Long-term outcomes of proton beam therapy in patients with previously untreated hepatocellular carcinoma.

Long-term efficacy of proton beam therapy (PBT) remains unclear for patients with previously untreated hepatocellular carcinoma (HCC). We aimed to study the long-term outcomes of PBT according to Barcelona Clinic Liver Cancer (BCLC) staging classifications in patients with previously untreated HCC. The major eligibility criteria of this observational study were an Eastern Cooperative Oncology Group performance status (PS) 0-2, Child-Pugh grade A or B, previously untreated HCC covered within an irradiation field, and no massive ascites. A total of 66.0-77.0 GyE was administered in 10-35 fractions. Local tumor control (LTC), defined as no progression in the irradiated field, progression-free survival (PFS), and overall survival (OS) were assessed according to BCLC staging. From 2002 to 2009 at our institution, 129 patients were eligible. The 5-year LTC, PFS, and OS rates were 94%, 28%, and 69% for patients with 0/A stage disease (n = 9/21), 87%, 23%, and 66% for patients with B stage disease (n = 34), and 75%, 9%, and 25% for patients with C stage disease (n = 65), respectively. The 5-year LTC and OS rates of 15 patients with tumor thrombi in major vessels were 90% and 34%, respectively. Multivariate analyses revealed that PS (0 versus 1-2) was a significant prognostic factor for OS. No grade 3 or higher adverse effects were observed. PBT showed favorable long-term efficacies with mild adverse effects in BCLC stage 0 to C, and can be an alternative treatment for localized HCC especially when accompanied with tumor thrombi. This study was registered with UMIN Clinical Trials Registry (UMIN000025342).

Identification of a unique hepatocellular carcinoma line, Li-7, with CD13(+) cancer stem cells hierarchy and population change upon its differentiation during culture and effects of sorafenib.

BACKGROUNDS: Cancer stem cell (CSC) research has highlighted the necessity of developing drugs targeting CSCs. We investigated a hepatocellular carcinoma (HCC) cell line that not only has CSC hierarchy but also shows phenotypic changes (population changes) upon differentiation of CSC during culture and can be used for screening drugs targeting CSC. METHODS: Based on a hypothesis that the CSC proportion should decrease upon its differentiation into progenitors (population change), we tested HCC cell lines (HuH-7, Li-7, PLC/PRF/5, HLF, HLE) before and after 2 months culture for several markers (CD13, EpCAM, CD133, CD44, CD90, CD24, CD166). Tumorigenicity was tested using nude mice. To evaluate the CSC hierarchy, we investigated reconstructivity, proliferation, ALDH activity, spheroid formation, chemosensitivity and microarray analysis of the cell populations sorted by FACS. RESULTS: Only Li-7 cells showed a population change during culture: the proportion of CD13 positive cells decreased, while that of CD166 positive cells increased. The high tumorigenicity of the Li-7 was lost after the population change. CD13(+)/CD166(-) cells showed slow growth and reconstructed the bulk Li-7 populations composed of CD13(+)/CD166(-), CD13(-)/CD166(-) and CD13(-)/CD166(+) fractions, whereas CD13(-)/CD166(+) cells showed rapid growth but could not reproduce any other population. CD13(+)/CD166(-) cells showed high ALDH activity, spheroid forming ability and resistance to 5-fluorouracil. Microarray analysis demonstrated higher expression of stemness-related genes in CD166(-) than CD166(+) fraction. These results indicated a hierarchy in Li-7 cells, in which CD13(+)/CD166(-) and CD13(-)/CD166(+) cells serve as slow growing CSCs and rapid growing progenitors, respectively. Sorafenib selectively targeted the CD166(-) fraction, including CD13(+) CSCs, which exhibited higher mRNA expression for FGF3 and FGF4, candidate biomarkers for sorafenib. 5-fluorouracil followed by sorafenib inhibited the growth of bulk Li-7 cells more effectively than the reverse sequence or either alone. CONCLUSIONS: We identified a unique HCC line, Li-7, which not only shows heterogeneity for a CD13(+) CSC hierarchy, but also undergoes a "population change" upon CSC differentiation. Sorafenib targeted the CSC in vitro, supporting the use of this model for screening drugs targeting the CSC. This type of "heterogeneous, unstable" cell line may prove more useful in the CSC era than conventional "homogeneous, stable" cell lines.

