Efficacy of Lusutrombopag for Thrombocytopenia in Patients with Chronic Liver Disease Scheduled to Undergo Invasive Procedures.

Objective Lusutrombopag is a thrombopoietin receptor agonist that improves thrombocytopenia in patients with chronic liver disease scheduled to undergo invasive procedures. However, information on the efficacy of repeated lusutrombopag treatment and factors associated with the treatment is scarce. We analyzed the efficacy of repeated lusutrombopag treatment and the factors associated with a response to lusutrombopag. Methods Thirty-nine patients with chronic liver disease who received lusutrombopag treatment before undergoing invasive procedures were enrolled in this retrospective study. Of the 39 patients, 10 received lusutrombopag treatment multiple times for a total of 53 regimens of lusutrombopag treatment. Changes in platelet counts, the effects of repeated lusutrombopag treatment, and factors associated with response to lusutrombopag were analyzed. Results The median platelet count increased significantly from 4.5×104/µL before lusutrombopag treatment to 7.2×104/µL before the invasive procedure (p<0.01), and patients undergoing 49 of the 53 (92%) treatment regimens succeeded in undergoing invasive procedures without needing platelet transfusions. In patients who received lusutrombopag treatment repeatedly, the median platelet count significantly increased following the second administration of lusutrombopag, and the effects of lusutrombopag were similar between the first and second administration. A multivariate analysis identified the absence of diabetes mellitus (odds ratio, 5.56 for presence; p=0.04) as a significant and independent predictor of a response to lusutrombopag. Conclusion Lusutrombopag treatment significantly increased platelet counts in patients with chronic liver disease, making it possible to receive invasive procedures. The treatment produced identical effects when it was repeated. The efficacy of lusutrombopag might be decreased in patients with diabetes mellitus.

Real-world efficacy of sofosbuvir plus velpatasvir therapy for patients with hepatitis C virus-related decompensated cirrhosis.

AIM: Combination therapy with sofosbuvir (SOF) plus velpatasvir (VEL) is approved for patients with hepatitis C virus (HCV)-related decompensated cirrhosis. We analyzed the real-world efficacy of SOF/VEL therapy. METHODS: Thirty-three patients with HCV-related decompensated cirrhosis (25 and eight patients with Child B and C, respectively) were treated with SOF/VEL for 12 weeks. The HCV non-structural protein (NS)5A and NS5B drug resistance-associated substitutions (RASs) were determined by direct sequencing. RESULT: Thirty-two of 33 patients completed the treatment, but the remaining patient discontinued the therapy during third week of the treatment due to aggravation of hepatic encephalopathy. Serum HCV-RNA became negative during the treatment in all patients but relapsed after the end of therapy in five patients. In total, 28 out of 33 patients (85%) achieved sustained virological response 12 weeks following completion of treatment (SVR12). The SVR12 rate was 96% in patients with Child B, but significantly lower, at 50%, in patients with Child C (P < 0.05). In genotype 1b HCV-infected patients, all eight patients without baseline NS5A RASs, but only three of seven patients with RASs, achieved SVR12. Multivariate analysis identified Child B (odds ratio, 35.8 for Child C; P = 0.045) as an independent predictor of SVR12. Median serum albumin levels significantly increased only in patients who achieved SVR12. Child-Pugh scores improved in 16 of 28 patients (57%) following achievement of SVR12. CONCLUSION: The effect of SOF/VEL therapy is lower for patients with Child C. Improvement of hepatic function is expected after viral eradication with SOF/VEL therapy in patients with decompensated cirrhosis.

Cytokeratin 7-positive/cytokeratin 20-negative cecal adenocarcinoma metastatic to the uterine cervix: a case report.

