Studies of lipid profile, liver function and kidney function parameters of rat plasma after chronic administration of "Sulavajrini Vatika".

The successful use of Ayurvedic medicines is for many years but there is no guideline for studying the toxicity of these preparations through preclinical or clinical investigations. The present study was conducted to evaluate the effect of conventionally prepared Sulavajrini Vatika (SBB), an Ayurvedic formulation on various biochemical parameters of experimental animals after chronic administration. The animal used was albino rats (Rattus norvegicus: Sprague-Dawley strain) and SBB was administered orally at a single dose of 100 mg kg(-1) b.wt. day(-1), up to 62 days. During the study, forty rats, equally of both sexes, were randomly grouped into four where one male and one female group were used as control and other groups were used as test. Among the lipid components, Triglyceride (TG) was decreased very high significantly in both sexes of animal. The decrease of Total Cholesterol (TC), Very Low Density Lipoprotein (VLDL) and high-density lipoprotein (HDL) were also highly significant. Low Density Lipoprotein (LDL) decreased in all SBB treated group. In the liver function parameters, the total protein and albumin content were increased very high significantly in both sexes of rat. But the bilirubin was decreased insignificantly in male and female rats. Serum Glutamic Pyruvic Transaminase (GPT), Glutamic Oxaloacetic Transaminase (GOT) and Alkaline Phosphatase (ALP) were decreased in all treated animals and it was very high significant. In case of kidney function parameters, creatinine was increased very high significantly but the urea was decreased very high significantly in both sexes of rat. The decrease in uric acid was not significant in none of the sexes of rat. The present study confirms that SBB can be contributory for the complications in diabetics with hyperlipidemia and nephropathy as it lowers most of the lipids components and improves liver function and kidney function parameters.

Geochemistry of soil around a fluoride contaminated area in Nayagarh District, Orissa, India: factor analytical appraisal.

Fluoride contamination in soil was studied in the vicinity of a hot spring in Nayagarh district of Orissa. Both bulk soil from 0 to 30 cm depth and profile soils from 0 to 90 cm depth were analyzed for total fluoride (F(t)) and 0.01 M CaCl(2) extractable fluoride (F(ca)), major elements, pH, EC and Organic Carbon (OC). High concentrations of both F(t) and F(ca) were observed in the area surrounding the hot spring and the village of Singhpur. Principal factor analysis (PFA) on the parameters of the bulk soils suggests that two major chemical processes due to three factors, control the soil geochemistry of the area. Factor-1 contributes 37.11% of the total variance and is strongly loaded with Al, Si, Fe, F(t)and F(ca), and explains the fluoride enrichment of the soil, whereas the second and the third factors contribute 16.6 and 12.2%, respectively and explain the controlling process of carbonate precipitation and soil alkalinity. Multiple regression analysis of the scores of the factors was performed to derive a fluoride contamination index in soil. The magnitude of the factor effect on the contamination index follows the order of Factor-1 > Factor-2 > Factor-3. The spatial distribution of the contamination index is used to classify the area into highly contaminated, moderately contaminated and uncontaminated zones.

Frequency-dependent prey-selection of predacious water bugs on Armigeres subalbatus immatures.

BACKGROUND & OBJECTIVES: The predatory behaviour with reference to the frequency-dependent prey-selection of the water bugs Sphaerodema annulatum Fabricius and S. rusticum Fabricius was studied in the laboratory using the IV instar larvae and pupae of Armigeres subalbatus as prey to ascertain their efficacy as predator of mosquito immatures. METHODS: Field collected adult morphs of the water bugs were allowed to predate on larvae and pupae provided in different ratios and densities as per the model of Greenwood and Elton' for a fixed time period. The data obtained on their predation rate were analysed with respect to the model parameters, lnV--the frequency independent component and b--the frequency dependent component of selection. RESULTS: It was found that the prey-selection was dependent on the relative numbers of prey available, favouring apostatic selection. The b values and lnV values for S. annulatum were 0.54 +/- 0.01 and 0.92 +/- 1.04 respectively whereas the corresponding values for S. rusticum were 0.71 +/- 0.03 and 0.17 +/- 1.57 respectively. INTERPRETATION & CONCLUSION: The selection of preys by the water bugs was dependent on the relative number of the prey forms and thus they are expected to predate on the form more abundant in a heterogeneous prey population and adversely affect the adult emergence.