Proton beam therapy for unresectable intrahepatic cholangiocarcinoma.

BACKGROUND AND AIM: Treatment for unresectable intrahepatic cholangiocarcinoma (ICC) has not been established. The aim of the study was to evaluate the outcome of proton beam therapy (PBT) for patients with unresectable ICC. METHODS: Up to 2010, 20 patients (11 males, 9 females, median age 63 years old) with unresectable ICC (two, seven, seven, and four in stages II, IIIA, IIIC, and IV, respectively) were treated with PBT. The largest dimensions of the tumors ranged from 15 to 140 mm (median: 50 mm). The intrahepatic region and lymph nodes received median total proton doses of 72.6 GyE in 22 fractions and 56.1 GyE in 17 fractions, respectively. Four patients received concurrent chemotherapy (tegafur, gimeracil, and oteracil; TS-1) during PBT. Twelve patients were treated curatively, and eight were treated palliatively because tumors were present outside the irradiation field. RESULTS: In the curative group, nine tumors within the irradiated field were controlled in follow-up of 8.6-62.6 months (median: 20.8 months). Median survival rates in the curative and palliative groups were 27.5 and 9.6 months, respectively, and overall 1- and 3-year survival rates were 82% and 38%, and 50% and 0%, respectively. Eight patients survived for > 2 years, and there was no distant metastasis in five of these patients after 2 years. No severe side-effects occurred. CONCLUSIONS: The results suggest that long-term survival can be achieved using PBT for patients with unresectable ICC without distant metastasis. Further studies are required to determine the optimal treatment schedule and best combination of PBT and chemotherapy.

Clinical features of hepatitis E virus infection in Ibaraki, Japan: autochthonous hepatitis E and acute-on-chronic liver failure.

Hepatitis E caused by hepatitis E virus (HEV) is a serious public health concern in developing countries where HEV is mainly transmitted through contaminated water. Recently, in industrialized countries, autochthonous hepatitis E, a porcine zoonosis, has been increasingly recognized. In Japan, the number of national notifications of acute hepatitis E has increased since the introduction of anti-HEV IgA antibody measurement, covered by the national health insurance program, in 2011. In the past three years, we examined five patients of acute hepatitis or acute-on-chronic liver failure caused by HEV infection who presented various clinical courses in the southern area of Ibaraki prefecture in Japan. Of these patients, 78-year-old and 63-year-old male patients presented acute hepatitis E and recovered by only bed rest. The latter patient had a history of consuming grilled or undercooked pork and shellfish prior to the onset of hepatitis E. Among the five patients examined, the infection route was detected only in this patient. Of note, a 65-year-old female patient presented severe hepatitis associated with painless thyroiditis. The patient was diagnosed with probable autoimmune hepatitis and was successfully treated with prednisolone (40 mg/day). Lastly, 58-year-old and 62-year-old male patients, both of whom had a history of diabetes mellitus and alcoholic liver disease, developed acute-on-chronic liver failure, and the latter patient with pre-existing liver cirrhosis died due to liver failure. Thus, patients with clinical HEV infection who display multiple underlying diseases can develop acute-on-chronic liver failure. In conclusion, HEV infection manifests the diverse clinical courses.

Cyclophosphamide enhances antitumor efficacy of oncolytic adenovirus expressing uracil phosphoribosyltransferase (UPRT) in immunocompetent Syrian hamsters.

