Eradication of hepatocellular carcinoma xenografts by radiolabelled, lipiodol-inducible gene therapy.

The promoter region of the early-growth response-1(Egr-1) gene has been shown to be activated by external radiation, thus making a selective tumoricidal effect possible. A previous experiment showed that the Egr-1 promoter can be activated by internal radiation using radioisotopes as well as external radiation. Internal radiation using I-131 lipiodol (I-131-Lip) has been established as one of the most useful therapeutic strategies against hepatoma. We herein linked the Egr-1 promoter to the herpes simplex virus-thymidine kinase (HSV-TK) gene, and investigated its efficacy in hepatoma gene therapy in combination with I-131-Lip. A luciferase assay showed the Egr-1-promoter activity to be markedly increased in hepatoma tissue specimens in an I-131-dose-dependent manner, whereas a less than two-fold increase in this activity was observed in other organs. In addition, the radioactivity derived from I-131 was selectively accumulated in the tumor tissue specimens. To examine the efficacy of EgrTK/ganciclovir (GCV) gene therapy in vivo, subcutaneous hepatoma xenografts in nude mice were transfected using a hemagglutinating virus of Japan (HVJ)-liposome vector. Complete tumor regression was observed in all the EgrTK-transfected tumors following combination treatment with I-131-Lip and GCV 42 days after treatment without any side effects (n=8). In contrast, the tumors continued to grow in all control mice (n=10). Furthermore, the serum alpha-fetoprotein levels decreased in the combination therapy group, while they increased in the controls. In conclusion, these data indicate that Egr-1 promoter-based gene therapy combined with internal radiation has a selective effect on hepatoma tumors while also showing an improved in vivo efficacy. This combination therapy might, therefore, be an effective human hepatoma gene therapy, even in advanced multiple cases.

Attenuated familial adenomatous polyposis associated with advanced rectal cancer in a 16-year-old boy: report of a case.

We herein present a case of attenuated familial adenomatous polyposis (AFAP) with advanced rectal cancer in a 16-year-old boy. His mother and younger brother both had subcutaneous soft tissue tumors in the back and sparse-type colorectal polyposis. His mother also had dental anomalies and gastric fundic gland polyposis. The patient was admitted to our hospital for investigation of bloody stools. Barium enema and colonofiberscopy revealed advanced rectal cancer and sparse (<50) colorectal polyps. He also had dental anomalies, a subcutaneous soft tissue tumor in the back, and gastric fundic gland polyposis as extracolonic manifestations. A total proctocolectomy and ileoanal anastomosis were performed, and histological examination of the resected specimens confirmed moderately differentiated adenocarcinomas of the rectum with metastases to the regional lymph nodes. The other colorectal polyps were tubular adenomas with no evidence of malignancy. Germline mutations in the APC gene were observed in codons 486, 545, 1493, and 1556. This case serves to demonstrate that a total proctocolectomy with ileoanal anastomosis should be the procedure of choice for young patients found to have advanced rectal cancer associated with FAP.

MRI facilitated a diagnosis of endometriosis of the rectum.

A 51-year-old pre-menopausal Japanese woman suffering from chronic lower abdominal pain was referred to our hospital. A barium enema showed a stenotic lesion in the recto-sigmoid region, and a pelvic computed axial tomography (CAT) scan revealed a thickened rectal wall. A colonoscopic examination showed the rectum to be constrictive, but the mucosa appeared to be intact. Magnetic resonance imaging (MRI) with T1 high-intensity revealed a cystic lesion in the thickened wall of the rectum, which led us to suspect possible bowel endometriosis. Part of the biopsy specimen showed endometrial epithelium within the interstitial layer of histologically normal mucosa; finally, endometriosis of the rectum was diagnosed. The patient became asymptomatic after the initiation of hormonal treatment and later experienced spontaneous menopause. MRI was effective for diagnosis and the patient did not undergo unnecessary laparotomy. Although bowel endometriosis is generally diagnosed by means of resected specimens, in our patient, diagnosis was made using MRI and biopsy, and hormonal therapy had an effective role as a bridge to menopause.

The detection of colorectal cancer at an asymptomatic stage by screening is useful.

