A CERTain Role for Ceramide in Taxane-Induced Cell Death

An unexpected benefit of functional genomic screens is that at times they answer questions that they were not designed to ask. A siRNA screen reported by Swanton et al. in this issue of Cancer Cell reveals that silencing of spindle assembly checkpoint genes facilitates mitotic slippage, resulting in escape from taxane-induced cell death, aneuploidy, and chromosomal instability, hallmarks of taxane resistance. Unexpectedly, the screen disclosed that the sphingolipid ceramide is a key regulator of the taxane-mediated spindle assembly checkpoint and taxane-induced cell death. Ceramide metabolism thus serves as a legitimate target for modulation of taxane effect on tumors

Acid-Sphingomyelinase Triggered Fluorescently Labeled Sphingomyelin Containing Liposomes in Tumor Diagnosis after Radiation-Induced Stress

In liposomal delivery, a big question is how to release the loaded material into the correct place. Here, we will test the targeting and release abilities of our sphingomyelin-consisting liposome. A change in release parameters can be observed when sphingomyelin-containing liposome is treated with sphingomyelinase enzyme. Sphingomyelinase is known to be endogenously released from the different cells in stress situations. We assume the effective enzyme treatment will weaken the liposome making it also leakier. To test the release abilities of the SM-liposome, we developed several fluorescence-based experiments. In in vitro studies, we used molecular quenching to study the sphingomyelinase enzyme-based release from the liposomes. We could show that the enzyme treatment releases loaded fluorescent markers from sphingomyelin-containing liposomes. Moreover, the release correlated with used enzymatic activities. We studied whether the stress-related enzyme expression is increased if the cells are treated with radiation as a stress inducer. It appeared that the radiation caused increased enzymatic activity. We studied our liposomes' biodistribution in the animal tumor model when the tumor was under radiation stress. Increased targeting of the fluorescent marker loaded to our liposomes could be found on the site of cancer. The liposomal targeting in vivo could be improved by radiation. Based on our studies, we propose sphingomyelin-containing liposomes can be used as a controlled release system sensitive to cell stress

CD95 Rapidly Clusters in Cells of Diverse Origins

We have shown that CD95-mediated cell death requires a clustering of the receptor in distinct sphingolipid-rich domains of the cell membrane (Grassmé et al., 2000, Cremesti et al., 2000). These domains form in response to acid sphingomyelinase (ASM)-induced ceramide generation. However, recent studies challenged the finding of early CD95 clustering (Algeciras-Schimnich et al., 2002). Here, six independent groups tested clustering of CD95 in diverse cell type including primary cells ex vivo and established cell lines. The studies show clustering of CD95 within seconds to minutes in all cell types tested by the different groups. In addition, clustering of CD95 was detected after stimulation of cells using three agonistic anti-CD95 antibodies (CH11, APO-1-3 and JO2), CD95 ligand and stimuli that induce an upregulation and activation of the endogenous CD95/CD95 ligand system. The data confirm our previous studies and suggest rapid, i.e., within seconds to minutes, CD95 clustering as a general phenomenon occurring in many cell types

Endothelial Membrane Remodeling Is Obligate for Anti-Angiogenic Radiosensitization during Tumor Radiosurgery

While there is significant interest in combining anti-angiogenesis therapy with conventional anti-cancer treatment, clinical trials have as of yet yielded limited therapeutic gain, mainly because mechanisms of anti-angiogenic therapy remain to a large extent unknown. Currently, anti-angiogenic tumor therapy is conceptualized to either "normalize" dysfunctional tumor vasculature, or to prevent recruitment of circulating endothelial precursors into the tumor. An alternative biology, restricted to delivery of anti-angiogenics immediately prior to single dose radiotherapy (radiosurgery), is provided in the present study.Genetic data indicate an acute wave of ceramide-mediated endothelial apoptosis, initiated by acid sphingomyelinase (ASMase), regulates tumor stem cell response to single dose radiotherapy, obligatory for tumor cure. Here we show VEGF prevented radiation-induced ASMase activation in cultured endothelium, occurring within minutes after radiation exposure, consequently repressing apoptosis, an event reversible with exogenous C(16)-ceramide. Anti-VEGFR2 acts conversely, enhancing ceramide generation and apoptosis. In vivo, MCA/129 fibrosarcoma tumors were implanted in asmase(+/+) mice or asmase(-/-) littermates and irradiated in the presence or absence of anti-VEGFR2 DC101 or anti-VEGF G6-31 antibodies. These anti-angiogenic agents, only if delivered immediately prior to single dose radiotherapy, de-repressed radiation-induced ASMase activation, synergistically increasing the endothelial apoptotic component of tumor response and tumor cure. Anti-angiogenic radiosensitization was abrogated in tumors implanted in asmase(-/-) mice that provide apoptosis-resistant vasculature, or in wild-type littermates pre-treated with anti-ceramide antibody, indicating that ceramide is necessary for this effect.These studies show that angiogenic factors fail to suppress apoptosis if ceramide remains elevated while anti-angiogenic therapies fail without ceramide elevation, defining a ceramide rheostat that determines outcome of single dose radiotherapy. Understanding the temporal sequencing of anti-angiogenic drugs and radiation enables optimized radiosensitization and design of innovative radiosurgery clinical trials

