Neutron radiation can activate K-ras via a point mutation in codon 146 and induces a different spectrum of ras mutations than does gamma radiation.

Abstract

Neutron radiation is known to produce tumors in animals and cause ceH transformation. We Animal model systems have provided a means by which oncogene activation can be studied in tumors in which the inducing agent and genetic background are well controlled (8,16,21). We have been studying ras oncogene activation in murine thymic lymphomas induced either by the chemical carcinogen N-nitroso-N-methylurea or by gamma radiation and have found activated K-and N-ras genes in a significant proportion of the tumors (7,20). In this study, we wanted to determine the extent of ras oncogene activation in neutroninduced thymic lymphomas in RF/J mice and compare it with our previous analysis of gamma ray-induced thymic lymphomas in the same strain of mouse.Gamma and neutron radiation are two types of ionizing radiation that deliver energy to target molecules in very different ways (13 (23,24). As yet, however, activation of specific oncogenes in neutron-induced tumors or cell transformants has not been described. Biologically significant consequences of the different actions of neutron and gamma radiation can be studied by comparing the activating point mutations of ras oncogenes that result from tumors induced by the different radiation treatments.To determine an optimal dose of neutron radiation that would induce thymic lymphomas with high frequency, we irradiated female RF/J mice (The Jackson Laboratory, Bar Harbor, Maine) between 4 and 7 weeks old with a single dose of 0.8, 1.0, 2.0, or 3.0 Gy of whole-body 0.44-MeV neutron radiation. Neutrons were produced from a Van de Graaff particle accelerator. Uniform whole-body neutron irradiation was delivered to mice restrained in Plexiglas cylinders and rotated around the neutron source. To correct for possible fluctuations, after one-half of the dose was administered, the tubes were rotated by 1800. The complete dose was administered in a period of 2 to 6 h. Mice were sacrificed when they appeared visibly ill and had palpable lymph nodes. The only effective dose was 1.0 Gy. All of the mice given 2.0 and 3.0 Gy of neutrons died within 2 weeks from radiation sickness.

Activating K-Ras mutations outwith ‘hotspot’ codons in sporadic colorectal tumours – implications for personalised cancer medicine

“…Consistent with our own data, codon 146 was the most frequently mutated of the mutations described. K-Ras codon 146 mutations have been described in a human colorectal tumour cell line (Higashi et al, 1990), in mice with thymic lymphomas (Sloan et al, 1990) and in a single human colorectal tumour case report (Orita et al, 1991). In more detailed molecular profiling studies, codon 146 mutations were described in 4% of two independent colorectal tumour series from Hong Kong and the United States of America (Edkins et al, 2006) and in chronic myelomonocytic leukaemia (Gelsi-Boyer et al, 2008;Tyner et al, 2009).…”

Inactivation of the cyclin-dependent kinase inhibitor p15INK4b by deletion and de novo methylation with independence of p16INK4a alterations in murine primary T-cell lymphomas

“…Di erent types of evidence for the involvement of DNA methylation in tumorigenesis and tumor progression have been published. In recent years, molecular links between the ras signaling pathway, frequently activated in murine lymphomas (Guerrero et al, 1984a,b;Diamond et al, 1988;Sloan et al, 1990), and cell cycle regulation (Winston et al, 1996) or DNA methylation (MacLeod et al, 1995) have been described. DNA methyltransferase (DNA-MTase) activity has been found to increase over the progressive stages of human colon cancer and overexpression of the mammalian DNAMTase gene or the bacterial HhaI DNA-MTase results in transformation of NIH3T3 cells (Wu et al, , 1996.…”

“…Thymic lymphomas were induced by whole-body gamma irradiation (Diamond et al, 1988;Santos et al, 1996), neutron irradiation (Sloan et al, 1990) or intraperitoneal injection of MNU (Newcomb et al, 1988;Diamond et al, 1988). Brie¯y, ionizing gamma radiation was given in 4 weekly doses of 175 rads/dose, high LET ionizing neutron radiation was applied once with 1.0 Gy of 0.44 MeV neutrons, and MNU treatment consisted of 5 consecutive weekly injections of 30 mg/kg doses.…”

“…All these agents are known to be tumorigenic in mouse systems producing monoclonal T-cell lymphomas (Newcomb et al, 1988), where K-ras and N-ras genes are commonly mutated (Guerrero et al, 1984a,b: Diamond et al, 1988Sloan et al, 1990). In this study, we show that although the homozygous deletions and base substitutions in these INK4 genes are not very frequent, the murine p15 INK4b and, to less extent, p16 INK4a genes are frequently inactivated by methylation and allelic losses in the g-and neutronradiation-induced lymphomas.…”

ras activation in human tumors and in animal model systems.

“…Prior to assaying for a ras mutation, the exon containing the possible ras mutation was amplified by the polymerase chain reaction (PCR) (8)(9)(10). In the first assay, the amplified samples were then affixed to nylon membranes using a slot blot apparatus, and the membranes were hybridized to 32P-labeled oligonucleotides specific for the relevant point mutations.…”