BACKGROUND: The vast majority of uterine cervical malignancies are primary carcinomas, and secondary neoplasms that metastasize to the uterine cervix from a distant organ are uncommon. Although relatively rare, metastases to the uterine cervix from a primary colon cancer have been reported. We report a rare case of metastatic carcinoma originating from a cecal adenocarcinoma with an unusual cytokeratin 7/cytokeratin 20 immunophenotype. CASE PRESENTATION: A 74-year-old postmenopausal Japanese woman was referred to our hospital for the evaluation of a uterine tumor. She had a past medical history of cecal cancer and had undergone laparoscopically assisted right hemicolectomy at the age of 69 years. During follow-up, she was found to have elevated levels of the tumor markers carbohydrate antigen 19-9 (179.7 IU/mL) and carcinoembryonic antigen (26.9 µg/L). Positron emission tomography/computed tomography showed a focus of high 18F-fluorodeoxyglucose uptake in her uterus. Examination of a cervical biopsy found a poorly differentiated adenocarcinoma that was immunopositive for cytokeratin (CK)7 and caudal-related homeobox 2 (CDX2) expression and immunonegative for cytokeratin 20 expression. The patient underwent radical hysterectomy and bilateral salpingo-oophorectomy. Histopathological examination found invasive growth of irregular and atypical ductal hyperplasia. Immunohistochemical staining of the tumor specimen revealed the same immunophenotype as the biopsy specimen. The cecal cancer specimen from her previous surgery was also examined and found to have the same immunophenotype. The histopathological diagnosis was cecal adenocarcinoma metastatic to the uterine cervix. The patient is currently receiving adjuvant chemotherapy and to date is without evidence of recurrent disease. CONCLUSIONS: Our report illustrates the importance of immunohistochemistry for the correct diagnosis of the origin of a uterine cervical adenocarcinoma in a patient with a medical history of colorectal cancer. Re-examination of a previous oncological specimen is critical for cases with a uterine lesion that is difficult to identify as primary or metastatic cancer.

Clinical outcome of esophageal varices after hepatic arterial infusion chemotherapy for advanced hepatocellular carcinoma with major portal vein tumor thrombus.

AIM: To analyze the clinical outcome of esophageal varices (EV) after hepatic arterial infusion chemotherapy (HAIC) in patients with advanced hepatocellular carcinoma (HCC) and major portal vein tumor thrombus (Vp3/4). METHODS: The study subjects were 45 consecutive patients who received HAIC for HCC with Vp3/4 between January 2005 and December 2009. HAIC comprised the combination therapy of intra-arterial 5-FU with interferon-α (5-FU/IFN) in 23 patients and low-dose cisplatin plus 5-FU (FP) in 22. Radiotherapy (RT) was also provided in 19 patients for portal vein tumor thrombosis. Aggravation rate for EV and overall survival rate were analyzed. RESULTS: The aggravation rates for EV were 47% and 64% at 12 and 24 months, respectively. The survival rates were 47% and 33% at 12 and 24 months, respectively. The response rates to 5-FU/IFN and FP were 35% and 41%, while the disease control rates in these two groups were 57% and 50%, respectively. There were no significant differences in the objective response and disease control between 5-FU/IFN and FP. Multivariate analysis identified size of EV (F2/F3) (HR = 7.554, P = 0.006) and HCC disease control (HR = 5.948, P = 0.015) as significant and independent determinants of aggravation of EV, and HCC disease control (HR = 12.233, P < 0.001), metastasis from HCC (HR = 11.469, P = 0.001), ascites (HR = 8.825, P = 0.003) and low serum albumin (HR = 4.953, P = 0.026) as determinants of overall survival. RT for portal vein tumor thrombosis tended to reduce the aggravation rate for EV in patients with these risk factors. CONCLUSIONS: Hepatocellular carcinoma disease control was the most significant and independent factor for aggravation of EV and overall survival in HCC patients with major portal vein tumor thrombosis treated with HAIC.

[Problems of adjuvant chemotherapy with S-1 for gastric cancer].

From results of ACTS-GC,postoperative adjuvant chemotherapy,administration of S-1 for one year has become the standard for gastric cancer of Stage II and III except T1. We inspected problems of adjuvant chemotherapy by S-1 by dose rate, an adverse event,and compliance. For the period from July 2006 to December 2008,among 41 cases of stage II/stage III gastric cancer, S-1 was as started as adjuvant therapy by for 28 cases (68.3%). Among 14 cases (63.6%) considered able to complete S-1 treatment for one year, 7 cases (31.8%) had to have their dose reduced or their administration schedule changed. No adverse event of grade 3/4 was found, but cancellation or reduced dose was necessary due to anorexia, malaise, diarrhea, severe skin reaction, and leukopenia resulting from myelosuppression. Thirteen patients took no S-1, and two (4.9%) of them took UFT, while 11 cases (26.8%) became a no-treatment follow-up group for reasons of age, coexisting symptoms and other reasons. The problem in the future is to improve compliance, and to establish a treatment strategy for patients who do not meet administration criteria and for patients for whom continuation of drug administration is impossible.

