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A recombinant Eimeria protozoan protein antigen (rEA) has been shown to have antitumor and antiviral activity. The purpose of this study was to determine the effect of rEA treatment alone or in combination with an agonist cocktail consisting of granulocyte macrophage colony stimulating factor (GM-CSF), interferon gamma (IFN-γ), interleukin 4 (IL-4), and anti CD-40 antibody, in the treatment of Banzi virus (BV) disease in BALB/c mice. Treatment with rEA resulted in a significant increase in survival, weight gain, and mean day to death in BV-infected mice and resulted in a significant decrease in brain virus titer. Treatment with rEA, in combination with a 4-agonist cocktail, improved disease parameters to a greater degree than rEA treatment alone. The effect of treatment with a reduced concentration of agonist cocktail or fewer components of the agonist cocktail, in combination with rEA, on disease outcome in BV-infected mice was also investigated. Treatment with rEA, alone or in combination with agonist cocktail, 24 hours after virus challenge did not improve disease. Treatment with rEA, alone or in combination with an agonist cocktail, is efficacious for the prophylaxis of BV infection in mice.

The distributions for human disease-specific mortality exhibit two striking characteristics: survivorship curves that intersect near the longevity limit; and, the clustering of best-fitting Weibull shape parameter values into groups centered on integers. Correspondingly, we have hypothesized that the distribution intersections result from either competitive processes or population partitioning and the integral clustering in the shape parameter results from the occurrence of a small number of rare, rate-limiting events in disease progression. In this report we initiate a theoretical examination of these questions by exploring serial chain model dynamics and parameteric competing risks theory. The links in our chain models are composed of more than one bond, where the number of bonds in a link are denoted the link size and are the number of events necessary to break the link and, hence, the chain. We explored chains with all links of the same size or with segments of the chain composed of different size links (competition). Simulations showed that chain breakage dynamics depended on the weakest-link principle and followed kinetics of extreme-values which were very similar to human mortality kinetics. In particular, failure distributions for simple chains were Weibull-type extreme-value distributions with shape parameter values that were identifiable with the integral link size in the limit of infinite chain length. Furthermore, for chains composed of several segments of differing link size, the survival distributions for the various segments converged at a point in the S(t) tails indistinguishable from human data. This was also predicted by parameteric competing risks theory using Weibull underlying distributions. In both the competitive chain simulations and the parametric competing risks theory, however, the shape values for the intersecting distributions deviated from the integer values typical of human data. We conclude that rare events can be the source of integral shapes in human mortality, that convergence is a salient feature of multiple endpoints, but that pure competition may not be the best explanation for the exact type of convergence observable in human mortality. Finally, while the chain models were not motivated by any specific biological structures, interesting biological correlates to them may be useful in gerontological research.

Toll-like receptor (TLR) ligands are attractive adjuvant candidates in vaccine development. Eimeria tenella profilin-like protein has recently been shown to be a potent agonist of the innate immune system through its recognition by Toll-like receptor-11. In this report, we studied the systemic and mucosal adjuvant activity of Eimeria profilin-like protein within a vaccinal strategy against Toxoplasma gondii in mice. Using intraperitoneal (i.p.) immunization, we observed that coadministration of the recombinant Eimeria antigen (rEA) with T. gondii antigen (TAg) effectively elevates plasma levels of IL-12p70 and consequently induced both enhanced specific humoral and Th1 cellular immune responses. The co-administration of TAg plus rEA by i.p route significantly enhanced the protection against T. gondii infection (62% brain cyst reduction) in comparison with control mice and with mice immunized with TAg alone (only 36% brain cyst reduction). After intranasal immunization, humoral and cellular responses were weak. However mice immunized nasally with TAg plus rEA were significantly protected with 50% of brain cyst reduction, conversely TAg immunized mice did not present any brain cyst reduction.These results indicate that Eimeria profilin-like protein would serve as an efficacious systemic and mucosal adjuvant inducing protective immune response against chronical stage of T. gondii infection through TLR11 activation.

A periodic pattern was discovered among the relative number of deaths per year for the major age-related diseases in humans. This pattern was observed among 178 causes-of-death composed of male and female. White (U.S.A.), Black (U.S.A.) and Japanese (Japan) populations. Both descriptive (Fourier transform) and statistical analysis procedures revealed evidence that the pattern resulted from the presence of an underlying power progression between disease prevalences, with a factor of approximately 2.0 between sizes. This periodicity was highly unlikely to have occurred by chance (P much less than 0.01). The presence of such a power progression suggests that the sizes of the various fractions of the total population, that are afflicted with these mortal diseases, are determined by forces intrinsic to the population and probably result from a common mechanism.

