Acid ceramidase promotes nuclear export of PTEN through sphingosine 1-phosphate mediated Akt signaling.

The tumor suppressor PTEN is now understood to regulate cellular processes at the cytoplasmic membrane, where it classically regulates PI3K signaling, as well as in the nucleus where multiple roles in controlling cell cycle and genome stability have been elucidated. Mechanisms that dictate nuclear import and, less extensively, nuclear export of PTEN have been described, however the relevance of these processes in disease states, particularly cancer, remain largely unknown. We investigated the impact of acid ceramidase on the nuclear-cytoplasmic trafficking of PTEN. Immunohistochemical analysis of a human prostate tissue microarray revealed that nuclear PTEN was lost in patients whose tumors had elevated acid ceramidase. We found that acid ceramidase promotes a reduction in nuclear PTEN that is dependent upon sphingosine 1-phosphate-mediated activation of Akt. We were further able to show that sphingosine 1-phosphate promotes formation of a complex between Crm1 and PTEN, and that leptomycin B prevents acid ceramidase and sphingosine 1-phosphate mediated loss of nuclear PTEN, suggesting an active exportin-mediated event. To investigate whether the tumor promoting aspects of acid ceramidase in prostate cancer depend upon its ability to export PTEN from the nucleus, we used enforced nuclear expression of PTEN to study docetaxel-induced apoptosis and cell killing, proliferation, and xenoengraftment. Interestingly, while acid ceramidase was able to protect cells expressing wild type PTEN from docetaxel, promote proliferation and xenoengraftment, acid ceramidase had no impact in cells expressing PTEN-NLS. These findings suggest that acid ceramidase, through sphingosine 1-phosphate, promotes nuclear export of PTEN as a means of promoting tumor formation, cell proliferation, and resistance to therapy.

Radiation-induced acid ceramidase confers prostate cancer resistance and tumor relapse.

Escape of prostate cancer (PCa) cells from ionizing radiation-induced (IR-induced) killing leads to disease progression and cancer relapse. The influence of sphingolipids, such as ceramide and its metabolite sphingosine 1-phosphate, on signal transduction pathways under cell stress is important to survival adaptation responses. In this study, we demonstrate that ceramide-deacylating enzyme acid ceramidase (AC) was preferentially upregulated in irradiated PCa cells. Radiation-induced AC gene transactivation by activator protein 1 (AP-1) binding on the proximal promoter was sensitive to inhibition of de novo ceramide biosynthesis, as demonstrated by promoter reporter and ChIP-qPCR analyses. Our data indicate that a protective feedback mechanism mitigates the apoptotic effect of IR-induced ceramide generation. We found that deregulation of c-Jun induced marked radiosensitization in vivo and in vitro, which was rescued by ectopic AC overexpression. AC overexpression in PCa clonogens that survived a fractionated 80-Gy IR course was associated with increased radioresistance and proliferation, suggesting a role for AC in radiotherapy failure and relapse. Immunohistochemical analysis of human PCa tissues revealed higher levels of AC after radiotherapy failure than those in therapy-naive PCa, prostatic intraepithelial neoplasia, or benign tissues. Addition of an AC inhibitor to an animal model of xenograft irradiation produced radiosensitization and prevention of relapse. These data indicate that AC is a potentially tractable target for adjuvant radiotherapy.

Interdiction of sphingolipid metabolism to improve standard cancer therapies.

Non-surgical therapies for human malignancies must negotiate complex cell signaling pathways to impede cancer cell growth, ideally promoting death of cancer cells while sparing healthy tissue. For most of the past half century, medical approaches for treating cancer have relied primarily on cytotoxic chemotherapeutics that interfere with DNA replication and cell division, susceptibilities of rapidly dividing cancer cells. As a consequence, these therapies exert considerable cell stress, promoting the generation of ceramide through de novo synthesis and recycling of complex glycosphingolipids and sphingomyelin into apoptotic ceramide. Radiotherapy of cancer exerts similar geno- and cytotoxic cell stresses, and generation of ceramide following ionizing radiation therapy is a well-described feature of radiation-induced cell death. Emerging evidence now describes sphingolipids as mediators of death in response to newer targeted therapies, cementing ceramide generation as a common mechanism of cell death in response to cancer therapy. Many studies have now shown that dysregulation of ceramide accumulation-whether by reduced generation or accelerated metabolism-is a common mechanism of resistance to standard cancer therapies. The aims of this chapter will be to discuss described mechanisms of cancer resistance to therapy related to dysregulation of sphingolipid metabolism and to explore clinical and preclinical approaches to interdict sphingolipid metabolism to improve outcomes of standard cancer therapies.

Hypoxia and radiation therapy: past history, ongoing research, and future promise.

Tumor hypoxia influences the outcome of treatment with radiotherapy, chemotherapy and even surgery, not only for the treatment of large bulky tumors with extensive necrosis, but also in the treatment of very small primary tumors and recurrences, micrometastases, and surgical margins with microscopic tumor involvement. Because hypoxic tumor cells are resistant to radiation and to many anticancer drugs, many approaches to circumventing the therapeutic resistance induced by hypoxia have been examined in laboratory studies and clinical trials. In this review, these approaches and the results of past laboratory and clinical studies are described and the limitations of the past agents and their testing are discussed. We describe the importance of new technologies for measuring hypoxia in human tumors, which allow assessment of pretreatment tumor oxygen levels and changes in hypoxia over the course of prolonged treatment regimens. These offer the possibility of improving the design of clinical trials and the selection of patients who will benefit from hypoxia-directed therapies, as well as the possibility of facilitating the development of better agents and regimens for use in hypoxia-directed therapy. We also discuss how the improved understanding of the abnormal vascular beds in solid tumors and of the effects of hypoxia and related microenvironmental insults, resulting from recent and ongoing research, offers the potential for finding new therapeutic targets, that may lead to the development of new agents and novel therapeutic approaches for selectively targeting cells in the adverse microenvironments within solid tumors.