Human Primary Liver Cancer Research Articles

Overexpression of glutamine synthetase in human primary liver cancer

Background/Aims: We have identified several clones specifically expressed during malignant cell proliferation by screening a complementary DNA library constructed from a human primary liver cancer with subtractive probes. One clone was identified as the glutamine synthetase (GS) transcript. Its expression is tightly regulated during development, especially in the hepatic lobule. Because this enzyme is involved in nitrogen homeostasis, it might contribute to tumor development/progression in primary liver cancer. Methods: We compared the expression of GS messenger RNA (mRNA) and protein in tumorous and nontumorous liver from 34 patients with primary liver cancers, using a combination of Northern blot, dot blot, western blot, and determination of GS enzyme activity. Results: GS mRNA was higher in tumors versus nontumors in 23 of 34 primary liver cancer samples. GS activity was higher in 6 of 8 selected primary liver cancer samples with high RNA levels. GS protein levels were proportional to enzyme activity. A major GS transcript of 2.8 kilobase was detected by Northern blotting and sequencing. This comprised the minor 1.8-kb transcript and a long 3′ untranslated region; the latter contained an AT-rich zone, fully conserved in the chicken, mouse, and rat, which might be important for stability. Conclusions: Our results show an overexpression of GS in human primary liver cancers and, thus, point to its potential involvement in hepatocyte transformation.

MDR1 (multidrug resistance) gene expression in human primary liver cancer and cirrhosis

MDR1 RNA levels were analysed in patients with hepatocellular carcinoma (HCC), benign liver tumors or cirrhosis. None of the patients with HCC had received chemotherapy. MDR1 RNA levels were increased relative to a normal liver specimen in 15/26 liver cancers, 2/6 benign liver tumors and 0/8 tumor-free cirrhotic livers. MDR1 was also overexpressed in cirrhotic non-tumorous liver tissues of some patients with HCC. The results indicate that the MDR1 gene is overexpressed in liver tumors and some preneoplastic lesions and that might account for the very poor response of primary liver cancers to chemotherapy.

Expression of insulin-like growth factor II (IGF-II) and IGF-II, IGF-I and insulin receptors mRNAs in isolated non-parenchymal rat liver cells.

The insulin-like growth factor II (IGF-II) is involved in embryonic growth. Modifications of its expression might play a role in the development of primary liver cancer in humans and woodchucks. In the liver, little information is available on the cell types involved in its synthesis. We have investigated the expression of IGF-II as well as IGF-II, IGF-I and insulin receptor mRNAs in non parenchymal liver cell preparations in rats of various ages. The results indicate that Kupffer cells, endothelial cells and fat-storing cells express both IGF-II and the three different receptor mRNAs. Furthermore, a switch from a fetal to an adult IGF-II mRNA profile was obtained in the different cell preparations. Therefore, our results indicate that regulation of IGF-II gene expression can be analyzed through these isolated liver cell preparations. These results might also be important in investigating the potential role of IGF-II in liver carcinogenesis.

Aflatoxins in animal and human health.

