture and longer residence time as compared to AZD4547. In vivo, inhibition of FGFR is highly effective against 4TO7 tumors in the pulmonary microenvironment. Importantly, FIIN4 showed the least toxicity. This result suggested that covalent inhibition of FGFR is a promising cancer therapy for patients with metastatic breast cancer. Citation Format: Hang Lin, Saeed S. Akhand, Michael K. Wendt. Inhibition of FGFR in breast cancer metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4782.

Using size-controlled multicellular spheroids of murine adenocarcinoma cells to efficiently establish pulmonary tumors in mice.

Previous studies demonstrated that multicellular spheroids developed using polydimethylsiloxane-based microwells exhibited superior functions, such as insulin secretion from pancreatic cells, over suspended cells. To successfully apply these spheroids, the effect of spheroid size on cellular functions must be determined. In this study, using murine adenocarcinoma colon26 cells, the authors examined whether such spheroids were useful for developing tumor-bearing animal models, which requires the efficient and stable engraftment of cancer cells at implanted sites and/or metastatic sites. The authors prepared microwells with widths of 360, 450, 560, and 770 μm through a micromolding technique, and obtained colon26 spheroids with average diameters of 169, 240, 272, and 341 μm, respectively. Small and medium spheroids were subsequently used. mRNA levels of integrin β1, CD44, and fibronectin, molecules involved in cell adhesion, increased with increasing colon26 spheroid size. Approximately 1.5 × 104 colon26 cells in suspension or in spheroids were intravenously inoculated into BALB/c mice. At 21 days after inoculation, the lung weight of both colon26 spheroid groups, especially the group injected with small spheroids, was significantly higher than that of mice in the suspended colon26 cell group. These results indicate that controlling cancer cell spheroid size is crucial for tumor development in tumor-bearing mouse models.

Dimethylarsine likely acts as a mouse-pulmonary tumor initiator via the production of dimethylarsine radical and/or its peroxy radical

Aims Recent animal experiments have indicated that dimethylarsinic acid (DMA), a main metabolite of inorganic arsenic, is a complete carcinogen in the lung of mice and the urinary bladder of rats, nevertheless, no ultimate-active metabolite from DMA has been identified thus far. We have proposed that dimethylarsine ((CH 3) 2AsH), an ultimate reductive metabolite of DMA, is excreted in the expired air of mice administered DMA, and furthermore, was easily converted into dimethylarsine radical ((CH 3) 2As•) and dimethylarsine peroxy radical ((CH 3) 2AsOO•) by its reaction with O 2. The aim of the present study was to elucidate the possible mode of the tumorigenic action by dimethylated arsenic. Main methods In vitro experiments using GSH reductase as a two-electron donor of dimethylarsenic-glutathione conjugate ((CH 3) 2As-SG) and DNA adduct assay via a photochemical approach were performed. A lung tumorigenicity assay of (CH 3) 2AsH suspended in argon-atmospheric olive oil for 5 days was also conducted in mice. Key findings The results indicated that (CH 3) 2AsH was easily produced enzymatically from (CH 3) 2As-SG and showed tumor-initiating action in mouse lung via the production of (CH 3) 2As• and (CH 3) 2AsOO• by its reaction with O 2, and that these radicals have the ability to form DNA adducts. Significance The carcinogenicity of DMA, at least in mouse lung, could be explained based on the proposal that oral administration of DMA induces pulmonary tumors in mice, and arises from the arsine radicals produced through (CH 3) 2AsH, which was enzymatically reduced from (CH 3) 2As-SG.

An Assessment of the Carcinogenic Potential of Trichloroethylene in Humans

ABSTRACT Controversy surrounds the assessments of carcinogenic potential associated with human exposure to trichloroethylene (TCE). The American Conference of Governmental Industrial Hygienists states that TCE is “not suspected to be a human carcinogen.” In contrast, the International Agency for Research on Cancer has classified TCE as a probable human carcinogen, based primarily on the results of animal toxicity studies. Chronic high-dose TCE exposures cause hepatic and pulmonary tumors in mice and renal tumors in rats. Human epidemiology studies, however, do not support a causal association between exposure to TCE at environmentally relevant levels and cancers of the lung, liver, or kidney. The apparent discrepancy between the animal data and the human data can be explained by (1) differences in TCE exposure levels between laboratory animals and humans, (2) species-specific differences in TCE metabolism, and (3) other species-specific mechanisms involved in the development of cancer in rodents. This paper critically assesses the experimental and epidemiological data relevant to the carcinogenic potential of TCE. From the analysis, we conclude that TCE exposure at concentrations likely to be encountered in most environmental media is not likely to cause liver, lung, or kidney cancers in humans.

