SUMMARY CD95 (Fas/APO-1), when bound by its cognate ligand CD95L, induces cells to die by apoptosis. We now show that elimination of CD95 or CD95L results in a form of cell death that is independent of caspase-8, RIPK1/MLKL, and p53, is not inhibited by Bcl-xL expression, and preferentially affects cancer cells. All tumors that formed in mouse models of low-grade serous ovarian cancer or chemically induced liver cancer with tissue specific deletion of CD95 still expressed CD95, suggesting that cancer cannot form in the absence of CD95. Death induced by CD95R/L elimination (DICE) is characterized by an increase in cell size and production of mitochondrial ROS, and DNA damage. It resembles a necrotic form of mitotic catastrophe. No single drug was found to completely block this form of cell death, and it could also not be blocked by the knockdown of a single gene, making it a promising new way to kill cancer cells.

Purpose: The clinical success of arsenic trioxide (As 2 O 3 ) in hematologic malignancies has not been replicated in solid tumors due to poor pharmacokinetics and dose-limiting toxicity. We have developed a novel nanoparticulate formulation of As 2 O 3 encapsulated in liposomal vesicles or "nanobins" [(NB(Ni, As)] to overcome these hurdles. We postulated that nanobin encapsulation of As 2 O 3 would improve its therapeutic index against clinically aggressive solid tumors, such as triple-negative breast carcinomas.Experimental Design: The cytotoxicity of NB(Ni,As), the empty nanobin, and free As 2 O 3 was evaluated against a panel of human breast cancer cell lines. The plasma pharmacokinetics of NB(Ni,As) and free As 2 O 3 were compared in rats to measure drug exposure. In addition, the antitumor activity of these agents was evaluated in an orthotopic model of human triple-negative breast cancer.Results: The NB(Ni,As) agent was much less cytotoxic in vitro than free As 2 O 3 against a panel of human breast cancer cell lines. In contrast, NB(Ni,As) dramatically potentiated the therapeutic efficacy of As 2 O 3 in vivo in an orthotopic model of triple-negative breast cancer. Reduced plasma clearance, enhanced tumor uptake, and induction of tumor cell apoptosis were observed for NB(Ni,As).Conclusions: Nanobin encapsulation of As 2 O 3 improves the pharmacokinetics and antitumor efficacy of this cytotoxic agent in vivo. Our findings demonstrate the therapeutic potential of this nanoscale agent and provide a foundation for future clinical studies in breast cancer and other solid tumors. Clin Cancer Res; 16(14); 3607-17. ©2010 AACR.Breast cancer is the second leading cause of cancer mortality for women in the United States (1). Although preventive agents and targeted therapies directed at the estrogen receptor, progesterone receptor, and human epidermal growth factor 2 receptor (HER2/neu) have resulted in improved clinical outcomes for many women with breast cancer, formidable challenges remain in treating tumors that do not express these molecular targets. These "triple-negative" breast carcinomas represent 15% of newly diagnosed breast cancer cases and often exhibit a basal epithelial or basal-like gene expression profile that is associated with poor survival (2-4). Consistent with its aggressive nature, triple-negative breast cancer is characterized by high rates of distant recurrence, particularly in the lung and brain, within the first five years after diagnosis despite adjuvant chemotherapy (5, 6). Hence, development of new therapeutic agents for these clinically intractable tumors is highly desirable.Arsenic trioxide (As 2 O 3 ) is a Food and Drug Administration-approved treatment for refractory acute promyelocytic leukemia (APL) and has shown preliminary activity in patients with relapsed/refractory multiple myeloma (7-10). Several mechanisms of action have been proposed for As 2 O 3 activity, including induction of apoptosis mediated by reactive oxygen species, promotion of cellular differentiation, and ...

Risk biomarkers for estrogen receptor (ER)-negative breast cancer have clear value for breast cancer prevention. We previously reported a set of lipid metabolism (LiMe) genes with high expression in the contralateral unaffected breasts (CUBs) of ER-negative cancer cases. We now further examine LiMe gene expression in both tumor and CUB, and investigate the role of Pre-B-cell leukemia homeobox-1 (PBX1) as a candidate common transcription factor for LiMe gene expression. mRNA was extracted from laser-capture microdissected epithelium from tumor and CUB of 84 subjects (28 ER-positive cases, 28 ER-negative cases, 28 healthy controls). Gene expression was quantitated by qRT-PCR. Logistic regression models were generated to predict ER status of the contralateral cancer. Protein expression of HMGCS2 and PBX1 was measured using immunohistochemistry. The effect of PBX1 on LiMe gene expression was examined by overexpressing PBX1 in MCF10A cells with or without ER, and by suppressing PBX1 in MDA-MB-453 cells. The expression of DHRS2, HMGCS2, UGT2B7, UGT2B11, ALOX15B, HPGD, UGT2B28 and GLYATL1 was significantly higher in ER-negative versus ER-positive CUBs, and predicted ER status of the tumor in test and validation sets. In contrast, LiMe gene expression was significantly lower in ER-negative than ER-positive tumors. PBX1 overexpression in MCF10A cells up-regulated most LiMe genes, but not in MCF10A cells overexpressing ER. Suppressing PBX1 in MDA-MB-453 cells resulted in decrease of LiMe gene expression. Four binding sites of PBX1 and cofactor were identified in three lipid metabolism genes using ChIP-qPCR. These data suggest a novel role for PBX1 in the regulation of lipid metabolism genes in benign breast, which may contribute to ER-negative tumorigenesis.

The antibody-drug conjugate enfortumab-vedotin acts by targeting nectin-4, a protein that is nearly ubiquitously expressed in conventional urothelial cancer. However, expression of nectin-4 in morphologic variants of urothelial carcinoma and nonurothelial histotypes was unknown. Immunohistochemistry for nectin-4 using was performed on 169 patients including 83 with nonmuscle invasive bladder cancer and 86 patients with muscle invasive bladder cancer. Staining was scored for intensity (0 to 3) and extent (% positive cells) using the histological score system, where > 15 was considered positive. Overall, 72/83 (87%) samples of nonmuscle invasive urothelial carcinoma were positive, including 29/30 (97%) noninvasive papillary urothelial carcinomas, 7/8 (87.5%) carcinomas in situ, 36/45 (80%) papillary urothelial carcinomas invading the lamina propria. Overall, 50/86 muscle invasive tumors were positive, including 15/22 (68.2%) urothelial carcinomas, 7/10 (70%) squamous cell carcinomas, 3/11 (28%) micropapillary tumors, 4/6 (66%) adenocarcinomas, 2/4 (50%) nested carcinomas, 5/8 (63%) plasmacytoid, 1/10 (10%) sarcomatoid carcinomas, and 0/15 (0%) small cell carcinomas. Whole transcriptome RNA sequencing revealed that compared with conventional urothelial carcinomas, most sarcomatoid carcinomas and all but 2 small cell carcinomas expressed very low levels of nectin-4 mRNA but expressed significant levels of either trop2 or ERBB2, which are the molecular targets of 2 other antibody-drug conjugatessacituzumab gavitecan (trop2) or trastuzumab deruxtecan (ERBB2/HER2). In summary, our study demonstrates that there is heterogeneity of expression of nectin-4 in morphologic variants of urothelial cancer and nonurothelial histotypes, and suggests that testing expression of nectin-4 should be considered in morphologic variants or nonurothelial histotypes found to have lower expression.