Aflatoxins remain as a threat to the health of livestock as well as humans by their continuing intermittent occurrence in both feeds and foods. The finding that aflatoxin-contaminated feeds, and eventually purified aflatoxins, were carcinogenic in rats and trout initiated a multitude of studies in search of the role of these toxins in human liver disease, especially cancer. Although aflatoxins have caused acute liver disease in humans, epidemiologic evidence of the involvement of aflatoxins in PLC has not been clarified. Earlier studies did not consider that the hepatitis B virus (HBV) may have contributed to the PLC in the selected populations. Although later studies that did include measurement of the HBV antigen in serum provided conflicting evidence for the role of aflatoxin in PLC in these populations, the latest and most comprehensive study found no association between aflatoxin exposure and PLC mortality. The technological advances and findings of the chemical, immunologic, and metabolic activities of aflatoxins such as binding to DNA and protein to form adducts, development of monoclonal antibodies, and mutational specificity of the genotoxic compounds will, it is hoped, help to clarify the role of aflatoxin as a risk factor, among many others, in the development of primary liver cancer in humans. Aflatoxicosis of animals is usually manifested by pathologic changes in the liver, but they have been found to be carcinogenic and teratogenic as well as causing impaired protein formation, coagulation, weight gains, and immunity. The importance of the carcinogenic effect in livestock is diminished because they are not fed contaminated diets for a sufficient time prior to marketing for slaughter. Animals are variably susceptible to aflatoxins, depending on such factors as age, species, breed, sex, nutrition, and certain stresses. Swine, cattle, and poultry are the domestic species of greatest economic concern in terms of aflatoxicosis. In all species, the evidence of disease is a general unthriftiness and reduction in weight gains, feed efficiency, immunity, and production. More conclusive evidence of aflatoxin involvement in disease includes acute to chronic liver disease with concomitant increases in specific liver enzymes in the serum. In cattle, milk production is affected, but of greater significance is that the aflatoxins in feeds can be rather efficiently converted to toxic metabolites in milk, with even small amounts being readily detectable. The poultry industry probably suffers greater economic loss than any of the livestock industries because of the greater susceptibility of their species to aflatoxins than other species.(ABSTRACT TRUNCATED AT 400 WORDS)

Insulin-like growth factor II (IGF-II) mRNA expression during hepatocarcinogenesis in transgenic mice

Insulin-like growth factor II (IGF-II) mRNA expression is developmentally regulated in liver tissue. We previously observed the reexpression of fetal IGF-II mRNAs in human primary liver cancer and in surrounding cirrhotic tissue. In order to determine the steps of liver cancer progression where the activation of IGF-II fetal mRNAs occurs, we analyzed IGF-II mRNA expression during hepatocarinogenesis in transgenic mice carrying an antithrombin III-SV40 early region hybrid gene. The comparative analysis of mRNAs encoding IGF-II and other differentiation-associated proteins, as well as histological analysis, indicate that the reexpression of fetal IGF-II mRNAs takes place in specific steps of liver cancer progression, both in early pretumorous lesions and in well-differentiated hepatocellular carcinomas.

A preliminary report on the intervention trials of primary liver cancer in high-risk populations with nutritional supplementation of selenium in China

The purpose of this study was to evaluate the effect of selenium (Se) in the prevention of human primary liver cancer. Three intervention trials were conducted among the residents at high risk to primary liver cancer (PLC) in Qidong county, Jiang-su province, the People's Republic of China. This area has the second highest rate of PLC in China. One trial was undertaken among the general population in a township with supplement of table salt fortified with 15 ppm anhydrous sodium selenite (Se-salt) for 5 y and the other four townships with similar PLC incidence rate served as the controls using normal table salt. The second trial was undertaken among hepatitis B virus surface antigen carriers (HBVsAg+) receiving supplement of 200 micrograms Se in form of selenized yeast (Se-yeast) daily vs placebo for 4 y. The third trial was carried out in members of families with high PLC incidence using Se-yeast (200 micrograms of Se daily) vs placebo for 2 y. The results showed that nutritional supplement of Se could reduce the PLC incidence significantly.

Expression of insulin-like growth factor II, α-fetoprotein and hepatitis B virus transcripts in human primary liver cancer

Expression of insulin-like growth factor II, α-fetoprotein and hepatitis B virus transcripts in human primary liver cancer

Expression of insulin-like growth factor II, α-fetoprotein and hepatitis B virus transcripts in human primary liver cancer

