Schedule-dependent combined sensitivity testing of anti-cancer agents in human gastric carcinoma cell lines.

The efficacy of combination chemotherapy for gastric carcinoma has been unsatisfactory, although the prognosis of advanced and recurrent disease has improved with the introduction of cisplatin (CDDP). This study examines the effect of the anti-cancer therapies CDDP, doxorubicin (ADM) and etoposide (VP-16) on the cell cycle and their cytotoxicity against two gastric carcinoma cell lines: MKN-28 (well differentiated) and MKN-45 (poorly differentiated). The treatments have different cytocidal mechanisms, and they were studied in dual combinations. For all combinations studied, cytotoxicity against MKN-45 was higher than against MKN-28. For ADM plus CDDP, or ADM plus VP-16, cytotoxicity was higher in patients pretreated with ADM than other regimens. The highest anti-tumour activity against both cell lines was obtained with ADM followed by CDDP (we have obtained good clinical results with this regimen). Schedule-dependent combined sensitivity testing of anti-cancer agents will be useful for the clinical application of therapies.

Long-term interferon therapy for 1 year or longer reduces the hepatocellular carcinogenesis rate in patients with liver cirrhosis caused by hepatitis C virus: a pilot study.

BACKGROUND AND METHODS: In order to elucidate the influence of a long-term administration of interferon on the appearance rates of hepatocellular carcinoma (HCC) in hepatitis C virus (HCV)-related cirrhosis, we retrospectively analyzed 694 patients with cirrhosis. A total of 113 patients underwent interferon therapy, including 25 patients with a long-term administration of interferon for 1 year or more, and the other 581 patients received no antiviral drugs. RESULTS: Crude cumulative appearance rates of HCC in the interferon and the untreated groups were 14.1, and 28.4% at the end of the 5th year, and 36.7 and 52.5% at the end of the 10th year, respectively (P = 0.0028). As there was a waiting time between diagnosis and treatment (median 2.0 months, average 21.3 months) in the treated group, Cox proportional hazard analysis using a time-dependent covariate was introduced to evaluate the anticarcinogenic effect of interferon. Although male sex, higher alpha-fetoprotein, older age, lower albumin concentration, and lower platelet count significantly increased the carcinogenesis rate, interferon was not a significant contributing factor to the carcinogenesis rate as a whole (hazard ratio = 0.83, P= 0.32). When the patients with interferon were divided into two groups according to therapy duration, long-term interferon therapy significantly decreased the hepatocellular carcinogenesis rate after an adjustment by significant covariates (hazard ratio = 0.28, P= 0.0048). CONCLUSION: When interferon is administered for 12 months or longer, effective cancer prevention will be achieved, even in patients with HCV-related cirrhosis.

Interferon therapy for flare-up of hepatitis B virus infection after emergence of lamivudine-induced YMDD motif mutant.

A 61-year-old man with chronic hepatitis B was treated with interferon (IFN)-alpha for flare-up after the emergence of a lamivudine-induced YMDD motif mutant. The YMDD mutant emerged 13 months after the initiation of lamivudine therapy. Despite this, lamivudine therapy was continued. Acute exacerbation occurred 25 months after the emergence of the YMDD mutant. Treatment with IFN-alpha resulted in rapid loss of hepatitis, B virus DNA, resolution of hepatitis, and clinical recovery.

Lamivudine therapy for spontaneously occurring severe acute exacerbation in chronic hepatitis B virus infection: a preliminary study.

OBJECTIVE: The nucleoside analogue lamivudine, a potent inhibitor of hepatitis B virus replication, has shown notable results in treating chronic hepatitis B. However, lamivudine has not been specifically tested for effectiveness against spontaneously occurring severe acute exacerbations of hepatitis in patients chronically infected with this virus. We addressed this issue in a pilot study. METHODS: Ten patients with chronic hepatitis B developed severe acute exacerbation spontaneously during follow-up; 3 of them developed hepatic failure shortly before entering the trial. Lamivudine was administered long-term to the 10 patients at a daily oral dose of 100 or 300 mg. RESULTS: All 3 patients with hepatic failure at initiation of treatment recovered dramatically. Of the remaining 7 patients, 5 recovered rapidly with lamivudine, but 2 progressed quickly to hepatic failure despite treatment. One died of sepsis and the other of multiorgan failure. In the 8 survivors, serum alanine transaminase activity decreased rapidly to normal with lamivudine therapy, and serum hepatitis B virus DNA level declined rapidly to undetectable levels. Serum total bilirubin concentrations normalized somewhat later. Prothrombin time improved steadily and gradually. Hepatitis B e antigen elimination or seroconversion was achieved in 3 survivors. No adverse effects were noted in any patient. All survivors had good quality of life with long-term lamivudine monotherapy. CONCLUSIONS: Lamivudine is effective, safe, and well tolerated by patients with spontaneous, severe, acute exacerbation complicating chronic hepatitis B virus infection, even in the presence of hepatic failure. Lamivudine appears to be an attractive therapeutic option and may represent the best choice.

