Establishment of a New Scirrhous Gastric Cancer Cell Line with FGFR2 Overexpression, OCUM-14.

BACKGROUND: The prognosis of scirrhous gastric carcinoma (SGC), which is characterized by rapid infiltration and proliferation of cancer cells accompanied by extensive stromal fibrosis, is extremely poor. In this study, we report the establishment of a unique SGC cell line from a gastric cancer patient in whom an autopsy was performed. METHODS: A new SGC cell line, OCUM-14, was established from malignant ascites of a male patient with SGC. A postmortem autopsy was performed on the patient. Characterization of OCUM-14 cells was analyzed by microscopic examination, reverse transcription polymerase chain reaction, fluorescence in situ hybridization analysis, immunohistochemical examination, CCK-8 assay, and in vivo assay. RESULTS: OCUM-14 cells grew singly or in clusters, and were floating and round-shaped. Most OCUM-14 cells had many microvilli on their surfaces. The doubling time was 43.1 h, and the subcutaneous inoculation of 1.0 × 107 OCUM-14 cells into mice resulted in 50% tumor formation. mRNA expressions of fibroblast growth factor receptor 2 (FGFR2) and human epidermal growth factor receptor 2 (HER2) were observed in OCUM-14 cells. FGFR2, but not HER2, overexpression was found in OCUM-14 cells. The heterogeneous overexpression of FGFR2 was also found in both the primary tumor and metastatic lesions of the peritoneum, lymph node, bone marrow, and lung of the patient. The FGFR2 inhibitors AZD4547 and BGJ398 significantly decreased the growth of OCUM-14 cells, while paclitaxel and 5-fluorouracil significantly decreased the proliferation of OCUM-14 cells, but cisplatin did not. CONCLUSION: A new gastric cancer cell line, OCUM-14, was established from SGC and showed FGFR2 overexpression. OCUM-14 might be useful for elucidating the characteristic mechanisms of SGC and clarifying the effect of FGFR2 inhibitors on SGC.

[A Hematogenous Metastasis from Scirrhous Gastric Cancer to the Uterus].

A 46-year-old woman was referred to our hospital because of nausea. Endoscopy revealed scirrhous gastric cancer, and abdominalcomputed tomography revealed peritonealdissemination. She was diagnosed with Stage IV gastric cancer and treated with S-1 plus CDDP combination chemotherapy. After 4 courses of chemotherapy, the primary tumor and peritoneal dissemination were considered clinically stable, but the uterus grew rapidly. She was diagnosed as having uterine metastasis based on cervicaland endometrialsmear class V cytology. As the chemotherapy was not effective for the uterine lesions, totalhysterectomy and bilateralsal pingo-oophorectomy were performed. Histological findings showed a poorly differentiated cancer with vascular emboli. Uterine metastases are an important consideration in women with scirrhous gastric cancer, and we recommend palliative hysterectomy for chemotherapy-resistant metastases if the primary tumor and other metastases are controlled.

Effects of 5-fluorouracil, adriamycin and irinotecan on HSC-39, a human scirrhous gastric cancer cell line.