Insulin-like growth factor II is a fetal growth factor structurally and functionally related to insulin and insulin-like growth factor I. Its mRNA expression is developmentally regulated in human liver, the reexpression of insulin-like growth factor II fetal transcripts being often observed in primary liver cancer. Insulin-like growth factor II and alpha-fetoprotein mRNAs were studied in 16 human primary liver cancers, most of which were highly differentiated. Hepatitis B virus transcripts were also analyzed in the tumors from hepatitis B virus chronic carriers. alpha-Fetoprotein mRNA was detected in only four tumors and in one nontumorous cirrhotic tissue; all these samples also displayed insulin-like growth factor II fetal transcripts. Furthermore, fetal insulin-like growth factor II mRNAs were observed in five tumors and six nontumorous cirrhotic areas not expressing alpha-fetoprotein mRNA. The presence of hepatitis B virus RNA was only observed in tissues not expressing alpha-fetoprotein or fetal insulin-like growth factor II mRNA. In conclusion, fetal insulin-like growth factor II transcripts are more frequently observed than alpha-fetoprotein mRNA in highly differentiated liver cancers and in surrounding cirrhotic areas. The reexpression of fetal insulin-like growth factor II transcripts might then be a marker of early steps of liver cell transformation.

Expression of insulin-like growth factor II (IGF-II) in human primary liver cancer: mRNA and protein analysis

Insulin-like growth factor II (IGF-II) is a polypeptide growth factor thought to be involved in fetal tissue development. We previously showed an increased expression of IGF-II mRNA in human primary liver cancer. The present investigation was undertaken to characterize the overexpressed IGF-II transcripts and to determine whether they are translated into protein. Two cDNAs with distinct 5' untranslated regions, corresponding to IGF-II transcripts expressed in fetal liver, were isolated from a primary liver cancer. Complete nucleotide sequence analysis showed an identical open reading frame of 540 bp, encoding a predicted polypeptide identical to the IGF-II isolated from serum. An increased synthesis of IGF-II protein was demonstrated by a protein-binding assay in tumorous liver samples, the highest levels being found in primary liver cancers with the highest IGF-II steady state level. By contrast, serum IGF-II content was low in most of primary liver cancer cases analyzed. Altogether, the results indicate reexpression of IGF-II both at the mRNA and protein levels in primary liver cancer. This finding is consistent with IGF-II being a marker of liver cell differentiation. In addition, this growth factor might be involved in liver cancer progression by an autocrine and/or paracrine mechanism.

Hepatitis B virus integration in a cyclin A gene in a hepatocellular carcinoma

Hepatitis B virus (HBV) DNA frequently integrates into the genome of human primary liver cancer cells, but the significance of this integration in liver carcinogenesis is still unclear. Here we report the cloning of a single HBV integration site in a human hepatocellular carcinoma at an early stage of development, and of its germline counterpart. The normal locus was found to be transcribed into two polyadenylated messenger RNA species of 1.8 and 2.7 kilobases. We have isolated a complementary DNA clone from a normal adult human liver cDNA library which has an open reading frame with a coding capacity for a protein of 432 amino acids and relative molecular mass 48,536. The strong homology of the C-terminal half of the protein to the A-type cyclins of clam and Drosophila identifies it as a human cyclin A. The cyclin A gene has several exons, and the HBV integration occurs within an intron. As cyclins are important in the control of cell division, the disruption of a cyclin A gene by viral insertion might contribute to tumorigenesis.

Insulin-like growth factor II (IGF-II) mRNA as a marker of differentiation in human primary liver cancer

Insulin-like growth factor II (IGF-II) mRNA as a marker of differentiation in human primary liver cancer

肝動脈塞栓術施行或は非施行ヒト肝癌組織の初代培養と株化

肝動脈塞栓術施行或は非施行ヒト肝癌組織の初代培養と株化

New abnormal isoenzyme of 5'-nucleotide phosphodiesterase in the serum of human hepatoma.

5'-nucleotide phosphodiesterase (5'-NP) isoenzymes were separated from the serum of human primary liver cancer (PLC) patients by 4-20% tubular polyacrylamide gel gradient electrophoresis. Electrophoresis showed 11 isoenzymes in which bands I, V and VI (from anode to cathode) were all absent in 125 normal control sera; bands V and VI were detectable in 31(73.8%) of 42 patients with PLC, but absent in 70 cases of other diseases. Bands V and VI may represent specific isoenzyme bands of PLC, comprising a new addition to the markers used in the diagnosis of this disease. By the same method, it was also found that the abnormal serum isoenzyme bands of PLC correspond to those found in the cord blood sera of the newborn and fetus. These abnormal isoenzymes diminish gradually after birth and all disappear when children reach the age of 10.