Efficacy of IFN therapy based on duration period of negative HCV-RNA during IFN administration.

We assessed the relationship between the duration period of negative hepatitis C virus (HCV)-RNA during interferon (IFN) therapy and the efficacy after prolonged IFN therapy in patients with HCV-genotype 1b and high virus load of more than 1 mega equivalents/ml (Meq/ml) retrospectively. A total of 100 patients who had HCV-genotype 1b and a high virus load of more than 1 Meq/ml and were treated with natural IFN-alpha for more than 12 months were enrolled in this trial. These patients were given 6 MU of IFN daily for 8 weeks, followed by three times weekly for another more than 44 weeks. The HCV-RNA pattern during IFN therapy according to negative or positive of the serum HCV-RNA by reverse transcription nested polymerase chain reaction (RT-nested PCR) from 2 months after the initiation of IFN to the termination of IFN were classified as follows: group 1: constant negative HCV-RNA (n=41 cases), group 2: constant positive HCV-RNA (n=35 cases), group 3: HCV-RNA pattern except for group 1 or group 2 (n=24 cases). A complete response (CR) was defined as negative HCV-RNA by RT-nested PCR at two points, 3 and 6 months after the completion of IFN therapy. CR rate was 58.5% (24 cases) in group 1, but CR rate in group 2 or group 3 was 0%. In group 1, the CR rate was 100% (10/10) in patients with negative HCV-RNA constantly for period of more than 24 months during IFN therapy. On the other hand, all patients who had positive HCV-RNA 2 months after the initiation of IFN did not get CR. In conclusion, it seems to us that the attainment of constantly negative HCV-RNA for the period of more than 24 months during IFN therapy is highly related to CR.

Distinction between chronic hepatitis and liver cirrhosis in patients with hepatitis C virus infection. Practical discriminant function using common laboratory data.

In order to distinguish patients with cirrhosis from those with chronic hepatitis, multivariate discriminant analysis was performed using common laboratory data. A total of 205 consecutive patients were diagnosed by peritoneoscopy and biopsy as having chronic liver disease caused by hepatitis C virus (HCV), 168 with chronic hepatitis and 37 with cirrhosis. Twenty variables and their natural logarithmic transformation were employed in the multivariate analysis. After stepwise variable selection, the following function was finally obtained to discriminate the disease severity, z=0.120xgamma-globulin (%)+0.423xln (hyaluronate) (µg l(-1))-0.059xplatelet (x10(4) counts per mm(3))-0.364xsex (male, 1; female, 2)-3.953. Since the function contained an expression of logarithm and was slightly troublesome to apply, we prepared another discriminant function composed of usual figures without logarithmic transformation, z=0.124x(gamma-globulin (%))+0.001x(hyaluronate) (µg l(-1))-0.075x(platelet (x10(4) counts per mm(3)))-0.413xgender (male, 1; female, 2)-2.005. When a positive result is calculated in the latter equation, the diagnosis of the liver disease indicates cirrhosis, and negative result chronic hepatitis. Accuracy of the two discriminant functions was 90.3 and 91.2%, respectively. A concise linear discriminant function could successfully differentiate liver cirrhosis from chronic hepatitis with an accuracy of 91.2%.

Progressive and sufficient decrease of hepatitis B core antibody can predict the disappearance of hepatitis B virus DNA in Japanese patients with hepatitis B surface antigen clearance.

