Search

Background/aims: Hepatocellular carcinoma is the main type of primary liver cancer. Macroscopic vascular invasion is usually identified during imaging, whereas microvascular invasion is usually determined by histopathological evaluation. We aim to identify the association between microvascular invasion and other markers of tumor aggressiveness and to identify the role of microvascular invasion in the prognosis of patients who were treated by liver transplantation for hepatocellular carcinoma., Materials and Methods: This is a single-center retrospective analysis of prospectively collected data. Patients who received liver transplantation for hepatocellular carcinoma were included in the study. Data were collected regarding sociodemographic variables, criteria of selection for liver transplantation, pretransplant alpha-fetoprotein, presence or absence of microvascular invasion, presence or absence of recurrence, overall survival, and disease-free survival. Data were analyzed using Statistical Package for the Social Sciences., Results: Sociodemographic laboratory values and radiologic tumor characteristics were found to be similar in patients with or without microvascular invasion. Our study revealed that microvascular invasion is associated with increased recurrence, decreased diseasedfree survival, and decreased overall survival, only for patients with hepatocellular carcinoma beyond Milan criteria at the time of liver transplantation., Conclusion: For patients beyond Milan criteria, but not within Milan criteria, microvascular invasion plays a significant role in predicting recurrence and shorter survival after liver transplantation.

Background: Macroscopic portal vein thrombosis (PVT) is a major poor prognosis factor in patients with hepatocellular carcinoma (HCC). Inflammation is increasingly recognized to be part of the hepatocarcinogenic process and its markers are also prognostically useful., Aims: To examine the relationship of inflammation biomarkers to the presence of PVT and to survival in PVT patients with HCC., Methods: A large HCC cohort was examined for the presence of PVT and analyzed retrospectively., Results: Blood levels of NLR, PLR, ESR, CRP, AFP and GGTP were significantly related to the presence of PVT, but not the Glasgow Index. For patients with low alpha-fetoprotein levels, blood ESR and GGTP levels were also significantly increased in patients with PVT compared with those in patients without PVT. In a Cox regression model, serum GGTP levels had a significantly increased hazard ratio on death (1.52, p = 0.008). Kaplan-Meier analysis showed that PVT patients with low serum GGTP levels had significantly longer survival than PVT patients with high GGTP levels (p = 0.0041)., Conclusions: Indices of inflammation, especially serum GGTP levels, related significantly to the presence of PVT and to survival in HCC patients with PVT.

Cdc25A, a dual specificity protein phosphatase, is well-recognized as a critical regulator for cell cycle progression. We recently found that it also regulates mitogen-activated protein kinase (MAPK) signal transduction pathway. Inhibition of Cdc25A activity by a K vitamin analog Compound 5 (Cpd 5) can induce a strong and prolonged activation of epidermal growth factor receptor (EGFR)-MAPK pathway, which leads to suppression of transcription factors CREB and c-Myc, resulting in decreased expression of Cdc25A and cyclin D1 levels. Our investigations suggest that Cdc25A plays a central role in regulating and linking cell cycle progression and MAPK signal transduction pathways. Several other recently synthesized K vitamin analogs also affect this pathway, including the non-quinone PM20 and fluoro-Cpd 5. Thus, searching for new and efficient small molecules to inhibit Cdc25A activity may provide new means to control cancers of the liver and other sites.

Liver transplantation (LT) which is currently an established therapy for sma1l, early stage hepatocellular carcinoma (HCC) in patients with cirrhosis requires in most cases long waiting period. Tumor development during the waiting period may be associated with vascular invasion which is a strong factor of postoperative recurrence. Therefore, local treatment of the tumor including trans-arterial chemoembolization (TACE), percutaneous radiofrequency (RF) or partial liver resection can be used before transplantation. In the present paper we reviewed the efficacy of these treatments prior to LT. Although, TACE induced complete tumor necrosis in some patients there is no convincing arguments showing that this treatment reduces the rate of drop out before LT, nor improves the survival after LT. Although, RF can induce complete necrosis in the majority of small tumors (<2.5 cm), there is no data demonstrating that this treatment reduce the rate of drop out before LT, nor improves the survival after LT. It has been showed that both short and long term survival after LT was not compromised by previous partial liver resection of HCC. However, there is no data demonstrating that liver resection before LT, which can be used either as a bridge treatment or as a primary treatment, improves the survival after LT. The current data suggest that there is no role for pre-transplant therapy for HCC within Milano criteria transplanted within six months. On the opposite, if the waiting time is predicted to be prolonged, the risk of tumor progression and either drop-off from the list or interval dissemination with post-transplant tumor recurrence is recognized. In this setting, bridge therapy can reduce that risk but its efficacy has to be determined.

Most patients with hepatocellular carcinoma (HCC) also have cirrhosis, an independent cause of death. We considered an alternative definition of tumor-related death in patients with HCC and attempted to validate our definition. Two hundred thirty-seven HCC patients were diagnosed, followed, and died over a 12-year period and were evaluated every 2 months, including their last 6 months of life. We defined death by cancer if there was, in the last 6 months of life, a CT scan increase of >25% in the sum of tumor index lesions' cross-sectional areas or new onset of, or increase in, either vascular invasion or metastatic disease (Group 1). Patients with stable cancer were considered to have died from any other cause (Group 2). We found that 135 (57%) patients died from cancer progression (Group 1), whereas 102 (43%) patients did not (Group 2). There was a statistically significant difference between Group 1 and Group 2 patients in percentage with bilobar disease (P = 0.03), more than one tumor (P = 0.01), an increase in AFP (P = 0.04), vascular invasion (P = 0.001), and the presence of metastases (P = 0.01). We conclude that 57% of patients with unresectable HCC died as a direct result of cancer progression, but 43% did not. The latter died from complications of their cirrhosis, including sepsis, GI bleeds, and renal failure.

Hepatocellular carcinoma (HCC) is estimated to be responsible for 250,000 deaths worldwide yearly. Aggressive surgical resection or liver transplantation still remain the only viable curative options for patients suffering the disease despite the multitude of emerging therapies for HCC. However, even with the most aggressive surgical intervention, survival varies widely within each particular stage of HCC. In order to improve utilization of available therapeutic modalities, a number of outcome prognostic models have been developed. This manuscript reviews the prognostic models most commonly utilized in clinical practice and the statistical methodologies on which these models are based. A multitude of statistical and mathematical techniques can be used for prognostic model development. The most common methodologies used for HCC prognostic model development can be generally divided into four groups: survival, artificial neural networks, analysis of variance, and cluster analysis. Survival methodologies (such as Cox proportional hazard model) are commonly employed for estimation of relative significance of risk factors for patient survival or cancer recurrence. Artificial neural networks (such as back-propagation network) can be supreme approximation tools for any continuous or binary function, and as such can be employed for prognostication of HCC recurrence (death). Analysis of variance and cluster analysis are the most common statistical tools of recently evolved microarrays technology, which, in turn, is one of the most promising tools available to the cancer researcher.