The Hydrazine Derivative Aminoguanidine Inhibits the Reaction of Tetrahydrofolic Acid with Hydroxymethylarginine Biomolecule

Aminoguanidine (AG), a hydrazine derivative is known to inhibit the formation of Advanced Glycosylation Endproducts (AGE) and AG has been proposed as an agent in prophylaxis of diabetic complications. However, treatment with hydrazine produced liver and lung tumors by formation of N7- and O6-methylguanine in the DNA of rodents. The hydrazine derivative, isonicotinic acid hydrazide induced pulmonary tumors in mice. N(G)-hydroxymethyl-arginine (HMA) was synthesized by our research group and it showed anticancer effect against experimental tumors. HMA was found earlier in human blood and urine, and recently in many plants (in fruits and vegetables). We could demonstrate a reaction (pH = 7.5, 37 degrees C, 1h) between HMA and tetrahydrofolate (THF) producing N5,N10-methylene-tetrahydrofolate (CH2-THF), the coenzyme of thymidylate synthase (TS). In model experiments AG proved to react with formaldehyde (HCHO) and to eliminate the C1-fragment of HMA, but not that of CH2-THF. In the presence of AG burst chemiluminescence and a higher speed of the formylation and methylation reactions were found in the AG, HCHO, hydrogen peroxide (H2O2) and L-lysine system than without AG. HMA as a biomolecule is one of the compounds which are responsible for the endogenous HCHO level. The biochemical function of HMA may be the direct supply of C1-fragment for the folate cycle. AG can disturb the above function of HMA. The reaction between AG and HCHO seems to be dangerous for biological systems because of the possible presence of L-lysine and H2O2. The burst chemiluminescence indicates excited molecules with extreme high energy producing uncontrolled formylation and methylation reactions. Considering the results of the experiments with AG its use as a medicament seems to be questionable.

Induction of benign and malignant pulmonary tumours in mice with benzo(a)pyrene: Km SH, Lee CS. Department of Anatomy, College of Veterinary Medicine, Chonnam National University, 300 Youngbong-dong, Puk-ku, Kwanju 500–757. Anticancer Res 1996;16:465–470

Most of the pulmonary tumors induced by chemical carcinogens in mice are pulmonary adenomas. Because of the morphological and biological characteristics, the site of origin and the spontaneous occurrence of this type of tumour, the pulmonary adenoma system is generally not considered to be an adequate model for studies designed to elucidate the pathogenesis of human bronchogenic carcinoma. The present study was carried out to observe the histopathological changes in the lung of 4 strains of mice intratracheally instilled with benzo(a)pyrene(BP). On the other hand, for the carcinogenesis experiments using animals, a method studying the incidence of pulmonary adenoma in newborn mice has been generally adopted, but this method still needs more than 24 weeks. This experiment was one of the attempts to make such an experimental period shorter. For the latter experiment, both inbred A/J and noninbred N:GP(s) newborn mice were used. In the group given intratracheal instillation, squamous cell carcinomas and adenocarcinomas were induced. Tumous were induced in high incidence in the lung of A/J and C57Bl/6 mice. Squamous cell carcinomas especially were well differentiated in the A/J mice with a higher incidence than C57BL/6. In the group given subcutaneous injection, adequate combinations at week 9 for the application following anticarcinogenesis assay were the groups of A/J treated with 40 micrograms (71.0 % adenoma incidence) and N:GP(s) mice treated with 500 micrograms (49.4 % adenoma incidence) of BP. These findings suggest that mice appear adequate for studies on the pathogenesis of malignant lung tumours and detecting anticarcinogens.

The effects of single versus triple intravenous injections of B16 melanoma cells on the development of pulmonary tumors in mice.