BACKGROUND/AIMS: The validity of mass screening using fecal occult blood testing remains controversial. In addition, no controlled clinical study has yet been performed to show the usefulness of sigmoidoscopy. The purpose of the present study was to compare the surgical results achieved in asymptomatic patients with colorectal cancer detected by screening with those in symptomatic individuals. METHODOLOGY: A total of 285 patients underwent a surgical resection of colorectal cancer between 1991 and 1997 at our institution. Among them, 233 patients had complaints related to cancer at the time of diagnosis. In contrast, 52 were asymptomatic. In those 52 patients, colorectal cancer had been suspected based on routine screening including fecal occult blood testing, colonoscopy and/or elevated serum levels of carcinoembryonic antigen. RESULTS: Early stage of colorectal cancer was more frequently seen in asymptomatic patients than in symptomatic patients P < 0.01. The survival rates for asymptomatic patients was also superior to those of symptomatic patients P < 0.05. CONCLUSIONS: Screening using fecal occult blood testing, colonoscopy and tumor markers is thus considered to be beneficial for the early detection of colorectal carcinoma, which also tends to demonstrate good surgical results.

Transcatheter arterial embolization for advanced hepatocellular carcinoma resulting in a curative resection: report of two cases.

Transcatheter hepatic arterial embolization and lipiodolization have been reported to be effective palliative treatments for patients with unresectable hepatocellular carcinoma. We experienced 2 patients with advanced hepatocellular carcinoma which were initially considered to be unresectable due to the extreme extension of the primary lesions. Therefore, transcatheter hepatic arterial embolization with lipiodolization were selected as the treatments of choice. Thereafter, these tumors markedly decreased in size and, as a result, curative resections could subsequently be performed. The pathological examination of the resected specimens revealed necrosis and hyaline degeneration in the main tumors. Viable tumor cells, however, still remained adjacent to the main tumors. Such evidence indicated the limited efficacy of transcatheter hepatic arterial embolization with lipiodolization and the necessity of performing surgical treatment in combination with transcatheter hepatic arterial embolization with lipiodolization. Based on these findings, transcatheter hepatic arterial embolization with lipiodolization both appear to be a good mode of therapy for advanced hepatocellular carcinoma, and in selected patients, subsequent surgery can also be considered.

Three amino acid residues determine selective binding of FK506-binding protein 12.6 to the cardiac ryanodine receptor.

FK506-binding protein (FKBP12) has been found to be associated with the skeletal muscle ryanodine receptor (RyR1) (calcium release channel), whereas FKBP12.6, a novel isoform of FKBP, is selectively associated with the cardiac ryanodine receptor (RyR2). For both RyRs, the stoichiometry is 4 FKBP/RyR. Although FKBP12.6 differs from FKBP12 by only 18 of 108 amino acids, FKBP12.6 selectively binds to RyR2 and exchanges with bound FKBP12.6 of RyR2, whereas both FKBP isoforms bind to RyR1 and exchange with bound FKBP12 of RyR1. To assess the amino acid residues of FKBP12.6 that are critical for selective binding to RyR2, the residues of FKBP12.6 that differ with FKBP12 were mutated to the respective residues of FKBP12. RyR2 of cardiac sarcoplasmic reticulum, prelabeled by exchange with [35S]FKBP12.6, was used as assay system for binding/exchange with the mutants. The triple mutant (Q31E/N32D/F59W) of FKBP12.6 was found to lack selective binding to the cardiac RyR2, comparable with that of FKBP12.0. In complementary studies, mutations of FKBP12 to the three critical amino acids of FKBP12.6, conferred selective binding to RyR2. Each of the FKBP12.6 and FKBP12 mutants retained binding to the skeletal muscle RyR1. We conclude that three amino acid residues (Gln31, Asn32, and Phe59) of human FKBP12.6 account for the selective binding to cardiac RyR2.

Isolation of 10 differentially expressed cDNAs in differentiated Neuro2a cells induced through controlled expression of the GD3 synthase gene.

Recently, we showed that transfection of GD3 synthase cDNA into Neuro2a cells, a mouse neuroblastoma cell line, causes cell differentiation with neurite sprouting. In a search for the genes involved in this ganglioside-induced Neuro2a differentiation, we used a tetracycline-regulated GD3 synthase cDNA expression system combined with differential display PCRs to identify mRNAs that were differentially expressed at four representative time points during the process. We report here the identification of 10 mRNAs that are expressed highly at the Neuro2a differentiated stage. These cDNAs were named GDAP1-GDAP10 for (ganglioside-induced differentiation-associated protein) cDNAs. It is interesting that in retinoic acid-induced neural differentiated mouse embryonic carcinoma P19 cells, GDAP mRNA expression levels were also up-regulated (except that of GDAP3), ranging from three to >10 times compared with nondifferentiated P19 cells. All the GDAP genes (except that of GDAP3) were developmentally regulated. The GDAP1, 2, 6, 8, and 10 mRNAs were expressed highly in the adult mouse brain, whereas all the other GDAP mRNAs were expressed in most tissues. Our results suggested that these GDAP genes might be involved in the signal transduction pathway that is triggered through the expression of a single sialyltransferase gene to induce neurite-like differentiation of Neuro2a cells.