Background: To test the reliability, sensitivity to change in biomarkers associated with disease progression and response to treatment, and clinical meaningfulness of the Functional Assessment of Cancer Therapy-Hepatobiliary (FACT-Hep) in patients with hepatobiliary carcinoma., Patients and Methods: One hundred and fifty-eight patients diagnosed with hepatobiliary carcinoma were prospectively studied. Health-related quality of life (HRQL) was assessed at baseline (prior to treatment), 3-month follow-up (n=55) and 6-month follow-up (n=27)., Results: The internal consistency of all the scales of the FACT-Hep were adequate at all time points (>0.75). The FACT-Hep was found to be sensitive to changes in clinical indicators (alkaline phosphate, alpha-fetoprotein, hemoglobin and survival) that reflect disease progression and response to treatment. Combined results from distribution-based and cross-sectional anchor-based analyses provide the following minimally important difference (MID) estimates: FACT-General (FACT-G) subscales=2-3; FACT-G=6-7; Hepatobiliary Cancer Subscale=5-6; FACT-Hep=8-9; Trial Outcome Index=7-8; and FACT-Hepatobiliary Symptom Index=2-3 points., Conclusions: The FACT-Hep is a reliable instrument that is responsive to clinical indicators of disease progression and response to treatment. The MID estimates can aid interpretation of HRQL data and facilitate sample size calculation in clinical trials.

Background: The aims of the study were to test the difference in health-related quality (HRQL) of life and survival in patients diagnosed with primary hepatocellular carcionma (HCC) and treated with either hepatic arterial infusion (HAI) of Cisplatin or 90-Yttrium microspheres (Therasphere)., Method: The design of the study was a non-randomized parallel cohort study. Twenty-eight patients participated in the present study. HRQL was assessed by administration of the Functional Assessment of Cancer Therapy-Hepatobiliary. Survival was measured using Kaplan Meier methods., Results: The results of present study suggest treatment with Therasphere) had an advantage in regard to HRQL and survival when compared to Cisplatin. At 3-month follow-up, patients who were treated with Therasphere had a higher level of functional well-being as well as overall quality of life when compared to patients treated with Cisplatin. At 6-month follow-up patients (treated with Therasphere) continued to have better functional well-being when compared to patients being treated with HAI of Cisplatin. At 6-month follow-up, survival was found to be similar for patients treated with Therasphere when compared to patients being treated with Cisplatin., Conclusions: Preliminary data suggest that treatment with Therasphere has a modest advantage in regard to HRQL when compared patients treated with HAI of Cisplatin. Future research with Therasphere, that includes a larger sample size and longer follow-up, is necessary to make definitive conclusions regarding the efficacy and effect on HRQL., (Copyright 2003 John Wiley & Sons, Ltd.)

The number of deaths per year from hepatocellular carcinoma (HCC) exceeds 250000, placing it sixth as the cause of death from cancer worldwide. The primary etiology of most cases of HCC in the US is hepatitis B and/or C. Extensive research has demonstrated that the relationship between hepatitis B infection and the progression to HCC is mediated by the immune system. A substantial, but unrelated literature, describes the relationship between psychosocial factors (e.g., stress, psychiatric morbidity), immune system reactivity, and disease progression in patients with cancer. However, the role of these factors in the progression of HBV-HCC has not been explored. An understanding of the relationship among virology, immunology, and behavior in the development and recurrence of HCC may provide alternative methods for secondary prevention of HCC (e.g., behavioral) until a vaccine and/or pharmacological treatments are developed, feasible, and affordable.

We previously found that K vitamin analogues caused cell growth inhibition in Hep3B hepatoma cells in vitro, which was associated with their inhibitory effects on protein tyrosine-phosphatases. In this study, we show that Cdc25A, a protein phosphatase, was inactivated by novel arylating K vitamin analogues. The inactivation of Cdc25A correlated with their effects on cell growth inhibition. Cyclin-dependent kinase (Cdk) 4, an important regulator for G(1) progression, was found to be tyrosine-phosphorylated by the arylating analogues, and this phosphorylation was correlated with the inhibitory effects of the analogues on Cdc25A activity. Furthermore, Cdk4 dephosphorylation experiments showed that Compound (Cpd) 5, a prototype arylating analogue, inhibited Cdc25A-mediated Cdk4 dephosphorylation, whereas Cpd 26, a nonarylating vitamin K analogue, had no effect on this event. We also examined Cdk4 kinase activity using retinoblastoma protein as a substrate and found that Cpd 5 inhibited retinoblastoma protein phosphorylation in a concentration-dependent manner, indicating that Cdk4 activity was inhibited by Cpd 5 treatment. Moreover, the thiol-antioxidants glutathione and N-acetyl-L-cysteine antagonized the Cpd 5-induced Cdk4 tyrosine phosphorylation, whereas the nonthiol-antioxidants catalase and superoxide dismutase did not. These results suggest that Hep3B cell growth inhibition by these K vitamin analogues may be related in part to inactivation of Cdc25A activity and support the hypothesis that Cdc25A is an attractive target for drugs designed to inhibit cancer cell growth.

The gross and histopathologic characteristics of 212 nonfibrolamellar hepatocellular carcinomas (HCCs) discovered in native livers removed at the time of liver transplantation were correlated with features of invasive growth and tumor-free survival. The results show that most HCCs begin as small well-differentiated tumors that have an increased proliferation rate and induce neovascularization, compared with the surrounding liver. But at this stage, they maintain a near-normal apoptosis/mitosis ratio and uncommonly show vascular invasion. As tumors enlarge, foci of dedifferentiation appear within the neoplastic nodules, which have a higher proliferation rate and show more pleomorphism than surrounding better-differentiated areas. Vascular invasion, which is the strongest predictor of disease recurrence, correlates significantly with tumor number and size, tumor giant cells and necrosis, the predominant and worst degree of differentiation, and the apoptosis/mitosis ratio. In the absence of macroscopic or large vessel invasion, largest tumor size (P <.006), apoptosis/mitosis ratio (P <.03), and number of tumors (P <.04) were independent predictors of tumor-free survival and none of 24 patients with tumors having an apoptosis/mitosis ratio greater than 7.2 had recurrence. A minority of HCCs (<15%) quickly develop aggressive features (moderate or poor differentiation, low apoptosis/mitosis ratio, and vascular invasion) while still small, similar to flat carcinomas of the bladder and colon. In conclusion, hepatic carcinogenesis in humans is a multistep and multifocal process. As in experimental animal studies, aggressive biologic behavior (vascular invasion and recurrence) correlates significantly with profound alterations in the apoptosis/mitosis ratio and with architectural and cytologic alterations that suggest a progressive accumulation of multiple genetic abnormalities.

Background/aims: To understand the mechanisms of liver regeneration or hepatoma apoptosis, it is important to estimate the turning point of the signal transduction by growth factor receptor. Since 2-(2-hydroxyethylsulfaryl) 3-methyl-1,4-naphthoquinone or CPD 5 has been shown to mediate the phosphorylation of epidermal growth factor (EGF) receptor in Hep3B hepatoma cells, the differences between EGF and CPD 5-mediated signal transduction were studied., Methods: DNA content was measured by Hoechst fluorescent assay. Phosphorylated proteins were described with Western blots or two-dimensional electrophoresis., Results: CPD 5-induced EGFR phosphorylation was functional to stimulate Ras pathway. However, CPD 5-mediated extracellular signal-regulated kinase (ERK) phosphorylation was not antagonized by inhibition of upstream activation with PD153035. CPD 5 inhibited ERK dephosphorylation in cell lysate, suggesting that ERK phosphorylation by CPD 5 was depending on kinase activity and phosphatase inhibition. Two-dimensional electrophoresis showed extra phospho ERK spot, which was indicated to have close association with CPD 5-induced growth inhibition, since U0126 antagonized growth inhibition and appearance of this spot., Conclusions: The turning point of EGFR pathway was proved to have close association with the expressed level of phosphorylated ERK. ERK phosphorylation was suggested to play a critical role in growth factor-induced signal transduction.