During natural metastasis, cancer cells are released continuously into the bloodstream and arrested in target organs; the process is repetitive rather than a single event. In this communication, the effect of preceding on successive "pulses" of cancer cells is discussed in relation to the arrest, retention and tumor formation of B16 melanoma cells in the lungs of mice receiving 3 separate tail-vein injections at 2-hr intervals. When animals received the same total number of cells in either 1 or 3 injections, no significant differences were detected in the initial arrest and subsequent retention in the lungs. However, 15 days after injection, mice which had received 3 pulses of cancer cells were found to have fewer pulmonary tumors than those which had received a single pulse of cells. The results show that interactions between 2 successive waves of B16 melanoma cells and their host did not affect the initial arrest and "short-term" retention of a third wave of radiolabelled cells. However, the "longer-term" experiments on tumor formation show that preceding pulses of cancer cells resulted in the lungs becoming more hostile to a succeeding pulse than was the case in appropriate controls. The results indicate that the relative metastatic capacities of cancer cells arrested in microvascular beds during hematogenous metastasis could well be diminished by antecedent interactions of this type.

DNA damage and repair in mouse tissues following procarbazine administration

Summary DNA fragmentation elicited in various organs of Swiss mice by i.p. treatment with procarbazine (PCZ) was studied by means of the alkaline elution assay modified to allow the analysis of DNA damage in vivo . A decreasing amount of DNA damage was detected in liver, lung, bone marrow, kidney, spleen, and thymus tissues. In the liver, dose-related increase in the DNA elution rate was observed to be from 50 mg/kg to 400 mg/kg, and the greatest amount of DNA damage was found six hr after treatment. DNA repair, as measured by return of the DNA elution rate to the control level, resulted, after the peak damage period, in a half-time of 10.5 hr. The high degree of DNA damage found in the lung correlated with PCZ specificity in inducing only pulmonary tumors in mice given a high single dose. This type of correlation was absent in liver tissue.

Combination adriamycin-radiation treatment of pulmonary tumors in mice.

Different schedules of thoracic irradiation (1,000 rad single dose) and adriamycin (3.0 mg/kd/day for 3 consecutive days given i.v.) were used to treat mice bearing pulmonary microcolonies of the Dunn osteosarcoma. Lung colony numbers and volumes were scored in mice sacrificed 21--25 days after tumor cell inoculation. Significantly greater levels of tumor control were achieved when the 2 agents were given concurrently or when the irradiation preceded adriamycin by 7 days, compared to lower levels of tumor control when the adriamycin preceded the irradiation by 7 days. The sequence of administering these 2 agents appears to have little effect, however, in reducing peripheral leukocyte count.

The use of immunologically competent cells from pig mesenteric lymph nodes to treat pulmonary tumors in mice

Pulmonary tumors were produced in A strain mice by intravenous injection of 1×106 A strain mammary carcinoma cells. The mice were killed on day 14, their lungs fixed in Bouin's fluid, and the number of tumors counted.

A new model of the mechanism of induction of pulmonary tumors in mice

A new model of the mechanism of induction of pulmonary tumors in mice

Der Lungentumor der Maus: ein progressiver Tumor

By subcutaneous serial transplantations of induced pulmonary tumors of mice a tumor progression has previously been correlated with the age of the tumors. The progression showed alterations from an alveolar-lining-type adenoma through papillary structures and different carcinomas to a spindle cell carcinoma in the final stage. Based on this report, 448 spontaneous pulmonary tumors in conventional and specific-pathogen-free Han: NMRI mice in different stages of life were classified. Six different types of pulmonary tumors were identified and compared with the age of the host. With increasing age of the host the pulmonary tumor tends to develop towards ultimate malignancy. Such a time-dependent transformation occurs regularly, as shown by a statistical analysis. A progression of primary spontaneous pulmonary tumors in mice is indicated as well.