Therapies for patients with scirrhous gastric cancer remain ineffective. Current treatments for gastric cancer based on systemic therapy, such as the combination of S-1 with cisplatin or docetaxel, show good clinical response rates. S-1 plus cisplatin is the standard treatment for HER2-negative advanced scirrhous gastric cancer in Japan. In spite of recent advances in the treatment of gastric cancer, a standard chemotherapy regimen is yet to be established for scirrhous gastric cancer. To develop new therapeutic approaches based on characteristic biological features of cancer cells, we examined the mechanisms underlying the cytotoxicity of anticancer drugs and reactive oxygen species (ROS) toward a human scirrhous cancer cell line, HSC-39, in vitro. Anticancer drugs such as 5-fluorouracil (5-FU), adriamycin (ADR) and irinotecan (CPT-11), as well as ROS, were previously shown to have important cytotoxic effects on these tumor cells. We demonstrated that 5-FU effectively induced apoptosis in HSC-39 cells in a dose­dependent manner, while ADR and CPT-11 induced necrosis and/or aponecrosis. 5-FU effectively inhibited WST-1 decrease in the MTT viability assay, even at low doses where little LDH release was observed, while ADR and CPT-11 only inhibited WST-1 decrease at high doses where LDH release was induced. Moreover, HSC-39 cells showed high sensitivity to H2O2 and NOC-18, but less sensitivity to other ROS, suggesting a link between cell damage and membrane permeability changes induced by H2O2 and NOC-18 or related oxygen radical species such as OH· or ·O2. These results suggest that combination treatment of chemotherapeutics with a fluoropyrimidine such as 5-FU is effective chemotherapy for scirrhous gastric cancer.

[A Patient Who Underwent Surgery after CR to Chemotherapy for Scirrhous Gastric Cancer Suspected because of Lymphangitis Carcinomatosa].

A 58-year-old woman was diagnosed with scirrhous gastric cancer suspected because of lymphangitis carcinomatosa. She was treated with 10 courses of S-1+CDDP chemotherapy. After 3 years and 6 months, CR was obtained and she underwent curative total gastrectomy with D2 lymph node dissection plus resection of the spleen and transverse colon. The pathological results of the resected specimen were tub2>por, pT1a, N1 (No.7), M0, CY0, P0, and HER2 (3+). After surgical treatment, supraclavicular lymph node metastasis occurred, and the patient underwent trastuzumab+capecitabine therapy, which resulted in CR for 1 year and 6 months. Thus, for unresectable scirrhous gastric cancer, multidisciplinary therapy such as longterm chemotherapy including trastuzumab and surgery is useful.

Impact of S-1 plus Cisplatin Neoadjuvant Chemotherapy on Scirrhous Gastric Cancer.

OBJECTIVE: This retrospective study aimed to address the therapeutic outcome for scirrhous gastric cancer patients by evaluating the effect of neoadjuvant chemotherapy prior to gastrectomy. METHODS: Two cycles of a 3-week regimen of fluoropyrimidine S-1 (40 mg/m(2), orally, twice daily), together with cisplatin (60 mg/m(2), intravenously, day 8), were administered to patients, separated by a 2-week rest period. Surgery was performed 3 weeks later in the neoadjuvant group (n = 27). We retrospectively evaluated overall survival and prognostic factors in these patients. RESULTS: Univariate analysis showed that positive lavage cytology indicated significantly worse prognoses. In the 15 patients who also underwent curative gastrectomies after S-1 plus cisplatin chemotherapy, the pathological response grade was a significant prognostic factor for 5-year survival. Additionally, lymph node metastasis tended to be an adverse prognostic factor. CONCLUSION: After S-1 plus cisplatin neoadjuvant chemotherapy, a grade 2-3 pathological response may predict favorable outcomes in scirrhous gastric cancer patients receiving curative gastrectomy, but further studies are needed to confirm these results.

[A case of stage IV breast cancer successfully treated over 1 year with eribulin as fourth-line systemic chemotherapy].

A 59-year-old woman with an exudative, reddish bilateral breast tumor and dyspnea visited our hospital. She was diagnosed as having scirrhous carcinoma with metastasis to the liver and pleural effusion, designated as Stage IV, hormone receptor positive, and human epidermal growth factor receptor-2 (HER2) negative. Systemic chemotherapy (3 regimens) and endocrine therapy (5 regimens) were administered for a total of 5 years 7 months. Eribulin was administered as fourth line systemic chemotherapy. The pleural effusion reduced and dyspnea improved. Her status was maintained for 1 year 3 months. This case suggests that eribulin may provide long-term survival and maintenance of quality of life (QOL) in metastatic breast cancer patients.

