Targeting N-cadherin increases vascular permeability and differentially activates AKT in melanoma.

OBJECTIVE: We investigate the mechanism through which N-cadherin disruption alters the effectiveness of regional chemotherapy for locally advanced melanoma. BACKGROUND: N-cadherin antagonism during regional chemotherapy has demonstrated variable treatment effects. METHODS: Isolated limb infusion (ILI) with melphalan (LPAM) or temozolomide (TMZ) was performed on rats bearing melanoma xenografts after systemic administration of the N-cadherin antagonist, ADH-1, or saline. Permeability studies were performed using Evans blue dye as the infusate, and interstitial fluid pressure was measured. Immunohistochemistry of LPAM-DNA adducts and damage was performed as surrogates for LPAM and TMZ delivery. Tumor signaling was studied by Western blotting and reverse-phase protein array analysis. RESULTS: Systemic ADH-1 was associated with increased growth and activation of the PI3K (phosphatidylinositol-3 kinase)-AKT pathway in A375 but not DM443 xenografts. ADH-1 in combination with LPAM ILI improved antitumor responses compared with LPAM alone in both cell lines. Combination of ADH-1 with TMZ ILI did not improve tumor response in A375 tumors. ADH-1 increased vascular permeability without effecting tumor interstitial fluid pressure, leading to increased delivery of LPAM but not TMZ. CONCLUSIONS: ADH-1 improved responses to regional LPAM but had variable effects on tumors regionally treated with TMZ. N-cadherin-targeting agents may lead to differential effects on the AKT signaling axis that can augment growth of some tumors. The vascular targeting actions of N-cadherin antagonism may not augment some regionally delivered alkylating agents, leading to a net increase in tumor size with this type of combination treatment strategy.

SRC family kinase inhibition as a novel strategy to augment melphalan-based regional chemotherapy of advanced extremity melanoma.

BACKGROUND: Src kinase inhibition has been shown to augment the efficacy of chemotherapy. Dasatinib, a dual Src/Abl kinase inhibitor approved for the treatment of CML, is under investigation as monotherapy for tumors with abnormal Src signaling, such as melanoma. The goal of this study was to determine if Src kinase inhibition using dasatinib could enhance the efficacy of regionally administered melphalan in advanced extremity melanoma. METHODS: The mutational status of c-kit and patterns of gene expression predictive of dysregulated Src kinase signaling were evaluated in a panel of 26 human melanoma cell lines. The effectiveness of dasatinib was measured by quantifying protein expression and activation of Src kinase, focal adhesion kinase, and Crk-associated substrate (p130(CAS)), in conjunction with in vitro cell viability assays using seven melanoma cell lines. Utilizing a rat model of regional chemotherapy, we evaluated the effectiveness of systemic dasatinib in conjunction with regional melphalan against the human melanoma cell line, DM443, grown as a xenograft. RESULTS: Only the WM3211 cell line harbored a c-kit mutation. Significant correlation was observed between Src-predicted dysregulation by gene expression and sensitivity to dasatinib in vitro. Tumor doubling time for DM443 xenografts treated with systemic dasatinib in combination with regional melphalan (44.8 days) was significantly longer (p = 0.007) than either dasatinib (21.3 days) or melphalan alone (24.7 days). CONCLUSIONS: Systemic dasatinib prior to melphalan-based regional chemotherapy markedly improves the efficacy of this alkylating agent in this melanoma xenograft model. Validation of this concept should be considered in the context of a regional therapy clinical trial.

[A case of pelvic recurrence of rectal cancer successfully treated by surgical resection using the extraperitoneal approach].

Here, we report a case of recurrent rectal cancer successfully treated with resective surgery using the extraperitoneal approach. A 73-year-old man underwent the Miles operation for advanced rectal cancer (Rb-P, tub1, pMP, pN0, Stage I). At 20 months after the initial operation, computed tomography( CT) and magnetic resonance imaging( MRI) scans and 2-[fluorine-18] fluoro-2-deoxy-D-glucose (FDG)-positron emission tomography (PET) images showed a recurrent pelvic tumor( 20 mm in diameter) located in the lower presacral region. The tumor was surgically excised. The presacral space was easily exposed by the surgical procedure using the extraperitoneal approach, and the tumor was easily dissected and resected. We conclude that a surgical procedure using the extraperitoneal approach might be effective for the treatment of pelvic recurrence of rectal cancer.