Oncolytic viruses (OVs) are novel cancer therapeutics with great promise, but host antiviral immunity represents the hurdle for their efficacy. Immunosuppression by cyclophosphamide (CP) has thus been shown to enhance the oncolytic efficacy of many OVs, but its effects on OVs armed with therapeutic genes remain unknown. We have previously reported on the efficacy of AxE1CAUP, an oncolytic adenovirus (OAd) expressing uracil phosphoribosyltransferase (UPRT), an enzyme that markedly enhanced the toxicity of 5-fluorouracil (5-FU), in immunodeficient, Ad-nonpermissive nude mice. Here we explored the efficacy and safety of intratumoral (i.t.) AxE1CAUP/5-FU therapy and of its combination with CP for syngenic HaP-T1 pancreatic cancers in immunocompetent, Ad-permissive Syrian hamsters. AxE1CAUP infected, replicated, expressed UPRT, and increased the sensitivity to 5-FU in HaP-T1 cells in vitro. I.t. AxE1CAUP/5-FU treatment inhibited the growth of subcutaneous HaP-T1 allografts. The combination with high-dose CP inhibited serum Ad-neutralizing antibody formation, increased intratumoral AxE1CAUP replication and UPRT expression, and resulted in further enhanced therapeutic effects with 5-FU. Neither body weight nor histology of the liver and lung changed during these treatments. A clinically-approved, intermediate-dose CP also enhanced the efficacy of i.t. AxE1CAUP/5-FU treatment in these hamsters, which was not affected by preexisting immunity to the vector. These data demonstrate the excellent antitumor efficacy and safety of an OAd armed with a suicide gene in combination with CP for treating syngenic tumors in immunocompetent, Ad-permissive animals, indicating the efficacy of CP in overcoming the hurdle of antiviral immunity for effective OV-mediated gene therapy.

Impact of solute carrier family 29 member 1 (SLC29A1) single nucleotide polymorphisms on mRNA expression in peripheral blood mononuclear cells.

The effects of solute carrier family 29 member 1 (SLC29A1) single nucleotide polymorphism (SNP), rs6932345 and rs747199, on SLC29A1 mRNA expression were examined. The expression levels of SLC29A1 mRNA in peripheral blood mononuclear cells (PBMCs) isolated from 46 healthy subjects (28 males and 18 females) was compared between wild-type and mutant carriers. The mRNA levels in the rs6932345 wild-type (AA genotype) was 1.71 times that in the mutation carriers (AC/CC genotype) (p<0.05). Similar results were observed for rs747199, because rs747199 was linked with rs6932345 at a frequency of 84.8%. It was confirmed that wild-type for rs6932345 and rs747199 showed higher SLC29A1 mRNA expression in PBMCs.

EpCAM- and EGFR-targeted selective gene therapy for biliary cancers using Z33-fiber-modified adenovirus.

A critical issue in adenovirus (Ad)-based cancer gene therapy is to improve the specificity of gene delivery to cancer cells for better efficacy and safety. We explored methods of retargeting Ad vectors for selective gene therapy of human biliary cancers using the Ad incorporating an IgG Fc-binding motif (Z33) from the Staphylococcus protein A (Ad-FZ33) combined with tumor-specific antibodies. Flow cytometry analysis revealed high-expression levels of epithelial cell adhesion molecule (EpCAM) and epidermal growth factor receptor (EGFR) on human biliary cancer cells. Ad-FZ33 expressing LacZ combined with antibodies against EpCAM or EGFR, followed by ß-gal assay, demonstrated highly efficient gene transduction in these biliary cancer cells, compared to the treatment with control antibody or without antibody. Ad-FZ33 expressing uracil phosphoribosyl transferase (UPRT), an enzyme which greatly enhances the toxicity of 5-fluorouracil (FU), combined with antibodies against EpCAM or EGFR, remarkably enhanced the sensitivity of biliary cancer cells to 5-FU. By contrast, the treatment did not affect the 5-FU sensitivity of the cells not expressing EpCAM or EGFR including normal hepatocytes. Finally, treatments with the UPRT-expressing Ad-FZ33 with antibodies against EpCAM or EGFR, followed by 5-FU administration, significantly suppressed the growth of biliary cancer xenografts in nude mice. These results indicate that the gene therapy mediated by the Z33 fiber modified Ad with anti-EpCAM or anti-EGFR antibodies offers a potentially effective therapeutic modality against biliary cancers.

Potent in vitro and in vivo antitumor effects of MDM2 inhibitor nutlin-3 in gastric cancer cells.