Neoadjuvant chemotherapy for gastric cancer in Japan: a standing position by comparing with adjuvant chemotherapy.

Adjuvant chemotherapy aims to eradicate micrometastatic tumor cells before and after curative surgery. Many Phase III trials have been conducted to study the efficacy of postoperative adjuvant chemotherapy; however, most trials have failed to show any survival benefit because of their low statistical power and/or poor patient compliance. Since 2000, two pivotal Phase III trials, the ACTS-GC and the CLASSIC, have demonstrated the efficacy of postoperative adjuvant chemotherapy following D2 gastrectomy. Although treatment with S-1 for 1 year or combination therapy with capecitabine and oxaliplatin for 6 months is effective, more intensive chemotherapy is necessary to further improve the survival rates. In Europe, two Phase III trials, the MAGIC and the FNCLCC/FFCD, have produced results that strongly suggest that neoadjuvant chemotherapy is beneficial. The advantages of neoadjuvant chemotherapy include a high rate of R0 resection, tumor regression, high compliance and the avoidance of unnecessary surgery. The disadvantage of neoadjuvant chemotherapy is over-diagnosis. In Japan, the Japan Clinical Oncology Group has conducted several clinical trials using neoadjuvant chemotherapy to target extensive nodal disease and/or scirrhous carcinomas. The optimal courses and regimens of neoadjuvant chemotherapy should, therefore, be clarified in the future.

[Synchronous male bladder cancer and breast cancer - a case report].

A 74-year-old man complained of blood in his urine over a 1-week period beginning in early October 2013, and was examined in the urology department of our hospital. A thorough examination revealed bladder cancer, and surgery was planned after two cycles of preoperative gemcitabine plus cisplatin chemotherapy. A chest computed tomography (CT) performed to evaluate the response to chemotherapy revealed a mass in the right breast. The patient had previously complained about the same site, and mammography and ultrasonography had suggested the possibility of a malignant mammary gland tumor. The results of aspiration cytology were Class V, and based on that finding, a diagnosis of cancer of the right breast was made. In February 2014, we performed a mastectomy, while preserving the pectoral muscles, along with sentinel node biopsy, total cystectomy, urethrectomy, pelvic lymph node dissection, and ureteroileal anastomosis. The histopathological diagnosis of the right breast tumor was invasive ductal carcinoma[scirrhous carcinoma, ly (+), v (-), g (+), f (+), s (+), nuclear grade 1=atypia 2+mitosis 1, EIC (-), ICT (-), NCAT (-)]. A micrometastatic tumor measuring approximately 1mm was observed in the sentinel lymph node. The breast disease was classified as pT1N1mi(sn)M0, Stage IIA, and the tumor was ER (+), PgR (+), HER2/neu (2+), and FISH (-). The bladder cancer was diagnosed as urothelial carcinoma, non-papillary, invasive G2>G3, pT2a; no pelvic lymph node metastases were detected, and it was classified as pT2aN0M0, Stage II. Synchronous male breast cancer and bladder cancer is a very rare condition, and we report the case with a review of the literature.

[A case report of scirrhous gastric cancer with peritoneal dissemination treated effectively with various chemotherapy regimens after laparoscopic distal subtotal gastrectomy].

We report a 44-year-old male patient who was diagnosed with scirrhous gastric cancer with peritoneal dissemination using laparoscopy. The patient underwent a non-curative resection with laparoscopic distal subtotal gastrectomy. In addition, we placed a port into the patient's abdomen for intraperitoneal chemotherapy administration. Postoperatively, we administered capecitabine (per os)+ paclitaxel (intraperitoneally) and, after 2 cycles, the oral anticancer agent 5-FU was given. The patient died of peritonitis carcinomatosa 25 months after the operation. The combined therapies contributed to improve the quality of life, specifically oral ingestion, for 2 years.

Paracrine activation of MET promotes peritoneal carcinomatosis in scirrhous gastric cancer.