Optimizing the HIV/AIDS informed consent process in India.

BACKGROUND: While the basic ethical issues regarding consent may be universal to all countries, the consent procedures required by international review boards which include detailed scientific and legal information, may not be optimal when administered within certain populations. The time and the technicalities of the process itself intimidate individuals in societies where literacy and awareness about medical and legal rights is low. METHODS: In this study, we examined pregnant women's understanding of group education and counseling (GEC) about HIV/AIDS provided within an antenatal clinic in Maharashtra, India. We then enhanced the GEC process with the use of culturally appropriate visual aids and assessed the subsequent changes in women's understanding of informed consent issues. RESULTS: We found the use of visual aids during group counseling sessions increased women's overall understanding of key issues regarding informed consent from 38% to 72%. Moreover, if these same visuals were reinforced during individual counseling, improvements in women's overall comprehension rose to 96%. CONCLUSIONS: This study demonstrates that complex constructs such as informed consent can be conveyed in populations with little education and within busy government hospital settings, and that the standard model may not be sufficient to ensure true informed consent.

Predatory efficiency of the water bug Sphaerodema annulatum on mosquito larvae (Culex quinquefasciatus) and its effect on the adult emergence.

The daily number of IV instar larva of Culex quinquefasciatus killed, rate of pupation and adult emergence was noted in presence of the predatory water bug Sphaerodema annulatum for a period of seven consecutive days, experimentally, in the laboratory. The rate of IV instar larva killed by the water bugs on an average was 65.17 per day. The rate of pupation ranged between 7.6 and 48 in control while in presence of water bugs it ranged between 6 and 35. The rate of adult emergence in control experiments varied between 1.4 and 4.8 per day, which was reduced to only 0.4-28.8 per day in case of the water bugs. The results clearly indicate that the water bugs on its way of predation reduces the rate of pupation and adult emergence of Cx. quinquefasciatus significantly which calls for an extensive field trials.

A series of three (E)-2-alkylidene-1,4-di-p-tosyl-1,2,3,4-tetrahydroquinoxaline compounds.

The three title quinoxaline derivatives, (E)-2-(4-methylbenzylidene)-1,4-di-p-tosyl-1,2,3,4-tetrahydroquinoxaline, C30H28N2O4S2, (II), (E)-2-(4-methoxybenzylidene)-1,4-di-p-tosyl-1,2,3,4-tetrahydroquinoxaline, C30H28N2O5S2, (III), and (E)-2-(3-chlorobenzylidene)-1,4-di-p-tosyl-1,2,3,4-tetrahydroquinoxaline, C29H25ClN2O4S2, (IV), were synthesized by palladium-catalyzed hetero-annulation. The E configuration of the exocyclic double bond in the three compounds has been established by the present X-ray study. The saturated part of the quinoxaline moiety in all three compounds assumes a distorted chair conformation. The numerical descriptors indicate a high degree of isostructurality between compounds (II) and (III), but no isostructurality with compound (IV).

Increased cyclooxygenase-2 (cox-2) expression and activity in a murine model of metastatic breast cancer.

Elevated prostaglandin E(2) (PGE(2)) production is a common feature of human malignancies. This activity has often been attributed to increased metabolic activity of the cyclooxygenase enzymes, although a direct comparison of these 2 parameters i.e., prostaglandin production and cox protein expression, is rarely performed in the same malignant tissue. Using a murine model of metastatic breast cancer, we show that PGE(2) levels are positively correlated with increased tumorigenic and metastatic potential. Because prostaglandin synthesis is a product of 2 isoforms of the cyclooxygenase enzyme, we examined the expression and activity of both isoforms. All tumor cell lines examined, regardless of phenotype, express both cox-1 and cox-2 proteins in vitro. In contrast to the uniform cox-2 expression in vitro, only tumors resulting from the transplantation of metastatic cell lines express cox-2 in vivo. Cox-1 is detected in both metastatic and nonmetastatic tumors. Thus, this is the first evidence that, in the tumor milieu, cox-2 expression can be regulated differently in metastatic vs. nonmetastatic lesions. Examination of PGE(2) synthesis in vitro reveals that nearly complete inhibition of prostaglandin synthesis occurs in the presence of either indomethacin, which inhibits both isoforms, or NS398, which is selective for the cox-2 isoform. Thus, even though cell lines express both isoforms, the majority of the prostaglandin synthesis stems from the activity of the inducible, cox-2 isoform. Likewise, cell growth is inhibited by both indomethacin and NS398 in a dose-dependent manner, albeit at higher drug concentrations than required to ablate PGE(2) synthesis. Despite the inhibition of prostaglandin synthesis, the cox-2 enzyme levels (protein and mRNA) were increased by either indomethacin or NS398.

