Prevention of intrahepatic metastasis of liver cancer by suicide gene therapy and chemokine ligand 2/monocyte chemoattractant protein-1 delivery in mice.

BACKGROUND: The prognosis of patients with hepatocellular carcinoma (HCC) remains poor, largely as a result of intrahepatic metastasis. Using a mouse model of intrahepatic metastasis, we investigated whether chemokine ligand 2/monocyte chemoattractant protein-1 (CCL2/MCP-1) could potentiate the antitumor effects of the herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV) system. METHODS: Mouse hepatoma cells infected with recombinant adenovirus vectors expressing HSV-tk, CCL2/MCP-1 and LacZ at multiplicities of infection of Ad-tk/Ad-MCP1 = 3/0.03 (T/M(Low)), 3/3 (T/M(High)) and Ad-tk/Ad-LacZ = 3/3 (T/L) were injected into BALB/c mice. RESULTS: Intrahepatic tumor growth was significantly lower in T/M(Low) mice. By contrast, no tumor suppression was observed in T/M(High) mice. The tumor-specific cytolytic activities of splenocytes from T/M(Low) and T/M(High) mice were comparable. Immunohistochemical analysis of liver tissues showed similar infiltration by Mac-1(+) and T cells in these animals, whereas the proportions of classical activated (M1) monocytes/macrophages were significantly higher in T/M(Low) mice. In addition, interleukin-12 production was elevated in these tissues. Vascular endothelial growth factor-A expression and CD31(+) microvessels were increased in T/M(High) mice. CONCLUSIONS: Collectively, these results demonstrate that an adequate amount of CCL2/MCP-1, together with the HSV-tk/GCV system, may induce T helper 1-polarized antitumor effects without inducing tumor angiogenesis in the microenvironment of intrahepatic HCC progression.

Advanced hepatocellular carcinoma treated effectively with irinotecan via hepatic arterial infusion followed by proton beam therapy.

We report a 48-year-old man with hepatocellular carcinoma (HCC) treated with hepatic arterial infusion (HAI) chemotherapy followed by proton beam therapy. The HCC lesion in this patient was 88 mm in diameter, with portal vein tumor thrombosis in the right lobe of the liver. He was first treated with 5-fluorouracil, cisplatin, and isovorin, administered by HAI, combined with interferon-alpha, and he was subsequently treated with epirubicin and mitomycin-C administered by HAI. However, no definite efficacy of either of these treatments was observed. Then, after 3 weeks' continuous administration of irinotecan by HAI, the tumor size decreased to 68 mm in diameter. However, 3 months after reduction of the tumor, the tumor had become enlarged to 100 mm in diameter and intrahepatic metastases were prominent. Angiographic findings indicated that the HCC was fed not only from the right hepatic artery but also from the left gastric and right and left subphrenic arteries. After rearrangement of the arteries, and 3 months' continuous HAI chemotherapy with irinotecan, plus hyperthermia, the tumor size had decreased to 50 mm in diameter. The reduction rate of the main tumor according to the Response Evaluation Criteria in Solid Tumors was 43%; therefore, the efficacy of this treatment was judged as a partial response. Two months after reduction of the tumor, the patient's serum alpha-fetoprotein (AFP) level was elevated, and so docetaxel was administered by HAI instead of irinotecan. The liver tumors showed gradual enlargement during the administration of docetaxel, although the AFP level was suppressed. Proton beam therapy was instituted and the liver tumors showed necrosis after this therapy. The patient died of hepatic failure and distant metastases 6 years after the onset of HCC. As far as we know, this is the first case report of HCC treated effectively with irinotecan administered by HAI followed by proton beam therapy in which tumor suppression and the long-term survival of the patient were observed.

Cancer of unknown primary site in which tumor marker-oriented chemotherapy was effective and pancreatic cancer was finally confirmed at autopsy.

We report a 47-year-old man with cancer of unknown primary site in whom pancreatic cancer was confirmed at autopsy. Although a primary lesion was not confirmed, we planned to perform tumor marker-oriented chemotherapy because pancreatic cancer was suspected as the primary lesion based on tumor markers and pathological findings from metastatic lymph node. Neither S-1 nor gemcitabine was effective. However, gemcitabine combined with low-dose cisplatin therapy resulted in a marked decrease in the size of tumors. Microscopic examination at autopsy revealed poorly differentiated adenocarcinoma in the pancreatic head, although a pancreatic mass was not clear macroscopically.