To study the Hoxa-13 gene in the liver, we examined its expression by RT-PCR in various liver cell lines, rat livers under different conditions, and human primary hepatocellular carcinomas (HCCs). The gene was found to be expressed in cell lines originating from liver stem-like cells, but not in cell lines originating from hepatocytes and bile duct epithelia. Expression was induced in rat livers after treatment with d-galactosamine, which is known to induce oval cell proliferation, but not after a two-thirds partial hepatectomy (2/3 PH) where induction of oval cell proliferation is thought not to occur. Expression of the gene correlated with human HCC samples associated with Hepatitis B or C virus infection in this small series. These results suggest that the Hoxa-13 gene may provide a potentially useful tool for elucidation of mechanisms involved in lineage-specific differentiation and carcinogenesis of liver stem cells., (Copyright 2001 Academic Press.)

Objective: The purpose of this study was to use hepatic arterial phase helical CT to assess tumor vascularity and predict the likelihood of response to transcatheter arterial chemoembolization in patients with hepatocellular carcinoma., Materials and Methods: Helical CT findings for 57 patients with hepatocellular carcinoma were classified into one of three patterns of vascularity on the basis of the degree of tumor or liver enhancement during the hepatic arterial phase. Cases in which hypervascular lesions predominated were classified as a type 1 pattern. Cases in which hypovascular lesions predominated were classified as a type 2 pattern. Patients were classified as responders or nonresponders on the basis of the changes of tumor size revealed on CT after three transcatheter arterial chemoembolization treatments., Results: We classified the 57 patients as 37 responders (65%) and 20 nonresponders (35%). A statistically significant correlation between the type 1 hypervascular pattern and response to transcatheter arterial chemoembolization was seen; conversely, the type 2 hypovascular pattern correlated with nonresponse to transcatheter arterial chemoembolization (chi-square = 7.85, p = 0.02). Patients classified as responders lived significantly longer than those classified as nonresponders with 12-, 24-, and 36-month survival rates of 90%, 67%, and 36%, respectively, for responders and 70%, 17%, and 10%, respectively, for nonresponders., Conclusion: We found that patients who responded to transcatheter arterial chemoembolization had prolonged survival (p < 0.01). Response correlated closely with tumor vascularity as shown on hepatic arterial phase helical CT.

Objective: To determine the effectiveness of a new multidisciplinary approach using neoadjuvant intrahepatic chemoembolization (IHCE) and liver transplant (OLTx) in patients with unresectable hepatic tumors who have failed systemic chemotherapy., Materials and Methods: From November 1989 to April 1998, 14 children (2-15 years old) were treated with 50 courses of intra-arterial chemotherapy. Baseline and post-treatment contrast-enhanced CT and alpha-fetoprotein levels were performed. Seven had hepatoblastoma, and 7 had hepatocellular carcinoma (1 fibrolamellar variant). All patients had subselective hepatic angiography and infusion of cisplatin and/or adriamycin (36 courses were followed by gelfoam embolization). The procedure was repeated every 3-4 weeks based on hepatic function and patency of the hepatic artery., Results: Six of 14 children received orthotopic liver transplants (31 courses of IHC). Pretransplant, 3 of 6 showed a significant decrease in alpha-fetoprotein, while only 1 demonstrated a significant further reduction in tumor size). Three of 6 patients are disease free at this time. Three of 6 patients died of metastatic tumor 6, 38, and 58 months, respectively post-transplant. One of 14 is currently undergoing treatment, has demonstrated a positive response, and is awaiting OLTx. Three of 14 withdrew from the program and died. Four of 14 patients developed an increase in tumor size, developed metastatic disease, and were not transplant candidates. Two hepatic arteries thrombosed, and one child had a small sealed-off gastric ulcer as complications of intrahepatic chemoembolization., Conclusion: The results of intrahepatic chemoembolization are promising and suggest that some children who do not respond to systemic therapy can be eventually cured by a combination of intrahepatic chemoembolization orthotopic liver transplant. Alpha-fetoprotein and cross-sectional imaging appear to be complementary in evaluating tumor response. IHCE does not appear to convert an anatomically unresectable lesion to a candidate for partial hepatectomy.

Background: The current staging system of hepatocellular carcinoma established by the International Union Against Cancer and the American Joint Committee on Cancer does not necessarily predict the outcomes after hepatic resection or transplantation., Study Design: Various clinical and pathologic risk factors for tumor recurrence were examined on 344 consecutive patients who received hepatic transplantation in the presence of nonfibrolamellar hepatocellular carcinoma to establish a reliable risk scoring system., Results: Multivariate analysis identified three factors as independently significant poor prognosticators: 1) bilobarly distributed tumors, 2) size of the greatest tumor (2 to 5 cm and > 5 cm), and 3) vascular invasion (microscopic and macroscopic). Prognostic risk score (PRS) of each patient was calculated from the relative risks of multivariate analysis. The patients were grouped into five grades of tumor recurrence risk: grade 1: PRS = 0 to < 7.5; grade 2: PRS = 7.5 to < or = 11.0; grade 3: PRS > 11.0 to 15.0; grade 4: PRS > or = 15.0; and grade 5: positive node, metastasis, or margin. The proposed PRS system correlated extremely well with tumor-free survival after liver transplantation (100%, 61%, 40%, 5%, and 0%, from grades 1 to 5, respectively, at 5 years), but current pTNM staging did not., Conclusions: 1) Patients with grades 1 and 2 are effectively treated with liver transplantation, 2) patients with grades 4 and 5 are poor candidates for liver transplantation, and 3) patients with grade 1 do not benefit from adjuvant chemotherapy.

Purpose: To determine the relative frequency, incidence, and locations of metastases of hepatocellular carcinoma (HCC), correlate extrahepatic metastatic disease with intrahepatic tumor staging, and determine the computed tomographic (CT) manifestations of HCC metastases., Materials and Methods: CT findings in 403 consecutive patients with HCC at our institution since 1992 were reviewed retrospectively. One hundred forty-eight patients with extrahepatic metastatic HCC were identified, and the locations, sizes, and attenuation and enhancement characteristics of the lesions were recorded., Results: A majority (128 [86%] of 148) of patients with extrahepatic HCC foci had either intrahepatic stage IVA tumor (112 [76%] patients) or an intrahepatic stage III tumor (16 [11%] patients) at the occurrence of metastases. The most frequent site of the first detectable metastasis was the lung (58 [39%] patients). Tabulation of all extrahepatic metastatic sites showed the most common to be the lung in 81 (55%) patients, the abdominal lymph nodes in 60 (41%) patients, and the bone in 41 (28%) patients., Conclusion: The lung, abdominal lymph nodes, and bone are the most common sites of extrahepatic metastatic HCC. Most extrahepatic HCC occurs in patients with advanced intrahepatic tumor stage (stage IVA). Incidental extrahepatic lesions at CT in patients with stage I or II intrahepatic HCC are unlikely to represent metastatic HCC.