Evaluation of portosystemic collaterals by MDCT-MPR imaging for management of hemorrhagic esophageal varices.

OBJECTIVE: To study the correlation between changes in portosystemic collaterals, evaluated by multidetector-row computed tomography imaging using multiplanar reconstruction (MDCT-MPR), and prognosis in patients with hemorrhagic esophageal varices (EV) after endoscopic treatment. METHODS: Forty-nine patients with primary hemostasis for variceal bleeding received radical endoscopic treatment: endoscopic injection sclerotherapy (EIS) or endoscopic variceal ligation (EVL). Patients were classified according to the rate of reduction in feeding vessel diameter on MDCT-MPR images, into the narrowing (n=24) and no-change (n=25) groups. We evaluated changes in portosystemic collaterals by MDCT-MPR before and after treatment, and determined rebleeding and survival rates. RESULTS: The left gastric and paraesophageal (PEV) veins were recognized as portosystemic collaterals in 100 and 80%, respectively, of patients with EV on MDCT-MPR images. The rebleeding rates at 1, 2, 3, and 5 years after endoscopic treatment were 10, 15, 23, and 23%, respectively, for the narrowing group, and 17, 24, 35, and 67%, respectively, for the no-change group (P=0.068). Among no-change group, the rebleeding rate in patients with large PEV was significantly lower than that with small PEV (P=0.027). The rebleeding rate in patients with small PEV of the no-change group was significantly higher than that in the narrowing group (P=0.018). There was no significant difference in rebleeding rates between the no-change group with a large PEV and narrowing group (P=0.435). CONCLUSION: Changes in portosystemic collaterals evaluated by MDCT-MPR imaging correlate with rebleeding rate. Evaluation of portosystemic collaterals in this manner would provide useful information for the management of hemorrhagic EV.

Evaluation of patients with esophageal varices after endoscopic injection sclerotherapy using multiplanar reconstruction MDCT images.

OBJECTIVE: The purpose of our study was to assess the relationship between hemodynamic changes in portosystemic collaterals and the prognosis of patients with esophageal varices after endoscopic injection sclerotherapy using multiplanar reconstruction (MPR) MDCT images. SUBJECTS AND METHODS: The subjects of this prospective study were 53 patients who underwent endoscopic injection sclerotherapy for esophageal varices. We evaluated the reconstructed MPR images of portosystemic collaterals before and after endoscopic injection sclerotherapy. Patients were divided into three groups based on the rate of change in the diameter of the feeding vessel into complete eradication (group A), narrowing (group B), and no change (group C). We analyzed the relationship between hemodynamic change in portosystemic collaterals and prognosis. RESULTS: The left gastric vein, posterior gastric vein, and left gastric vein plus posterior gastric vein were the main feeding vessels (n=44 [83%] of patients, n=5 [9%], and n=4 [8%], respectively). The proportions of patients of groups A, B, and C were 19% (n=10), 24% (n=13), and 57% (n=30), respectively. The relapse-free rates at 2 years after endoscopic injection sclerotherapy were 100%, 65%, and 52% in groups A, B, and C, respectively (p<0.05). For group C, the relapse-free rate at 2 years after endoscopic injection sclerotherapy of patients with a large-diameter paraesophageal vein (>or= 3 mm, 63%) was significantly higher than in those with a small-diameter paraesophageal vein (<3 mm, 36%; p<0.05). However, there were no significant differences in the survival rate among the three groups. CONCLUSION: MPR MDCT images on portosystemic collaterals can accurately predict relapse of esophageal varices after endoscopic injection sclerotherapy.

Etiology and outcome of acute liver failure: retrospective analysis of 50 patients treated at a single center.

BACKGROUND AND AIM: Acute liver failure (ALF) remains a devastating disease carrying considerable mortality. Since deceased donor liver transplantation is rarely performed in Japan, the artificial liver support system (ALS) and living donor liver transplantation (LDLT) are the main modalities used for treatment of ALF. The aim of this study was to analyze the outcome of ALF patients and to evaluate therapies for ALF according to etiology. METHODS: Fifty consecutive patients with ALF were treated between January 1990 and December 2006. Prior to 1997, patients received ALS only. After 1997, ALS and/or LDLT were applied. LDLT was performed in 10 patients. RESULTS: Four of 15 (27%) pre-1997 ALF patients survived, and 16 of 35 (46%) post-1997 ALF patients survived, including eight who underwent LDLT. The causes of ALF were acute hepatitis B virus (HBV) infection in 18%, severe acute exacerbation (SAE) of chronic HBV infection in 18%, autoimmune hepatitis (AIH) in 8%, and cryptogenic hepatitis in 44%. In total, 67% of the patients with ALF caused by acute HBV infection and AIH were cured without LDLT; only 11% of patients with ALF caused by SAE of HBV and 24% of cryptogenic hepatitis were successfully treated without LDLT. Notably, 80% of patients with cryptogenic hepatitis who underwent LDLT survived. CONCLUSION: Since 1997, the survival rate of ALF patients has increased, mainly due to the introduction of LDLT. Liver transplantation should be performed especially in patients with ALF caused by SAE of HBV and cryptogenic hepatitis.

