Innate Toxicity Research Articles

Analysis of adenovirus sequestration in the liver, transduction of hepatic cells, and innate toxicity after injection of fiber-modified vectors.

After intravenous administration, adenovirus (Ad) vectors are predominantly sequestered by the liver. Delineating the mechanisms for Ad accumulation in the liver is crucial for a better understanding of Ad clearance and Ad-associated innate toxicity. To help address these issues, in this study, we used Ad vectors with different fiber shaft lengths and either coxsackievirus-Ad receptor (CAR)-interacting Ad serotype 9 (Ad9) or non-CAR-interacting Ad35 fiber knob domains. We analyzed the kinetics of Ad vector accumulation in the liver, uptake into hepatocytes and Kupffer cells, and induction of cytokine expression and release in response to systemic vector application. Immediately after intravenous injection, all Ad vectors accumulated equally efficiently in the liver; however, only genomes of long-shafted Ads were maintained in the liver tissue over time. We found that Kupffer cell uptake of long-shafted Ads was mediated by the fiber knob domain and was CAR independent. The short-shafted Ads were unable to efficiently interact with hepatocellular receptors and were not taken up by Kupffer cells. Moreover, our studies indicated that Kupffer cells were not the major reservoir for the observed accumulation of Ads (used in this study) in the liver within the first 30 min after virus infusion. The lower level of liver cell transduction by short-shafted Ads correlated with a significantly reduced inflammatory anti-Ad response as well as liver damage induced by the systemic administration of these vectors. This study contributes to a better understanding of the biology of systemically applied Ad and will help in designing safer vectors that can efficiently transduce target tissues.

A Tumor-Targeted and Conditionally Replicating Oncolytic Adenovirus Vector Expressing TRAIL for Treatment of Liver Metastases

We have constructed a new capsid-modified adenovirus (Ad) vector that specifically replicates in tumor cells and expresses TNF-related apoptosis-inducing ligand (TRAIL). The Ad capsid contains short-shafted fibers derived from Ad serotype 35, which allow for efficient infection of malignant tumor cells, and largely avoids innate toxicity after intravenous application. Replication-dependent homologous recombination in Ad genomes was used to achieve tumor-specific expression of Ad E1a (to mediate viral replication) and TRAIL (to mediate apoptosis and enhance release of progeny virus from infected cells). We demonstrated that our oncolytic vector (Ad5/35.IR-E1A/TRAIL) induced apoptosis in human tumor cell lines derived from colorectal, lung, prostate, and liver cancer. Both in vitro and in vivo tumor models showed efficient intratumoral spread of this vector. In a model for metastatic colon cancer, tail vein infusion of Ad5/35.IR-E1A/TRAIL resulted in elimination of preestablished liver metastases. Intravenous injection of this vector caused a transient elevation of serum glutamic pyruvic transaminase in tumor-bearing mice, which we attributed to factors released from apoptotic tumor cells. Liver histology analyzed at day 14 after virus injection did not show signs of hepatocellular damage. This new oncolytic vector represents a potentially efficient means for gene therapy of metastatic cancer.

Clinical reversal of drug resistance

Although combination chemotherapy has had a significant impact on survival for malignancies such as Hodgkin's disease, testicular cancer, and childhood acute leukemias, the majority of cancers are either initially resistant to chemotherapy (renal, colon, ect.) or are initially chemosensitive but acquire resistance during treatment, such as lymphoma and breast cancer. Resistance to chemotherapy remains an obstacle to the successful treatment of human cancer and has been the subject of numerous investigations aimed at identifying the molecular mechanisms of resistance in cancer cells. An improved understanding of the mechanisms by which tumor cells develop resistance to chemotherapy may not only enhance the activity of cytotoxic therapy in advanced malignancies but may ultimately improve the impact of adjuvant therapy, potentially resulting in prolonging disease-free intervals and survival. In this review, therefore, we discuss our current understanding of the MDR1 gene, encoding P-glycoprotein, which is responsible for one mechanism of multidrug resistance (MDR). We also review the evidence supporting the clinical relevance of the MDR1 gene and clinical trials aimed at reversing MDR-mediated resistance. Although MDR-mediated drug resistance has been well characterized in preclinical models, its role in clinical drug resistance is not as well characterized and requires further investigation. Prospective studies are necessary to establish the role of MDR1 gene expression in the clinical resistance. The ability to identify tumors with increased MDR1 gene expression has several potential applications (for example, the prediction of response to chemotherapy and the design of studies aimed at reversal of resistance with agents that inhibit MDR-mediated drug efflux). The initial goal of such trials is to demonstrate the ability to reverse MDR1-mediated drug resistance in the appropriate advanced refractory malignancies. Ultimately, it will be important to incorporate these reversal strategies in the treatment of early-stage disease, at which time the tumor burden is smaller and fewer mechanisms of resistance may be present. Prospective phase I, II, and III clinical trials using reversing agents in conjunction with chemotherapy in malignancies that express the MDR1 gene, such as the hematologic malignancies and breast cancer, are necessary before routine use of agents such as verapamil, quinidine, and cyclosporine, which carry innate toxicities. MDR is a mechanism of drug resistance that provides the potential for an alteration in drug efflux, which may have a significant impact on response and possibly result in improved survival for some cancer, patients.