[A case of mucosa-associated lymphoid tissue lymphoma of the rectum].

We report a case of mucosa-associated lymphoid tissue (MALT) lymphoma of the rectum in a 67-year-old woman who was admitted to our hospital owing to bowel abnormalities. Colonoscopic examination revealed a submucosal tumor (SMT) in the lower rectum. However, no malignancy was found on rectal mucosa biopsy. After providing informed consent, the patient underwent transanal surgery for the SMT. Rectal MALT lymphoma was diagnosed based on results of histological and immunohistochemical examinations. According to the Lugano International Conference classification system, the present case was classified as stage I MALT lymphoma. After the operation, Helicobacter pylori infection occurred, for which eradication therapy was performed, but no further complications or recurrence occurred.

[A case of rapidly progressing anorectal malignant melanoma].

We report here a case of rapidly progressing anorectal malignant melanoma. A 66-year-old man was admitted to our hospital due to bowel abnormalities and anal pain. Detailed gastrointestinal examination revealed a nigrities-like type 1 tumor that occupied a semicircle in the intestinal lumen from the lower rectum to the anatomical anal canal. We diagnosed anorectal malignant melanoma from a biopsy of the tumor. For the first time, we performed abdominoperineal resection. DAV-feron chemotherapy was administered from 18 days after the operation. However, multiple liver metastases, multiple lung metastases, and multiple skin metastases appeared in the early phase after the operation. Metastases increased rapidly and the patient died 138 days after the operation.

[A case of rectal cancer with interstitial pneumonia successfully treated with preoperative radiation therapy].

We report here a case of rectal cancer with interstitial pneumonia was successfully treated with preoperative radiation therapy. An 81-year-old man with complaints of constipation and melena was admitted to our hospital for the purpose of close inspection and medical treatment. In colonoscopic examination, we found a type-3 9 cm tumor in mainly occupied lower rectum (Rb), which developed all circumference-related stenosis. We diagnosed the tumor as Group V (adenocarcinoma) in biopsy. In abdominal computed tomography (CT) scan, the rectal tumor was directly invaded to the prostate and left internal obturator muscle. We diagnosed it to be cStage II rectal cancer (Rb) from various image findings. In addition, chest CT showed interstitial pneumonia. At first therapy, we did not perform pelvic evisceration nor chemotherapy because the patient was aged having interstitial pneumonia. We performed loop colostomy and preoperative radiation therapy (total 50 Gy). After the radiation therapy, there was a notable reduction in tumor, and a direct invasion to the prostate and left internal obturator muscle was not identified upon imaging. After the one year course of radiotherapy, we performed Mile's operation. After the operation, we did not perform adjuvant chemotherapy, but there has been no recurrence observed.

Inhibition of poly(ADP-ribose) polymerase enhances the effect of chemotherapy in an animal model of regional therapy for the treatment of advanced extremity malignant melanoma.

BACKGROUND: Poly(ADP-ribose) polymerase (PARP) is an important regulator of programmed cell death in response to alkylating agents such as temozolomide (TMZ). The goal of this study was to determine if a systemically administered PARP-inhibitor (INO-1001) could augment the efficacy of TMZ in a rat model of extremity malignant melanoma. MATERIALS AND METHODS: PARP activity was measured in vitro across a panel of 5 human malignant melanoma-derived cell lines. To evaluate tumor response to PARP inhibition in combination with regional isolated limb infusion (ILI) therapy with TMZ, two TMZ-resistant malignant melanoma cell lines were grown as xenografts in the hind limb of rats. INO-1001 (400 mg/kg) was injected intraperitoneally 7 times every 8 hours prior to ILI. Tumor volume was measured for up to 40 days. RESULTS: In vitro inhibition of PARP activity by INO-1001 ranged from 25.5% to 65.6%. In a mismatch repair (MMR)-deficient xenograft, treatment with INO-1001 prior to ILI significantly (P < .04) increased the efficacy of TMZ. The increase in tumor volume at day 40 following TMZ-ILI with INO-1001 was only 22.6% compared with 322.8% with TMZ-ILI alone. In a xenograft that was MMR-proficient and had high levels of O(6)-methylguanine-DNA methyltransferase (MGMT) activity, there was little improvement in TMZ efficacy with INO-1001 treatment. CONCLUSION: The PARP-inhibitor, INO-1001, can enhance the response of TMZ-resistant, MMR-deficient, malignant melanoma xenografts to intra-arterially administered TMZ in a regional treatment model of advanced extremity malignant melanoma.