The tumor suppressor gene p53 is the most frequently mutated gene in human cancers. However, its mutation rate is relatively low in gastric cancer compared with other cancers. In this study, we investigated the mechanisms underlying the antitumor effects of nutlin-3, an inhibitor of human homolog of murine double minute 2 (MDM2). MDM2 is a negative regulator of p53. Four gastric cancer cell lines with wild-type p53 (wt p53) and three with mutant-type p53 (mt p53) were analyzed for MDM2 and MDM4 expression by immunoblotting, and for their gene amplification by quantitative real-time PCR. Moreover, the viability of cells exposed to nutlin-3 was examined by WST-8 assay, and the expression of p53 and its downstream genes was analyzed by immunoblotting. Nutlin-3 stabilized p53 and increased the expression of p21(WAF1) and Noxa, and cleaved poly (ADP)-ribose polymerase regardless of the pre-expression levels of MDM2 and MDM4 in gastric cancer cells with wt p53. Flow cytometry revealed that nutlin-3 arrested the cell cycle in G(1) phase and induced apoptosis in the cell lines. These nutlin-3 effects were not observed in the cell lines with mt p53. Nutlin-3 exerted additive or synergistic cytotoxicity in combination with 5-fluorouracil or cisplatin in most cell lines with wt p53. An in vivo antitumor effect of nutlin-3 alone and its additive augmentation by 5-fluorouracil were confirmed in an MDM2 overexpressed xenograft tumor model. Nutlin-3 showed potent antitumor activity against human gastric cancer cells with wt p53 and shows promise as a single agent and in combination with conventional anticancer drugs.

Proton-beam therapy for hepatocellular carcinoma associated with portal vein tumor thrombosis.

BACKGROUND AND PURPOSE: The prognosis of patients with advanced hepatocellular carcinoma (HCC) with portal vein tumor thrombosis (PVTT) is extremely poor, as effective treatment options are limited. The authors performed a retrospective review to evaluate the efficacy of proton-beam therapy (PBT) for patients presenting with PVTT in the setting of HCC. PATIENTS AND METHODS: Between February 1991 and September 2005, 35 patients with HCC and tumor thrombi in the main trunk or major branches of the portal vein presented for consideration of PBT. Their tumor sizes ranged from 25 mm to 130 mm (median, 60 mm). A median total dose of 72.6 GyE in 22 fractions was delivered over 31 days to a target volume that encompassed both the primary hepatic lesion and the PVTT. RESULTS: 32 patients were progression-free during a median follow-up period of 21 months (range, 2-88 months) and three patients experienced disease progression. Local progression-free survival rates were 46% at 2 years and 20% at 5 years, and the median local progression-free survival was 21 months. Acute toxicity > or = grade 3 was observed in three patients, and no patient experienced late toxicity > or = grade 3. None of the patients had to discontinue treatment as a result of toxicity. CONCLUSION: PBT improved local control and significantly prolonged survival in HCC patients with PVTT.

Safety margin of radiofrequency ablation for hepatocellular carcinoma: a prospective study using magnetic resonance imaging with superparamagnetic iron oxide.

PURPOSE: In radiofrequency ablation (RFA) for hepatocellular carcinoma (HCC), it is difficult to assess the ablative margin (AM) precisely by comparing pre- and post-RFA CT images. We prospectively studied the AMs using magnetic resonance imaging (MRI) with pre-administered superparamagnetic iron oxide (SPIO). SPIO is safe for kidney disease patients. MATERIALS AND METHODS: Hepatocellular carcinoma patients were treated with RFA within 8 h of SPIO administration. On T2*-weighted MRI performed 4-7 days later, AM was visualized as a hypointense rim. The ablation status was classified as AM(+) if the rim completely surrounded the tumor, AM(0) if the rim was partly discontinuous without tumor protrusion, and AM(-) if the rim was partly discontinuous with tumor protrusion. The minimal thickness of AM was measured. AM(-) tumors were re-treated consecutively. RESULTS: In total, 85 HCCs ablated in 76 patients were evaluated. The local recurrence rate at 3 years was 2% for AM(+) tumors and 34% for AM(0) tumors (p < 0.01). In addition, no local recurrence was seen in the tumors with an AM of ≥ 2 mm. CONCLUSION: MRI with pre-administered SPIO is useful for determining the AM precisely, and an AM of ≥ 2 mm is recommended for curative RFA. TRIAL REGISTRATION NUMBER: This study was registered with UMIN Clinical Trials Registry (UMIN 000025406).