Rapid generation of human T cell hybridomas

Using the concept of a selectable surface marker we have generated screenable human T cell hybridomas within 4–7 days of fusion. OKT4 + T cell blasts were fused with MOLT4, an OKT4 − T cell line, and OKT4 + fusion products were obtained by indirect rosetting technique. Supernatants of this heterogeneous population of hybrids were screened and those with activities of interest were subjected to cloning of soft agar and re-screening to dissect out various activities. Fusion efficiency, by staining, ranges from 6 to 80% depending on the state of activation of the T blasts and the growth phase of the MOLT4 line. This method allows for immoryalization of T cell subpopulations, clones and malignant cells to study factor production and potentially messenger RNA for these factors. Additionally selectable surface markers may be used in human B cell and rodent: human fusions, avoiding the innate toxicity of various selection media.

Toxicity to Boophilus microplus of formamidine acaricides and related compounds, and modification of toxicity by certain insecticide synergists.

Twenty-nine formamidines and related compounds displayed activity toward engorged female southern cattle ticks, as judged by reduction in oviposition and egg viability, but only the most toxic adulticides showed activity against larvae. Certain methylenedioxyphenyl compounds markedly antagonized the toxicity of the aryl N, N-dimethylformamidines to larvae, but in most cases they synergized the toxicity of the aryl N-methylformamidines. This fact suggested that the aryl N, N-dimethylformamidines possessed little if any innate toxicity to cattle tick larvae. The high toxicity of 2 diaryl methyl triazapentadiene compounds to engorged females and larvae was attributed in part to their ability to hydrolyze to their respective aryl N-methylformamidine derivatives.

Age-dependent variations in the response of several species of diptera to insecticidal chemicals

The toxicity of carbaryl to three species of fleshflies Sarcophaga bullata Parker, S. crassipalpis Macquart, and S. argyrostoma (Robineau-Desvoidy) varied considerably with age and sex. In contrast, the susceptibility of a blowfly, Phormia regina (Meigen) to carbaryl decreased with age and that of two muscid flies, Musca autumnalis DeGeer and Stomoxys calcitrans (L.), remained relatively constant. The synergistic activity of piperonyl butoxide varied inversely with the innate toxicity of carbaryl to each species suggesting that the observed age- and sex-dependent variations in carbaryl toxicity result mainly from differences in detoxifying capability. This was supported by in vitro measurements of oxidative microsomal metabolism. It was further established that differences in the rates of penetration and excretion of carbaryl and in the susceptibility of the head cholinesterase to carbaryl inhibition were of little importance in determining the susceptibility of the flies to this insecticide.

Selenium and tellurium in mice. Effects on growth, survival, and tumors.

In order to ascertain innate toxicities of selenium and tellurium, 427 mice were given selenite, selenate, tellurite, and tellurate in drinking water for life, at 3 ppm selenium and 2 ppm tellurium, in an environment controlled as to contaminating trace elements. There were 209 controls. No signs of toxic effects in terms of growth, survival, or longevity appeared in males. Selenite-fed males were heavier and females were lighter than their controls at various ages, and female longevity was decreased. No toxic effects were observed in the groups fed tellurate, but in females fed tellurite longevity was decreased. Selenium level was increased in tissues when selenium was fed. None of the four compounds affected the incidence of spontaneous tumors significantly. Amyloidosis occurred more frequently in mice fed selenium than in controls or those fed tellurium.

Toxic effects of trace elements on the reproduction of mice and rats.

Breeding mice and rats were exposed to low doses of six trace elements in drinking water in an environment controlled as to contaminating trace metals. Each group was carried through three generations. Compared to control mice given only doubly deionized wafer, selenafe (3 ppm selenium) resulted in excess deaths before weaning, runts, and failures to breed. Lead (25 ppm) and cadmium (10 ppm) resulted in loss of the strain in two generations with many abnormalities. Molybdate (10 ppm molybdenum) was slightly toxic in this respect, and arsenic resulted only in elevated ratios of males to females. In rats, lead was very toxic, and titanium and nickel moderately toxic, resulting in many early deaths and runts. This method provides fairly rapid estimates of innate toxicities of trace elements in doses tolerable for growth and survival.