The aim of this study was to elucidate the relationships among serum levels of hepatitis B virus (HBV) DNA, periods after hepatitis B surface (HBs) antigen clearance, and the titer of hepatitis B core (HBc) antibody in 200-fold diluted serum. Twelve patients who had clearance of HBs antigen from serum were studied. Five patients had not received any treatment (group A), and seven had received prednisolone withdrawal therapy. The patients in groups A and B were followed up for 86 months and 108 months (median), respectively. Serum HBV was measured by the nested polymerase chain reaction method. In both groups, serum HBV tended to become gradually undetectable after HBs antigen clearance. The positive rate of HBV in the sera 5 years or more after HBs antigen clearance was significantly lower than that in the sera at less than 5 years, both in group A (P = 0.004) and group B (P = 0.010). In both groups, the titer of HBs tended to decline every year after HBs antigen clearance. HBV was still detectable in the sera of some patients for a long period of time after they showed seroconversion to HBs antibody. The results suggest that detection of HBV was difficult in sera with an HBc titer of 30% or lower and at more than 5 years after HBs antigen clearance in both groups. It is important to note that HBV DNA rarely exists in the serum, even when HBs antigen and HBc are both negative.

Association of amino acid sequence in the PKR-eIF2 phosphorylation homology domain and response to interferon therapy.

Hepatitis C virus (HCV) genotype 1b and high pretreatment virus load are well known predictive factors of poor response to interferon (IFN) therapy. In addition, a sparsity of amino acid substitutions in the interferon sensitivity determining region (ISDR) is also predictive of a poor response to IFN in patients with genotype 1b, although this issue is still controversial. Recently, a 12 amino acid domain in the E2 protein of HCV (PKR-eIF2 alpha phosphorylation homology domain [PePHD]) has been reported to bind with and block the virus replication inhibition ability of PKR, suggesting that the interaction of E2 and PKR may be one mechanism by which HCV circumvents the antiviral effect of IFN. To clarify the significance of amino-acid sequences in this domain in predicting the effect of IFN therapy, we analyzed 82 patients with genotype 1b. Eleven patients (13.4%) responded to treatment whereas the remaining 71 patients (86.6%) were nonresponders. Multivariate analysis showed that only HCV load and amino-acid substitutions in the ISDR were predictive of sustained response to IFN. Amino-acid substitutions in the PePHD were detected in only eight of 82 patients (9.8%), and did not correlate with the therapeutic effect of IFN. However, amino-acid-sequence analyses of quasispecies before and after 1 week of IFN therapy showed elimination of clones with substitutions in this domain. Our results suggest that amino-acid sequences of the PePHD domain may be related to viral resistance to IFN but do not predict the outcome of IFN therapy as amino-acid substitutions in this domain are rare.

Increased response rate to interferon therapy after a second course in hepatitis C patients who show relapse after the initial course.

Some patients with chronic hepatitis C become HCV-RNA seronegative during interferon (IFN) therapy. However, about one-half of these patients show a relapse, evident by high serum alanine aminotransferase (ALT) level. In some patients with biochemical relapse, the serum HCV-RNA level becomes low immediately after the ALT relapse. Here, we assessed the changes in serum HCV-RNA level in patients with ALT relapse after IFN therapy, and evaluated the efficacy of a second course of IFN, started at the recovery stage after ALT relapse. Two hundred and seventy-seven patients who showed HCV-RNA seronegativity by reverse transcription nested-polymerase chain reaction (RT nested-PCR) and normalization of ALT during the initial IFN therapy, and had positive HCV-RNA with ALT relapse (> 100 IU/l) within 3 months after completion of the initial IFN course were enrolled in this retrospective study. Two hundred and sixty patients were followed-up without further IFN retreatment after the ALT relapse (group 1), and 17 patients received another 6-month course of IFN after the ALT relapse (group 2). The median level of serum HCV-RNA, determined with a branched DNA probe assay (version 1; Chiron-Dai-ichi Kagaku Tokyo, Japan), in group 1 was 3.1 Meq/ml before IFN therapy, 1.3 Meq/ml at the time point of the ALT peak after the completion of IFN therapy, and 0.7 and 2.6 Meq/ml at 2-4 and 6-8 weeks after the ALT peak, respectively. The serum HCV-RNA level at 2-4 weeks after the ALT peak was lower than that before IFN therapy. The eradication rate of HCV-RNA (complete response; CR) in group 2 (47.1%; 8/17) was significantly higher than that in group 1 (1.5%; 4/260; P < 0.001). In conclusion, our data suggested that: (1) patients who showed biochemical relapse after initial IFN therapy had a significantly lower serum HCV-RNA level at recovery after ALT relapse compared with that before initial IFN therapy. (2) A high response rate was noted after a second course of IFN administered at the recovery stage of the ALT relapse, compared with patients without IFN retreatment.