Prospective study of short-term peginterferon-alpha-2a monotherapy in patients who had a virological response at 2 weeks after initiation of interferon therapy.

BACKGROUND AND AIMS: Long-term interferon (IFN) therapy is effective in eliminating hepatitis C virus (HCV). However, it carries the risk of adverse effects and reduced quality of life. To assess whether short-term IFN therapy effectively eliminates HCV, we performed a prospective pilot study of pegylated (peg)IFN-alpha-2a therapy for 8 or 24 weeks. METHODS: After excluding patients with high titers of genotype-1, 55 HCV patients received pegIFN-alpha-2a. Patients who became negative for HCV-RNA at week 2 were allocated to either an 8-week (n = 19) or 24-week (n = 15) course of IFN. We evaluated the efficacy of and tolerance to IFN therapy. RESULTS: The sustained virological response rate was excellent in the two groups (8 weeks, 89.5% [17/19]; 24 weeks, 100% [15/15], respectively,). IFN dose reduction was required in one patient of the 8-week group, but in six patients of the 24-week group (P = 0.028). Treatment was completed by all patients of the 8-week group, but discontinued in five patients of the 24-week group (P = 0.011). CONCLUSIONS: The 8-week IFN therapy is more tolerable than the 24-week therapy and had similar outcomes. Excluding the patients with high titers of genotype-1, we recommend switching to an 8-week course of pegIFN-alpha monotherapy once patients show an ultra rapid virological response at week 2 from the start of IFN therapy.

[Side effects of FEC100].

PURPOSE: The purpose was to investigate side effects of FEC100 treatment, particularly bone marrow suppression, nausea and vomiting. PATIENTS AND METHODS: We investigated side effects of FEC100 at the time of initial administration in 49 breast cancer patients (including 3 with recurrence). We used a 5-HT3 receptor antagonist as an antiemetic agent, steroids and Haloperidol, an antidopamine agent. Antibiotics were also administered for five days from day 10. RESULTS: The mean nadir of white blood cell count was 1,492+/-575 microL, and the mean day on which the nadir was reached was the 13.0+/-1.45th day. Leukopenia of less than 1,000 microL (grade 4) was seen in 8 (16.3%) of the 49 patients, and the incidence of leukopenia was particularly high (3/6, 50%) in patients with multiple bone metastases. Febrile neutropenia occurred in 4 (8.1%) of the 49 patients, 2 of those 4 patients having multiple bone metastasis. Fifteen (30.6%) of the 49 patients had nausea, and 3 patients (6.1%) had vomiting. CONCLUSION: The results indicate that side effects of FEC100 treatment are tolerable. However, care is needed for patients with multiple bone metastases because of the strong bone marrow suppression in such patients.

Similar effects of recombinant interferon-alpha-2b and natural interferon-alpha when combined with intra-arterial 5-fluorouracil for the treatment of advanced hepatocellular carcinoma.

AIM: Intra-arterial 5-fluorouracil (5-FU) plus interferon (IFN) combination therapy is effective against advanced hepatocellular carcinoma (HCC) with portal vein tumour thrombosis. In this study, we compared the efficiency and safety of recombinant IFN-alpha-2b with natural IFN-alpha as components of the combination therapy. METHODS: Consecutive HCC patients (n=31) with portal vein tumour thrombosis were enrolled in this prospective study. They received combination therapy of 5-FU and either recombinant IFN-alpha-2b (R group, n=15) or natural IFN-alpha (N group, n=16). We compared the two groups for the early response rate, adverse reactions, time to progression (TTP) and survival rates. In addition, we assessed the cost-effectiveness of each protocol. RESULTS: The early response rate (R: 26.7%, N: 31.2%), median TTP (R: 5.8 months, N: 5.6 months) and median survival time (R: 7.5 months, N: 6.5 months) were not significantly different between the R and N groups. There were no differences in adverse reactions between the two groups. The estimated cost-effectiveness ratio of recombinant IFN-alpha-2b was better than natural IFN-alpha. CONCLUSIONS: In our protocol of combination therapy, there were no significant differences between recombinant IFN-alpha-2b and natural IFN-alpha with regard to early response to therapy, adverse effects, TTP and survival rates. 5-FU could be combined with either recombinant IFN-alpha-2b or natural IFN-alpha, although the cost-effectiveness of the former warrants its use clinically.