Palladium-copper catalyzed synthesis of benzofused heterocycles with two heteroatoms: novel and highly regio- and stereoselective syntheses of (E)-2-(2-arylvinyl)-3-tosyl-2,3-dihydro-1,3-benzothiazoles and (E)-2-alkyl(aryl)idene-3,4-dihydro-2H-1,4-benzothiazines.

A highly novel, general, and convenient palladium and copper-catalyzed procedure has been developed for the synthesis of (E)-2-(2-arylvinyl)-3-tosyl-2,3-dihydro-1,3-benzothiazoles 28-40. 3-(2-Aminophenylthio)prop-1-yne 1 reacts with aryl iodides 2-14 under palladium-copper catalysis to yield the disubstituted alkynes 15-27 which after tosylation undergo a novel cyclization with CuI in the presence of triethylamine in THF to (E)-2-(2-arylvinyl)-3-tosyl-2,3-dihydro-1,3-benzothiazoles 28-40 rather than to the expected 3-alkylidene-4-tosyl-3,4-dihydro-2H-1,4-benzothiazines 41. The reaction is highly regio- and stereoselective. The synthesis of 2-(2-arylethyl)-3-tosylbenzothiazolines 42-47, 2-(2-arylvinyl)benzothiazoles 48-54, and a novel 5-substituted uracil derivative 55 of potential biological importance is also being reported. Similarly, the palladium-copper-catalyzed arylation of S-[2-(N-prop-2'-ynyl)aminophenyl]-N,N-dimethylthiocarbamate 58 with aryl iodides yields the disubstituted alkynes 59 which on cyclization with KOH in methanol leads to (E)-2-(2-aryl)methylidene-3,4-dihydro-2H-1,4-benzothiazines 61. The reaction of the diiodo compounds 12-14a, however, with 58 under palladium-copper-catalyzed reactions involves the participation of only one of the iodo groups in the heteroannulation process giving compounds 61i and 61j. These are amenable to further palladium-catalyzed reactions and afford polyunsaturated heteroaromatic compounds 62 and 63.

(Z)-7-Chloro-3-[(3-chlorophenyl)-methylidene]-4-p-tosyl-3,4-dihydro-2H-1,4-benzoxazine.

In the title compound, C22H17Cl2NO3S, the molecule is a substituted 3,4-dihydro-2H-1,4-benzoxazine compound which has three phenyl rings which are essentially planar. The 3,4-dihydro-2H-oxazine part of the molecule is fused to the benzo ring and has a half-boat conformation; the dihedral angle between the planar part of the oxazine ring and the benzo ring is 10.2 (2) degrees. The (3-chlorophenyl)methylidene substituent has a Z configuration in relation to the ring N atom of the oxazine moiety. Interestingly, the p-toluenesulfonyl (p-tosyl) substituent on the ring N atom protrudes away from the 3-chlorophenyl substituent thus avoiding any steric interaction.

Palladium-catalyzed heteroannulation leading to heterocyclic structures with two heteroatoms: a highly regio- and stereoselective synthesis of (Z)-4-alkyl-2-alkyl(aryl)idene-3,4-dihydro-2H-1,4-benzoxazines and (Z)-3-alkyl(aryl)idene-4-tosyl-3,4-dihydro-2H-1,4-benzoxazines.