Proton beam therapy for hepatocellular carcinoma adjacent to the porta hepatis.

PURPOSE: To evaluate the efficacy and safety of proton beam therapy (PBT) for patients with hepatocellular carcinoma (HCC) located adjacent to the porta hepatis. METHODS AND MATERIALS: Subjects of the study were 53 patients with HCC located within 2 cm of the main portal vein. All patients had tumor confined to the radiation field with no evidence of metastatic disease. All patients had hepatic function levels of a Child-Pugh score of 10 or less, Eastern Cooperative Oncology Group performance status of 2 or less, and no uncontrolled ascites. Patients underwent PBT of 72.6 GyE in 22 fractions from Sept 2001 to Dec 2004. RESULTS: After 3 years, the actuarial survival rate was 45.1% and local control rate was 86.0%. Prognostic factors for survival included Child-Pugh score, number of tumors, and alpha-fetoprotein levels. No late treatment-related toxicity of Grade 2 or higher was observed. CONCLUSIONS: The PBT delivering 72.6 GyE in 22 fractions appears to be effective and safe for HCC adjacent to the porta hepatis.

E1A, E1B double-restricted adenovirus with RGD-fiber modification exhibits enhanced oncolysis for CAR-deficient biliary cancers.

PURPOSE: Cancers of biliary system represent highly malignant diseases of dismal prognosis. We have previously introduced AxdAdB3, an E1A, E1B double-restricted oncolytic adenovirus, which showed excellent oncolytic efficacy for approximately half of the biliary cancer lines with an enhanced safety to normal cells. The purpose of this study was to evaluate whether RGD-fiber modification (AxdAdB3-F/RGD), which enables integrin-dependent infection, can improve the infectivity and efficacy of AxdAdB3 for biliary cancers. EXPERIMENTAL DESIGN: Expressions of adenoviral receptors, coxsackievirus adenovirus receptor (CAR) and integrins (alpha(v)beta(3) and alpha(v)beta(5)), were compared with the level of infectivity of LacZ-expressing replication-defective adenoviruses with wild-type fibers or RGD-modified fibers in a panel of biliary cancer cell lines in vitro. Viral replication and cytotoxicity in vitro of AxdAdB3-F/RGD, a novel E1A, E1B double-restricted replication-selective adenovirus with RGD-modified fibers, were compared with those of its parent virus, AxdAdB3, in various biliary cancer cells and in normal cells. In vivo antitumor effects of these oncolytic viruses were compared in a xenograft tumor model. RESULTS: Expression of CAR significantly correlated with the adenovirus infectivity, whereas integrin alpha(v)beta(5) was abundantly expressed in almost all biliary cancer cells. Whereas AxdAdB3 effectively replicated and lysed only the biliary cancer cells with a preserved expression of CAR, AxdAdB3-F/RGD exhibited efficient replication and potent oncolysis in both CAR-positive and CAR-negative biliary cancer cells. AxdAdB3-F/RGD showed attenuated replication and little cytopathy in human normal cells (i.e., hepatocytes, WI-38 cells) as well as AxdAdB3. Furthermore, in nude mice with s.c. xenografts of CAR-deficient human biliary cancer, i.t. AxdAdB3-F/RGD therapy caused a marked inhibition of tumor growth. CONCLUSIONS: The RGD-fiber modification strategy enhanced the infectivity, replication, and oncolytic effects of the E1A, E1B double-restricted oncolytic adenovirus for CAR-deficient biliary cancers. In addition, it preserved the merit of excellent safety of the double-restricted virus for normal cells. These results suggest a potential use of this agent for the treatment of biliary cancers.

