Ronald L. Melnick

Hexavalent chromium is carcinogenic to F344/N rats and B6C3F1 mice after chronic oral exposure.

Background Hexavalent chromium [Cr(VI)] is a human carcinogen after inhalation exposure. Humans also ingest Cr(VI) from contaminated drinking water and soil; however, limited data exist on the oral toxicity and carcinogenicity of Cr(VI). Objective We characterized the chronic oral toxicity and carcinogenicity of Cr(VI) in rodents. Methods The National Toxicology Program (NTP) conducted 2-year drinking water studies of Cr(VI) (as sodium dichromate dihydrate) in male and female F344/N rats and B6C3F1 mice. Results Cr(VI) exposure resulted in increased incidences of rare neoplasms of the squamous epithelium that lines the oral cavity (oral mucosa and tongue) in male and female rats, and of the epithelium lining the small intestine in male and female mice. Cr(VI) exposure did not affect survival but resulted in reduced mean body weights and water consumption, due at least in part to poor palatability of the dosed water. Cr(VI) exposure resulted in transient microcytic hypochromic anemia in rats and microcytosis in mice. Nonneoplastic lesions included diffuse epithelial hyperplasia in the duodenum and jejunum of mice and histiocytic cell infiltration in the duodenum, liver, and mesenteric and pancreatic lymph nodes of rats and mice. Conclusions Cr(VI) was carcinogenic after administration in drinking water to male and female rats and mice.

Developmental neurotoxicity of perfluorinated chemicals modeled in vitro.

Background The widespread detection of perfluoroalkyl acids and their derivatives in wildlife and humans, and their entry into the immature brain, raise increasing concern about whether these agents might be developmental neurotoxicants. Objectives We evaluated perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), perfluorooctane sulfonamide (PFOSA), and perfluorobutane sulfonate (PFBS) in undifferentiated and differentiating PC12 cells, a neuronotypic line used to characterize neurotoxicity. Methods We assessed inhibition of DNA synthesis, deficits in cell numbers and growth, oxidative stress, reduced cell viability, and shifts in differentiation toward or away from the dopamine (DA) and acetylcholine (ACh) neurotransmitter phenotypes. Results In general, the rank order of adverse effects was PFOSA > PFOS > PFBS ≈ PFOA. However, superimposed on this scheme, the various agents differed in their underlying mechanisms and specific outcomes. Notably, PFOS promoted differentiation into the ACh phenotype at the expense of the DA phenotype, PFBS suppressed differentiation of both phenotypes, PFOSA enhanced differentiation of both, and PFOA had little or no effect on phenotypic specification. Conclusions These findings indicate that all perfluorinated chemicals are not the same in their impact on neurodevelopment and that it is unlikely that there is one simple, shared mechanism by which they all produce their effects. Our results reinforce the potential for in vitro models to aid in the rapid and cost-effective screening for comparative effects among different chemicals in the same class and in relation to known developmental neurotoxicants.

Fundamental flaws of hormesis for public health decisions.

Hormesis (defined operationally as low-dose stimulation, high-dose inhibition) is often used to promote the notion that while high-level exposures to toxic chemicals could be detrimental to human health, low-level exposures would be beneficial. Some proponents claim hormesis is an adaptive, generalizable phenomenon and argue that the default assumption for risk assessments should be that toxic chemicals induce stimulatory (i.e., “beneficial”) effects at low exposures. In many cases, nonmonotonic dose–response curves are called hormetic responses even in the absence of any mechanistic characterization of that response. Use of the term “hormesis,” with its associated descriptors, distracts from the broader and more important questions regarding the frequency and interpretation of nonmonotonic dose responses in biological systems. A better understanding of the biological basis and consequences of nonmonotonic dose–response curves is warranted for evaluating human health risks. The assumption that hormesis is generally adaptive is an oversimplification of complex biological processes. Even if certain low-dose effects were sometimes considered beneficial, this should not influence regulatory decisions to allow increased environmental exposures to toxic and carcinogenic agents, given factors such as interindividual differences in susceptibility and multiplicity in exposures. In this commentary we evaluate the hormesis hypothesis and potential adverse consequences of incorporating low-dose beneficial effects into public health decisions.