[A case of recurrent esophageal cancer successfully treated by combined therapies].

We report here a case of recurrent esophageal cancer successfully treated by cervical lymph node dissection and chemo-radiation therapy. A 66-year-old woman received esophagectomy for advanced esophageal cancer in February 2003. Cervical CT and ultrasonography revealed multiple lymph node metastases in February 2008. Surgical excision was performed to the lymph node recurrence. After the operation, she received radiation therapy combined with cisplatin and 5-FU. She is alive without recurrence for two years after the treatment. We conclude that surgery and chemo-radiation therapy may be effective for cervical lymph node recurrence of esophageal cancer.

[A case of rectal cancer with synchronous multiple liver metastases successfully treated with combined chemotherapy of modified FOLFOX6 and bevacizumab].

We report here a case of rectal cancer with synchronous multiple liver metastases successfully treated with a combined chemotherapy of modified FOLFOX6 (mFOLFOX6) and bevacizumab. A 49-year-old man was admitted to our hospital due to constipation and anorexia. Abdominal ultrasonography and abdominal computed tomography (CT) scan revealed a rectal tumor (Rs) and abdominal abscess and 11 mm hepatic nodular lesion in S3 and 21 mm and 14 mm hepatic nodular lesions in S4. We diagnosed the patient had penetrated rectal cancer (Rs) and multiple liver metastases. We underwent a low anterior resection with D3 lymphadenectomy for the first time. After the operation, an 8 mm new liver metastasis in S6 appeared. We performed a combined therapy of mFOLFOX6 and bevacizumab 16 days after post operation. After the 8th course, there were a notable reduction in S3 and S4 liver metastases, and S6 liver metastasis was unidentifiable upon imaging. At this point, we underwent liver left lobectomy using the Liver Hanging Maneuver Method. After the second operation, there has been no recurrence and same chemotherapy is being continued.

Molecular features linked to the growth-inhibitory effects of gemcitabine on human pancreatic cancer cells.

Although gemcitabine (GEM) is widely used in the treatment of pancreatic cancers, the molecular mechanisms that underlie its anti-tumor effects are not fully understood. To clarify the anti-tumor mechanism(s) of GEM, we studied a human pancreatic cancer cell line, YPK-1, that showed a 50% inhibitory concentration (IC50) of GEM of 6.3 x 10(-3) microg/ml after 72 h of exposure. Cell proliferation was perturbed by 6 to 72 h of exposure to GEM concentrations equal to one-half or one-quarter of the IC50. We used cDNA microarrays containing 2976 genes to identify genes with expression affected by exposure to GEM. The self-organizing map identified nine clusters, including 85 and 87 genes, that showed differential expression in response to exposure to one half and one quarter IC50 GEM, respectively. Of these, 24 genes were common to cells exposed to the two different concentrations of GEM. Most are signal transduction or transcription-related genes. The microarray data for two of these genes, SPARC and RPS8, were validated by RT-PCR. Although further studies are needed to examine whether the changes in expression profiles of these genes are specific to cells exposed to GEM, the present data provide insights into the anti-tumor effects of GEM on pancreatic cancers.

[A resected case of effective treatment with TS-1 and CDDP for advanced gastric cancer with carcinomatous ascites].