Inhibition of the growth of human hepatocellular carcinoma in vitro and in athymic mice by a quinazoline inhibitor of thymidylate synthase, CB3717.

Two human primary liver cancer cell-lines, PLC/PRF/5 and Hep 3B, grown both in vitro and as xenografts in nude mice were used to evaluate the chemotherapeutic potential of a new quinazoline antifolate CB3717. Xenograft growth rate was followed both by serial serum alphafoetoprotein (AFP) measurement and direct volume estimation. A dose regime of 200 mg CB3717 kg-1 body wt day-1 for 5 days caused a significant reduction in growth rate, as measured by relative serum AFP, of both xenografts; PLC/PRF/5 derived xenograft growth was also inhibited by 125 mg CB 3717 kg-1 day-1 for 5 days. Cell culture experiments showed that the ID50 for the cell lines fell within the range of serum CB3717 concentration achieved by a dose of 300 mg m-2 given to patients. Treatment with CB3717 stimulated the incorporation of exogenous thymidine into DNA by the tumour cells, presumably because of inhibition of the de novo pathway and reduction of endogenous thymidine triphosphate pools. These results suggest that CB3717 may be a useful new therapeutic agent in human primary liver cell carcinoma and that blocking the salvage pathway may further increase efficacy.

Oncogenes in human primary hepatic cancer

Transfection assay of NIH 3T3 cells was performed with DNAs isolated from ten human PHC (primary hepatic cancer) specimens and a hepatoma 7402 cell line. Positive results were obtained in 7402 and in six out of ten PHC DNAs. Human N-ras gene was identified in transfectants from 7402 DNA and transformed cells from three PHC DNA samples, which had passed more than two cycles of transfection. The expression of N-ras was also remarkably enhanced in six out of nine poly(A)+RNA samples isolated from PHC tissues. P21 synthesis was elevated in 7402 cells as well as in transfectants derived from 7402 cells and PHC DNA. In analysis of PHC DNA, rearrangement and amplification of N-ras gene was observed in two PHC samples. The discrepancy of results of the transfection assay and mRNA expression was discussed. Furthermore, c-myc was also highly expressed in most PHC tissues. It implied that the cooperating activity of N-ras and c-myc might be responsible for the malignant phenotypic alteration in some or most cases in human primary liver cancer.

Distribution of fibronectin and laminin in human liver tumors

Two extracellular matrix and basement-membrane components, fibronectin (Fn) and laminin, were studied by the indirect immunoperoxidase technique in ten primary human liver cancers. Similar distributions of both Fn and laminin were detected in the well differentiated hepatocellular carcinomas with trabecular and tubular pattern. Two moderately differentiated hepatomas contained Fn only. Neither Fn nor laminin were present, however, in the parenchyma of one poorly differentiated hepatoma. In three cases of cholangiocarcinoma, laminin surrounded the tumorous ducts, while Fn appeared mainly in the reactive connective tissue stroma. The present findings indicate that bile-duct cancers synthesize laminin, and not Fn while differentiated hepatocellular carcinomas produce both Fn and laminin in vivo. The presence of Fn even in moderately differentiated types of liver cancer is in contrast to the findings for carcinomas developing from other organs and it may serve as a marker for primary hepatocellular carcinomas in the differential diagnosis.