Recombinant Eimeria antigen (rEA) has been shown to have potent anticancer and antiviral activity in respective mouse disease models, presumably through robust immune stimulation that occurs via TLR11, a pattern recognition receptor that recognizes profilin-like proteins expressed on apicomplexan protozoans. Comparable immunostimulatory activity in other species has yet to be demonstrated. Since rEA is known to be highly effective in treating Punta Toro virus (PTV) infection in mice, its ability to elicit protective immunity in the hamster PTV infection model was investigated. rEA was given alone, or in combination with IL-18 or IL-2, and virally challenged hamsters were observed for mortality. Cytokine transcript profiles for IL-12p40, IL-21, IFN-gamma and TNF-alpha were assessed to evaluate the induction of these inflammatory mediators known to be induced in mice following exposure to rEA. A dose of 100 microg of rEA, given once 4 h prior to viral challenge, and a second time on day 3 of the infection, was found to be the most effective prophylactic therapy protecting 60% of treated hamsters from mortality, compared to only 5-10% observed in animals receiving placebo. Increased expression of IFN-gamma and IL-12p40 was evident following treatment with rEA. The data suggest that rEA does induce host antiviral responses in hamsters that result in significant protection from death, although determining the most appropriate dose for intervention in other species, including humans, will likely be challenging.

Human mortality distributions were analyzed for 29 disease-specific causes-of-death in male and female, White (U.S.A.), Black (U.S.A.) and Japanese (Japan) populations, constituting a total of 162 separate cohorts. For each cohort distribution, the curve moments and the parameter values for fits to model equations were determined. The differences between cohort distributions were characterized by two degrees of freedom, related to distribution position and shape, respectively. A form of the Weibull function was shown to contain two parameters that mapped to these two degrees of freedom. Parametric analysis on 136 best-fitting cohorts yielded periodic clustering in the set of values for both Weibull parameters as quantitated using a Fourier transform method and an independent statistical method. This periodicity was unlikely to have occurred by chance (P less than 0.01). We have combined these results into a Law of Mortality, based on a Weibull function containing only integer parameters and constants, which is valid for all human age-related disease mortality. We show that the life expectancy differences between races and sexes is completely described by this formalism. We conclude that human mortality is controlled by discrete events, which are manifested in the appearance of only allowed mortality curve shapes and positions.

A protein antigen from an Eimeria protozoan has recently been reported to induce antitumor activity in mice. This activity most likely results from the strong induction of interkeukin-12 (IL-12) and gamma interferon (IFN-γ), which are also essential factors in the establishment of protective immunity against viral infection. We evaluated recombinant Eimeria antigen (rEA) as a potential immunotherapeutic agent in mouse and hamster models of acute phleboviral disease. Punta Toro virus (PTV) was highly sensitive to a single dose of nanogram quantities of rEA in the mouse infection model. Intraperitoneal treatment with rEA also reduced virus load and liver damage associated with PTV infection. IL-12 was elicited following exposure of uninfected mice to quantities of rEA of 10 ng or greater, and the levels peaked at between 3 and 8 h postexposure. IFN-γ release was induced more slowly and required less rEA (1 ng) to produce a significant rise in systemic levels. The induction of IL-12 and IFN-γ involved in the coordination of innate and adaptive immune responses to microbial pathogens required myeloid differentiation factor 88, a signaling adaptor shared by most members of the Toll-like receptor (TLR) family. Despite encouraging results in the murine system, rEA failed to protect hamsters challenged with PTV. Our findings suggest that hamsters may lack functional TLR11, which has recently been shown to recognize a profilin-like protein homologous to rEA from the protozoan Toxoplasma gondii . Further investigation into the immunostimulatory capacity of rEA in other mammalian systems is necessary.

The small intestine (SI) of vertebrates exhibits low tumorigenesis and rarely supports metastatic growth from distant tumors. Many theories have been proposed to address this phenomenon, but none has been consistently supported. One candidate mechanism is that the vast immunologic compartment of the SI provides a heightened level of tumor immunosurveillance. Consistent with this, we have identified a molecule of low abundance from bovine SI that has the hallmarks of a potent immunostimulant and may be associated with the natural suppression of cancer in the intestinal tract. The protein originates from an endemic gut protozoan, Eimeria spp., and is homologous to the antigen 3-1E previously isolated from the avian apicomplexan E. acervulina. We show here that it is a very potent stimulator of IL-12 release from dendritic cells, upregulates inflammatory modulators in vivo (IL-12, MCP-1, IL-6, TNF-alpha and INF-gamma) and has antitumor properties in mice. In addition, it is synergistic in vitro with anti-CD40 antibody, IFN-gamma, IL-4 and GM-CSF; is active across species barriers in vivo; and has no observable toxicity. Based on these activities, we speculate that it is an inducer of protozoan-targeted innate immunity, which may explain its potential benefit to the intestinal tract and potency as an agent in cancer immunotherapy.