We encountered a resected case of advanced gastric cancer after successful chemotherapy. The patient remained alive in good condition without any signs of recurrence at 2 years and 8 months after surgery. The 40-year-old woman initially complained of low abdominal pain, which was diagnosed as gastric cancer with metastasis of ovaries and carcinomatous ascites. We administered TS-1 (100 mg/body/day) for 3 weeks followed by 2-week rest and CDDP (60 mg/body) on the 8th day of one cycle. After 3 cycles of treatment, carcinomatous ascites disappeared and the ovarian lesion was reduced. The patient underwent total gastrectomy, ileocecal resection, cholecystectomy and radical hysterectomy. The antitumor efficacy of the treatment was histologically Grade 2. We administered 17 cycles of TS-1 treatment, and the patient is alive in good condition at this writing. This shows that TS-1 and CDDP may have potent therapeutic efficacy in advanced gastric cancer patients. For the future, the safety and efficacious administration of TS-1 and CDDP should be examined further.

Prediction of nodal metastasis by comparative genomic hybridization in biopsy specimens from patients with superficial esophageal squamous cell carcinoma.

PURPOSE: Selection of appropriate protocols for treatment of superficial esophageal squamous cell carcinoma (SESCC) is dependent on lymph node metastasis status. Therefore, it is important to know whether lymph node metastasis is present before treatment. EXPERIMENTAL DESIGN: In this study, we examined the relation between DNA sequence copy number aberrations detected by comparative genomic hybridization and lymph node metastasis in 26 surgically resected SESCCs (training samples). We then assessed whether the genetic information is predictive for nodal status in biopsy specimens from eight newly enrolled patients with SESCC (blinded samples). RESULTS: Pathological examination revealed that 17 of 26 training samples (65.4%) did not have associated lymph node metastasis. Gains of 8q24 and/or 20q12-qter were observed in 12, including all (nine of nine) with nodal metastasis. Fourteen training samples did not have gain of either 8q24 or 20q12-qter. Of the blinded samples, two showed no gain of 8q24 or 20q12-qter, and as anticipated the postoperative pathological examination revealed no nodal metastasis. The remaining six blinded samples had gains of 8q24 and/or 20q12-qter, and lymph node metastasis was detected by postoperative examination in four of these tumors. CONCLUSIONS: Absence of gains of 8q24 and/or 20q12-qter appears to be associated with absence of lymph node metastasis in patients with SESCC; therefore, less invasive surgery can be chosen.

Gain of 5p15 detected by comparative genomic hybridization as an independent marker of poor prognosis in patients with esophageal squamous cell carcinoma.

Comparative genomic hybridization was applied to 51 primary esophageal squamous cell carcinomas (ESCCs) to clarify the relation between DNA sequence copy number aberrations (DSCNAs) and the clinicopathology of the disease. The average number of DSCNAs was 10.9 DSCNAs/tumor (6.1 gains and 4.9 losses), ranging from 1-30 DSCNAs/tumor. Gain of 3q26-qter and loss of 18q22-qter were detected in >60% of stage I tumors and considered to play an important role in the development of ESCC. Whereas gain of 8q24-qter was observed in 82.6% (19 of 23) of stage III and IV cancers, it was seen in 27.3% (3 of 11) of stage I tumors. It is suggested that gain of 8q24-qter plays an important role in tumor progression. Gains of 8q24-qter and 20q12-qter and loss of 11q22-23 were linked to nodal metastasis (P = 0.0006, 0.002, and 0.02, respectively). Gains of 5p15 and 14q21 were associated with distant organ metastasis after surgery (P = 0.006 and 0.02, respectively). These observations suggest that nodal and distant organ metastases involve different genes. Gains of 5p15, 8q24-qter, and 14q21 were significantly associated with unfavorable prognosis (P = 0.0002, 0.007, and 0.04, respectively). Multivariate analysis revealed the 5p15 gain to be an independent prognostic marker with a higher significance than that of nodal status (risk ratio = 5.95; P = 0.001). The present findings indicate that comparative genomic hybridization analysis may be used to predict the likelihood of a poor or favorable outcome in cases of ESCC.

Molecular signature linked to acquired resistance to cisplatin in esophageal cancer cells.