Aortoesophageal fistula following nasogastric tube placement.

A 78-year-old woman was admitted to our hospital because of fresh cerebral infarction. She had been diagnosed as having rheumatoid arthritis, but had not been treated for 50 years. She could not take in sufficient food. Upper gastrointestinal endoscopy revealed no esophageal or gastric lesions, but the procedure was difficult because of her stiff neck from severe rheumatoid degenerative changes of the cervical spine. A nasogastric (NG) tube was placed, and enteral nutrition was initiated. On the 15th day from initiation of enteral nutrition, she presented hematemesis, and suddenly went into a state of shock and died. An autopsy revealed two esophageal ulcers, one of which penetrated into the descending thoracic aorta. The patient was diagnosed with hemorrhagic shock due to aortoesophageal fistula. We suspect that the NG tube compressed the esophageal wall, and ischemia caused the ulcers.

Optimal amount of monocyte chemoattractant protein-1 enhances antitumor effects of suicide gene therapy against hepatocellular carcinoma by M1 macrophage activation.

Suicide gene therapy combined with chemokines provides significant antitumor efficacy. Coexpression of suicide gene and monocyte chemoattractant protein-1 (MCP-1) increases antitumor effects in murine models of hepatocellular carcinoma (HCC) and colon cancer. However, it is unclear whether the doses administered achieved the maximum antitumor effects. We evaluated antitumor effects of various amounts of recombinant adenovirus vector (rAd) expressing MCP-1 in the presence of a suicide gene in a murine model of HCC. HCC cells were transplanted subcutaneously into BALB/c nude mice, and transduced with a fixed amount of Ad-tk harboring the suicide gene, HSV-tk, and various doses of Ad-MCP1 harboring MCP-1 (ratios of 1:1, 0.1:1, and 0.01:1 relative to Ad-tk). Growth of primary tumors was suppressed when treated with Ad-tk plus Ad-MCP1 (1:1 and 1:0.1) as compared with Ad-tk alone. The antitumor effects against tumor rechallenge tended to be high in the Ad-tk plus Ad-MCP1 group (1:0.1). The effects were dependent on production of Th1 type-cytokines. Delivery of an optimal amount of rAd expressing MCP-1 enhanced the antitumor effects of suicide gene therapy against HCC by M1 macrophage activation, suggesting that this is a plausible form of cancer gene therapy to prevent HCC progression and recurrence.

[An autopsy case of pancreatic cancer with poor performance status due to peritonitis carcinomatosa treated successfully with continuous infusion of low-dose 5-FU with octreotide].

A 60-year-old man with pancreatic cancer was admitted due to massive ascites in the course of gemcitabine treatment. Cachexic condition progressed due to peritonitis carcinomatosa. Continuous infusion of low dose 5-FU with octreotide was carefully started. Almost all of ascites disappeared after 4 courses of treatment and his general condition markedly improved. This patient died of pneumonia about 13 months after diagnosis of peritonitis carcinomatosa. Autopsy was undergone, and the effect of chemotherapy was confirmed.

A case of pancreatic cancer with formation of a mass mimicking alcoholic or autoimmune pancreatitis in a young man. Possibility of diagnosis by hypermethylation of pure pancreatic juice.

CONTEXT: Mass-forming pancreatitis can be divided into two distinct types: alcoholic and autoimmune. There have been some cases of an ambiguous diagnosis although care was taken to differentiate between alcoholic mass-forming pancreatitis, focal type autoimmune pancreatitis and pancreatic cancer. CASE REPORT: We report a case of pancreatic cancer mimicking alcoholic or autoimmune pancreatitis with the formation of a mass in a 32-year-old man with a history of heavy drinking. Although both serum immunoglobulin G and immunoglobulin G4 levels were normal, many serum auto-antibodies, including the antinuclear antibody, were detected. After he stopped drinking, abdominal computed tomography showed a pancreatic head mass 28 mm in diameter with little and weak enhancement in the early and delayed phases, respectively. Endoscopic retrograde cholangiopancreatography showed an obstruction of the main pancreatic duct in the pancreatic head and marked stenosis of the lower common bile duct. Although a percutaneous ultrasound-guided pancreatic biopsy demonstrated no evidence of autoimmune pancreatitis, he was treated with prednisolone to test the efficacy of steroid therapy. However, the pancreatic mass became enlarged after steroid therapy, and he underwent surgery during which the mass was found to be pancreatic cancer. Although the patient was treated with gemcitabine, he died 5 months after surgery. We retrospectively assessed DNA hypermethylation in the patient's pure pancreatic juice obtained on admission. We observed hypermethylation of the cancer-specific gene tissue factor pathway inhibitor 2 (TFPI2). CONCLUSION: This finding suggests that if the DNA hypermethylation of pure pancreatic juice had been assayed before steroid therapy, it would have supported the diagnosis of pancreatic cancer, and steroid therapy could have been avoided.