Scirrhous gastric cancer is associated with abundant stroma and frequently develops into peritoneal carcinomatosis with malignant ascites. Although malignant ascites is among the most deadly diseases worldwide, its molecular pathogenesis is poorly understood. We investigated the role of hepatocyte growth factor (HGF) in the production of peritoneal carcinomatosis with malignant ascites. We examined three scirrhous and three non-scirrhous human gastric cancer cell lines for the production of peritoneal carcinomatosis in vivo and responses to HGF in vitro. Furthermore, clinical scirrhous gastric cancer specimens were examined for HGF production. Among the six cell lines examined, only two scirrhous cell lines (NUGC4 and GCIY) produced peritoneal carcinomatosis with massive ascites after intraperitoneal injection in nude mice. Their proliferation was stimulated by exogenous HGF in vitro. On the other hand, a non-scirrhous cell line, MKN45, with MET amplification generated peritoneal tumors but not ascites. MET tyrosine kinase inhibitors, crizotinib and TAS-115, inhibited HGF-stimulated proliferation of NUGC4 and GCIY as well as constitutive proliferation of MKN45. Furthermore, crizotinib and TAS-115 prolonged the survival of mice bearing established tumors by NUGC4 or MKN45. In clinical specimens, HGF was markedly produced by stromal fibroblasts. Malignant ascitic fluids from patients with peritoneal carcinomatosis contained high levels of HGF. Our results strongly suggest that paracrine HGF-induced activation of MET-mediated signaling pathways plays an important role in the pathogenesis of peritoneal carcinomatosis in scirrhous gastric cancer. Thus, MET signaling pathway may be a potential therapeutic target for peritoneal carcinomatosis of gastric cancer, even without MET amplification.

Hepatitis B virus reactivation in adjuvant chemotherapy for breast cancer.

Immunosuppressive therapy, such as chemotherapy or the use of corticosteroids, may stimulate reactivation of hepatitis B virus (HBV). Most of these episodes occur in patients whose hepatitis B surface antigens are positive (HBsAg+). We report a case of HBV reactivation in a patient with negative HBsAg during chemotherapy for breast cancer, in spite of avoiding corticosteroids. A 68-year-old woman received adjuvant chemotherapy for breast cancer. Her serological examinations showed that HBsAg, HBeAg, and HBV-DNA were all negative. Her chemotherapy consisted of CAF (cyclophosphamide 500 mg/m(2), doxorubicin 50 mg/m(2), fluorouracil 500 mg/m(2)) without administration of corticosteroids. She received six cycles of CAF. At day 27 after her sixth CAF, she was admitted to the hospital with acute hepatitis B virus (HBV) reactivation. She received glycyrrhizinic acid by intravenous injection (80 mg/day), ursodeoxycholic acid (300 mg/day), and entecavir (0.5 mg/day). Then she received interferon by intravenous injection (3 million units/day), prednisolon by intravenous injection (45 mg/day), and plasma exchange therapy. However, she died of liver failure. This is a rare case in which HBV reactivation occurred in an HBsAg negative patient during chemotherapy without using corticosteroids. This episode suggests that HBV reactivation may occur during chemotherapy in any patient with a history of HBV infection. Therefore, we recommend checking HBsAg, HBsAb, and HBcAb before starting chemotherapy. Moreover, with positive HBsAb or/and HBcAb patients, HBV-DNA should be checked before starting chemotherapy and monitored during chemotherapy by a sensitive PCR method.

[A case of acute appendicitis which occurred during chemotherapy for breast cancer].