Long-term administration of sterigmatocystin and Penicillium viridicatum to mice

The long-term toxic effects and carcinogenicity of pure sterigmatocystin (S-p), of sterigmatocystin administered in the form of a rice culture of Aspergillus versicolor (S-rc) and of a toxigenic species of Penicillium viridicatum were studied by feeding mice on a diet containing 5 ppm sterigmatocystin (S-p or S-rc) and/or a 75% content of a rice culture of P. viridicatum (P-rc) for periods of 2 wk alternating with 2-wk periods on the control diet, for a total of 54–58 wk. A control group was fed only the purified diet. In mice fed S-p or S-rc alone, the incidence of pulmonary tumours was, respectively, 73 and 49% higher than that in the control group. The incidence of pulmonary tumours in mice fed P-rc was about 16% higher than that in the controls. In the groups fed P-rc with S-p or S-rc, the incidence of pulmonary neoplasms exceeded that in the controls by 54 and 52%, respectively. The tumours were classified as adenomas or adenocarcinomas arising from the bronchial and alveolar epithelium. Adenomatosis was more common in mice fed diets containing sterigmatocystin, the incidence being 27–47% greater than that in controls. The incidence of other neoplasms was not affected by the feeding of the experimental diets. The increased incidence of pulmonary neoplasms in the mice fed S-p, S-rc or P-rc appeared to be directly related to the feeding of these test materials. The possible presence of certain common mycotoxins in the P-rc was eliminated by chemical analysis. It was concluded that P-rc and sterigmatocystin (S-p or S-rc) each had a carcinogenic effect when fed in the diet to mice but no additive effect was found when sterigmatocystin was combined with the P-rc diet.

Intracisternal A-type and C-type particles observed in pulmonary tumors in mice.

Intracisternal A-type and C-type particles observed in pulmonary tumors in mice.

Progression of pulmonary tumor in mice. 1. Histological studies of primary and transplanted pulmonary tumors.

Histological and biological observations were carried out on pulmonary tumors in mice which arose in the following four series of experiments: 1) pulmonary tumors in A/Jax and DD strain of mice which developed spontaneously or were induced by a single injection of 3‐methylcholanthrene or 4‐nitroquinoline‐n‐oxide, 2) transplanted pulmonary tumors in the first and second transfer generations, 3) tumorous growth which developed in subcutaneously transplanted lung tissues, and 4) serially transplanted pulmonary tumors in five established lines.The pulmonary tumor in mice was considered as a progressive tumor and the varieties of histology were classified into five or eight types on the basis of relationship between the histological features and the biological activities. The idea of the progression from benign to malignant stage was presented in the pulmonary tumors in mice by the author for the first time.

INDUCTION OF PULMONARY TUMOURS IN MICE BY MITROSONORNICOTINE, A POSSIBLE CONSTITUENT OF TOBACCO SMOKE.

NITROSAMINES could be formed in cigarette smoke by reactions between secondary amines and oxides of nitrogen1,2. Several secondary amines, including anabasine, nornicotine, dimethylamine, diethylamine, pyrrolidine, piperidine and proline, have been detected in cigarette smoke, which also contains about 200 parts per million of oxides of nitrogen (as the sum of NO and NO2) (refs. 3 and 4). The nitrosamine derivatives of some of these amines have already been shown to be carcinogenic5,6. We have reported that nitrosoanabasine administered in the drinking water induced benign and malignant tumours of the œsophagus of rats1. In parallel with these biological investigations, attempts have been made to detect nitrosoanabasine and nitrosonornicotine in cigarette smoke. So far these attempts have not been successful, although methods capable of detecting small amounts (for example, 5 µg) of specific nitrosamines have been developed1. Lack of success may be due either to instability of nitrosamines under the conditions prevailing in cigarette smoke or to interference in their detection by other constituents of the smoke. Nitrosoanabasine could not be detected in cigarette smoke condensates to which it had been deliberately added nor in the smoke of cigarettes which had been impregnated with the substance before smoking. If these negative results are due to instability of the nitrosamines, the possibility of their biological importance is only ruled out if their half-lives are very short. Further investigations into these problems are in progress.

Pulmonary tumours in mice induced by oral isoniazid and its metabolites.

Juhasz et al.1 induced solitary or multiple pulmonary adenomas in 7 of 45 albino mice which received intraperitoneal injections of isoniazid in water for three months and survived for periods up to 7½ months. Mori et al.2 incorporated the same compound in varying doses in the diet of dd mice for periods of 2, 3 or 4 months, obtaining 100 per cent of pulmonary ‘tumours’ in mice surviving the largest dose for 7½ months. Subcutaneous injection was also effective.

Experimental induction of pulmonary tumors in mice.

Experimental induction of pulmonary tumors in mice.

Elimination of the Effect of the <italic>A<sup>y</sup></italic> Gene on Pulmonary Tumors in Mice by Alteration of Its Effect on Normal Growth

Elimination of the Effect of the <italic>A<sup>y</sup></italic> Gene on Pulmonary Tumors in Mice by Alteration of Its Effect on Normal Growth