Masaharu Ohgaki

Overexpression of bax sensitizes human breast cancer MCF‐7 cells to radiation‐induced apoptosis

Resistance to apoptosis plays an important role in tumors that are refractory to chemotherapy and ionizing radiation (IR). bax, which forms a heterodimer with bcl‐2 and accelerates apoptosis, is not, or only weakly, expressed in most human breast cancer cells, and weak bax expression is considered to be related to the resistance of breast cancer cells to apoptosis. bax expression vector was introduced to human breast cancer MCF‐7 cells, which exhibit weak expression of bax, to demonstrate its role of modulating radiation‐induced apoptosis. bax overexpression in MCF‐7 cells by stable transfection does not affect viability by itself, but each stable transfectant was more sensitive to IR than the parental MCF‐7 cells. The degree of enhancement in radiosensitivity was dependent on the expression level of bax. IR upregulated p53 and p21WAFI about 5‐ to 10‐fold and downregulated bcl‐2 and bcl‐XL by 80–90% at 6 hr in both parent and bax stably transfected MCF‐7 cells to the same degree. FACS analysis and DNA electrophoresis revealed that this sensitization was due to apoptosis. We suggest that exogenous bax expression might be one of the factors determining cellular radiosensitivity in MCF‐7 breast cancer cells and may have therapeutic applications for enhancing radiation sensitivity in breast cancer cells.

OVEREXPRESSION OF BAX ENHANCES THE RADIATION SENSITIVITY IN HUMAN BREAST CANCER CELLS

Bax-a, a splice variant ofbax which promotes apoptosis, is expressed in many kinds of untransformed cell lines and breast tissue, whereas only weak or no expression could be detected in breast cancer cell lines and malignant breast tissue. Human breast cancer MCF-7 cells, which have a weakbax gene expression, were stably transfected with pCX2neobax encoding humanbax and neomycin-resistant genes, and two unique clones (MCF-7/bax-1 and MCF-7/bax-2) were thus generated which expressed different levels ofbax-α. Sensitivity to ionizing radiation (IR) was examined and each was more sensitive to IR than the parental MCF-7 cells. The degree of enhancement in radiosensitivity was dependent on the expression level ofbax, and IR was found to induce intranucleosomal DNA fragmentation in stable transfectant but not in parent cells, thus suggesting that this sensitization is due to apoptosis. We suggest that exogenousbax-α expression might therefore be one of the factors determining cellular radiosensitivity in MCF-7 breast cancer cells and may potentially have a therapeutic application by enhancing radiation sensitivity in breast cancer cells.

Role of milky spots as selective implantation sites for malignant cells in peritoneal dissemination in mice

We investigated the significance of milky spots for malignant cells in peritoneal dissemination using three mouse carcinomatous peritonitis models. P388 leukemia and Colon 26 cancer cells were labeled with bromodeoxyuridine (BrdU) and mice were inoculated intraperitoneally. After 24 h the greater omentum and the mesenterium were removed and stained immunohistochemically with anti-BrdU antibody. The labeled cells were found to have preferentially infiltrated into the milky spots in these specimens. Next, using B16 PC melanoma cells, which can be easily distinguished from the other cells by the intrinsic black melanin, the distribution of the melanoma cells was observed macro- and microscopically following intraperitoneal inoculation. The melanoma cells were similarly found to have selectively infiltrated into the milky spots in the omentum and mesenterium after 1 day. Moreover, the melanoma cells were growing and forming distinct metastic lesions within the milky spots 1 week later.

Acute Peritonitis Caused by Intraperitoneal Rupture of an Infected Urachal Cyst: Report of a Case

Abstract.Embryologically, the urachus is the tubular structure that connects the dome of the bladder to the umbilicus. Incomplete obliteration of the urachal lumen results in several anomalies. The most common urachal abnormality is the urachal cyst and, while intraperitoneal rupture of an infected urachal cyst is very rare, acute peritonitis resulting from intraperitoneal rupture is the most dangerous of all complications associated with urachal anomalies. We report the case of an 80-year-old woman who underwent an emergency laparotomy for lower abdominal pain and signs of acute peritonitis, which revealed intraperitoneal rupture of an infected urachal cyst. Infected urachal cysts with intraperitoneal rupture are often misdiagnosed as a common acute abdomen and result in emergency exploratory laparotomy. These patients should be managed by complete excision of the urachal remnant to prevent any malignant change occurring, as malignant changes have been reported.

Selective drug delivery to peri-tumoral region and regional lymphatics by local injection of aclarubicin adsorbed on activated carbon particles in patients with breast cancer???a pilot study

ACR-CH, which consists of aclarubicin (ACR) adsorbed onto activated carbon particles, was developed for locoregional chemotherapy for breast cancer. Thirty patients with breast cancer received an ACR (10 mg) injection intra- and peritumorally, either as ACR-CH or as ACR aqueous solution (ACR-AQ) 5 min before the operation for breast cancer. The ACR concentrations were significantly higher in the peritumoral regions and regional lymph nodes, and were also significantly lower in the blood plasma in patients given ACR-CH versus patients given ACR-AQ

Pharmacological effects of 5-fluorouracil microspheres on peritoneal carcinomatosis in animals.