Temperature Relationships in Organophosphorus Poisoning in Boll Weevils12

A laboratory culture or organophosphorus-susceptible boll weevils was exposed at different temperatures to cotton leaves or filter-paper discs previously treated with insecticide or defoliant. All materials examined showed a positive temperature coefficient of toxicity with methyl parathion and malathion being the most toxic at all temperatures. The toxicities of azinphosmethyl and EPN® (O-ethyl O-p-nitrophenyl phenylphosphonothioate) were severely decreased at 10°C while DEF® (S, S, S-tributyl phosphorotrithioate) showed appreciable toxicity only at 32°C. Studies involving different exposure and holding temperatures indicated that a higher exposure temperature results in increased toxicity, even though the holding temperature is the same for all treatments. Although transferring the insects to a low holding temperature from a high exposure temperature decreased the effectiveness of the toxicant, the corrected mortality of the insects exposed at 32°C and held at 10°C was more than twice that of those exposed and held at 10°C. These data suggest that temperature may affect both the innate toxicity and the rate of absorption of the insecticide into the insect.

Zirconium, Niobium, Antimony, Vanadium and Lead in Rats: Life term studies

To evaluate innate effects of the trace elements zirconium, niobium, antimony and vanadium, and to reevaluate those of lead, 603 rats of the Long-Evans strain were fed a diet containing relatively small amounts of these elements in an environment reasonably free of trace contaminants. Groups of 100 or more divided as to sex were given 5 ppm (as metal) either zirconium, niobate, antimonite, or vanadyl ions, and 25 ppm lead (males only) in drinking water from the time of weaning until natural death, and compared with an equal number of controls. Chromium 1 ppm was in the water. These doses were tolerable for growth which was enhanced in the male niobium group. Innate toxicity in terms of life span and longevity occurred in the antimony groups. In rats given antimony, nonfasting serum glucose levels were lower than fasting, an unusual finding. Increased incidences of glycosuria occurred in the zirconium, niobium and lead groups. Serum cholesterol was abnormal in the antimony and vanadium groups. Antimony and lead accumulated in soft tissues, the former with age. No element was tumorigenic. Lead-fed males lost weight from 24 to 30 months of age and their coats were poor, the only signs of toxicity. A previous series of chromium-deficient rats given the same dose of lead showed early mortality, shortened life span and decreased longevity. Chromium may be antagonistic to lead toxicity.

Germanium, Tin and Arsenic in Rats: Effects on growth, survival, pathological lesions and life span

To evaluate innate effects of the trace elements germanium, tin and arsenic, 455 rats of the Long-Evans strain were fed a diet containing small amounts of these elements in an environment relatively free of trace contaminants. Groups of 100 or more, divided as to sex, were given 5 ppm germanate, arsenite or stannous ions in drinking water from weaning until natural death and compared with an equal number of controls. These levels were tolerable for growth. Innate toxicity in terms of life span and longevity occurred in females given tin and both sexes given germanium; increased incidences of fatty degeneration of the liver were observed for both these elements. Animals fed tin also showed an increased incidence of vacuolar changes in their renal tubules. Large amounts of arsenic accumulated in tissues, especially aorta and red blood cells, with no signs of toxicity. Tin accumulated to some extent and germanium less so. No element was tumorigenic or carcinogenic, there being somewhat fewer tumors in rats fed germanium than in the controls.

Zirconium, Niobium, Antimony and Fluorine in Mice: Effects on growth, survival and tissue levels

To evaluate innate effects of certain trace elements, 540 mice were fed a diet of rye, corn oil and dried skim milk containing moderate amounts of zirconium and niobium and no detectable antimony or fluorine, in an environment relatively free of trace contaminants. Groups of 108, divided as to sex, were given 5 ppm zirconium, niobate or antimony or 10 ppm fluoride in drinking water from weaning until natural death. Females given fluoride grew somewhat larger at older ages and both sexes survived as long as their controls. Inherent toxicity, manifest by decreased median life spans and longevity and by some suppression of growth of older animals, was observed in females given niobium and antimony. The feeding of niobium was associated with an increased incidence of hepatic fatty degeneration. No element was carcinogenic or tumorigenic. Fluoride did not accumulate in soft tissues, but increments of niobium and antimony were found. Zirconium occurred in both controls and experimental groups, and showed slight toxicity. Of 15 trace elements studied in this manner, chromium, fluorine and nickel showed no demonstrable innate toxicity, whereas tellurium, arsenic, tin and vanadium showed the most.