Oncolytic Virotherapy and Gene Therapy Strategies for Hepatobiliary Cancers.

Advanced liver cancers and biliary cancers represent diseases with dismal prognosis because of frequent local invasion and metastasis. Effective therapeutic agents for these cancers have not been established. Oncolytic viruses (OVs) constitute a novel class of promising, selective anticancer agents and recent studies have elucidated their unique features. Moreover, clinical trials are demonstrating promising results. Numerous OVs are being tested in preclinical models of hepatocellular carcinoma (HCC). The lead agent Pexa-Vec (pexastimogene devacirepvec, JX-594), a recombinant Wyeth strain vaccinia virus, has demonstrated preliminary evidence of safety and efficacy for HCC in clinical trials. Few other OVs have entered clinical testing. Relatively few preclinical studies and clinical trials exist for biliary cancers. In this review, we introduce various approaches using OVs to treat the intractable hepatobiliary cancers.

Oncolytic Viruses: The Best is Yet to Come.

Oncolytic viruses are a promising anti-cancer platform, achieving significant pre-clinical and clinical milestones in recent years. A full arsenal of selective, safe, and effective viruses has been developed with some emerging pre-clinical research focusing on optimizing these therapies in the face of remaining challenges, both in the bloodstream and in the tumour microenvironment. Herein we discuss the recent progress in pre-clinical virotherapy research to address these challenges, with special focus on innovative strategies that seek to complement the current strengths of virotherapy, ensuring an optimal multi-faceted attack on cancer. This review highlights the research areas that we believe provide the most potential to increase the efficacy of this exciting biotherapy platform: cell carriers, tumour vascular destruction, microenvironment modulation, combination therapies, and virus-mediated anti-tumour immune responses.

Proton beam therapy for aged patients with hepatocellular carcinoma.

PURPOSE: To investigate the safety and efficacy of proton beam therapy for aged patients with hepatocellular carcinoma (HCC). METHODS AND MATERIALS: Twenty-one patients aged > or =80 years with HCC underwent proton beam therapy. At the time of irradiation, patient age ranged from 80 to 85 years (median, 81 years). Hepatic tumors were solitary in 17 patients and multiple in 4. Tumor size ranged from 10 to 135 mm (median, 40 mm) in maximum diameter. Ten, 5, and 6 patients received proton beam irradiation with total doses of 60 Gy in 10 fractions, 66 Gy in 22 fractions, and 70 Gy in 35 fractions, respectively, according to tumor location. RESULTS: All irradiated tumors were controlled during the follow-up period of 6-49 months (median, 16 months). Five patients showed new hepatic tumors outside the irradiated volume, 2-13 months after treatment, and 1 of them also had lung metastasis. The local progression-free and disease-free rates were 100% and 72% at 3 years, respectively. Of 21 patients, 7 died 6-49 months after treatment; 2 patients each died of trauma and old age, and 1 patient each died of HCC, pneumonia, and arrhythmia. The 3-year overall, cause-specific, and disease-free survival rates were 62%, 88%, and 51%, respectively. No therapy-related toxicity of Grade > or = 3 but thrombocytopenia in 2 patients was observed. CONCLUSIONS: Proton beam therapy seems to be tolerable, effective, and safe for aged patients with HCC. It may contribute to prolonged survival due to tumor control.

Effective gene therapy of biliary tract cancers by a conditionally replicative adenovirus expressing uracil phosphoribosyltransferase: significance of timing of 5-fluorouracil administration.