Hyperbaric oxygen therapy for intestinal obstruction in children: an exceptional experience in a compromised child.

Intestinal obstruction in a 2-year-old girl with a histologically proven diagnosis of retroperitoneal yolk sac carcinoma developed after the second course of anticancer chemotherapy. Nonoperative treatment was not effective. Because the patient had fallen into a state of chemotherapy-induced myelosuppression, surgery was ruled out. Thus, hyperbaric oxygen therapy was the next treatment of choice. It was performed twice under hyperbaric oxygen conditions at 2.8 atmospheric pressure for 111 minutes. After the procedure, her general status recovered well. The air-fluid level disappeared on the radiograph, and no adverse effects were observed. Later, a surgical removal of the primary tumor was performed successfully, but an intestinal resection was not required. This is the first instance in which we performed hyperbaric oxygen therapy on a child in the management of an intestinal obstruction. Based on the successful outcome in this case, hyperbaric oxygen therapy is suggested to be a useful adjunct to nonoperative therapy for intestinal obstruction when a patient's overall state does not allow operative intervention.

A new inositol 1,4,5-trisphosphate binding protein similar to phospholipase C-delta 1.

We have reported that two inositol 1,4,5-trisphosphate binding proteins, with molecular masses of 85 and 130 kDa, were purified from rat brain; the former protein was found to be the delta 1-isoenzyme of phospholipase C (PLC-delta 1) and the latter was an unidentified novel protein [Kanematsu, Takeya, Watanabe, Ozaki, Yoshida, Koga, Iwanaga and Hirata (1992) J. Biol. Chem. 267, 6518-6525]. Here we describe the isolation of the full-length cDNA for the 130 kDa Ins(1,4,5)P3 binding protein, which encodes 1096 amino acids. The predicted sequence of the 130 kDa protein had 38.2% homology to that of PLC-delta 1. Three known domains of PLC-delta 1 (pleckstrin homology and putative catalytic X and Y domains) were located at residues 110-222, 377-544 and 585-804 with 35.2%, 48.2% and 45.8% homologies respectively. However, the protein showed no PLC activity to phosphatidylinositol 4,5-bisphosphate and phosphatidylinositol. The 130 kDa protein expressed by transfection in COS-1 cells bound Ins(1,4,5)P3 in the same way as the molecule purified from brain. Thus the 130 kDa protein is a novel Ins(1,4,5)P3 binding protein homologous to PLC-delta 1, but with no catalytic activity. The functional significance of the 130 kDa protein is discussed.

A comparative study on effects of doxorubicin alone or mixed with lipiodol given through the hepatic artery or portal vein for liver tumors in rats.

Vascular feeding of metastatic liver tumors at early stage is uncertain. It is controversial whether anticancer agents should be given through the hepatic artery or portal vein. In order to clarify this point, a rat model of liver metastases generated by an intraportal injection of syngeneic tumor cells was used to determine the optimal regional chemotherapeutic modality for early hepatic metastases. The rats given the tumor cells through the portal vein were placed into 5 groups: In groups I and II, Adriamycin (ADR) 4 mg/kg alone was given either into the hepatic artery or into the portal vein, respectively, 24 h after the inoculation of tumor cells. In groups III and IV, ADR mixed with lipiodol (lipiodolized ADR) 4 mg/kg was given into the hepatic artery or into the portal vein, respectively, 24 h after inoculation. For the group V rats, no treatment was given after inoculation of the tumor. When a comparison was made with regard to the forms of anticancer drug administered, statistically significant differences in survival rates were recognized between groups I and III (p < 0.001), and groups II and IV (p < 0.05). The anticancer agent not mixed with lipiodol and given through the hepatic artery had a more preventive effect than that given through the portal vein. Thus, we conclude that administration of ADR not mixed with lipiodol and given through the hepatic artery is the preferred modality for treating early metastatic liver tumors.

Hyperbaric oxygen therapy in the treatment of refractory peripancreatic abscess associated with severe acute pancreatitis.