Interferon beta prevents recurrence of hepatocellular carcinoma after complete resection or ablation of the primary tumor-A prospective randomized study of hepatitis C virus-related liver cancer.

Because hepatocellular carcinoma often recurs after surgical resection or ethanol injection therapy, we conducted a prospective randomized controlled trial of interferon (IFN) in patients with chronic liver disease caused by hepatitis C virus (HCV). Twenty eligible patients with cirrhosis were randomized into two groups: 10 patients treated with 6 million units of natural IFN-beta twice a week for 36 months and 10 patients without IFN therapy. One patient within the treatment group discontinued interferon therapy after 19 months of treatment because of a mild degree of retinopathy. None of the patients in either group lost HCV-RNA until the end of the observation. Although 7 (70.0%) of 10 patients in the nontreatment group showed tumor recurrence, only 1 (10.0%) of 10 patients with IFN therapy developed tumor recurrence during a median observation period of 25.0 months. Cumulative recurrence rates of the treated and untreated groups were 0% and 62.5% at the end of the first year, and 0% and 100% at the second year, respectively (log-rank test, P =.0004). In conclusion, intermittent administration of IFN suppressed tumor recurrence after treatment with surgery or ethanol injection in patients with HCV-related chronic liver disease.

[A case of nonresectable scirrhous type gastric cancer successfully treated by low-dose cisplatin (CDDP), 5-fluorouracil (5-FU) and pirarubicin (THP)].

There have been few effective chemotherapeutic regimens for scirrhous type gastric cancer. Recently, the usefulness of combined cancer agent chemotherapy based on the concept of biochemical modulation has been reported. For example sequential MTX and 5-FU therapy, low-dose CDDP plus 5-FU, and the like. In this paper, we report the usefulness of low-dose CDDP plus 5-FU therapy in combination with pirarubicin (THP) for inoperable scirrhous type gastric cancer. A 32-year-old man who was suffering from scirrhous type gastric cancer with pyloric stenosis was treated with this regimen. Eight weeks after the start of therapy, his gastric capacity and lumen diameter had clearly increased, and he was taking ordinary meals. Ascites had also completely disappeared. CR has now been continued about 7 months. This regimen is considered to be promising for scirrhous type gastric cancers with a poor prognosis.

Hepatic vascular side effects of styrene maleic acid neocarzinostatin in the treatment of hepatocellular carcinoma.

Styrene-maleic acid neocarzinostatin (SMANCS) sometimes causes hepatic vascular side effects, including arterial stricture, obstruction, and arterio-portal shunt. A total of 128 intra-arterial SMANCS injection treatments, performed for 89 patients with hepatocellular carcinoma, were analyzed to determine the relationship between angiographic findings and subsequent hepatic vascular injuries. After SMANCS therapy, hepatic arterial stricture or obstruction occurred in 5 patients (5/128; 3.9%), arterio-portal shunting in 12 (12/128; 9.4%), liver shrinkage in 4 (4/128; 3.1%), and cholangitis or biloma in 2 (2/128; 1.6%). Among 23 patients whose plain abdominal X-ray films just after SMANCS injection showed Lipiodol retention in the hepatic artery, 5 patients developed arterial obstruction, 10 developed arterio-portal shunt, and 2, cholangitis or biloma. Among 26 patients with Lipiodol retention in the portal vein, 4 developed hepatic lobe atrophy with aggravation of liver function. Among 3 patients with Lipiodol retention in both the hepatic artery and the portal vein, 1 developed arterio-portal shunt. In 76 treatments without excessive Lipiodol retention, only 1 of the patients developed arterio-portal shunt. Excessive retention of Lipiodol in hepatic vascular beds just after SMANCS therapy was significantly associated with future vascular side effects (22/52 vs 1/76; P < 0.0001). Lipiodol retention in arteries just after SMANCS injection was closely associated with subsequent arterial obstruction or arterio-portal shunt, and Lipiodol retention in the portal vein was related to subsequent hepatic lobe atrophy.