To clarify the molecular basis of acquired cisplatin (CDDP) resistance in esophageal squamous cell carcinoma (ESCC), we used cDNA microarray technology. A CDDP-resistant cell line (YES-2/CDDP), which shows a 7.5-fold increase in resistance to CDDP and a 3-fold decrease in CDDP accumulation compared with the parental YES-2 ESCC cell line, was generated from YES-2 by exposure to increased concentrations of CDDP. By cDNA microarray analysis, we identified 44 genes with significantly different expression levels between YES-2/CDDP and YES-2 cells. Interestingly, 15 of these 44 genes encoded ribosome-related proteins, almost all of which were underexpressed in YES-2/CDDP cells. Our present data suggest that many ribosome-related genes may be involved in the acquired resistance to CDDP in ESCC and that such information may allow us to better understand the mechanism of CDDP resistance.

Bevacizumab-induced alterations in vascular permeability and drug delivery: a novel approach to augment regional chemotherapy for in-transit melanoma.

PURPOSE: To investigate whether the systemically administered anti-VEGF monoclonal antibody bevacizumab could improve regional chemotherapy treatment of advanced extremity melanoma by enhancing delivery and tumor uptake of regionally infused melphalan (LPAM). EXPERIMENTAL DESIGN: After treatment with systemic bevacizumab or saline, changes in vascular permeability were determined by spectrophotometric analysis of tumors infused with Evan's blue dye. Changes in vascular structure and tumor hemoglobin-oxygen saturation HbO(2) were determined by intravital microscopy and diffuse reflectance spectroscopy, respectively. Rats bearing the low-VEGF secreting DM738 and the high-VEGF secreting DM443 melanoma xenografts underwent isolated limb infusion (ILI) with melphalan (LPAM) or saline via the femoral vessels. The effect of bevacizumab on terminal drug delivery was determined by immunohistochemical analysis of LPAM-DNA adducts in tumor tissues. RESULTS: Single-dose bevacizumab given three days before ILI with LPAM significantly decreased vascular permeability (50.3% in DM443, P < 0.01 and 35% in DM738, P < 0.01) and interstitial fluid pressure (57% in DM443, P < 0.01 and 50% in DM738, P = 0.01). HbO(2) decreased from baseline in mice following treatment with bevacizumab. Systemic bevacizumab significantly enhanced tumor response to ILI with LPAM in two melanoma xenografts, DM443 and DM738, increasing quadrupling time 37% and 113%, respectively (P = 0.03). Immunohistochemical analyses of tumor specimens showed that pretreatment with systemic bevacizumab markedly increased LPAM-DNA adduct formation. CONCLUSIONS: Systemic treatment with bevacizumab before regional chemotherapy increases delivery of LPAM to tumor cells and represents a novel way to augment response to regional therapy for advanced extremity melanoma.

Sorafenib, a multikinase inhibitor, enhances the response of melanoma to regional chemotherapy.

Melanoma responds poorly to standard chemotherapy due to its intrinsic chemoresistance. Multiple genetic and molecular defects, including an activating mutation in the BRaf kinase gene, are associated with melanoma, and the resulting alterations in signal transduction pathways regulating proliferation and apoptosis are thought to contribute to its chemoresistance. Sorafenib, a multikinase inhibitor that targets BRaf kinase, is Food and Drug Administration approved for use in advanced renal cell and hepatocellular carcinomas. Although sorafenib has shown little promise as a single agent in melanoma patients, recent clinical trials suggest that, when combined with chemotherapy, it may have more benefit. We evaluated the ability of sorafenib to augment the cytotoxic effects of melphalan, a regional chemotherapeutic agent, and temozolomide, used in systemic and regional treatment of melanoma, on a panel of 24 human melanoma-derived cell lines and in an animal model of melanoma. Marked differences in response to 10 micromol/L sorafenib alone were observed in vitro across cell lines. Response to sorafenib significantly correlated with extracellular signal-regulated kinase (ERK) downregulation and loss of Mcl-1 expression (P < 0.05). Experiments with the mitogen-activated protein kinase/ERK kinase inhibitor U0126 suggest a unique role for ERK downregulation in the observed effects. Sorafenib in combination with melphalan or temozolomide led to significantly improved responses in vitro (P < 0.05). In the animal model of melanoma, sorafenib in combination with regional melphalan or regional temozolomide was more effective than either treatment alone in slowing tumor growth. These results show that sorafenib in combination with chemotherapy provides a novel approach to enhance chemotherapeutic efficacy in the regional treatment of in-transit melanoma.