A highly convenient method has been developed for the synthesis of (Z)-4-alkyl-2-alkyl(aryl)idene-3,4-dihydro-2H-1,4-benzoxazines 9 and (Z)-3-alkyl(aryl)idene-4-tosyl-3,4-dihydro-2H-1,4-benzoxazines 34-38 through palladium-copper-catalyzed reactions. Aryl halides 7 reacted with 2-[N-alkyl(benzyl)-N-prop-2'-ynyl]aminophenyl tosylate 6 in the presence of (PPh3)2PdCl2 (3 mol %), CuI(5 mol %) in triethylamine at room temperature to yield 2-[N-alkyl(benzyl)-N-(3-aryl-prop-2'-ynyl)]-aminophenyl tosylates 8 in extremely good yields (72-96%). The latter could then be cyclized with KOH in ethanol-water to Z-9 in a highly regio- and stereoselective manner. Similarly, palladium-copper-catalyzed reaction of 2-(prop-2'-ynyloxy)aniline (21) with aryl iodides 7 led to 22-26 which after tosylation and cyclization with cuprous iodide in CH3CN in the presence of K2CO3 and Bu4-NBr led to the (Z)-3-alkyl(aryl)idene-4-tosyl 3,4-dihydro-2H-1,4-benzoxazines 34-38 in good overall yields. The Z-stereochemistry of the products was established from 1H NMR spectra, 3JCH values (between vinylic proton and methylenic carbon of the heterocyclic ring), NOE experiments, and X-ray analysis. The method was also found to be suitable for the synthesis of bis(benzoxazinylated) derivatives 17, 39, and 2-alkyl-3,4-dihydro-2H-1,4-benzoxazines 18. Our method for the synthesis of 3,4-dihydro-2H-1,4-benzoxazines is highly efficacious, using easily available starting materials under very mild conditions. Also the synthesis of some novel 5-substituted uracil derivatives 40 and 41 containing the benzoxazinyl moiety and of potential biological interest is being reported.

Expression of the chemokines IP-10 and Mig in IL-10 transduced tumors.

Several laboratories have reported marked tumor inhibition when the cytokine interleukin-10 (IL-10) is overexpressed as a transgene in a variety of tumor cells. To identify critical effector molecules, we compared the expression of the chemokine crg-2, the murine homolog of human inducible protein 10 (human IP-10) in murine mammary tumors derived from the transplantation of six IL-10 expressing clones of tumor cell line 66.1, parental 66.1, or 66-neo-cells. We observed increased levels of IP-10 mRNA in all IL-10-expressing tumors examined in comparison to 66-neo. IP-10 mRNA was not detected in parental 66.1 tumors. The closely related chemokine Mig (monokine induced by interferon-gamma [IFN-gamma]) was also induced in all IL-10-expressing tumors. Studies of cultured tumor cells in vitro show that mammary epithelial tumor cells, in the absence of host elements, can express IP-10 and Mig in response to induction with either lipopolysaccharide (LPS) or IFN-gamma alone. The combination of LPS plus IFN-gamma resulted in even greater induction of IP-10 RNA. The kinetics of induction differ somewhat for the two chemokines, with IP-10 showing slower induction and less rapid decline. Because both Mig and IP-10 are chemotactic for tumor-infiltrating lymphocytes, we examined the presence of CD4+ and CD8+ lymphocytes in these tumors. Consistent with the upregulation of Mig and IP-10, we saw significantly increased numbers of CD8+ cells and a lesser increase in CD4+ cells in tumors with elevated levels of both chemokines.

(E)-2-[2-(1-Naphthyl)vinyl]-3-tosyl-2,3-dihydro-1,3-benzothiazole.

The title compound, C(26)H(21)NO(2)S(2), which consists of a benzothiazole skeleton with alpha-naphthylvinyl and tosyl groups at positions 2 and 3, respectively, was prepared by palladium-copper-catalyzed heteroannulation. The E configuration of the molecule about the vinyl C=C bond is established by the benzothiazole-naphthyl C-C-C-C torsion angle of 177.5 (4) degrees. The five-membered heterocyclic ring adopts an envelope conformation with the Csp(3) atom 0.380 (6) A from the C(2)NS plane. The two S-C [1.751 (4) and 1.838 (4) A] and two N-C [1.426 (5) and 1.482 (5) A] bond lengths in the thiazole ring differ significantly.

Essential role of nitric oxide and interferon-gamma for tumor immunotherapy with interleukin-10.