Low-dose intermittent interferon-alpha therapy for HCV-related liver cirrhosis after curative treatment of hepatocellular carcinoma.

AIM: To assess the efficacy of low-dose intermittent interferon (IFN) therapy in patients with hepatitis C virus (HCV)-related compensated cirrhosis who had received curative treatment for primary hepatocellular carcinoma (HCC). METHODS: We performed a prospective case controlled study. Sixteen patients received 3 MIU of natural IFN-alpha intramuscularly 3 times weekly for at least 48 wk (IFN group). They were compared with 16 matched historical controls (non-IFN group). RESULTS: The cumulative rate of first recurrence of HCC was not significantly different between the IFN group and the non-IFN group (0% vs 6.7% and 68.6% vs 80% at 1- and 3-year, P = 0.157, respectively). The cumulative rate of second recurrence was not also significantly different between the IFN group and the non-IFN group (0% vs 6.7% and 35.9% vs 67% at 1- and 3-year, P = 0.056, respectively). Although the difference in the Child-Pugh classification score between the groups at initial treatment of HCC was not significant, the score was significantly worse at the time of data analysis in the non-IFN group than IFN group (7.19 +/- 1.42 vs 5.81 +/- 0.75, P = 0.0008). The cumulative rate of deviation from objects of any treatment for recurrent HCC was also higher in the non-IFN group than IFN group (6.7% and 27% vs 0 and 0% at 1- and 3-year, P = 0.048, respectively). CONCLUSION: Low-dose intermittent IFN-alpha therapy for patients with HCV-related compensated cirrhosis after curative HCC treatment was effective by making patients tolerant to medical or surgical treatment for recurrent HCC in the later period of observation.

Effects of a 24-week course of interferon-alpha therapy after curative treatment of hepatitis C virus-associated hepatocellular carcinoma.

AIM: To assess whether a 24-wk course of interferon (IFN) could prevent hepatocellular carcinoma (HCC) recurrence and worsening of liver function in patients with hepatitis C virus (HCV)-infected patients after receiving curative treatment for primary HCC. METHODS: Outcomes in 42 patients with HCV infection treated with IFN-alpha, after curative treatment for primary HCC (IFN group), were compared with 42 matched curatively treated historical controls not given IFN (non-IFN group). RESULTS: Although the rate of initial recurrence did not differ significantly between IFN group and non-IFN group (0%, 44%, 61%, and 67% vs 4.8%, 53%, 81%, and 87% at 1, 3, 5, and 7 years, P = 0.153, respectively), IFN group showed a lower rate than the non-IFN group for second recurrence (0%, 10.4%, 28%, and 35% vs 0%, 30%, 59%, and 66% at 1, 3, 5 and 7 years, P = 0.022, respectively). Among the IFN group, patients with sustained virologic response (SVR) were less likely to have a second HCC recurrence than IFN patients without an SVR, or non-IFN patients. Multivariate analysis identified the lack of SVR as the only independent risk factor for a second recurrence, while SVR and Child-Pugh class A independently favored overall survival. CONCLUSION: Most intrahepatic recurrences of HCV-related HCC occurred during persistent viral infection. Eradication of HCV is essential for the prevention of HCC recurrence and improvement of survival.

The long-term outcome of patients with bleeding gastric varices after balloon-occluded retrograde transvenous obliteration.