Mutations of ras protooncogenes and p53 tumor suppressor gene in cardiac hemangiosarcomas from B6C3F1 mice exposed to 1,3-butadiene for 2 years

1,3-Butadiene is a multisite carcinogen in rodents. Incidences of cardiac hemangiosarcomas were significantly increased in male and female B6C3F1 mice that inhaled 1,3-butadiene (BD) for 2 years. Eleven BD-induced cardiac hemangiosarcomas were examined for genetic alterations in ras protooncogenes and in the p53 tumor suppressor gene. Nine of 11 (82%) BD-induced hemangiosarcomas had K-ras mutations and 5 of 11 (46%) had H-ras mutations. All of the K-ras mutations were G→C transversions (GGC→CGC) at codon 13; this pattern is consistent with reported results in BD-induced lung neoplasms and lymphomas. Both K- ras codon 13 CGC mutations and H-ras codon 61 CGA mutations were detected in 5 of 9 (56%) hemangiosarcomas. The 11 hemangiosarcomas stained positive for p53 protein by immunohistochemistry and were analyzed for p53 mutations using cycle sequencing of polymerase chain reaction (PCR) amplified DNA isolated from paraffin-embedded sections. Mutations in exons 5 to 8 of the p53 gene were identified in 5 of 11 (46%) hemangiosarcomas, and all of these were from the 200- or 625-ppm exposure groups that also had K-ras codon 13 CGC mutations. Our data indicate that K-ras, H- ras, and p53 mutations in these hemangiosarcomas most likely occurred as a result of the genotoxic effects of BD and that these mutations may play a role in the pathogenesis of BD-induced cardiac hemangiosarcomas in the B6C3F1 mouse.

Toxicities of ethylene glycol and ethylene glycol monoethyl ether in Fischer 344/N rats and B6C3F1 mice.

The toxicities of ethylene glycol (EG) and ethylene glycol monoethyl ether (EGEE) were studied in Fischer 344/N rats and B6C3F1 mice. In a 13-week study, EG was administered in feed to groups of 10 rats and 10 mice of both sexes at dose levels of 0 (control), 0.32, 0.63, 1.25, 2.5 and 5.0%. Kidney/body weight ratios were elevated in the 2.5 and 5.0% dose groups of male and female rats relative to controls, while serum urea nitrogen and serum creatinine levels were elevated in the two highest dose groups of male rats. Toxic nephrosis and crystal deposits in renal tubules were observed in the 2.5 and 5.0% dose groups of male rats. Crystals were also observed in brains of male rats in the 5.0% dose group. Nephrosis was the only lesion observed in female rats (5.0% dose group). Mild, compound-related lesions were seen in kidneys (nephrosis) and livers (centrilobular degeneration) of male mice in the 2.5 and 5.0% dose groups. There were no adverse effects observed in female mice. Groups of 50 rats and 50 mice of both sexes were administered EGEE by gavage in a 2-year study at dose levels of 0 (control), 0.5, 1.0 and 2.0 g/kg body weight. Testicular atrophy was observed in male rats that died early in this study and in the medium- and high-dose male mouse groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Weight of evidence versus weight of speculation to evaluate the alpha2u-globulin hypothesis.

whereas the severity of spontaneous chronic nephropathy in treated male rats relative to controls increased from 52 and 104 weeks. As a result of the current discussion in the literature (3), the kidney specimens were reanalyzed for the hallmarks of x2u-globulin nephropathy sequelae, namely the presence of granular casts and linear mineralization (8). These changes were not observed in male rats receiving 250, 1,000, or 2,000 mg/kg of gabapentin for 2 years. In terms of proliferative effects, there were 3 foci of atypical hyperplasia of renal tubules: 2/50 were found in male rats at 1,000 mg/kg and 1/50 male rats at 2,000 mg/kg of gabapentin. These kidney examinations in the 2-year study included four sections (two from each kidney) from every animal in the group (50/sex/group), amounting to 1,600 kidney sections in the 2-year rat study. This standard procedure seems to adequately sample the kidney. Reference was made to renal accumulation of gabapentin and to its ability to bind a2u-globulin. Metabolic disposition data revealed no differences in the accumulation of gabapentin in the kidney of male and female rats. Gabapentin distributes rapidly to the kidney in both sexes, and approximately 10-12% of the dose is found at 2hr post-dose. The elimination decay is rapid, and 0.20-0.14 gabapentin radioequivalents in micrograms per gram are found at 12 hr post-dose. Full profile toxicokinetic studies have shown linear kinetics without accumulation, and elimination follows rapid clearance from the plasma and organ compartments. Recoveries in the mass balance studies yielded 99% or more of the administered dose. Why such a diverse series of chemicals causes a putatively similar microglobulin nephropathy with (i.e., unleaded gasoline) and without (i.e., gabapentin) nephrocarcinogenesis is intriguing. Evaluation of the gabapentin data does not prove or disprove that a2u-globulin accumulation precedes, promotes, or causes renal cancer in rats over a 2-year constant exposure at toxicity-limiting doses. We propose that although morphofunctional features may appear similar for all documented cases of this xenobioticinduced nephropathy, different mechanisms may be operative. Alternatively, a structural comparison of the x2u microglobulin species would assert this microglobulin not to be the same in all cases when induced by the diverse chemicals. The work by Hildebrand et al. (9) would indicate some support to this notion. In the case of gabapentin, in which we have substantial data on hand, it is difficult to advocate existing mechanisms because of the drugs lack of metabolism or biotransformation, lack of serum protein binding, or lack of significant organ accumulation. It remains unanswered whether data on cell proliferation or specific 4x2uglobulin binding to renal tubule proteins would substantiate current mechanisms. Clearly new data, rather than new assumptions, will give fresh insight to our understanding of the a2u nephropathy puzzle.

