Deborah R. Neal

Cell damage and death by autoschizis in human bladder (RT4) carcinoma cells resulting from treatment with ascorbate and menadione.

A human bladder carcinoma cell line RT4 was sham-treated with buffer or treated with ascorbate (VC) alone, menadione alone (VK3), or a combination of ascorbate:menadione (VC+VK3) for 1, 2, and 4 h. Cytotoxic damage was found to be treatment-dependent in this sequence: VC+VK3>VC>VK3>sham. The combined treatment induced the greatest oxidative stress, with early tumor cell injury affecting the cytoskeletal architecture and contributing to the self-excisions of pieces of cytoplasm freed from organelles. Additional damage, including a reduction in cell size, organelle alterations, nuclear damage, and nucleic acid degradation as well as compromised lysosome integrity, is caused by reactivation of DNases and the redox cycling of VC or VC+VK3. In addition, cell death caused by VC+VK3 treatment as well as by prolonged VC treatment is consistent with cell demise by autoschizis, not apoptosis. This report confirms and complements previous observations about this new mode of tumor cell death. It supports the contention that a combination of VC+VK3, also named Apatone, could be co-administered as a nontoxic adjuvant with radiation and/or chemotherapies to kill bladder tumor cells and other cancer cells without any supplementary risk or side effects for patients.

Cell death by autoschizis in TRAMP prostate carcinoma cells as a result of treatment by ascorbate: menadione combination.

A prostate carcinoma cell line derived from the transgenic murine prostate cancer model (TRAMP) was treated with ascorbate (VC) alone, menadione (VK3) alone, or a combination of ascorbate:menadione (VC + VK3) for 1, 2, and 4 h. Cytotoxic cell alterations examined by light and electron microscopy were treatment-dependent with VC + VK3 > VC > VK3. Induced by oxidative stress, these alterations included cytokeletal changes conducive to cytoplasmic blebbing, self-excisions, and progressive nuclear alterations. While the excised parts contained ribosomes, they were devoid of nuclear fragments or other organelles. The organelle-free self-excisions caused an extreme reduction in cell size as well as chromatolysis and karyolysis that were consistent with cell death by autoschizis, but not with apoptosis.

Xenotransplanted Human Prostate Carcinoma (DU145) Cells Develop into Carcinomas and Cribriform Carcinomas: Ultrastructural Aspects

Androgen-independent, human prostate carcinoma cells (DU145) develop into solid, carcinomatous xenotransplants on the diaphragm of nu/nu mice. Tumors encompass at least two poorly differentiated cell types: a rapidly dividing, eosinophilic cell comprises the main cell population and a few, but large basophilic cells able to invade the peritoneal stroma, the muscular tissue, lymph vessels. Poor cell contacts, intracytoplasmic lumina, and signet cells are noted. Lysosomal activities are reflected by entoses and programmed cell deaths forming cribriform carcinomas. In large tumors, degraded cells may align with others to facilitate formation of blood supply routes. Malignant cells would spread via ascites and through lymphatics.

Abstract 2076: Oxidative stress induced by sodium ascorbate:menadione sodium-bisulfate combination results in lysosomal and mitochondrial changes that lead to prostate cancer cell death

Prostate cancer is a leading cause of death in American males and therapy targeting hormone refractory prostate cancer is urgently needed. Treatment of prostate cancer cell lines (LNCaP, LNCaP-C42, DU-145 and PC3) with either sodium ascorbate or menadione sodium-bisulfate alone induced cell killing (as measured by the MTT cell proliferation assay) only at high concentrations, while the combination (apatone) potentiated cytotoxicity 4- to 20-fold with exposure times as short as 1hr. Apatone treatment induces the reactivation of DNase I and II resulting in autoschizis, a caspase independent form of cell death. Cell cycle arrest (G1/S and G2/M blocks), increased hydrogen peroxide and lipid peroxidation levels, decreased cellular thiol levels, and increased intracellular Ca2+ levels were observed following apatone treatment. To further understand the mechanism(s) of action of this cytotoxic drug, the effects of apatone treatment on the lysosomal and mitochondrial membranes in DU-145 prostate cancer cells were measured by live cell fluorescence microscopy. Examination of lysosomal membranes using the metachromatic fluorochrome, acridine orange, revealed rapid permeabilization of the lysosomal membrane. The effect of apatone on mitochondrial membrane potential was also examined, using JC-1 staining, and revealed de-polarization of the mitochondria membrane, which occurred on a delayed time scale when compared to lysosomal activity. The in vivo anti-tumoral activity of apatone was evaluated in nude mice inoculated subcutaneously with DU-145 cells. Oral apatone significantly reduced the growth rate of the solid tumors (p Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2076. doi:1538-7445.AM2012-2076