Interleukin-22 Protects Intestinal Stem Cells from Immune-Mediated Tissue Damage and Regulates Sensitivity to Graft versus Host Disease

SummaryLittle is known about the maintenance of intestinal stem cells (ISCs) and progenitors during immune-mediated tissue damage or about the susceptibility of transplant recipients to tissue damage mediated by the donor immune system during graft versus host disease (GVHD). We demonstrate here that deficiency of recipient-derived IL-22 increased acute GVHD tissue damage and mortality, that ISCs were eliminated during GVHD, and that ISCs as well as their downstream progenitors expressed the IL-22 receptor. Intestinal IL-22 was produced after bone marrow transplant by IL-23-responsive innate lymphoid cells (ILCs) from the transplant recipients, and intestinal IL-22 increased in response to pretransplant conditioning. However, ILC frequency and IL-22 amounts were decreased by GVHD. Recipient IL-22 deficiency led to increased crypt apoptosis, depletion of ISCs, and loss of epithelial integrity. Our findings reveal IL-22 as a critical regulator of tissue sensitivity to GVHD and a protective factor for ISCs during inflammatory intestinal damage

Down-regulation of ATM protein sensitizes human prostate cancer cells to radiation-induced apoptosis

Treatment with the protein kinase C activator 12-O-tetradecanoylphorbol 12-acetate (TPA) enables radiation-resistant LNCaP human prostate cancer cells to undergo radiation-induced apoptosis, mediated via activation of the enzyme ceramide synthase ( CS) and de novo synthesis of the sphingolipid ceramide (Garzotto, M., Haimovitz-Friedman, A., Liao, W. C., White-Jones, M., Huryk, R., Heston, D. W. W., Cardon-Cardo, C., Kolesnick, R., and Fuks, Z. ( 1999) Cancer Res. 59, 5194-5201). Here, we show that TPA functions to decrease the cellular level of the ATM ( ataxia telangiectasia mutated) protein, known to repress CS activation ( Liao, W.-C., Haimovitz-Friedman, A., Persaud, R., McLoughlin, M., Ehleiter, D., Zhang, N., Gatei, M., Lavin, M., Kolesnick, R., and Fuks, Z. ( 1999) J. Biol. Chem. 274, 17908 - 17917). Gel shift analysis in LNCaP and CWR22-Rv1 cells demonstrated a significant reduction in DNA binding of the Sp1 transcription factor to the ATM promoter, and quantitative reverse transcription-PCR showed a 50% reduction of ATM mRNA between 8 and 16 h of TPA treatment, indicating that TPA inhibits ATM transcription. Furthermore, treatment of LNCaP, CWR22-Rv1, PC-3, and DU-145 human prostate cells with antisense-ATM oligonucleotides, which markedly reduced cellular ATM levels, significantly enhanced radiation-induced CS activation and apoptosis, leading to apoptosis at doses as a low as 1 gray. These data suggest that the CS pathway initiates a generic mode of radiation- induced apoptosis in human prostate cancer cells, regulated by a suppressive function of ATM, and that ATM might represent a potential target for pharmacologic inactivation with potential clinical applications in human prostate cancer

The life and work of Dr Robert Bittman (1942-2014)

With the passing of Dr. Robert Bittman on the 1st Oct 2014 from cancer, the lipid field lost its most distinguished lipid chemist researcher. Herein, we provide a brief account of Dr. Robert Bittman’s research achievements over a 50 year career. Dr. Robert Bittman’s lasting legacies to the fields of lipid membrane biophysics, cholesterol trafficking and cellular functions of phospholipids and sphingolipids are truly inestimable