Five patients with peripancreatic abscesses associated with severe acute pancreatitis were treated by hyperbaric oxygen therapy (HBO). In 3 patients, the course after surgical mobilization of the pancreas and drainage of the pancreas bed was complicated by peripancreatic abscesses. HBO was conducted under a pressure of 2.8 atmospheres for two hours daily. Four of the 5 patients showed a progressive improvement in their condition. In one patient who failed to respond despite seven sessions of HBO, Pseudomonas aeruginosa was isolated from the discharge, and resection of necrotic tissue and drainage were performed. The main effects of HBO were the alleviation of high spiking fever, the improvement of white blood cell count and serum amylase levels, and the reduction of the abscess size. We recognized HBO to be a successful treatment for peripancreatic abscess associated with severe acute pancreatitis and better results were obtained than in cases that did not receive HBO.

Effect of 2-buten-4-olide, an endogenous suppressant of feeding, on reproductive function in rats.

The effects of 2-buten-4-olide, an endogenous feeding suppressant, on the estrous cycle and LH secretion were studied to determine the influence of this compound on reproductive function. Estrous cycling female Wistar rats were treated ip with 2-buten-4-olide (0, 30 or 100 mg.kg-1.day-1) for 2 weeks. Treatment with 100 mg.kg-1.day-1 delayed the estrous cycle. 2-Buten-4-olide increased the pituitary content of LH (1651.3 +/- 164.4 vs 927.7 +/- 65.1 ng/pituitary; p less than 0.01), and decreased the serum LH level compared with the control level in diestrus (0.16 +/- 0.01 vs 0.26 +/- 0.03 microgram/l; p less than 0.05). However, it did not affect the GnRH content of the mediobasal hypothalamus. The direct effects of 2-buten-4-olide on the pituitary response to GnRH was examined by perifusing the pituitary. Medium containing 2-buten-4-olide (10(-4) mol/l) suppressed the pituitary response to GnRH (2 micrograms/l) (percent increase at 50 min after start of GnRH stimulation: 180 +/- 47 vs 406 +/- 66%; p less than 0.05). These findings suggest that 2-buten-4-olide is involved in the regulation of pituitary gonadotropin secretion directly by suppressing the pituitary responsiveness to GnRH, and that 2-buten-4-olide may play an important role in starvation-induced anestrus.

Lipiodol drug targeting in the treatment of primary hepatocellular carcinoma.

When Lipiodol Ultra-Fluid (lipiodol) is administered through arteries feeding the tumor, lipiodol remains selectively in the neoplastic tissue for an extended period of time. This characteristic of the oily contrast medium has been utilized for regional targeting of chemotherapy in patients with hepatocellular carcinoma, which has been termed "lipiodolization". The current status of lipiodolization is reviewed herein.

Lipiodolization prolongs survival rates in postoperative patients with a recurrent hepatocellular carcinoma.

Curative resections were performed in 200 patients with primary hepatocellular carcinoma (HCC) during the period from 1971 to 1988, at Kyushu University Hospital. Patients seen before 1982 numbered 73 (group I), and the remaining 127 underwent hepatic resection after 1983 (group II). Thirty-nine (53.4%) of the 73 patients in group I had a recurrence during the follow-up period, as did 61 (48.0%) in group II. HCC resections done after 1983 (group II) were associated with (1) a low incidence of operative deaths (I 15.1% vs. II 2.4%) and hospital deaths (I 30.1% vs. II 5.5%), (2) relatively well-preserved hepatocellular function (indocyanine green test, I 24.5 +/- 12.6% vs. II 17.4 +/- 9.2%), (3) a low incidence of histologically verified concomitant cirrhosis (I 87.7% vs. II 68.5%) (4), and smaller HCC nodules (I 6.4 +/- 4.4 cm vs. II 4.1 +/- 3.1 cm). The survival rates of patients seen after 1983 were significantly better than those of patients resected prior to 1982. With respect of the recurrence-free rates achieved by curative hepatic resection, there was no statistically significant difference to be seen between the two groups. In the case of patients submitted of lipiodolization treatment for a recurrent HCC, the survival rates were better than in patients on systemic chemotherapy. Since lipiodolization prolongs survival time after a recurrence of HCC, this form of treatment should be given due consideration.

[Eight cases of non-resectable hepatocellular carcinoma surviving for more than 3 years after Lipiodol administration].