In order to enhance the efficacy of conditionally replicating adenoviruses (CRAd) in the treatment of cancers of the biliary tract, we studied the efficacy in vitro and in vivo of AxE1CAUP, a CRAd vector that carries a gene for uracil phosphoribosyltransferase (UPRT), which converts 5-fluorouracil (5-FU) directly to 5-fluorouridine monophosphate and greatly enhances the cytotoxicity of 5-FU. AxE1CAUP replicated and induced an increased UPRT expression in biliary cancer cells more efficiently than AxCAUP, a nonreplicative adenovirus carrying the UPRT gene. Whereas AxCAUP and AxE1AdB, a CRAd without the UPRT gene, modestly increased the sensitivity of BC cells to 5-FU, AxE1CAUP markedly increased the sensitivity, especially when the timing of 5-FU administration was appropriately chosen. AxE1CAUP replicated much less efficiently in normal WI-38 fibroblasts without any change in the sensitivity to 5-FU. In nude mice with s.c. biliary cancer xenografts, i.t. AxE1CAUP/5-FU therapy inhibited tumor growth significantly more strongly than AxCAUP/5-FU or AxE1AdB/5-FU therapy. Furthermore, in mice with peritoneally disseminated biliary cancer, i.p. AxE1CAUP efficiently proliferated in the tumors, decreased the tumor burden, and prolonged the survival of the mice when 5-FU was started 10 or 15 days after the vector inoculation, whereas earlier initiation of 5-FU resulted in early eradication of the vector and no survival benefit. The present study shows that the CRAd expressing UPRT was a more potent sensitizer of biliary cancer to 5-FU, than was a nonreplicative UPRT-encoding vector or a CRAd without UPRT gene, even at a lower dose of the vector, and that timing of 5-FU administration was a key factor to maximize the efficacy. This gene therapy with appropriately timed administration of 5-FU should be useful in overcoming the resistance of biliary cancers to 5-FU.

Analysis of repeated proton beam therapy for patients with hepatocellular carcinoma.

PURPOSE: To evaluate the feasibility and efficacy of repeated proton beam therapy (PBT) and to analyze the dose-volume relationship. PATIENTS AND METHODS: A retrospective analysis was performed in 83 patients who received definitive repeated PBT. Proton beams were delivered with expiratory gating. The numbers of treatment courses were 2, 3, and 4 in 68, 12, and 3 patients, respectively. MIM software was used for dose analysis. RESULTS: The planned median total dose was 70.5 GyE for all tumors and the median doses for the 1st, 2nd, 3rd and 4th treatments were 71.0, 70.0, 70.0, and 69.3 GyE, respectively. There was no severe acute toxicity, and no radiation-induced liver dysfunction (RILD) was observed. The median overall survival (OS) period from the first PBT was 61months (95% CI: 50-71months), and the 2- and 5-year OS rates were 87.5% (95%CI: 80.2-94.8%) and 49.4% (95%CI: 37.6-61.2%), respectively. The maximal delivered dose to the liver ranged from 66.7 to 248.1 GyE (mean: 124.93 GyE) and the mean liver dose ranged from 5.4 to 66.5 GyE (mean: 24.23 GyE). CONCLUSION: Repeated PBT was well tolerated and safe, even though the liver doses in many patients deviated substantially from well-known critical levels for RILD.

Proton beam therapy for hepatocellular carcinoma: a retrospective review of 162 patients.

PURPOSE: We present results of patients with hepatocellular carcinoma (HCC) treated with proton beam therapy. EXPERIMENTAL DESIGN: We reviewed 162 patients having 192 HCCs treated from November 1985 to July 1998 by proton beam therapy with or without transarterial embolization and percutaneous ethanol injection. The patients in the present series were considered unsuitable for surgery for various reasons, including hepatic dysfunction, multiple tumors, recurrence after surgical resection, and concomitant illnesses. The median total dose of proton irradiation was 72 Gy in 16 fractions over 29 days. RESULTS: The overall survival rate for all of the 162 patients was 23.5% at 5 years. The local control rate at 5 years was 86.9% for all 192 tumors among the 162 patients. The degree of impairment of hepatic functions attributable to coexisting liver cirrhosis and the number of tumors in the liver significantly affected patient survival. For 50 patients having least impaired hepatic functions and a solitary tumor, the survival rate at 5 years was 53.5%. The patients had very few acute reactions to treatments and a few late sequelae during and after the treatments. CONCLUSIONS: Proton beam therapy for patients with HCC is effective, safe, well tolerable, and repeatable. It is the useful treatment mode for either cure or palliation for patients with HCC irrespective of tumor size, tumor location in the liver, insufficient feeding of the tumor with arteries, presence of vascular invasion, impaired hepatic functions, and coexisting intercurrent diseases.