Several laboratories have reported anti-tumor activity for high levels of interleukin-10 (IL-10) expressed as a transgene or administered as recombinant protein. The authors have reported a positive correlation for nitric oxide production and anti-tumor activity of IL-10 in a murine model of breast cancer. In the current study, they sought evidence of a mechanistic role for nitric oxide in IL-10-mediated tumor growth inhibition. They wanted to determine whether pharmacologic inhibition of nitric oxide synthase (NOS) activity reverses the therapeutic effect of IL-10. Administration of either of two NOS inhibitors, aminoguanidine (AG) or L-lysine,N6-1-iminoethyl-dihydrochloride, appears to abrogate in part the tumor growth inhibition observed when IL-10 is overexpressed as a transgene in two murine mammary tumor cell lines. Nitric oxide levels were assessed at the tumor site by measuring nitrosylated heme levels by electron spin resonance spectroscopy. Nitric oxide hemoglobin levels were lower in tumors from aminoguanidine-treated mice, indicating that effective inhibition of nitric oxide production occurred at the tumor site. Previous investigations showed that the inducible form of NOS protein (iNOS), but not constitutive NOS, was expressed at higher levels in IL-10-expressing tumors. Because iNOS is regulated at the transcriptional level, the authors compared iNOS mRNA levels in IL-10 and control tumors. Northern analysis revealed strong iNOS message expression in all six IL-10-expressing tumors examined, whereas message was faintly detected in parental or 66-neo tumors. The inducible form of NOS is responsive to induction by interferon-gamma (IFN-gamma). The role of IFN-gamma in IL-10-mediated tumor inhibition and iNOS mRNA induction was determined. When tumors were transplanted to IFN-gamma mutant mice, the tumor-inhibitory activity of IL-10 was lost. Furthermore, iNOS mRNA was no longer induced in the absence of host expression of IFN-gamma. These data indicate that nitric oxide contributes to the anti-tumor activity of IL-10 and that expression of iNOS in this context depends on IFN-gamma.

Interleukin-10 gene transfer activates interferon-gamma and the interferon-gamma-inducible genes Gbp-1/Mag-1 and Mig-1 in mammary tumors.

Expression of IL-10 as a transgene inhibits murine mammary tumor growth and metastasis. Using differential display methodology, we sought genes whose expression was modulated by IL-10. We compared mRNA isolated from parental murine mammary 66.1 tumors, as well as tumors derived from neo(r)-transfected cells and 6 different IL-10-expressing cell lines. We identified 2 cDNA products that were up-regulated in all 6 IL-10-expressing tumors in comparison to parental and 66-neo tumors. One cDNA corresponds to the murine guanylate-binding protein gene Gbp-1/Mag-1. The other cDNA corresponds to the chemokine Mig-1 (monokine induced by IFN-gamma). Both genes were originally identified in IFN-gamma-activated macrophages or macrophage cell lines. We now report that cultured mammary epithelial tumor cell lines also express both genes in response to treatment with IFN-gamma and LPS. Furthermore, IFN-gamma mRNA is elevated in IL-10-expressing tumors in comparison with parental or neo-transfected tumors. Thus, high-level expression of IL-10 as a transgene results in activation rather than suppression of IFN-gamma as well as 2 IFN-gamma-inducible genes. Up-regulation of host IFN-gamma is critical to anti-tumor activity since IL-10 no longer inhibits tumor growth in hosts with a deletion in the IFN-gamma gene. Additionally, Gbp-1/Mag-1 and Mig-1 gene induction no longer occur in IFN-gamma mutant mice.

Interleukin-10 gene transfer inhibits murine mammary tumors and elevates nitric oxide.

Transfection of cDNA for IL-10 into line 66.1 murine mammary tumor cells results in marked suppression of tumor growth and metastasis. Others have reported that nitric oxide has potent antitumor activity and IL-10 is known to regulate the inducible isoform of nitric oxide synthase (iNOS) expressed in macrophages. We identified nitric oxide production in mammary tumors as indicated by electron paramagnetic resonance detection of nitric oxide-hemoglobin (NO-Hb). IL-10 expression resulted in elevated levels of NO-Hb in mammary tumors. Immunohistochemical examination of mammary tumors for iNOS protein revealed few positively staining cells in parental or control neo-transfected tumors but strong iNOS staining in all IL-10 transfected tumors, consistent with the NO-Hb data. To determine if mammary epithelial tumor cells themselves, express nitric oxide synthase activity, cultured tumor cells were treated with pro-inflammatory cytokines and nitrite accumulation was assessed in the conditioned medium. All IL-10 producing cell lines accumulated uM concentrations of nitrite in response to short term (24 hr) cytokine stimulation. Cells not expressing IL-10 (parental and neo-transfectants) accumulated no nitrite under similar culture conditions. After longer stimulation (48 hr), parental and 66-neo cells accumulated lower amounts of nitrite. IL-10 gene transfer is associated with increased iNOS protein expression and enzymatic activity detected both in vitro and in vivo. Our findings suggest that the antimetastatic and antitumor activity of IL-10 is related to enhanced production of nitric oxide.