A new delivery formulation (5FU-MS) of 5-fluorouracil (5FU), 5FU incorporated in microspheres composed of poly(glycolide-co-lactide) matrix, has been developed for the treatment of peritoneal carcinomatosis, and is designed to slowly release the incorporated 5FU for 3 weeks. Intraperitoneal 5FU-MS distributed higher concentrations of 5FU to the intraperitoneal tissues, such as the omentum and the mesentery, for a longer period with lower blood plasma concentrations than did the aqueous 5FU solution in rats. In experiments using mice, the lethal toxicity, determined by the probit method, in 5FU-MS was reduced to less than half that in aqueous 5FU solution. We evaluated the therapeutic effects on peritoneal carcinomatosis induced by the intraperitoneal inoculation of B-16 PC melanoma cells. The therapeutic effects of 5FU-MS were enhanced when compared with both the equivalent doses and same toxicity doses of the aqueous 5FU solution.

Therapeutic effects of the angiogenesis inhibitor TNP-470 against carcinomatous peritonitis in mice.

The therapeutic effects of the new anti-angiogenesis factor TNP-470 were examined against carcinomatous peritonitis in mice. In the first experiment using carcinomatous peritonitis caused by i.p. inoculation of 10(6) M5076 tumor cells, TNP-470 solution was injected i.p. in a bolus of 50 mg/kg body weight into two groups of 10 mice either 1 or 8 days after the i.p. inoculation. The administration of TNP-470 on day 1 extended the survival time of the mice compared with 10 control mice receiving no treatment, whereas TNP-470 given on day 8 did not affect the survival time. In the next experiment on the M5076 tumor, TNP-470 solutions at 100 or 300 mg/kg were injected i.p. in a bolus into two groups of 20 mice 1 day after the inoculation 10(6) tumor cells, respectively. The administration of TNP-470 at 100 mg/kg also had an inhibitory effect. However, TNP-470 at 300 mg/kg caused toxic death in half of the mice. Next, we examined the effects of TNP-470 on another type of carcinomatous peritonitis model, which was caused by i.p. inoculation of 10(6) B16 melanoma cells. In this experiment, TNP-470 solutions in a bolus of 150 mg/kg were injected i.p. into six groups of 10 mice each on day 1 only (group 1), on days 1 and 4 (group 2), on days 1, 4 and 7 (group 3), on day 8 only (group 4), on days 8 and 11 (group 5), or on days 8, 11 and 14 (group 6), respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Prevention of peritoneal metastasis of cancer with dextran sulfate???an experimental study in mice

Peritoneal metastases occur most often in the greater omentum, where tumor implantation sites are densely distributed. We used dextran sulfate (S-Dex) as an anti-cell-adherence agent to prevent i.p. seeded malignant cells from causing peritoneal metastases. S-Dex was tested for its anti-adherent activity against B-16 melanoma cells on plastic, and was examined for its ability to prevent implantation in the omentum and to improve survival in mice after B-16 melanoma was inoculated i.p. S-Dex prevented B-16 melanoma cells from adhering to the plastic wall. S-Dex reduced the number of B-16 melanoma cells implanted into the greater omentum and improved the survival of mice inoculated with B-16 melanoma cells. We conclude that S-Dex attenuated peritoneal metastases when B-16 melanoma cells were seeded i.p.

Methotrexate bound to carbon particles used for treating cancers with lymph node metastases in animal experiments and a clinical pilot study

A new dosage formulation, methotrexate adsorbed onto activated carbon particles (MTX‐CH), was developed for treating cancer patients with lymph node metastases.

Endoscopic local injection of a new drug delivery formulation, anticancer drug bound to carbon particles, for digestive cancers: Pilot study

A new dosage formulation consisting of an anticancer drug bound to activated carbon particles was developed for the treatment of digestive cancer in patients in whom operation is contraindicated. The new formulation is designed to distribute higher levels of anticancer drug to the regional lymph nodes and at the injection site compared to distribution of the drug in aqueous solution. In 12 patients with histologically proven carcinoma (7 with superficial esophageal cancer and 5 with early or proper muscle layer-infiltrating gastric cancer), an anticancer drug bound to carbon particles (total dose, 40–100mg peplomycin or 250–500mg methotrexate per person) was injected endoscopically into the primary lesions. Eleven of the 12 patients are currently alive, 12–64 months after therapy, or they died without evidence of cancer 12–98 months after the treatment. One patient has remained cancer-free for 32 months after a second course of the new formulation therapy given to treat a recurrence detected 26 months after the first treatment. Endoscopic injection of this new dosage formulation seems to control these digestive cancers in patients in whom operation is contraindicated.