Incidence of primary cholangiocellular carcinoma of the liver in japanese patients with hepatitis C virus-related cirrhosis.

BACKGROUND: Hepatitis C virus (HCV) infection is a major risk factor for the development of hepatocellular carcinoma. However, the risk factors for primary cholangiocellular carcinoma of the liver (PCC-L) have not been fully investigated. The authors determined the incidence of PCC-L in patients with HCV-related cirrhosis. METHODS: Between 1980 and 1997, the authors prospectively studied 600 consecutive patients for the appearance of PCC-L; these patients were positive for HCV and later developed cirrhosis. The follow-up period ranged from 0 to 18.5 years (median, 7.2 years). RESULTS: During the observation period, PCC-L developed in 14 patients (2.3%). Among these, 11 (1.8%) had cholangiocellular carcinomas and the other 3 (0.5%) had a combined type of hepatocellular and cholangiocellular carcinoma. Within the same period, hepatocellular carcinoma (HCC) developed in 206 patients (34.3%). The cumulative rates of newly diagnosed PCC-L were 1.6% at 5 years and 3.5% at 10 years, which was about 1000 times higher than the estimated incidence of PCC-L in the general population of Japan. PCC-L was treated by surgical resection in 3 patients who survived for > 3 years. However, the other 11 patients received palliative therapy or chemotherapy. The survival rates among PCC-L patients were 39.3%, 23. 6%, and 16.5% at the end of 1, 3, and 5 years, respectively, and were significantly lower than those of HCC (P = 0.0001). CONCLUSIONS: The results of this study show a relatively high incidence of PCC-L in patients with HCV-related cirrhosis, and also show that this type of liver cancer is associated with a relatively poor prognosis. These results indicate that HCV-related cirrhosis is a major risk factor for PCC-L in Japanese patients.

Fluctuation patterns of HCV-RNA serum level in patients with chronic hepatitis C.

The serum level of hepatitis C virus (HCV)-RNA is clinically important as a predictor of the response to interferon (IFN) therapy in patients with chronic hepatitis C. If serum HCV-RNA levels fluctuate during follow-up, and IFN therapy is begun at the time of a low HCV-RNA level, the IFN therapy may be more effective. We evaluated the fluctuation of HCV-RNA serum levels for 2 years in 212 patients with chronic hepatitis C, untreated with IFN who had HCV genotype 1b and an HCV-RNA level of 10 Meq/ml or more at first consultation. The HCV-RNA level was measured monthly for 2 years with an HCV branched DNA probe assay (b DNA probe assay). We classified HCV-RNA patterns into three types by the ratio of maximum HCV-RNA level (a) to minimum HCV-RNA level (b). In pattern 1 (constant type, 151 patients; 71.2%) the a/ b ratio was 1-5. In pattern 2 (slight fluctuation type, 46 patients; 21.7%) the a/b ratio was 5-10. In pattern 3 (severe fluctuation type, 15 patients; 7.1%), the a/b ratio was 10 or more. Next, we evaluated the factors associated with the three patterns. Acute exacerbation of chronic hepatitis was regarded as an increase in serum alanine aminotransferase (ALT) level to more than 250 IU/l. The incidence of acute exacerbation for a 2-year follow-up was 13.9% (21/151) in pattern 1, 19.6% (9/46) in pattern 2, and 53.3% (8/15) in pattern 3. Multivariate analysis showed that acute exacerbation was the most important factor in the manifestation pattern 3. In conclusion, we found that: (1) about 70% of patients had a constant HCV-RNA levels for 2 years. (2) A few patients had severe fluctuation of serum HCV-RNA level after acute exacerbation of chronic hepatitis.

Chronic liver disease in the extremely elderly of 80 years or more: clinical characteristics, prognosis and patient survival analysis.