The patient was a 66-year-old woman with left breast cancer who underwent left segmental mastectomy with sentinel lymph node biopsy. The histopathological diagnosis was estrogen receptor-positive (ER+), progesterone receptor-positive( PgR+), human epidermal growth factor receptor-2-equivocal( HER2()2+)( with no HER2 gene amplification by fluorescence in-situ hybridization analysis) invasive ductal carcinoma (scirrhous carcinoma) with Ki-67 expression of less than 10% (pathological T1c, N0, M0, stage I). The patient requested chemotherapy, and 4 cycles of docetaxel plus cyclophosphamide (TC) were scheduled. Fever and epigastric pain developed on day 13 of cycle 2. On day 22, the patient was examined before the third cycle of TC, and right lower abdominal pain was reported. Computed tomography revealed appendicitis and an intraperitoneal abscess. She was admitted to the hospital and underwent partial ileocecal resection. The patient was discharged on the 12th postoperative day with no further complications. Acute abdomen during chemotherapy for malignant tumors has been reported sporadically in patients with leukemia. A diagnosis of acute abdomen in patients undergoing cancer treatment requires careful assessment of gastrointestinal symptoms such as nausea and vomiting during chemotherapy, fever associated with granulocytopenia, and findings indicative of local inflammation. The patient in this case recovered uneventfully because imaging studies and surgery were performed promptly after presentation.

[A case of metastatic breast cancer with bilateral hydronephrosis effectively treated with capecitabine].

A 54-year-old female was diagnosed with invasive ductal carcinoma (pT3N1M0, Stage IIIA, estrogen receptor positive [ER (+)], progesterone receptor positive [PgR (+)], human epidermal growth factor receptor type 2 [HER2] score 0) and was treated by preoperative chemotherapy with weekly paclitaxel followed by 5-fluorouracil(5-FU) plus epirubicin plus cyclophosphamide regimen(FEC). Partial mastectomy with axillary dissection was performed. The pathological examination disclosed that the tumor was scirrhous carcinoma, and a pathological partial response was achieved by chemotherapy. Multiple bone metastases were detected 18 months after the surgery during treatment with letrozole as adjuvant therapy. Retroperitoneal metastases with hydronephrosis and a lung metastasis were detected 28 months after the surgery, even though exemestane and zoledronate were administrated after detection of the bone metastases. Chemotherapy with capecitabine was started and she recovered from hydronephrosis 4 months after the start of treatment. After 32 months from the first treatment with capecitabine, the patient is presently alive without hydronephrosis due to continued chemotherapy.

[A complete response of scirrhous gastric carcinoma treated with trastuzumab combination therapy].

Trastuzumab is the first molecular-target medicine for gastric cancer in acknowledgment of the effectiveness and safety in a randomized controlled trial for the HER2-positive gastric cancer. The HER2-positive rate of gastric cancer tends to be high for a well-differentiated adenocarcinoma. We report a case of HER2-positive scirrhous gastric carcinoma we treated, for whom a complete response was obtained by trastuzumab combination therapy. He was a 62-year-old man. In April, 2006, he was diagnosed with metastatic scirrhous gastric carcinoma(mainly poorly-differentiated adenocarcinoma). We become clear with HER2 strong positive in a pathology tissue and started capecitabine+cisplatin+trastuzumab therapy. We confirmed the disappearance of the lesion at the 10th cycle and judged it to be a complete response. The HER2 of advanced gastric cancer must be screened immediately without asking patients for their background or their clinical and pathologic features.

[Mechanisms responsible for the progression of scirrhous gastric cancer].

Scirrhous gastric carcinoma is characterized by rapid cancer cell infiltration and proliferation accompanied by extensive stromal fibrosis. The proliferative and invasive ability of scirrhous gastric cancer cells are closely associated with the growth factors, FGF7 and TGFbeta produced by organ-specific fibroblasts. Peritoneal fibroblasts morphologically change mesothelial cells, and stimulate the migratory capability of cancer cells. A FGFR2 phosphorylation inhibitor prolongs the survival of mice with peritoneal metastasis of scirrhous gastric cancer. A TGFbetaR inhibitor decreases the growth of fibroblast, and invasion-stimulating activity of fibroblasts on cancer cells. A FGFR2 phosphorylation inhibitor or TGFbetaR inhibitor appears therapeutically promising in scirrhous gastric carcinoma.