Background/aim: Both p16INK4a and p53 proteins are negative regulators of the cell cycle. In human hepatocellular carcinomas (HCC), the loss of function of p53, retinoblastoma (pRb) and pl6INK4a genes by different mechanisms has been largely documented, but their hepatocellular effects are poorly known. We compared the growth-inhibitory effects of p16INK4a and p53 proteins in Hep3B cell line-derived clones., Methods: Cells were transfected with inducible p16INK4a and p53 expression vectors, and stable clones were analyzed for transgene expression by Western blotting and immunoperoxidase staining. Effects on cell growth were analyzed by in vitro growth assay, thymidine incorporation and flow cytometry. Biochemical effects of p53 were tested by Northern blotting of p21Cip1 transcripts and by Western blotting of p21Cip1, mdm-2, bax, cyclin-dependent kinase 2 and cyclin E proteins. The pRb protein was studied by Western blotting and immunoprecipitation assays., Results: The induction of p16INK4a protein expression did not affect in vitro growth of cells. In contrast, p53 protein in its wild-type conformation provoked a growth arrest accompanied by transactivation of p21Cip1 gene and accumulation of p21Cip1, bax and mdm-2 proteins. p53-induced growth arrest was due to a cell cycle arrest at the G1/S transition, probably mediated by p21Cip1 protein, which inhibits cyclin-dependent kinase 2/cyclin E complexes., Conclusions: The lack of detectable pRb protein and resistance of cells to p16TNK4a strongly suggest that p53 is able to arrest the growth of HCC cells by a mechanism independent of "p53-retinoblastoma pathway". These findings are applicable to HCC with abberrations of both p53 and pRb genes, and may not represent the universal effects of p53 in hepatic cells.

Objective: To describe our experience with total hepatectomy and liver transplantation as treatment for primary hepatoblastoma (HBL) and hepatocellular carcinoma (HCC) in children., Study Design: A retrospective analysis of the perioperative course of 31 children with unresectable primary HBL (n = 12) and HCC (n = 19) who underwent transplantation between May 1989 and December 1998. Systemic (n = 18) and intraarterial (n = 7) neoadjuvant chemotherapy were administered; follow-up ranged from 1 to 185 months., Results: For HBL, 1-year, 3-year, and 5-year posttransplantation survival rates were 92%, 92%, and 83%, respectively. Intravenous invasion, positive hilar lymph nodes, and contiguous spread did not have a significant adverse effect on outcome; distant metastasis was responsible for 2 deaths. Intraarterial chemotherapy was effective in all patients treated. For HCC, the overall 1-year, 3-year, and 5-year disease-free survival rates were 79%, 68%, and 63%, respectively. Vascular invasion, distant metastases, lymph node involvement, tumor size, and gender were significant risk factors for recurrence. Intraarterial chemotherapy was effective in 1 of 3 patients. Six patients died of recurrent HCC, and 3 deaths were unrelated to recurrent tumor., Conclusion: Liver transplantation for unresectable HBL and HCC can be curative. Risk factors for recurrence were significant only for HCC, with more advanced stages amenable to cure in the HBL group.

We recently found that a thioether analog of K vitamin (Cpd 5) inhibited the activity of protein-tyrosine phosphatases (PTPases) and induced protein-tyrosine phosphorylation in a human hepatoma cell line (Hep3B). We have now examined the structural requirements for induction of protein-tyrosine phosphorylation and PTPase inhibition by several K vitamin analogs. Thioether analogs with sulfhydryl arylation capacity, especially those with a hydroxy (Cpd 5) or a methoxy group at the end of the side chain, induced protein-tyrosine phosphorylation, but non-arylating analogs, such as those with an all-carbon or O-ether side chain, did not. Among the receptor-tyrosine kinases, epidermal growth factor receptors were tyrosine-phosphorylated by treatment with thioether analogs, whereas insulin and hepatocyte growth factor receptors were not. An increase in tyrosine-phosphorylated ERK2 mitogen-activated protein kinase was also observed. The activity of purified T cell PTPase was inhibited only by the thioether analogs, but not by non-arylating analogs. Furthermore, the epidermal growth factor receptor dephosphorylation activity of Hep3B cell lysates was inhibited by Cpd 5 treatment. A similar induction of protein-tyrosine phosphorylation by Cpd 5 was seen in other human hepatoma cell lines together with growth inhibition. However, one cell line (HepG2), which was relatively resistant to growth inhibition by Cpd 5, did not increase its phosphorylation levels upon Cpd 5 treatment. These results suggest that cell growth inhibition by thioether analogs is closely associated with inhibition of PTPases by sulfhydryl arylation and with tyrosine phosphorylation of selected proteins.

Jaundice in hepatocellular carcinoma (HCC) can be due to biliary obstruction. Endoscopic retrograde cholangiopancreatography (ERCP) can be both diagnostic and therapeutic. Biliary stenting can relieve jaundice and allow further chemotherapy, but at additional expense and potential morbidity. We sought to determine whether CT scan or ultrasound (US) could identify which patients with HCC and jaundice would benefit from endoscopic stenting. We retrospectively analyzed 26 patients with HCC and jaundice who underwent ERCP after CT or US. We compared biliary dilation on CT or US with the dominant biliary stricture seen on ERCP, and with response to biliary stenting. Eleven of 26 patients had dominant biliary stricture on ERCP; 11 underwent stenting. Six of 11 (55%) stented patients had a significant decline in bilirubin; three became eligible for further chemotherapy. All six responders to stenting had biliary dilation on prior CT or US. Procedure-related complications occurred in 1/11 (9%) who underwent stent placement. In conclusion, in selected patients, stenting can safely relieve jaundice and allow subsequent chemotherapy. CT or US accurately predicted lesions that responded to stenting. ERCP and stenting provided no benefit in the absence of biliary dilation on CT or US.

Primary hepatocellular carcinomas (HCCs) of different etiologies were studied to determine the rate of alteration of several genetic regions previously associated with the HCC phenotype. The focus of our study was to identify the frequency of genetic alterations within individual HCCs and their distribution among male and female cases. Genetic differences were evaluated between DNA isolated from tumor (T) and corresponding non-tumor (N) tissue using short tandem repeat (STR)-microsatellites and restriction fragment length polymorphism (RFLP) analyses. Twenty-eight HCC cases were studied with polymorphic markers from different parts of the genome. Three or more loci were identified with genetic alterations from 28 loci tested in 63% of HCC cases. The highest frequency of alteration occurred in the chromosome regions 1p36, 13q14, 17p13, and the 4q28 region identified in HCCs for the first time. High loss of heterozygosity (LOH) in 1p36, 4q28, 13q14, and 17p13 regions indicates important HCC suppressors within the regions. The study documents no association between cause and specific genetic change or their frequency in HCCs analyzed. The data document a significant gender distortion for genetic alteration in chromosome 13q14 and 17p13 regions and a concordant gender alteration rate in the 1p36 and 4q28 regions. An overall higher frequency of genetic alterations was identified in male cases. Future study with an extended number of HCC cases should substantiate the frequency of alteration between genders and identify tentative suppressors in the 1p36 and 4q28 regions.