A case of autoimmune pancreatitis associated with retroperitoneal fibrosis.

CONTEXT: Autoimmune pancreatitis is characterized by diffuse enlargement of the pancreas, diffuse irregular narrowing of the main pancreatic duct, severe lymphoplasmacytic infiltration and fibrosis of the pancreas. Retroperitoneal fibrosis may occasionally be associated with autoimmune pancreatitis. CASE REPORT: We report a 77-year-old man with autoimmune pancreatitis associated with retroperitoneal fibrosis. Abdominal ultrasonography and computed tomography demonstrated diffuse enlargement of the pancreas and a capsule-like rim. Furthermore, a retroperitoneal mass was recognized anterior to the abdominal aorta. Antinuclear antibody, IgG and IgG4 values were elevated. Therefore, this patient was diagnosed as having autoimmune pancreatitis associated with retroperitoneal fibrosis. We performed steroid therapy using prednisolone. After 4 weeks, both IgG and IgG4 values decreased and both the swelling of the pancreas and also the retroperitoneal mass were obviously diminished. CONCLUSION: This is a rare case of autoimmune pancreatitis associated with retroperitoneal fibrosis.

Prolonged, NK cell-mediated antitumor effects of suicide gene therapy combined with monocyte chemoattractant protein-1 against hepatocellular carcinoma.

Tumor recurrence rates remain high after curative treatments for hepatocellular carcinoma (HCC). Immunomodulatory agents, including chemokines, are believed to enhance the antitumor effects of tumor cell apoptosis induced by suicide gene therapy. We therefore evaluated the immunomodulatory effects of a bicistronic recombinant adenovirus vector (rAd) expressing both HSV thymidine kinase and MCP-1 on HCC cells. Using an athymic nude mouse model (BALB/c-nu/nu), primary s.c. tumors (HuH7; human HCC cells) were completely eradicated by rAd followed by treatment with ganciclovir. The same animals were subsequently rechallenged with HCC cells, tumor development was monitored, and the recruitment or activation of NK cells was analyzed immunohistochemically or by measuring IFN-gamma mRNA expression. Tumor growth was markedly suppressed as compared with that in mice treated with a rAd expressing the HSV thymidine kinase gene alone (p < 0.001). Suppression of tumor growth was associated with the elevation of serum IL-12 and IL-18. During suppression, NK cells were recruited exclusively, and Th1 cytokine gene expression was enhanced in tumor tissues. The antitumor activity, however, was abolished either when the NK cells were inactivated with anti-asialo GM1 Ab or when anti-IL-12 and anti-IL-18 Abs were administered. These results indicate that suicide gene therapy, together with delivery of MCP-1, eradicates HCC cells and exerts prolonged NK cell-mediated antitumor effects in a model of HCC, suggesting a plausible strategy to prevent tumor recurrence.

Diagnostic utility of aberrant methylation of tissue factor pathway inhibitor 2 in pure pancreatic juice for pancreatic carcinoma.