Arsenic, Germanium, Tin and Vanadium in Mice: Effects on growth, survival and tissue levels

Certain trace elements found in the human environment may produce adverse biological effects when fed in small amounts to rats and mice for their lifetime. Others may be inert or have favorable actions. Mice numbering 643 were fed a diet of rye, corn oil and dried skim milk containing small amounts of arsenic, germanium and tin and larger amounts of vanadium, in an environment designed to exclude trace contaminants. Groups of 52 to 108 or more divided as to sex were given 5 ppm of these elements as arsenite, germanate, vanadyl or stannous ions in drinking water from weaning until natural death. Chromium (1 ppm) was added to the water of those given arsenic and germanium and their controls. No element but chromium affected growth. Median life spans and longevity of the oldest 10% of males given germanium and of both sexes given arsenic were somewhat shortened. This innate toxicity was not observed in animals given tin and vanadium. All elements accumulated in one or more organs, germanium and tin in spleen with age. No element was carcinogenic. Life-term exposures of animals to trace elements may show inherent toxicities at levels tolerable to young animals.

Chromium, Lead, Cadmium, Nickel and Titanium in Mice: Effect on Mortality, Tumors and Tissue Levels

About 700 mice were given throughout their lives 5 ppm cadmium, lead, chromium, nickel or titanium in drinking water while fed a diet deficient in cadmium and low in other metals. Tissue concentrations comparable to those of man were observed. Sex differences appeared. Mortality of females was unaffected. Mortality of males on cadmium, lead and nickel was increased compared with that of the chromium group, and of those on cadmium and lead compared with the controls. Longevity of the oldest 10% of both sexes was less in lead and titanium groups, and of males in cadmium group, compared with controls. No metal was carcinogenic; incidence of tumors in males on cadmium and lead, and in females given nickel was decreased. Body weights at death were greater in titanium, chromium and lead groups. Large increments of titanium, moderate increments of cadmium and nickel, and lesser increments of chromium and lead were observed in 5 organs, compared with controls. Higher concentrations of cadmium and titanium occurred in younger mice and the other metals did not increase markedly in tissues with age. All metals except chromium exhibited one or more signs of innate toxicity.

The susceptibility of tsetse flies to topical applications of insecticides. II.—Young adults of Glossina morsitans Westw. and organophosphorus compounds, pyrethrins and Sevin

SummarySolutions of six organophosphorus compounds, Sevin, and pyrethrins (alone or synergised with piperonyl butoxide) were applied by microburette in drops of constant volume (0·0216 μl.) to the dorsum of the thorax of young adults of Glossina morsitans Westw., 2–5 days old that had taken their first blood-meal the previous day. The solvents used were decalin (decahydronaphthalene), toluene and lighting kerosene, respectively.Malathion, methyl-parathion, DDVP (dichlorvos) and Sevin were eliminated in preliminary tests as insufficiently toxic. Muscatox (coumaphos) was reasonably toxic but not readily soluble and was therefore not considered further. Diazinon and Baytex (fenthion) were fully evaluated; the former was about as lethal as DDT, the latter as γ BHC (LD50 about 0·004 μg.). The LD50 of diazinon for males (0·0115 μg.) was significantly smaller than that for females (0·016 μg.) but the LD95 was much the same for both sexes.Pyrethrins were about equitoxic with dieldrin (LD50, 0·002 μLg.); when synergised with 15 parts of piperonyl butoxide to one of pyrethrins the LD50 for males was reduced to less than half this value, and there was a significant difference in susceptibility of the sexes, the LD50 for females being 1·4 times that for males. However, the slopes of the regression lines were such that at LD95 the difference between synergised and plain pyrethrins was too small to be of any practical use.These results show that, judged by innate toxicity to young flies, none of these insecticides can compete with dieldrin or Telodrin for practical control, although Baytex is a useful reserve should Glossina acquire resistance to chlorinated hydrocarbons.

Synergism of Carbamate Insecticides Against Cabbage Looper and Beet Armyworm Larvae1

Piperonyl butoxide and Monsanto 16226 (bis (2,3,3,3-tetra-chloropropyl) ether) were tested as synergists in combination with Sevin® (1-naphthyl N -methylcarbamate), Hercules 5727 ( m -iso-propylphenyl N -methylcarbamate) and Zectran® (4-dimethylamino-8,5-xylyl N-methylearbamate). In 1: 1 mixtures with piperonyl butoxide, tested against larvae of the cabbage looper, Trichoplusia ni (Hubner), Sevin was potentiated 10.3-fold, Hercules 5727, 2.6-fold, and Zedran, not at all. Against larvae of the beet armyworm, Spodoptera exigua (Hubner), Sevin and Zeetran were potentiated 4.4-fold and 1.2-fold, respectively. An increase in the ratio of piperonyl butoxide to carbamate gencrally provided a corresponding increase of potentiation against the cabbage looper. Monsanto 16226, with a relatively high innate toxicity against both species, was not considered as efficient a synergist as piperonyl butoxide.