Techniques from cancer epidemiology and time series analysis were used to explore the hypothesis that cosmic radiation can induce germ cell changes leading to increases in future breast cancer mortality. A birth cohort time series for female breast cancer mortality was obtained using a model-independent, age-period-cohort analysis on age-specific mortality data for 1940–1990. The birth cohort series contained several oscillatory components, which were isolated and compared to the corresponding frequency components of a cosmic ray surrogate time series – Greenland ice-core 10Be concentrations. A technique, referred to as component wave-train alignment, was used to show that the breast cancer and cosmic ray oscillations were phase-locked approx. 25 years before the time of birth. This is consistent with the time of germ cell formation, which occurs during the fetal development stage of the preceding generation. Evidence is presented that the observable oscillations in the birth cohort series were residues of oscillations of much larger amplitude in the germ cell cohort, which were attenuated by the effect of the broad maternal age distribution. It is predicted that a minimum of 50% of breast cancer risk is associated with germ cell damage by cosmic radiation (priming event), which leads to the development of individuals with a higher risk of breast cancer. It is proposed that the priming event, by preceding other steps of carcinogenesis, works in concert with risk factor exposure during life. The priming event is consistent with epigenetic changes such as imprinting.

The Gompertz and Weibull functions are compared with respect to goodness-of-fit to human mortality distributions; ability to describe mortality curve intersections; and, parameter interpretation. The Gompertz function is shown to be a better descriptor for 'all-causes' of deaths and combined disease categories while the Weibull function is shown to be a better descriptor of purer, single causes-of-death. A modified form of the Weibull function maps directly to the inherent degrees of freedom of human mortality distributions while the Gompertz function does not. Intersections in the old-age tails of mortality are explored in the context of both functions and, in particular, the relationship between distribution intersections, and the Gompertz ln[R0] versus alpha regression is examined. Evidence is also presented that mortality intersections are fundamental to the survivorship form and not the rate (hazard) form. Finally, comparisons are made to the parameter estimates in recent longitudinal Gompertzian analyses and the probable errors in those analyses are discussed.

The dying off of human population cohorts due to different age related diseases rigorously follows stochastically derived rules. The existence of such rules suggests a common, fundamental mechanism for human mortality. We show here, by a simple quantitative analysis, that the progressive loss with age of genetic triplet code information (anticodons) in body cells, as described by the statistics of extremes in order theory, serves as such a mechanism.

An examination of past human mortality trends revealed that the mean longevity of birth cohorts from 1740 to 1900 for United States of America (U.S.) Congressional Representatives exhibited oscillations that coincided with the 9- to 12-year sunspot cycle. Cohort mean longevities were 2-3 years greater during times of low sunspot activity than at peak activity. This phenomenon was confirmed in data from members of the House of Commons of the United Kingdom Parliament and from University of Cambridge alumni. An additional longevity oscillation with a longer period was visible in the data and may also be related to sunspot cycles. The amplitude and frequency modulations in the longevity and sunspot oscillations aligned when a 20-year phase shift was incorporated. This shift requires the existence of a lag between solar changes and the affected birth cohorts. Several possible causes of the effect are discussed, in particular: radiation on primordial germ cells in developing embryos; influenza epidemics and pandemics; and weather. The size of the longevity oscillation requires that the solar effect must be considered in studies that examine longevity trends and risk estimation.

The synthesis and /sup 195/Pt chemical shifts of the hydroxo-bridged dimer and trimer, isolated form an aquated solution of bis(nitrato)(trans-1,2-diaminocyclohexane)platinum (I), at different pH are described. The chemical shifts of these three complexes are widely separated, and /sup 195/Pt NMR provides a convenient method for the investigation of dimer formation kinetics from I. Rate constants for the dimerization reaction calculated at different pD, temperature, and concentration agree well with the hypothesis that dimerization occurs with a rate-limiting bimolecular reaction of an intermediate hydroxo species formed by the loss of a proton from the parent complex.