BACKGROUND: The purpose of our study was to evaluate the long-term outcome and complications of balloon-occluded retrograde transvenous obliteration (B-RTO) in patients with hemorrhage from gastric fundal varices. METHODS: Thirty-four consecutive patients with bleeding from gastric varices who were treated with B-RTO were enrolled in this study between December 1994 and September 2005 (urgent cases, n = 12; elective cases, n = 22). The long-term outcome, complications, and various liver functions were evaluated. RESULTS: Complete obliteration was achieved in 31 of 34 (91%) patients with an acute bleeding episode. In one of the remaining patients, there was a technical failure, and the other two had only partial obliteration. The two patients with partial obliteration did not obtain hemostasis. Thus, the rate of hemostasis was 94% (31/33). Gastric varices disappeared in all patients with complete obliteration during the treatment. The rate of gastric variceal eradication was 91%. Variceal rebleeding from esophageal varices occurred in three patients. The rate of rebleeding was 10% (3/31). Rebleeding from gastric varices was not observed after complete obliteration. None of the patients showed worsening of their Child-Pugh score. Although the 5-year cumulative worsening rate of esophageal varices was 52%, neither portal hypertensive gastropathy nor ectopic varices were observed. The patients with worsening esophageal varices were successfully treated with an endoscopic procedure. The 5-year survival rate was 68%. CONCLUSIONS: B-RTO is useful for treatment of bleeding gastric varices, achieving high eradication of gastric varices, a low rebleeding rate, and a fairly good prognosis with improved hepatic function.

Clinical features and prognosis of patients with extrahepatic metastases from hepatocellular carcinoma.

AIM: To assess the clinical features and prognosis of 151 patients with extrahepatic metastases from primary hepatocellular carcinoma (HCC), and describe the treatment strategy for such patients. METHODS: After the diagnosis of HCC, all 995 consecutive HCC patients were followed up at regular intervals and 151 (15.2%) patients were found to have extrahepatic metastases at the initial diagnosis of primary HCC or developed such tumors during the follow-up period. We assessed their clinical features, prognosis, and treatment strategies. RESULTS: The most frequent site of extrahepatic metastases was the lungs (47%), followed by lymph nodes (45%), bones (37%), and adrenal glands (12%). The cumulative survival rates after the initial diagnosis of extrahepatic metastases at 6, 12, 24, and 36 mo were 44.1%, 21.7%, 14.2%, 7.1%, respectively. The median survival time was 4.9 mo (range, 0-37 mo). Fourteen patients (11%) died of extrahepatic HCC, others died of primary HCC or liver failure. CONCLUSION: The prognosis of HCC patients with extrahepatic metastases is poor. With regard to the cause of death, many patients would die of intrahepatic HCC and few of extrahepatic metastases. Although most of HCC patients with extrahepatic metastases should undergo treatment for the primary HCC mainly, treatment of extrahepatic metastases in selected HCC patients who have good hepatic reserve, intrahepatic tumor stage (T0-T2), and are free of portal venous invasion may improve survival.

Efficacy of percutaneous cementoplasty for bone metastasis from hepatocellular carcinoma.

OBJECTIVE: To assess the efficacy of percutaneous cementoplasty for painful bone metastasis from hepatocellular carcinoma (HCC). METHODS: Cementoplasty was performed for 22 metastatic bone tumors in 13 patients. All patients had intractable pain and were diagnosed as metastatic bone tumor from HCC. The ostyneedle was inserted in the center of the site of bone metastasis under CT guidance. VAS score, Tokuhashi score and Frankel score were used for assessment of the efficacy of cementoplasty. We also assessed the response to treatment, adverse events and prognosis. RESULTS: Only the VAS score, but not Tokuhashi and Frankel scores, improved after cementoplasty. Cementoplasty for painful bone metastasis provided relief of severe pain but did not improve prognosis, neurological function or survival. Eleven of 13 (85%) patients showed CR or PR and the mean pain-free period was 5 months, including a 10-month pain-free period in one case. No major complications were encountered. CONCLUSIONS: Percutaneous cementoplasty can provide pain relief and improvement of quality of life, though without survival benefits, for HCC patients with painful bone metastasis.

A highly effective and stable bispecific diabody for cancer immunotherapy: cure of xenografted tumors by bispecific diabody and T-LAK cells.