E1A, E1B double-restricted adenovirus for oncolytic gene therapy of gallbladder cancer.

New treatments, such as gene therapy, are necessary for advanced gallbladder cancer (GBC), but little has been studied. Recent studies have introduced mutant adenoviruses (Ads) with either defective E1B-55kD or mutated E1A, focusing on tumor-specific replication, and the results have been promising. To enhance the safety of this approach, we constructed AxdAdB-3, a double-restricted Ad with a mutant E1A and E1B-55kD deletion. We studied the effects of this Ad in vitro and in vivo on GBC, as well as its safety for normal human cells. We compared the replication and cytopathic effects of AxdAdB-3 in several lines of GBC and primary normal cells with those of wild-type Ad or of AxE1AdB, an E1B-55kD-deleted Ad. The efficacy in vivo was examined in nude mice with s.c. implanted or i.p. disseminated GBC. AxdAdB-3 replicated in and caused oncolysis of GBC cell lines (TGBC-44TKB and Mz-ChA2) as efficiently as wild-type Ad or AxE1AdB in vitro. By contrast, AxdAdB-3 replicated much less effectively in primary normal cells (e.g., epithelial cells, endothelial cells, and hepatocytes) than in GBC cells and had only mild cytopathic effects, unlike wild-type Ad. Furthermore, cytotoxicity of AxdAdB-3 in normal cells was milder than that of AxE1AdB. AxdAdB-3 significantly (P < 0.01) suppressed the growth of GBC (TGBC-44TKB) xenografts. AxdAdB-3 was also effective in the treatment of mice with peritoneally disseminated GBC (TGBC-44TKB), demonstrating tumor-selective replication and oncolysis that resulted in significantly (P < 0.05) prolonged survival. The present study shows that the E1 double-restricted Ad effectively and selectively replicates in and causes oncolysis of GBC in vitro and in vivo with reduced negative effects on normal cells, suggesting that this approach could be a promising tool for gene therapy of GBC.

Association between pretreatment retention rate of indocyanine green 15 min after administration and life prognosis in patients with HCC treated by proton beam therapy.

PURPOSE: The Child-Pugh score is often used to judge the outcome of radiotherapy for hepatocellular carcinoma (HCC). The retention rate of indocyanine green 15 min after administration (ICG R15) can also be used to predict prognosis after liver resection. We evaluated the utility of ICG R15 for prediction of outcomes after proton beam therapy (PBT) for HCC. METHODS AND MATERIALS: A retrospective evaluation was performed in 250 patients who received PBT between 2002 and 2007. The patients (178 males and 72 females) had a median age of 71 years (range: 43-88). Child-Pugh categories were A (score 5-6), B (7-9), and C (10-15) in 197, 51, and 2 patients, respectively. ICG scores were 0-<10, 10-<20, 20-<30, 30-<40 and ⩾40 in 27, 99, 59, 28 and 37 patients, respectively; including 26, 92, 45, 16 and 18 Child-Pugh A patients and 1, 8, 14, 11, and 17 Child-Pugh B patients, respectively. Survival times from the start of PBT were compared between Child-Pugh A and B patients, and among each ICG group. RESULTS: The median survival times were 61 months (95% CI: 50-72 months) in all patients, and 64 and 20 months in Child-Pugh A and B patients, respectively (p=0.001), The 3-year survival rates were 72%, 72%, 75%, 63%, and 26% in patients with ICG scores of 0-<10, 10-<20, 20-<30, 30-<40, and ⩾40 (p=0.001); 70%, 75%, 77%, 65%, and 38% in these respective groups in Child-Pugh A patients (p=0.02); and 100%, 57%, 67%, 36%, and 14% in Child-Pugh B patients (p=0.173, not significant). Multivariate analysis showed that low ICG R15 and the absence of portal vein tumor thrombus were associated with good survival. CONCLUSIONS: Pretreatment ICG R15 is a useful prognostic factor for prediction of outcome of PBT in HCC patients, especially in those with Child-Pugh A liver function.