From December 1982 to February 1986, selective regional cancer chemotherapy using Lipiodol plus anticancer drug (Lipiodolization) was prescribed for 87 patients with non-resectable hepatocellular carcinoma (HCC). Eight out of 87 cases survived for more than 3 years. The mean age of the 8 long survivors was 52.6. In the liver function, 2 cases were in Child A, 5 cases in Child B, and 1 case in Child C. The largest diameter of HCCs was 5.3 cm, and the number of the tumors was one in 6 cases, two in one case, and three in one case. No vascular invasion was detected on hepatic angiography. These findings suggested that long survivors in lipiodolization are not in far advanced HCCs. For these 8 cases, lipiodolization was repeatedly prescribed from two to 6 times. The largest amount of adriamycin was 192 mg/case. The longest survivor has lived for 5 years and 6 months after first lipiodolization. After lipiodolization, one tumor vanished, and three tumors decreased in size. Although seven tumors increased in size after lipiodolization, the tumor doubling time of seven tumors were 64, 265, 313, 317, 350, and 1943 days (average 539 +/- 639 days). It is possible that lipiodolization remarkably inhibited tumor growth. Nevertheless, in 5 out of 7 cases, the daughter lesions increased in size and number, although the main tumors grew slowly. Lipiodolization was less effective for newly arising daughter lesions than main lesions.

A 5-year experience of lipiodolization: selective regional chemotherapy for 200 patients with hepatocellular carcinoma.

Between 1982 and 1987, selective regional cancer chemotherapy using Lipiodol plus an anticancer drug (lipiodolization) was prescribed for 200 patients with hepatocellular carcinoma. One hundred forty-nine patients were given lipiodolization alone, and the remaining 51 underwent hepatic resection following lipiodolization. The grades of deposits of the oily contrast medium in the neoplastic tissue seen on the plain X-ray correlated well with the antitumor effect. In the resected specimens of 17 patients treated with lipiodolization prior to surgery, concentrations of adriamycin in the malignant liver tissues were 13.2 +/- 18.2 micrograms per gm, whereas the adjacent liver parenchyma contained 1.4 +/- 2.0 micrograms per gm of adriamycin; the difference was statistically significant (p less than 0.05). In the 149 patients treated with lipiodolization, 1-, 2-, 3- and 4-year survival rates were 56.1, 28.9, 17.3 and 7.4%, respectively. Thus, lipiodolization was considerably more effective, compared to the results achieved by hepatic artery ligation and cannulation into the hepatic artery for patients in Stages I and II. In this sequential nonrandomized study, the survival rates for patients undergoing hepatic resection were superior to those for patients in Stage I and treated with lipiodolization. The significant difference appeared to depend on incomplete killing of tumor cells, which are most often present in the fibrous capsule, by lipiodolization. We conclude from these data that lipiodolization is an effective treatment for hepatocellular carcinoma when the tumor is not curatively resectable. When the clinical status is good, then surgery is warranted.

Selective effect of doxorubicin suspended in lipiodol on VX2 carcinoma in rabbits.

When Lipiodol Ultra-Fluid (Lipiodol) was given through arteries feeding the tumor, this material was retained in the tumor. After mixing Lipiodol and doxorubicin (adriamycin; ADR), the effect of this antitumor agent on VX2 carcinoma inoculated into the hindlimb in 57 rabbits was investigated. Size of the tumor remarkably decreased in the rabbits treated with ADR 2 mg/kg suspended in Lipiodol and given through the femoral artery. Intraarterial injection of ADR 2 mg/kg alone led to a transient decrease in size of the tumor, but there was a regrowth. The result was comparable to a finding in a group in which half the dose of ADR (1 mg/kg) suspended in Lipiodol was given intraarterially. When ADR 2 mg/kg mixed with Lipiodol was injected directly into the tumor, growth of the tumor was not completely inhibited. Lipiodolized ADR has a prominent antitumor effect, and Lipiodol has the potential to be applied as a carrier of anticancer drugs.

Antitumor effect of lipiodolized doxorubicin on VX2 carcinoma in rabbits.

Twelve-day-old VX2 carcinoma was inoculated in the hind leg of 24 rabbits and, after 12 days, Doxorubicin (Adriamycin) suspended in Lipiodol Ultra-Fluid (Lipiodol) was then given through the femoral artery. A selective deposit of the contrast material in the tumor for an extended time was evident on the x-rays and the antitumorous effect was remarkable. Lipiodolized antitumor agents warrant further investigation for possible clinical application.