Any thermodynamic study of aging and death in laboratory mammals is dependent on techniques which would allow for the chronic manipulation of the core body temperature of these organisms. Accordingly, the hypothermic response after parenteral administration of chlorpromazine, L-dopa, reserpine, and p -chlorophenylalanine was investigated in the mouse. Mice injected at 24 hour intervals with chlorpromazine exhibited a diminished response (tolerance) with repeated administration. The degree of induced hypothermia and resistance to tolerance was greater in male mice than in female mice. L-dopa was unable to potentiate chlorpromazine hypothermia but did evoke a non-adapting hypothermic response when administered alone. Reserpine also evoked a non-adapting hypothermia. Further, animals treated with small doses of reserpine exhibited an increasing response with repeated injections. p -Chlorophenylalanine induced hypothermy that became intermittent with successive administration. The results presented here indicate that both reserpine and L-dopa would be of value as hypothermic agents in long term investigations.

The mortality kinetics of white humans of the United States were examined for 25 different age-related causes of death (22 male, 21 female). The survivorship distributions for these diseases clustered into groups, as defined by their position on the time axis. When the survivorship curves were linearized, by plotting as log(— log S(t)) vs. log t, this clustering was easily identified as well-defined intersections among the lines separated by 2-year intervals. The four largest groups had intersections at approximate time values of 101, 99, 95, and 93 years, with a small group having an intersection at 97 years. The distribution of diseases among the intersections was not random but was related to sex and disease type. The two-parameter Weibull and GDCP (Gamma Distribution raised to a Combinatoric Power) functions were fit to the individual cause of death survivorship curves and yielded parametric values for the shape (slope) and median time to death. These two parameters varied with disease type and exhibited a positive, linear regression. The regression slope between the shape (α) and time (τm) parameters of the GDCP was also equal to approximately 2 years. This suggested to us that the 2-year intervals in the median times of death with integral changes in the shape parameter and the 2-year time interval between the survivorship clusters may both arise from a similar process involving integral numbers of discrete steps.

Treatment of mixtures of cis and trans Pt(NH3)2Cl2 with thiourea (thu) gave Pt(thu)4Cl2 and trans [Pt(NH3)2(thu)2]Cl2 which were separated and quantified by high-performance liquid chromatography using a strong cation exchange column. Several platinum-thiourea compounds have been studied by NMR (1H, 13C, 195Pt), an increase in the coalescence temperature of the NH2 resonances being observed as a result of co-ordination. The NMR data has also been used to show the presence of interactions between the Pt(thu)2+4 and PtCl2−6 ions of [Pt(thu)4][PtCl6] in dimethylsulphoxide solution.

The kinetics of heat-induced lysis of a population of sheep red blood cells over the temperature range 42 degrees - 56 degrees C are shown to be similar in form to the survivorship curves of multicellular organisms and are describable by the power law function,--(1/N)(dN/dt) = At(n-1). The A parameter of the power law function is examined in this model system in an attempt to show its relationship to molecular events. Arrhenius-type plots of the A parameter are different for old populations of red blood cells compared to young populations. The plot for the old cells shows a high energy transition at 50 degrees C. For the young cells an activation enthalpy of 232 kcal/mole is obtained with no transition occurring at 50 degrees C. The parameter l/tau, defined as A1/n, is more directly related to the molecular basis of the temperature dependence of the lysis kinetics. The Arrhenius plots of 1/tau give activation enthalpies of 42.8 and 40.4 kcal/mole for young and old cells, respectively, and activation entropies of 57.6 and 50.3 cal/mole per degree. These activation enthalpies and activation entropies appear to be in accord with a compensation law for these qualities for protein denaturation, and support the suggestion that protein denaturation is the rate-limiting step in the lysis of sheep red blood cells.

The preparation of Pt(NH3)2(MeU)2·2H2O, 1. [(NH3)2Pt(MeU)2Pt(NH3)2](NO3)2, 2. Pt(NH3)2Cl(MeU), 3. and (in solution) [Pt(NH3)2(OH)(MeU)], 4. (MeU = 1-methyluracil monoanion) is reported. Levels of l-methyluracil and 1–4 in platinum-l-methyluracil blue (PtMeUB) have been assessed by high performance liquid chromatography (HPLC). This technique has also been used to show that in physiological saline or water, PtMeUB hydrolyzes to 3 or 4, respectively. Visible spectroscopy shows that the rate of hydrolysis of PtMeUB is much faster in fetal calf serum than in saline or water, with HPLC indicating that the product of hydrolysis in serum is 3. The precipitate obtained upon treatment of DNA solutions with PtMeUB has also been shown to hydrolyze to 3 or 4 when suspended in saline or water. Compounds 1–3 have been tested against the Ascites S-180J tumors, with 2 and 3 being active, while 4 has been shown to react readily with DNA. Possible mechanisms of antitumor action of PtMeUB that involve 3 and 4 are proposed.