We examined stage T1 to T4 hepatocellular carcinomas (HCC) to determine whether transforming growth factor alpha (TGFalpha) presence differed between early- and late-stage HCC and between tumors with low and high proliferative rates. Paraffin sections from 36 HCC were evaluated for TGFalpha and the proliferation markers Kiel 67 antigen (Ki67) or proliferating cell nuclear antigen (PCNA) by immunoperoxidase staining. In 12 cases, double staining for TGFalpha and Ki67 was also performed. Eighty-one percent of tumors and 94% of adjacent liver sections contained TGFalpha. A trend toward inverse correlation was seen between the percentage of TGFalpha-positive tumor cells and the proliferative rate as determined by Ki67 staining. No clear correlation of TGFalpha to either tumor stage or percentage of PCNA-positive cells was seen. This study confirms the presence of TGFalpha in the majority of early- and late-stage HCC. Positivity within tumor tissue is consistent with autocrine or paracrine stimulation. A trend toward inverse correlation between TGFalpha-producing cells and the number of cycling cells suggests that rapidly proliferating tumors may consume this growth factor at an accelerated rate. Alternatively, other hepatic mitogens may have more functional significance in these latter tumors. Finally, the presence of TGFalpha in peritumoral hepatocytes suggests these cells as potential sources of paracrine stimulation for HCC.

Background: Hepatocellular carcinoma in cirrhotic patients increases the risk of variceal bleeding. We sought to characterize bleeding in a cirrhotic patient population undergoing intrahepatic artery chemotherapy for hepatocellular carcinoma and to determine the possible influence of this treatment on gastrointestinal bleeding., Methods: We retrospectively reviewed 179 patients with hepatocellular carcinoma who underwent intrahepatic artery doxorubicin and cis-platinum chemotherapy to determine the incidence of gastrointestinal bleeding and compared them with 434 hepatocellular carcinoma historic controls not undergoing regional chemotherapy., Results: Of the 179 patients, 27 patients (15.1%) developed upper gastrointestinal bleeding over a mean follow-up of 15.2 months; 18 of the 27 (66.7%) bled from a variceal source and 9 (33.3%) bled from a nonvariceal source: ulcer (n = 6), gastropathy (n = 1), Mallory-Weiss (n = 1), erosive gastritis (n = 1). Twenty-one patients developed bleeding after initiation of chemotherapy (14 variceal and 7 nonvariceal). The number of chemotherapy sessions among patients with variceal and nonvariceal bleeding was similar (2.1 +/- 0.4 and 4.0 +/- 1.2; p = Not significant). Patients with variceal and nonvariceal bleeding were comparable with respect to Child-Pugh classification, pTNM stage, age, time to bleeding, and gender., Conclusions: Regional intra-arterial chemotherapy for hepatocellular carcinoma is associated with a low risk of variceal bleeding. Nonvariceal sources of upper gastrointestinal bleeding in this population account for a significant component of bleeding episodes.

Purpose: To assess the role of helical biphasic computed tomography (CT) versus CT during arterial portography (CTAP) in the detection of hepatocellular carcinoma (HCC)., Materials and Methods: Helical biphasic CT and CTAP examinations were performed in 33 patients with biopsy-proved HCC. Three blinded radiologists retrospectively evaluated portal venous phase images, portal venous phase plus hepatic arterial phase images (helical biphasic CT), and CTAP plus portal venous phase images at separate sittings. In 23 patients with pathologic proof of total tumor burden, the readers conducted a consensus matched-pair reading to determine the sensitivity of each method., Results: In the 23 patients with a proved tumor burden of 100 HCC lesions, the observers detected 74 tumors on portal venous phase images, 82 tumors on hepatic arterial phase images, 87 tumors on CTAP images, 87 tumors on helical biphasic CT images, and 96 tumors on portal venous phase plus CTAP images. Hepatic arterial phase images and CTAP images, respectively, depicted 13 and 23 tumors not seen on portal venous phase images in eight (35%) and 13 (56%) of 23 patients. Ten tumors were seen on portal venous phase images but not on CTAP images owing to the presence of central tumors or segmental portal venous thrombi that obstructed the flow of contrast material to more peripheral portions of the liver., Conclusion: The addition of hepatic arterial phase imaging to portal venous phase imaging (helical biphasic CT) provided an incrementally increased yield in HCC tumor detection. Helical biphasic CT should be used as the initial screening examination in patients with suspected or known HCC.

Orthotopic liver transplantation (OLTx) in the presence of hepatocellular carcinoma (HCC) has been complicated by high recurrence rates. The ability to determine the risk and timing of HCC recurrence on an individual basis would greatly aid in the candidate selection process resulting in a more efficient use of donated organs and allow the individualization and better evaluation of adjuvant chemotherapy. The 214 patients who underwent OLTx in the presence of HCC were analyzed. From the 178 patients who survived more than 150 days, 71 (40%) have suffered HCC recurrence. Based on five risk factors, that is, gender, tumor number, lobar tumor distribution, tumor size, grade of vascular invasion, artificial neural network models predicting the likelihood of HCC recurrence within 1, 2, and 3 consecutive years after transplantation were developed. Based on model predictions, those combinations of risk factors that should/should not lead to recurrence were generated, allowing stratification of patients into the following three groups: 1) patients who should not suffer HCC recurrence and who should not need adjuvant therapy, 2) patients who will suffer recurrence and for whom postoperative chemotherapy significantly prolonged survival (but did not prevent recurrence), and 3) patients who may or may not suffer HCC recurrence and whose recurrence may be prevented by adjuvant chemotherapy. The outcome of OLTx for patients with HCC can be prognosticated based on a number of clinical variables. If verified through multicenter trials, these models could be made available to transplantation programs performing OLTx in the presence of HCC.

Met and ron proto-oncogenes encode the cell surface receptors for hepatocyte growth factor (HGF) and hepatocyte growth factor-like (HLP) protein, respectively, and induce mitogenesis, motogenesis, morphogenesis, and metastatic activity in various cell types. Overexpression of met in human carcinoma has been reported by several groups including ours; however, the mechanisms that control met gene expression are thus far unclear. The present study focuses on the expression and regulation of the Met and Ron receptors in human hepatocellular carcinoma (HCC). We report here that abnormal expression of met and ron proteins occurs in some cases of human HCC. Using several HCC cell lines as a model system, we show that HGF, as well as other cytokines, such as epidermal growth factor (EGF), interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF-alpha), induce met and ron expression. Using several chimeric constructs consisting of various lengths of the met promoter region fused to the reporter gene of chloramphenicol acetyl transferase (CAT), and by performing transient transfection of these constructs into HepG2 cells, we show that induction of met gene expression by HGF and other cytokines is, at least in part, through up-regulation of met gene promoter activity. The DNA region conferring responsiveness to cytokine induction was located within 0.2 kb of the met core promoter. Interestingly, EGF did not stimulate met promoter activity in any of the met-CAT chimeric constructs. These results provide evidence that met and ron are modulated in the liver by a similar cytokine network. In the case of met expression, the 0.2-kb region in the met gene promoter may play an important role in mediating its gene induction in response to HGF and other cytokines. Our results also suggest that unregulated expression of met and ron may be associated with pathological conditions, such as HCC, in the liver.