The tissue factor pathway inhibitor 2 (TFPI-2) is a Kunitz-type serine proteinase inhibitor. Recently, the aberrant methylation of TFPI-2 was detected frequently in pancreatic carcinoma (PCa) tissues but not in normal pancreatic tissues. We analyzed the aberrant methylation of TFPI-2 in the pure pancreatic juice (PPJ) aspirated endoscopically from patients with various pancreatic diseases. Using the highly sensitive methylation-specific polymerase chain reaction (MSP) and quantitative MSP (Q-MSP) assay, we investigated the aberrant methylation of TFPI-2 in nine human PCa cell lines and in the PPJ from patients with PCa, intraductal papillary mucinous neoplasms (IPMN) and chronic pancreatitis (CP). The incidence of aberrant TFPI-2 methylation was seven (77.8%) of nine PCa cell lines by Q-MSP. In cell lines, the expression of TFPI-2 mRNA by quantitative reverse transcription-polymerase chain reaction showed an inverse correlation to the aberrant methylation of TFPI-2. The incidence of aberrant TFPI-2 methylation in the PPJ was 21 (58.3%) of 36 PCa patients, three (17.6%) of 17 IPMN and one (4.8%) of 21 CP by MSP assay. Using a suitable cut-off value of 2.5 according to the receiver operating characteristic curve, the incidence of aberrant TFPI-2 methylation in the PPJ by real-time MSP was 18 (62.1%) of 29 PCa patients, one (5.1%) of 17 IPMN and three (14.3%) of 21 CP, respectively. The incidence of quantitative TFPI-2 hypermethylation in the PPJ with PCa was significantly higher than that with IPMN (P < 0.001) or CP (P < 0.001). Moreover, the aberrant methylation rate of TFPI-2 in the PPJ was 100%, as observed (6/6) in the PCa patients with liver metastasis, and 86.7% (26/30) in stages IVa + IVb of PCa by Q-MSP assay. These results suggest that promoter methylation of TFPI-2 in the PPJ may be a useful marker in the diagnosis and progression of PCa using an endoscopically feasible approach.

Enhanced antitumor effects of a bicistronic adenovirus vector expressing both herpes simplex virus thymidine kinase and monocyte chemoattractant protein-1 against hepatocellular carcinoma.

The efficacy of the suicide gene therapy using the herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV) system for the treatment of cancer is limited because of the insufficient gene transfer and the low killing activity. To enhance the antitumor activity, we determined whether recombinant adenovirus vector (rAd)s expressing both HSV-tk and monocyte chemoattractant protein-1 (MCP-1) genes could potentiate the destruction of hepatocellular carcinoma (HCC). The rAd Ad-tk-MCP1 harboring HSV-tk and MCP-1 genes in sequence under the universal CAG promoter was constructed with a bicistronic unit including the encephalomyocarditis virus-internal ribosomal entry site. The levels of HSV-tk expression and GCV-sensitive tumoricidal activity of Ad-tk-MCP1 were comparable to those of rAd expressing HSV-tk alone. The growth of subcutaneous tumors in athymic nude mice was markedly suppressed when tumors were treated with Ad-tk-MCP1 as opposed to another bicistronic vector Ad-MCP1-tk, rAd expressing either HSV-tk or MCP-1, or both of these vectors. The antitumor effects of Ad-tkMCP1 may be dependent on the activation of macrophages, since the recruitment of macrophages was observed tumor necrosis factor-alpha production was enhanced in the tumor tissue. Furthermore, the enhanced antitumor effect was abolished by inactivating macrophages with carrageenan treatment. These results demonstrated that a bicistronic rAd harboring both suicide and chemokine genes in sequence exerted the enhanced, macrophage-dependent, antitumor effects in a model of HCC and support the use of this strategy for the treatment of HCC.

Tiny staining spots in liver cirrhosis associated with HCV infection observed by computed tomographic hepatic arteriography: follow-up study.

BACKGROUND: It is important to distinguish small lesions with increased arterial perfusion observed by computed tomographic arteriography (CT-A) from hepatocellular carcinoma (HCC). However, the clinical characteristics and prognosis of such lesions have not been clarified. METHODS: We retrospectively examined 200 patients with cirrhosis related to hepatitis C virus (HCV) infection who had undergone both CT-A and CT arterioportography between 1995 and 1998, and found 80 tiny staining spots (TSS)s, with a diameter of 5-10 mm, by CT-A (35 patients). The mean TSS observation period was 29.0 months. If the major axis was larger than 10 mm and showed a 1.5-fold or more increase, the lesion was regarded as tumor growth (TG). The TSS lesions were divided into two groups according to whether the patient had or did not have HCC. The prognosis of TSS was classified into three groups; HCC-suspected group, nontumor group, and unclassified group, in which TG was negative although transcatheter arterial embolization (TAE) had been performed. RESULTS: Of the 40 TSSs in 14 patients without HCC, 2 (5%) were suspected as HCC. Of the 40 TSSs in 21 patients with HCC, 13 (32.5%) were suspected as HCC. There were no significant differences in the size, position, and morphology of TSSs among the three prognostic groups. Of the 7 TSSs with a high signal intensity on T2-weighted magnetic resonance (MR) images, 5 were in the HCC-suspected group. CONCLUSIONS: We recommend early treatment of TSSs accompanying HCC or showing features of malignancy at the imaging workup.