Hepatitis B surface antigen and alpha-fetoprotein secreting human primary liver cell cancer in athymic mice

A human primary liver cancer cell line which retains the property of synthesizing hepatitis B surface antigen has been successfully transplanted into nude (athymic) mice. The morphology of the heterotransplanted tumor is similar to that of a well-differentiated human primary liver cell cancer. It produces hepatitis B surface antigen, but there is no evidence of hepatitis B virion production: Hepatitis B core antigen is not detected in the PLC tissue, and serum is negative for hepatitis B e antigen. The nude mouse exhibits a resistance to the transplantation of the human primary liver cancer cells which can be modified by sublethal total body irradiation, suggesting involvement of an immunologic rejection mechanism. The heterotransplanted primary liver cell cancer also produces alpha-fetoprotein, as did the original tumor in vivo, although this marker was not detected during in vitro cell culture. The serum level of alpha-fetoprotein rises exponentially, enabling quantitative evaluation of tumor growth. The human primary liver cell cancer in nude mice provides an in vivo model for determination of tumor response to chemotherapeutic agents.

Formes moléculaires fœtales d'enzymes dans l'hépatome

Summary The resurgence of fetal type aldolases in hepatoma offers a striking example of the fetal pattern of enzymes in some cancerous tissues. In slow growing hepatomas (transplantable or induced by 3'MeDAB) we have observed an increase of aldolase A (muscle type) which is relatively abundant in fetal liver but present only in small amounts in normal adult liver. This increase is proved by the elevation of the aldolase activity ratio, by the action of specific antiserum, and by electrophoresis. Electrophoresis on starch gel shows that aldolase A is hybridized with the normal liver type aldolase (aldolase B). This hybridization indicates a synthesis of the two aldolases by the same type of cells. In fast-growing hepatomas (ascitic or solid) aldolase B is absent or nearly absent ; aldolase A is present in large amounts. Moreover, the presence of a third type of aldolase, aldolase C (brain type), which is completely absent from normal adult rat liver, has been demonstrated in fetal and in cancerous liver. This was proved by (a) the aldolase activity ratio (about 30.0 instead of 1.0), (b) electrophoresis which shows in fetal liver and in hepatoma anodic isozymes similar to brain aldolase isozymes ; (c) the action of specific antiserum anti aldolase C. The degree of inhibition by each specific antiserum corresponds roughly to that expected from the electrophoretic pattern. Fetal liver contains the three types A, B and C ; fast-growing hepatomas contain aldolases A and C and very little aldolase B. Aldolase C from fast-growing hepatomas was partly purified. Similar observations were made in human primary liver cancer. A fetal liver pattern has also been found for some other enzymes in hepatomas by several authors ; mainly hexokinases, lactic dehydrogenases, glutaminases and branched chain amino acid transaminases ; it can be compared with the resurgence of fetal antigens in cancer tissues and sera. And although the placental type of alkaline phosphatase found in cancer by Fishman was not found in fetus, it is noteworthy that placenta is an embryonic tissue. S. Weinhouse has shown that the liver-type isozyme of several enzymes is replaced by non-hepatic types in fast-growing hepatomas. This « fetal patternis not specific for carcinogenesis : for example « fetal-typeof several muscle enzymes is present in atrophied and dystrophic muscle. In conclusion, it seems likely that in several pathological states the biosynthesis of the adult form of enzymes is preferentially « repressedwhile the fetal, or less differentiated molecular form would be derepressed.

Aldolase isozymes in cancer

There are two different types of aldolase in mammalian tissues, the main difference between these isozymes is the relative activity towards the second substrate of aldolase, fructose- 1-phosphate (F- 1-P). In normal liver the ratio of the activities towards fructose di-phosphate (F-d-P) and towards fructose- 1-phosphate, is near 1, because normal liver contains almost exclusively the type of aldolase characterized by a ratio F-d-P/F-1-P= 1. In human primary liver cancer (and also in experimental hepatoma), this ratio is raised to a mean value of 5.5. On the contrary, muscle contains almost exclusively the type of aldolase with a ratio greater than 50. The two types of aldolase exist in normal spleen, but principally the latter type. In spleen of normal mice, we have found an aldolase ratio of 8.97; in experimental cancer (reticulum cell sarcoma of spleen) we have found the ratio lowered to 4·51. These results are discussed in presenting the hypothesis of the repression, in cancer, of the most differentiated enzyme.