A monoclonal antibody highly specific for benign and malignant hepatocytes (HepPar 1) was evaluated as part of an antibody panel used to differentiate hepatocellular from nonhepatocellular neoplasms. Sixty-five liver tumors and two extrahepatic tumors from patients with documented liver tumors were studied. Twenty-two neoplasms were of hepatocellular origin, three were combined hepatocellular/cholangiocarcinomas, and the remainder were of nonhepatocellular origin. HepPar 1 alone had an 82% sensitivity and 90% specificity for the detection of hepatocellular neoplasms. The corresponding values for alpha-fetoprotein were 57% and 97%. Polyclonal antibody to carcinoembryonic antigen (canalicular pattern) had a sensitivity of 79% and specificity of 97% for these tumors. The use of antibody panels provided superior results when compared with individual antibodies. In summary, HepPar 1 monoclonal antibody is a useful reagent for the differential diagnosis of hepatocellular tumors. Its utility is enhanced when it is used as part of a diagnostic antibody panel.

Patients with advanced-stage unresectable hepatocellular carcinoma (HCC) were treated with intrahepatic arterial doxorubicin 30 mg/m2 plus escalating doses of cisplatin up to 100 mg/m2 in conjunction with rapid bolus injection of Spherex (degradable starch microspheres; Kabi Pharmacia, Lund, Sweden) into the hepatic artery, until slowing or reversal of blood flow. Treatments were repeated every 4 to 6 weeks until progression, or were continued indefinitely if there was disease stability or response. Thirty-five evaluable patients have so far been accrued to the study. Objective tumor responses have occurred in 22 patients (63%), of whom 20 had partial responses and two had complete responses. Four of the patients had reversal of tumor-induced portal vein thrombus. Toxicities included death, one patient (and a death of uncertain cause in an additional patient); hepatitis, two patients; pancreatitis, one patient; dyspnea/hypotension, two patients; and hepatic artery nontransient thrombosis in four patients. Six patients have survived 2 years and an additional 10 patients have survived 1 year. The addition of Spherex to intrahepatic arterial chemotherapy for advanced-stage HCC appears to be relatively safe and is well tolerated even in patients with portal vein thrombosis, which represent the majority of patients with advanced-stage HCC.

We screened genes responsive to transforming growth factor-beta (TGF-beta 1) protein in a human hepatoma cell line (Hep3B) using a PCR-mediated differential display technique, in order to investigate the mechanisms involved in TGF-beta-induced growth suppression. We found a gene that was down-regulated by TGF-beta 1 to be completely identical in an approx. 620 bp segment to the gene for the enzyme spermidine synthase, which mediates the conversion of putrescine into spermidine. Both spermidine synthase mRNA expression and its enzyme activity were decreased after TGF-beta 1 treatment of Hep3B cells. The inhibition of spermidine synthase gene expression by TGF-beta 1 protein was also observed in other hepatoma cell lines. The expression of genes for other biosynthetic enzymes in polyamine metabolism (ornithine decarboxylase and S-adenosylmethionine decarboxylase) was also inhibited to the same extent as for spermidine synthase, while the gene expression of spermidine/spermine N1-acetyltransferase, a catabolic enzyme, was relatively resistant to TGF-beta 1. Spermine levels in Hep3B cells were decreased by TGF-beta 1 treatment, although the levels of spermidine and putrescine were unchanged, probably due to compensation by remaining spermidine/spermine N1-acetyltransferase activity. Exogenously added spermidine or spermine, but not putrescine, partially antagonized the growth-inhibitor effects of TGF-beta 1 on Hep3B cells. Our data suggest that down-regulation of gene expression of the enzymes involved in polyamine metabolism, including spermidine synthase, may be associated with the mechanism of TGF-beta-induced growth suppression.

Purpose: To evaluate the added value of hepatic arterial-dominant phase (HAP) imaging to portal venous-dominant phase (PVP) imaging in patients with hepatocellular carcinoma (HCC) at computed tomography (CT)., Materials and Methods: Sixty-six patients with proved HCC underwent biphasic contrast-enhanced CT. HAP and PVP images were obtained at 20-50 and 60-100 seconds, respectively. PVP images were evaluated for the number of HCC foci. Then, HAP images were reviewed to determine whether any additional HCCs were seen., Results: Three hundred twenty-six tumor foci were seen. HAP images depicted 309 foci (95%) and PVP images 268 (82%). In seven patients (11%), tumor was visible only on HAP images. During the HAP, tumors were hyperattenuating compared with liver in 26 patients, of mixed attenuation in 26, and hypoattenuating in 14 without correlation with histologic appearance. Portal vein thrombosis was identified in 17 of 21 patients on HAP images; in 12 patients, the thrombosis was diagnosed as malignant with neovascularity within the thrombus or diffuse thrombus enhancement., Conclusion: Use of both HAP and PVP contrast-enhanced CT optimizes the evaluation of patients with or at risk for HCC.

Objective: To describe the fine needle aspiration (FNA) findings in metastatic hepatocellular carcinoma (HCC)., Study Design: The cytologic findings in 15 cases of extrahepatic metastatic HCC diagnosed by FNA biopsy were reviewed., Results: The anatomic sites of the FNAs were: musculoskeletal (four biopsies from 3 patients), adrenal (4 patients), regional lymph nodes (4 patients), pancreas (2 patients) and pelvic region (1 patient). The 15 aspirates came from 14 patients, 11 of whom had a biopsy-proven primary HCC. In two of the remaining patients, the FNA diagnosis of HCC in the metastatic site was the initial diagnosis. In one of these patients the diagnosis was strongly suspected on clinical grounds, but in the other case it was unsuspected. In the remaining patient the liver mass and massive retroperitoneal adenopathy were biopsied concurrently. A trabecular pattern was observed in the smears and/or cell block preparations in nine cases. Eleven cases were well to moderately differentiated. Three cases were of large cell pleomorphic type. The remaining case was poorly differentiated., Conclusion: Familiarity with the FNA cytology of HCC should allow its diagnosis in metastatic sites in most instances, even without a history of primary liver cancer.

Purpose: To correlate computed tomographic (CT) and serum tumor marker (alpha-fetoprotein [AFP] and des-gamma-carboxy-prothrombin [DGCP]) findings in recurrent hepatocellular carcinoma (HCC) after orthotopic liver transplantation (OLT)., Materials and Methods: At retrospective review of the cases in 124 patients, CT findings were recorded by consensus of at least two nonblinded observers and compared with levels of AFP and DGCP., Results: In 35 patients (68 sites), CT depicted recurrent HCC (most frequently in lungs [n = 18] and liver allograft [n = 16]) in a single site in 19 patients (54%) and in more than one site in 16 patients (46%). No stage I or II HCC recurred after 18-78 months (mean recurrence, 39 months). Stage IVA HCC recurred four times as often as stage III HCC (P < .001). Abnormally high serum AFP and DGCP levels indicated 69% and 43%, respectively, in patients with recurrent disease., Conclusion: HCC recurrence after OLT correlates with initial stage, and CT is more sensitive than serum tumor markers in its detection.