Some platinum coordination complexes are active anticancer drugs in animal and man. This new class of chemotherapeutics was discovered during the course of investigation of the electric field effect on bacterial growth. The platinum electrodes electrolyzed during the experiment, releasing a platinum complex which caused complete cessation of cell division in the bacterial rods. With this filamentation assay system, we were able to identify the specific chemical as cis-dichlorodiammineplatinum(II), a complex known since 1848. The bacterial studies with many such complexes suggested to us the generalizations that charged platinum complexes were bacteriocidal, while the neutral platinum complexes induced filamentation and, in lysogenic bacteria, lysis. The neutral complexes have ignificant activity against transplantable, virally induced, and chemically induced cancers in animals. They are synergistic with almost every other anticancer drug in current use. In man kidney toxicity is the dose limiting ...

Platinum-pyrimidine complexes have been used as the sole electron-dense stains for electron microscopy on 18 types of mammalian cells. These recently discovered complexes react avidly with nucleic acids in vitro, are antitumor agents, and are highly soluble in water. In cells, they are selective for regions rich in nucleic acid and make visible the cellular chromatin, nucleolus, and ribosomes. In addition, cells that are tumorigenic exhibit electron-dense patches at the plasma membrane; normal cells do not. The results of treatment of ascites sarcoma-180 cells with concanavalin A, nucleases, glycosidases, or a protease suggest that the electron-dense patches are DNA, with neuroaminic acid involved in the surface binding.

Publisher Summary Cisplatin (cis-dichlorodiammineplatinum II) is the first member of a new class of potent anticancer drugs, the metal coordination complexes, to enter general use in cancer chemotherapy. The drug has been proved to be effective alone or in combination for the treatment of a wide variety of solid cancers in man. Evidence suggests that it is curative for testicular cancers and ovarian cancers. It is also palliative in the treatment of cancers of the head and neck, bladder, prostate, lung, and cervix and in some pediatric cancers. The major toxic side effects of the drug are damage to the kidney, nausea, and vomiting. The hydration of the patient, with or without concomitant diuretics, ameliorates the kidney toxicity so well that in some clinical reports, it is no longer considered to be the dose-limiting side effect. The nausea and vomiting due to this drug are extreme, ubiquitous, and long lasting. Evidence has been reported indicating that nonstandard drugs such as marijuana, metaclopromide, and droperidol are effective in reducing the nausea and vomiting to an acceptable level.

We propose a new mathematical function for the analyses of age-related human deaths due to single causes. Like the earlier Gompertz and power law functions, it is a two parameter function. Unlike them, one of the parameters is an integer in the range of 5-13. The function is the integral gamma distribution raised to a combinatoric power (GDCP). The function has a deep relation to the power law, and this explains the past successes of the power law. The GDCP function generates highly accurate distributions for ages at death due to specific diseases, and also the means and variances for the distributions. It is possible now to assign unambiguously an integer to each cause of death. In model systems, the alpha integers are the number of "events" necessary to commit the organism to death. Different diseases with the same integer have the same age distribution at death. The major remaining problem is the relative sizes of the populations dying of each single cause. The solution to this must be model derived. The analysis of 24 single causes of death for males and females in the U.S.A., white population over the years 1968-1978 show: (1) the alpha integer is eight for almost all digestive organ carcinomas in both males and females; (2) cancers at other sites have various values for alpha; and (3) for vascular diseases females have alpha integers higher by one or two than males. If the combinatoric power is multiplied by a constant then the second number in the function, tau, the time constant, which is different for each disease, takes on an approximately constant value, which we call the "characteristic age" of the species. Thus, with one number, the "characteristic age" and the nine integers, we can predict, using the GDCP function, the relevant distributions of deaths due to all the different diseases.

Platinum coordination complexes form a new class of active anticancer agents in animals and man. Cis-dichlorodiammineplatinum(II), the most widely investigated drug, is now in experimental clinical use against a wide variety of cancers in man. The dose-limiting toxicity in man is renal tubular damage. Hydration of the patient and the use of osmotic diuretics have minimized this effect and allowed higher doses with largely improved responses. Combination chemotherapy with the drug has also produced significant response rates in a variety of cancers. The mode of action of the drug is not yet clear, but most likely involves a primary lesion on nuclear DNA and the stimulation of a host reaction to the cancer. So far, only square planar and octahedral complexes of platinum, with a variety of inorganic and organic ligands, have shown marked activity in animal studies.