Complement-dependent cytotoxicity (CDC) is an important antitumor mechanism of monoclonal antibodies (mAbs). However, trastuzumab, an anti-HER2 mAb, exerts only minor CDC. Overexpression of membrane-bound complement regulatory proteins (mCRPs), which suppress CDC, have been implicated in various malignant tumors. Here, we explored the predictive role of the expression levels of three mCRPs (CD55, CD59 and CD46) in the prognosis of breast cancer cases that underwent adjuvant trastuzumab treatment. We also studied the effect of mCRP downregulation on trastuzumab-induced CDC in vitro. Sixty-five HER2-positive breast cancer patients who received adjuvant therapy containing trastuzumab, were retrospectively analyzed. Levels of CD55, CD59 and CD46 expression were detected by immunohistochemistry. Chi-square test, Kaplan‑Meier survival analysis and a Cox proportional hazards model were used to analyze the association between CD55, CD59 and CD46 expression and prognosis. HER2-positive SK-Br3 and BT-474 breast cancer cells were pretreated with various drugs to reduce mCRP expression. Afterwards, trastuzumab‑mediated cytolytic effects were measured. Among the 65 patients, 46.2% had high expression of CD55, 44.6% had high expression of CD59 and 44.6% had high expression of CD46. The median follow-up duration was 47 months (range from 24 to 75 months). Patients with CD55 or CD59 overexpression had a higher relapse rate than those with low expression of CD55 (33.3 vs. 8.6%; P=0.013) or CD59 (31.0 vs. 11.1%; P=0.046). Similarly, mean disease-free survival of patients with CD55 or CD59 overexpression was significantly shorter than those with a low expression of CD55 (56 vs. 70 months; log-rank test, P=0.008) or CD59 (56 vs. 69 months; log-rank test, P=0.033). Multivariate analysis confirmed that CD55, but not CD59, was an independent risk factor of recurrence (HR=4.757; 95% CI, 0.985-22.974; P=0.05). In vitro, we found that tamoxifen inhibited both the protein and mRNA expression levels of CD55, but not CD59 or CD46 in SK-Br3 and BT-474 cells. After pretreatment of tamoxifen, trastuzumab-induced cytolysis was enhanced through CD55 downregulation. In conclusion, CD55 overexpression is an independent risk factor for recurrence in breast cancer patients receiving postoperative adjuvant therapy containing trastuzumab. Combined use of tamoxifen and trastuzumab for HER2-positive breast cancer treatment may enhance the antitumor effects of trastuzumab by elevated CDC, which warrants further study.

In consideration of budget and efficiency, we suggest OCT as the best storing method that not only preserves RNA quality during the freezing-thawing process well, but also ensures more secure and stable DNA and protein.

Y. CD55 limits sensitivity to complement-dependent cytolysis triggered by heterologous expression of ␣-gal xenoantigen in colon tumor cells. Am J Physiol Gastrointest Liver Physiol 306: G1056-G1064, 2014. First published April 24, 2014 doi:10.1152/ajpgi.00464.2013.-Engineering cancer cells to express heterologous antigen ␣-gal and induce the destruction of tumor cells depending on the complement cascade may be a promising strategy of tumor therapy. However, the feasibility and effect of using ␣-gal to induce colorectal adenocarcinoma cell line cytolysis is not yet known. In this study, we evaluated ␣-gal expression's ability to sensitize human colorectal adenocarcinoma cell lines to complement attack in cell lines LoVo, SW620, and Ls-174T. Nearly all ␣-gal-expressing LoVo and SW620 cells were killed by normal human serum (NHS), but ␣-gal-expressing Ls-174T cells showed no significant lysis. We analyzed the expression levels of membrane-bound complement regulatory proteins (mCRPs) on the three cell lines, and their protective role in ␣-gal-mediated activation of the complement. LoVo showed no expression of any of the three proteins. CD59 was strongly expressed by SW620 and Ls-174T. CD46 and CD55 varied between the two cell lines. CD46 on SW620 was only half the intensity of CD46 on Ls-174T. Ls-174T showed a notable expression of CD55, while expression of CD55 on SW620 was not detected. The sensitivity of Ls-174T expressing ␣-gal to NHS greatly increased following the downregulation of CD46 and CD55 with short hairpin RNA (shRNA). However, there is no increase in cell killing when CD59 expression was diminished. Our findings suggest that the use of ␣-gal as antigen to induce tumor cell killing may be a potential therapeutic strategy in colon cancer and that CD55 plays a primary role in conferring resistance to lysis. ␣-gal epitope; membrane-bound complement regulatory proteins; complement-dependent cytotoxicity; colorectal carcinoma COLORECTAL CANCER is the third most commonly diagnosed cancer in males and the second in females (28, 57). Surgery is the mainstay of treatment in most cases of colon cancer. Adjuvant therapies, primarily chemotherapy and radiotherapy, have improved the survival rate and quality of life in colorectal cancer patients. However, the 5-yr survival rate remains at ϳ50%, which explains the growing interest in alternative therapies that have no serious side effects and that have the potential to eradicate residual tumors after conventional treatment. A deeper understanding of the interaction between the host immune system and malignant tumors has enabled the development of effective clinical strategies that improve immune responses against tumors. We seek to develop a therapeutic strategy that induces a specific immune response resulting in direct tumor cell killing, depending on the activation of the complement system.

Cocrystallization of 2-amino-6-methy-1,3-benzothia­zole with octa­nedioic acid in a mixed methanol–water medium afforded the title 2:1 cocrystal, 2C8H8N2S·C8H14O4. The octa­nedioic acid mol­ecule is located on an inversion centre. In the crystal, inter­molecular N—H⋯O and O—H⋯O hydrogen bonds connect the components into a three-dimensional network.