Polymeric micelles incorporating (1,2-diaminocyclohexane)platinum (II) suppress the growth of orthotopic scirrhous gastric tumors and their lymph node metastasis.

Nano-scaled drug carriers have great potential for the treatment of solid tumors. Nevertheless, hypovascularity and fibrosis in some types of solid tumors have been demonstrated to reduce the penetration and accumulation of nano-scaled drug carriers. Diffuse-type scirrhous gastric cancers present such characteristics as well as frequent metastasis to the lymph nodes; therefore, it remains a great challenge to eradicate scirrhous gastric cancers based on the drug targeting using nanocarriers. Herein, we demonstrated that polymeric micelles with 30-nm diameter incorporating (1,2-diaminocyclohexane)platinum(II) (DACHPt), the parent complex of the anticancer drug oxaliplatin, efficiently penetrated and accumulated in an orthotopic scirrhous gastric cancer model, leading to the inhibition of the tumor growth. Moreover, the elevated localization of systemically injected DACHPt-loaded micelles in metastastic lymph nodes reduced the metastatic tumor growth. These results suggest DACHPt-loaded micelles as a promising nanocarrier for the treatment of scirrhous gastric cancers and their lymphatic metastases.

[A case report of scirrhous gastric carcinoma diagnosed by staging laparoscopy].

The case is a sixty-something man with a complication of epigastric abdominal pain. X-ray and endoscopic examination of upper gastrointestinal tract showed a rigidity of the gastric wall and the presence of giant folds on gastric body and fornix. Scirrhous type of gastric cancer was suspected and gastric forceps biopsy was performed at many points under the retreated endoscopic examinations. However, the histologic findings revealed no malignant features. We performed staging laparoscopy, and by peritoneal washing cytology and biopsy of a nodule of abdominal wall, he was diagnosed with advanced gastric carcinoma with peritoneal dissemination. In conclusion, staging laparoscopy is one of the useful methods for diagnosis and determination of the management of scirrhous type gastric carcinoma.

Efficacy of RAD001 (everolimus) against advanced gastric cancer with peritoneal dissemination.

Peritoneal dissemination occurs frequently in patients with unresectable advanced-stage gastric cancer. In this study, we tested the efficacy of the mTOR inhibitor RAD001 (everolimus) against advanced gastric cancer with peritoneal dissemination. Using the two cell lines, 58As1, a cell line exhibiting a high propensity for peritoneal metastasis, and its parental cell line, HSC-58, a human scirrhous gastric cancer cell line, we first examined the growth-inhibitory activity of everolimus in vitro. Methylene blue assay demonstrated a moderate inhibitory effect of the drug on both cell lines under normal culture conditions (maximal inhibitory effect: 50.5% at 1 µM, HSC-58, 65.3%, 58As1). However, under the hypoxic condition (1% O(2)), while the growth-inhibitory activity of everolimus was greatly reduced in the HSC-58 cell line, the degree of reduction of the inhibitory activity was much smaller in the 58As1 cell line. Western blotting revealed that the degree of phosphorylation of mTOR and its downstream signaling molecules, p70S6K and 4E-BP1, was decreased under hypoxic conditions in HSC-58. On the other hand, phospho-p70S6K and phospho-4E-BP1 remained active under hypoxic conditions in 58As1, the molecular activity was suppressed by everolimus. Cell-cycle analysis showed that hypoxia-induced G1 arrest was not manifested in the 58As1 cells, unlike in the HSC-58 cells. Separately, an in vivo orthotopic mouse model of 58As1 revealed that everolimus significantly reduced peritoneal dissemination as evaluated by the quantitative photon counting method. Taken together, our results suggest that everolimus may have favorable activity against gastric cancer, particularly in cases with peritoneal dissemination.