The growth characteristics of a newly established cell line, Hep40, derived from a human hepatoma are described. An absolute requirement was found for serum to mediate cell growth. Neither EGF, TGF-alpha, nor HGF altered cell growth in the presence or absence of serum. A partial suppression of cell growth was achieved by several TGF-beta family proteins. Affinity crosslinking gels using 125I-labeled TGF-beta showed a significant decrease in the TGF-beta cell-surface type II receptor in Hep40 cells, compared to the TGF-beta-sensitive Hep3B cell line. However, growth could be completely suppressed by addition of vitamins K to the culture medium in both Hep40 and several other hepatoma cell lines. Growth suppression by vitamins K was accompanied by an increased level of transcripts for c-myc, c-jun, and prothrombin genes, in contrast to the actions of TGF-beta 1 protein, which caused a decrease in the level of c-myc transcripts. These data show that this new human hepatoma cell line has partial resistance to growth inhibition by TGF-beta with a unique TGF-beta receptor defect. However, growth was completely suppressed by vitamins K. The differing gene expression patterns in response to TGF-beta as compared to vitamin K suggest that these two growth inhibitors act through differing pathways.

Plasma levels of endothelin-1 (ET-1) are markedly higher in patients with hepatocellular carcinoma (HCC) than in normal controls. In order to further investigate this, we evaluated ET-1 immunoreactivity and mRNA expression in human HCC tissue. 70% (14/20) of the tumor tissues immunostained positively with ET-1 antibody and a significant association was observed between immunostaining in cells lining the tumor feeding vessels and tumor vascularity as determined by hepatic angiography. Moreover, the neoplastic hepatocytes in the tumors also stained positively with ET-1 antibody. All of the HCC tissue samples which immunostained for ET-1 also expressed ET-1 mRNA, indicating that ET-1 is actively synthesized by the tumors.

We have studied the genetic profile of the host genome and hepatitis B virus (HBV) in HBV-associated primary hepatocellular carcinoma (HCC). Comparative analyses of HCC cell line Hep 40 and the original biopsy specimens showed the episomal and replicating form of HBV only in the biopsy specimen from nontumor (NT) cirrhotic liver tissue, where a molecular change in the 1p36 region was detected (NT tissue showed a normal 46XY karyotype). In contrast, only integrated HBV was detected in HCC tumor (T) tissue and Hep 40 cells. Two HBV integration sites were identical in HCC tissue and the hyperdiploid Hep 40 cell line, where genetic alteration in the 1p36 region was identified. These data indicate that viral replication is ongoing only in NT cirrhotic-hyperplastic, chromosomally normal tissue with evidence for genetic instability. Only the tumor cell with altered genotype has virus integrated

A characteristic defect occurs in rat and human hepatocellular carcinoma (HCC) resulting in a loss of function of the vitamin K-dependent enzyme gamma-glutamyl-carboxylase in the tumor but not in the underlying liver. This causes the secretion of elevated levels of the immature or des-gamma-carboxylated form of prothrombin, which is used as a marker of HCC. We investigated whether, using the defined conditions of growing HCC cell lines in tissue culture, addition of the naturally occurring vitamins K1 or K2 or the synthetic vitamin K3 could influence the secretion of immature prothrombin. We found that vitamins K1, K2 and K3 all suppressed the secretion of immature prothrombin into the tissue culture medium. Vitamins K2 and K3 were also found to inhibit growth of the HCC cell line, in an apparently nontoxic and reversible manner. The influence of the vitamins K on the expression of some genes related to vitamin K action was examined and compared with that of another growth inhibitor, TGF beta 1 protein. The vitamins K were found to increase the expression of prothrombin and carboxylase messenger RNA and c-myc messenger RNA, but had no effects on the expression of TGF beta 1 messenger RNA. By contrast, TGF beta 1 increased TGF beta 1 messenger RNA levels, but had no effects on the other genes, suggesting a different pathway. The previously studied vitamin K3-mediated inhibition of growth was antagonized by the addition of catalase to the culture medium, but the inhibitory effects of vitamin K2 were not antagonized.(ABSTRACT TRUNCATED AT 250 WORDS)

Background: Detection of portal vein tumor invasion in patients with hepatocellular carcinoma (HCC) is important in determining therapy and prognosis. Fine-needle aspiration (FNA) of a portal vein thrombus under ultrasound guidance facilitates the distinction of malignant from benign thrombus without resorting to laparotomy. In this study, the FNA findings of 46 patients who underwent this procedure are described., Methods: Cytologic findings of 48 aspirations from 46 patients who underwent percutaneous ultrasound-guided portal vein FNA were reviewed., Results: Twenty-nine of the 46 patients had a prior or concurrent biopsy-confirmed diagnosis of HCC at the time of portal vein FNA. On cytologic review, 39 of the aspirates were positive for malignancy, 6 were negative, and 3 were suspicious. Histologic follow-up of three of the six patients with negative aspirates confirmed bland thrombi in their portal veins. No complications resulted from the FNA procedure. Of the 39 aspirates positive for HCC, 22 were well differentiated, 5 were well to moderately differentiated, 9 were moderately differentiated, and 3 were poorly differentiated. In all except the poorly differentiated tumors, a trabecular cellular arrangement was detected in either smears or cell blocks. The cytologic findings in these aspirates, in general, mirror those found in aspirates of HCC in the liver proper., Conclusions: Portal vein FNA is an effective, well tolerated method for disease staging of patients with HCC. When used as the initial diagnostic procedure, in selected patients, it can provide the diagnosis and staging information simultaneously.

Single and double immunohistochemical staining for transforming growth factor (TGF)-alpha and epidermal growth factor-receptor (EGF-R) was done in order to identify the localization of TGF-alpha and EGF-R in human hepatocellular carcinoma (HCC). Single immunohistochemical staining for TGF-alpha showed immunoreactivity in the cytoplasm of hepatoma cells in 22 of 30 cases of HCC. The localization of TGF-alpha was heterogeneous from HCC cells to HCC cells. In the surrounding regenerative nodules, the hepatocytes were mildly to moderately positive for TGF-alpha. The proliferating bile ductules and peripheral nerves were also immunopositive for TGF-alpha. Single immunohistochemical staining for EGF-R demonstrated a linear localization of EGF-R along the cell membrane of the HCC cells in 21 of the 30 cases of HCC. In the regenerative nodules, the hepatocytes also showed linear staining along the cell membrane. Double staining for TGF-alpha and EGF-R in 12 cases of HCC showed a concurrent localization of TGF-alpha and EGF-R in some hepatoma cells and isolated localization of the two substances of other HCC cells. These combinations either abruptly moved around or intermingled with each other. These immunohistochemical results thus support the theory of an autocrine, paracrine, and endocrine mechanism of TGF-alpha and EGF-R on the proliferation of human hepatocellular carcinoma.