IL-10 gene transfer to intracranial 9L glioma: tumor inhibition and cooperation with IL-2.

This study examines the effects of interleukin-10 (IL-10) and combination IL-10 + IL-2 gene transfer on experimental brain tumor growth in vivo. 9L gliosarcoma cells were engineered to stably express murine IL-10 (9L-IL-10 cells) and implanted subcutaneously or to the caudate/putamen of syngeneic rats. The growth of tumors expressing IL-10 was substantially reduced compared to that of control tumors (p < 0.05). Intracranial tumors expressing IL-10 and IL-2 were established by co-implanting 9L-IL-10 cells with endothelial cells engineered to express IL-2. At 14 days post-implantation, tumors expressing IL-10 + IL-2 were 99% smaller than control-transfected tumors (p < 0.0001). This extent of anti-tumor effect could not be achieved by expression of IL-10 or IL-2 alone within tumors. Neither IL-10 nor a combination of IL-10 + IL-2 gene delivery inhibited tumor growth in severe combined immunodeficient (SCID-Beige) mice (p > 0.05). Immunohistochemical analysis revealed that IL-10 + IL-2 gene delivery markedly increased T-cell infiltration within the striatum ipsilateral to tumor cell implantation. These findings establish that IL-10 expression, particularly in combination with IL-2 expression, can have significant immune-dependent anti-tumor actions within intracranial gliomas.

Interleukin-10 inhibits tumor metastasis, downregulates MHC class I, and enhances NK lysis.

We have engineered highly aggressive murine mammary tumor cell line 410.4 to express interleukin-10 (IL-10) and compared the behavior in vivo of these cells to parental 410.4 and 410.4 transfected with the control plasmid (410.4-neo). Transplantation of parental 410.4 and 410.4-neo tumor cells to syngeneic mice resulted in progressive growth and death from pulmonary metastases. In contrast, both subcutaneous growth and metastatic disease were completely inhibited by IL-10 expression. We had shown previously that the antimetastatic activity of IL-10 is expressed in T-cell-deficient mice but is lost when NK activity is suppressed. This study confirms that IL-10 is dependent on NK activity, since no therapeutic effect is seen in C.B-17/IcrCrl-SCID/Beige mice which lack T, B, and NK cell function. We compared the sensitivity to NK lysis of four IL-10-expressing clones with 410.4 and 410.4-neo and found that IL-10 expression resulted in enhanced NK lysis of all four clones. Furthermore, IL-10 expression was correlated with decreased surface expression of MHC class I Kd, Ld, and Dd. Pretreatment of IL-10-expressing cell lines with IFN-gamma reversed the class I downregulation and reduced the sensitivity of these cells to NK lysis. Taken together, these studies in vitro and in vivo are consistent with a mechanism by which IL-10 expression downregulates class I expression, leading to enhanced NK lysis of tumor cells, resulting in control of metastatic disease.

Expression of the RET/PTC fusion gene as a marker for papillary carcinoma in Hashimoto's thyroiditis.

Hashimoto's thyroiditis is an inflammatory disease of the thyroid gland with autoimmune etiology. Patients afflicted with Hashimoto's have a higher risk of thyroid malignancies such as papillary thyroid carcinoma. In the present study, we investigated the frequency of papillary thyroid carcinoma specific genes in patients diagnosed with Hashimoto's disease. The newly identified oncogenes RET/PTC1 and RET/PTC3 provide useful and specific markers of the early stages of papillary carcinoma as they are highly specific for malignant cells. Using a sensitive and specific reverse transcriptase-polymerase chain reaction (RT-PCR) assay, we found messenger RNA (mRNA) expression for the RET/PTC1 and RET/PTC3 oncogenes in 95% of the Hashimoto's patients studied. All Hashimoto's patients presenting without histopathologic evidence of papillary thyroid cancer showed molecular genetic evidence of cancer. These data suggest that multiple, independent occult tumors exist in these patients at high frequency.