PURPOSE: In the field of cancer immunotherapy research, the targeting of effector cells with specific antibodies is a very promising approach. Recent advances in genetic engineering have made it possible to prepare immunoglobulin fragments consisting of variable domains using bacterial expression systems. METHODS: We have produced an anti-epidermal growth-factor receptor (EGFR) x anti-CD3 bispecific diabody (Ex3 diabody) in an Escherichia coli (E. coli) expression system with refolding method. The Ex3 diabody targets lymphokine-activated killer cells with a T-cell phenotype (T-LAK cells) to EGFR positive bile duct carcinoma cells with dramatic enhancement of cytotoxicity in vitro. This specific killing of EGFR-positive cells was completely inhibited by parental mAb IgGs directed to EGFR and the CD3 antigen. RESULTS: When T-LAK cells were cultured with EGFR-positive tumor cells in the presence of Ex3 diabody, they produced much higher levels of IFN-gamma, GM-CSF, and TNF-alpha than in its absence, this being a possible mechanism underlying specific antitumor activity. The Ex3 diabody showed good stability when tested at 37 degrees C for 48 h, and also markedly inhibited tumor growth of bile duct carcinoma xenografts in severe combined immunodeficient (SCID) mice. When Ex3 diabody (20 microg/mouse) was administrated intravenously, together with T-LAK cells and interleukin-2 (IL-2), complete cure of tumors were observed in three of six mice, and the other three showed marked retardation of tumor growth. CONCLUSION: The Ex3 diabody can be considered a highly promising reagent for study of specific targeting immunotherapy against bile duct and other EGFR-positive carcinomas.

A novel adenovirus expressing human 4-1BB ligand enhances antitumor immunity.

4-1BB ligand (4-1BBL), a member of the tumor necrosis factor (TNF) superfamily, interacts with 4-1BB (CDw137) expressed on activated T cells and delivers a costimulatory signal for T cell activation and growth. Various studies have demonstrated a role for murine 4-1BB in immune function, but relatively few investigations of human 4-1BB have been conducted. Here we report on the construction of a recombinant E1/E3-deleted adenovirus encoding human 4-1BBL (Ad4-1BBL) and its stimulation of antitumor immunity. Ad4-1BBL was able to efficiently infect several human adenocarcinoma cell lines and induce 4-1BBL expression on the cell surface within 24 h, this enhancing the antitumor activity not only of lymphokine-activated killer cells with a T cell phenotype (T-LAK) but also naive peripheral blood mononuclear cells (PBMC). This antitumor activity with T-LAK cells was further enhanced by addition of bispecific antibody (BsAb; anti-MUC1xanti-CD3). Cocultivation of Ad4-1BBL-infected tumor cells with either T-LAK cells or PBMC resulted in significant elevation of interferon-gamma (IFN-gamma), interleukin-2 (IL-2), and granulocyte-macrophage colony-stimulating factor (GM-CSF) production. Furthermore, remarkable tumor growth inhibition was observed in cholangiocarcinoma-grafted severe combined immunodeficient (SCID) mice to which Ad4-1BBL and T-LAK cells were administered when tumor size exceeded 5 mm in diameter. These results provide strong evidence in support of the efficacy of adenovirally delivered 4-1BBL for genetic immunotherapy of cancer.

Specific and effective targeting cancer immunotherapy with a combination of three bispecific antibodies.

For the purpose of establishing a new adoptive immunotherapy for bile duct carcinoma (BDC), we previously constructed two kinds of bispecific antibodies (bsAbs), anti-MUC1 x anti-CD3 (M x 3) and anti-MUC1 x anti-CD28 (M x 28), which activate T cells and form bridges between them and MUC1-expressing tumor cells. In our previous studies [Cancer Res. 56 (1996) 4205] specific targeting therapy (STT) consisting of i.v. administration of lymphokine activated killer cells with a T cell phenotype (T-LAK) sensitized with two kinds of bsAbs to human BDC-grafted severe combined immunodeficient (SCID) mice demonstrated remarkable inhibition of tumor growth. However, complete cures could not be obtained. In order to improve antitumor efficacy, we have paid attention to anti-CD2 monoclonal antibodies (mAbs), thought to play an important roles in signal transduction in T cell activation or control of T cell receptor (TCR)-driven activation. Therefore, we developed another bsAb, anti-MUC1 x anti-CD2 (M x 2), in order to examine if this would show synergism with the two previously described bsAbs. The combination of the three bsAbs (M x 3, M x 28 and M x 2 bsAbs) showed highest cytotoxicity against MUC1-expressing BDC cells when given simultaneously with peripheral blood mononuclear cells (PBMCs) or T-LAK cells in vitro. When 2 x 10(7) T-LAK cells sensitized with different combinations of bsAbs were administered four times i.v. to BDC-grafted SCID mice, the best therapeutic result was obtained with a combination of all three bsAbs. These results indicate usefulness of combination of three bsAbs for targeting cancer immunotherapy.