In the University of Pittsburgh experience, the most successful setting for human islet allografts is in patients undergoing upper abdominal exenteration with total pancreatectomy and liver transplantation for the indication of malignancy (cluster). In this group of patients 6/11 were insulin-independent for long periods. We report herein the metabolic course or the longest survivor (> 3 years). This patient has been free of exogenous insulin since the third postoperative month and has sustained her body weight without total parenteral nutrition since the 4th postoperative month. The patient has some postprandial hyperglycemia but average capillary glucoses are near-normal to normal as are glycosylated hemoglobin values. The clearance of glucose during the administration of an intravenous glucose load has been well preserved and is currently normal. C-peptide stimulates significantly in response to intravenously injected glucose. The absolute levels of stimulation during the test have declined possibly related to improvements in renal function, decreased immunosuppression or the natural history of cells transplanted into the portal site. The kinetics of the C-peptide response to intravenously injected glucose shows a persistent abnormality of first-phase insulin release and a prolonged second phase release. Basal glucagon levels are low but stimulate to a mixed meal. This patient's results demonstrate long-term function of islet cells from a single donor transplanted into the portal vein using FK506 as an immunosuppressant agent.

The pathway of growth inhibitory cellular signal transduction by transforming growth factor beta 1 is largely unknown. Although several cellular proteins have been shown to be involved in the pathway, others remain to be identified. To search for other involved proteins, differentially expressed genes were examined in two human hepatoma cell lines that were respectively sensitive and resistant to growth inhibition by TGF-beta 1. Two such genes (SB31 and SB16) were characterized and found to be 100% homologous to leucine-rich alpha 2-glycoprotein and the ribosomal protein S25, respectively. SB31 was coordinately expressed with the TGF-beta I type II receptor, implicating a possible interaction. Expression of both genes in different cell lines can be broadly correlated with the sensitivity of the cell lines to growth inhibition by TGF-beta 1. SB16 expression is strongly suppressed in rat liver regeneration after partial hepatectomy, suggesting that it may have a role in liver growth.

A large percentage of patients with advanced-stage hepatocellular carcinoma (HCC) have a recurrence of tumor in the liver or lung after primary resection and even after orthotopic liver transplantation. One reason for this may be the presence of small numbers of tumor cells circulating in the blood before surgery or the liberation of tumor cells into circulation during surgical manipulation. We tested this hypothesis by measuring messenger RNA (mRNA) for human albumin gene as a liver cell marker with the highly sensitive reverse transcriptase polymerase chain reaction (RT-PCR) technique. Albumin mRNA was not found in peripheral blood from normal humans (0 of 6), from patients with liver cirrhosis (0 of 10), from other tumors metastatic to liver (0 of 10), or during liver transplant surgery for cirrhosis (0 of 10). In patients with advanced-stage HCC (TNM stages III and IV), albumin mRNA was detected (16 of 17) in peripheral blood. After liver transplantation in the HCC patients, the level of mRNA decreased below the detectable limit (0 of 9). Three of these patients again had detectable mRNA levels when they had recurrence of HCC after liver transplantation. Patients with stage I HCC did not have detectable expression. These results suggest that circulating tumor cells are present in patients with advanced-stage HCC, which may be one of the reasons why these patients have a high incidence of tumor recurrence after apparently definitive surgical resection and even after liver transplantation.

Total hepatectomy plus liver transplantation was performed on 105 patients considered unsuitable for liver resection. Postoperative 5-year actuarial survivals correlated with the pathologic stage of the tumor: stage I 75%, stage II 68%, stage III 52.1%, and stage IVA 11%. The overall 5-year survival for all patients was 36%. Nodal disease, bilobar tumor, and macroscopic venous invasion were significant poor-prognosis features. In addition, 12 patients with pT4N1M0 lesions (also stage IVA) had hepatectomy plus more extensive en bloc regional resection (Whipple procedure or cluster resection) plus transplantation in an effort to prevent local recurrence. Only 2 of these 12 patients (16.7%) are alive and free of disease after 2 years. Seven patients (58%) have died from tumor recurrence usually originating from distant metastases an average of 10.6 months after transplantation. Successful transplantation for hepatoma depends on screening programs to identify early stage disease. Successful outcome of transplantation for late stage disease, which includes most of the patients in our series, awaits the development of neoadjuvant therapy to control distant microscopic metastases, which are almost certainly present though not apparent at the time of transplantation.

A new cell line (Hep 3B-TR), which is resistant to growth-inhibition by transforming growth factor beta 1 (TGF-beta 1) up to 10 ng/ml (400 pM), was isolated from parental Hep 3B human hepatoma cells, which are sensitive to growth-inhibition by TGF-beta 1. In the presence of TGF-beta 1 (1 to 10 ng/ml), the growth of the parental cell line (Hep 3B-TS) was inhibited by more than 95%. Under the same conditions, the growth rate of the resistant clone (Hep 3B-TR) however, was identical in the presence or absence of TGF-beta 1 and was almost the same as that of the Hep 3B-TS cells in the absence of TGF-beta 1. Affinity crosslinking with 5 pM 125I-labeled TGF-beta 1 showed that the TGF-beta 1 receptors type I (TGF-beta RI) and type II (TGF-beta RII) were not present on the cell surface of the Hep 3B-TR cells, whereas they were present on the sensitive HEP 3B-TS cells. Hep 3B-TS cells had detectable TGF-beta RII mRNA, which was not found in Hep 3B-TR cells. RNA analysis showed different effects on the expression of TGF-beta 1, c-fos, c-myc, and protein disulfide isomerase (PDI) genes in the two cell lines in response to TGF-beta 1 protein. Addition of TGF-beta 1 (1 ng/ml) strongly increased the expression of TGF-beta 1 mRNA in Hep 3B-TS cells, but not in Hep 3B-TR cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Differential gene expression between the normal human liver and a cell line derived from human hepatocellular carcinoma (HCC) was studied using the differential display polymerase chain reaction technique. One gene (mitochondria proteolipid like gene, MPL), whose expression was found to be repressed in the HCC cell line compared to normal liver, was cloned and sequenced. Amino acid sequence translated from the nucleotide sequence had a 73% homology with the carboxyl terminus of a mitochondria proteolipid (MPLP) isolated from beef heart. Northern blot analysis showed that the expression of the 3 kb MPL transcript was undetectable in 20 of 45 (44%) of human hepatocellular carcinomas, whereas only 1 of 14 (17%) of cirrhotic livers without HCC had undetectable expression when compared to normal livers. Hence MPL may be a candidate tumor suppressor gene for human HCC. This decrease in MPL expression was not due to gross alteration of its genomic DNA.

Human hepatoma cell lines (Hep 3B-TS, PLC/PRF/5, and Hep G2), sensitive to growth inhibition by transforming growth factor beta 1 (TGF-beta 1), express TGF-beta receptors type I, type II, and type III. We report that TGF-beta 1 protein selectively increased steady-state levels of the mRNA for the serine/threonine kinase receptor 1 (SKR1), a member of the TGF-beta superfamily receptor genes in these cells, whereas TGF-beta 1 had little effect on expression of the TGF-beta receptor type II gene. This increase of SKR1 mRNA in Hep 3B-TS cells could be detected by Northern blot analysis within 3 h of addition of TGF-beta 1 to the cells, and enhanced message levels peaked at 12 h as long as TGF-beta 1 was present in the culture medium. Hep 3B-TR cells which were resistant to TGF-beta 1 due to lack of both TGF-beta receptors type I and type II, expressed SKR1 mRNA, but it was not induced by TGF-beta 1 protein. The increased expression of SKR1 mRNA in the cells was actinomycin D-sensitive and was not dependent on new protein synthesis. The results indicate that TGF-beta 1 selectively induces SKR1 message at a transcriptional level by a positive regulator.