Lorenzo Tomatis and primary prevention of environmental cancer.

The leading 20th century proponent for primary prevention of environmental cancer was Dr. Lorenzo Tomatis, the former Director of the International Agency for Research on Cancer and founder of the IARC Monographs program. This paper is dedicated to the memory of Dr. Tomatis – eminent scientist, scholar, teacher, humanitarian, and public health champion - and includes many perspectives that he promoted throughout his career, with original quotations from some of his scientific writings on primary prevention of environmental cancer. Any attempt by us to simply summarize his views would only detract from the power and logic of his language.“Cancer still remains a mainly lethal disease. Primary prevention remains the most relevant approach to reduce mortality through a reduction in incidence”[1].

Chemically Exacerbated Chronic Progressive Nephropathy Not Associated with Renal Tubular Tumor Induction in Rats: An Evaluation Based on 60 Carcinogenicity Studies by the National Toxicology Program

Chronic progressive nephropathy (CPN) is a common age-related degenerative-regenerative disease of the kidney that occurs in both sexes of most strains of rats. Recently, claims have been made that enhanced CPN is a mode of action for chemically induced kidney tumors in male rats and that renal tubular tumors (RTTs) induced by chemicals that concomitantly exacerbate CPN are not relevant for human cancer risk assessments. Although CPN is an observable histopathological lesion that may be modified by diet, the etiology of this disease and the mechanisms for its exacerbation by chemicals are unknown, and it fails to meet fundamental principles for defining carcinogenic modes of action and human relevance. Our comprehensive evaluation of possible relationships between exacerbated CPN and induction of RTTs in 58 carcinogenicity studies, conducted by the National Toxicology Program, in male and 11 studies in female F344 rats using 60 chemicals revealed widespread inconsistency in the claimed association. Because the proposed hypothesis lacks evidence of biological plausibility, and due to inconsistent relationships between exacerbated CPN and kidney tumor incidence in carcinogenicity studies in rats, dismissing the human relevance of kidney tumors induced by chemicals that also exacerbate CPN in rats would be wrong.

War on Carcinogens: Industry Disputes Human Relevance of Chemicals Causing Cancer in Laboratory Animals Based on Unproven Hypotheses, Using Kidney Tumors as an Example

Abstract Evidence from studies in animals is essential for identifying chemicals likely to cause or contribute to many diseases in humans, including cancers. Yet, to avoid or delay the implementation of protective public health standards, the chemical industry typically denies cancer causation by agents they produce. The spurious arguments put forward to discount human relevance are often based on inadequately tested hypotheses or modes of action that fail to meet Bradford Hill criteria for causation. We term the industry attacks on the relevance of animal cancer findings as the “War on Carcinogens.” Unfortunately, this tactic has been effective in preventing timely and appropriate health protective actions on many economically important yet carcinogenic chemicals, including: arsenic, asbestos, benzene, 1,3-butadiene, formaldehyde, methylene chloride, phthalates, tobacco usage, trichloroethylene [TCE], and others. Recent examples of the “War on Carcinogens” are chemicals causing kidney cancer in animals. Industry consultants argue that kidney tumor findings in rats with exacerbated chronic progressive nephropathy (CPN) are not relevant to humans exposed to these chemicals. We dispute and dismiss this unsubstantiated claim with data and facts, and divulge unprofessional actions from a leading toxicology journal.

Environmental justice and primary prevention of cancer: the odyssey and legacy of lorenzo tomatis.

Lorenzo Tomatis [1929–2007] devoted his private and professional life to the betterment of mankind. As a physician, scientist, and humanitarian he championed against the plight of social injustice and promoted the obvious benefits of primary prevention of diseases compared to treatments that prevent or delay disease progression, especially occupational cancers. An avowed student and scholar of literature, the arts, the history of medicine and science, and chemical carcinogenesis, he believed in and wrote about these issues throughout his storied life. Some of his achievements, with excerpts from his writings, especially on primary prevention and on social injustice, are highlighted herein.