BACKGROUND/AIMS: This study aimed to elucidate the clinical characteristics of patients with chronic liver disease aged 80 years or more, especially the factors affecting prognosis and carcinogenesis. METHODS: A total of 135 patients aged 80 years or above were divided into chronic liver disease without cirrhosis (non-LC) and cirrhosis (LC) groups according to the severity of fibrosis, and the clinical characteristics and prognoses were evaluated. RESULTS: Seventy-three (54.1%) of 135 patients were in the LC group and 79 patients (58.5%) had hepatitis C virus. Various concomitant diseases were seen in 122 patients (90.4%). Liver-related deaths occurred in only 19 (36.5%) of 52 patients who died during observation, although 28 patients (53.8%) had liver cancer at the time of death. Cumulative survival rates in the non-LC and the LC groups were 85.7% and 58.8% at the 5th year, and 69.4% and 19.4% at the 9th year, respectively. Cumulative liver cancer appearance rates in the non-LC and the LC groups were 1.6% and 6.1% at the 1st year, 12.4% and 19.9% at the 5th year, and 12.4% and 32.0% at the 7th year, respectively. A multivariate Cox regression analysis revealed that the presence of liver cancer (p=0.0001), platelet count (p=0.0242), and fibrotic stage (p=0.0118) were independently associated with survival period, and alfa-fetoprotein (p=0.0194) and bilirubin (p=0.0282) were independently associated with carcinogenesis. CONCLUSIONS: Cirrhosis is the major risk factor affecting the prognosis. On the other hand, we must pay more attention to concomitant diseases specific to advanced age.

Time course of histological changes in patients with a sustained biochemical and virological response to corticosteroid withdrawal therapy for chronic hepatitis B.

OBJECTIVE: Although biochemical and virological responses to corticosteroid withdrawal therapy for chronic hepatitis B have been extensively studied, long term changes in liver histology have not been well documented. METHODS: We retrospectively analyzed 45 paired liver biopsy specimens taken before and after treatment from 40 patients who persistently showed biochemical remission and an absence of HBe antigen (RIA) for up to 20 yr. RESULTS: The grading scores for necroinflammatory and fibrotic activity in the liver specimens decreased significantly after corticosteroid withdrawal therapy. Histological scores graded according to Knodell's components improved significantly in every category after corticosteroid withdrawal therapy. However, inflammatory cell infiltrates remained within the liver for long periods. The disappearance rate of necroinflammation in the periportal, lobular, and portal regions of the liver were 25%, 7.4%, and 7.4%, respectively, at yr 5 after therapy, and were 84.4%, 78.2%, and 58.7%, respectively, at yr 10 after therapy. The cumulative disappearance rate, calculated using the Kaplan-Meier method, was significantly lower for portal inflammation than for periportal necroinflammation. CONCLUSIONS: Our results show that: 1) despite clinical remission of chronic hepatitis B virus infection, long periods are needed for histological resolution of necroinflammation in the liver; and 2) by a Cox proportional hazard analysis of the factors contributing to histopathological disappearance of disease-related inflammation, the degree of fibrosis of liver biopsy specimens from pretherapy patients was the most statistically significant factor (p = 0.049).

Combined ursodeoxycholic acid and glycyrrhizin therapy for chronic hepatitis C virus infection: a randomized controlled trial in 170 patients.

OBJECTIVE AND DESIGN: To assess the efficacy and safety of combination therapy using ursodeoxycholic acid with glycyrrhizin for chronic hepatitis C virus infection, we conducted a prospective randomized controlled trial of glycyrrhizin (group G) compared with glycyrrhizin plus ursodeoxycholic acid (group G+U) in 170 patients. METHODS: All patients had elevated serum aminotransferase levels over 6 months before entry into the trial. Glycyrrhizin was administered to both groups for 24 weeks, and in group G+U, ursodeoxycholic acid (600 mg/day) was administered orally as well. RESULTS: Serum aspartate transaminase and alanine transaminase concentrations significantly decreased during treatment in both groups, but serum gamma-glutamyl transpeptidase concentrations fell significantly only in group G+U. Concentrations of all three enzymes fell significantly more in group G+U than in group G, and had normalized in more cases when the trial ended at 24 weeks. However, levels of HCV viraemia did not change during the trial in either group. Multiple regression analysis linked only the treatment regimen, not HCV-related factors or liver histology, to the degree of serum enzyme reduction. No adverse effects were noted in either group. CONCLUSIONS: The combined therapy with ursodeoxycholic acid and glycyrrhizin is safe and effective in improving liver-specific enzyme abnormalities, and may be an alternative to interferon in chronic hepatitis C virus infection, especially for interferon-resistant or unstable patients.