Pil1, an eisosome organizer, plays an important role in the recruitment of synaptojanins and amphiphysins to facilitate receptor-mediated endocytosis in yeast.

The eisosome protein Pil1 is known to be implicated in the endocytosis of Ste3, but the precise biological function of it during endocytosis is poorly understood. Here, we present data to reveal Pil1's role in receptor-mediated endocytosis. Using live cell imaging, we show that endocytic patches carrying Abp1 and Las17 persisted much longer in PIL1-deficient cells.

Valspodar: current status and perspectives.

Valspodar (Amdray, SDZ PSC 833) is derived from cyclosporin, but lacks the immunosuppressive and most of the collateral activities of cyclosporin A (CsA, Sandimmune, Neoral); it exhibits an enhanced capacity to chemosensitise tumour cells showing the classical type multiple drug-resistance (MDR) associated with MDR1 P-glycoprotein (Pgp) overexpression. This valspodar-mediated chemosensitisation of MDR tumour cells is reviewed with regard to its mechanism of inhibition on Pgp flippase function, and its potential inhibition of anticancer drug (ACD) metabolisation by CYP3A enzymes is discussed. Potent inhibition of the membranous and cytoplasmic detoxification mechanisms expressed by cells at the absorption and clearance borders in the body by valspodar results in the many pharmacokinetic interactions with other drugs that are substrates of either, or both, Pgp and CYP classes of detoxifying enzyme.

Cyclosporins: structure-activity relationships for the inhibition of the human FPR1 formylpeptide receptor.

The human formylpeptide receptor (FPR) is a seven-transmembranous G-protein-coupled receptor (7TM-GPCR) for chemotactic peptides of bacterial origins, possibly involved in the recruitment and activation of neutrophils in various inflammatory diseases of mucosal epithelia. Mutational analyses suggest that interactions of formylated peptides with FPR occur on the outer exoplasmic leaflet/domains of the plasma membrane. The immunosuppressive and antifungal antibiotic cyclic undecapeptide cyclosporin A (CsA; cyclo-[MeBmt(1)-Abu(2)-MeGly(3)-MeLeu(4)-Val(5)-MeLeu(6)-Ala(7)-D-Ala(8)-MeLeu(9)-MeLeu(10)-MeVal(11)]) and some tested analogues such as [Ala(2)]-CsA, [Thr(2)]-CsA, [Val(2)]-CsA, and [Nva(2)]-CsA were able of inhibiting the binding of formylpeptides to the FPR, with [D-MeVal(11)]-CsA (CsH) being much more active than the other analogues.

Cyclosporins: structure-activity relationships for the inhibition of the human MDR1 P-glycoprotein ABC transporter.

Cyclic undecapeptide cyclo-[MeBmt(1)-Abu(2)-MeGly(3)-MeLeu(4)-Val(5)-MeLeu(6)-Ala(7)-D-Ala(8)-MeLeu(9)-MeLeu(10)-MeVal(11)], the immunosuppressive and antifungal antibiotic cyclosporin A (CsA), was reported to interfere with the MDR1 P-glycoprotein (Pgp), a transmembranous adenosine 5'-triphosphate binding cassette (ABC) transporter with phospholipid flippase or "hydrophobic vacuum cleaner" properties that mediate multidrug resistance (MDR) of cancer cells. By use of photoaffinity-labeled cyclosporins and membranes from Pgp-expressing cells, it was recently shown that in vitro, Pgp molecules could bind a large cyclosporin domain involving residues 4-9 as well as the side chain of residue 1. Tumor cell MDR can also be reversed by a product more distantly related to cyclosporin with the structure [Thr(2), Leu(5), D-Hiv(8), Leu(10)]-CsA (SDZ 214-103).

The potent immunosuppressive cyclosporin FR901459 inhibits the human P-glycoprotein and formyl peptide receptor functions.

By sequestering cytosolic calcineurin into a molecular complex with cyclophilin and its consequent T-cell dysfunction, some cyclosporins, such as CsA and FR901459 ([Thr2-Leu5-Leu10]-CsA), display potent immunosuppressive activity. Independently on this property, cyclosporins may display one or more other biological activities mediated by interaction with cell surface glycoproteins. Several cyclosporins inhibit the function of human MDRI-encoded P-glycoprotein (Pgp), a flippase known to cause cancer multidrug resistance, but also expressed by some normal immunocompetent cells and by normal epithelial cells which control drug bioavailability in vivo.

Aureobasidins: structure-activity relationships for the inhibition of the human MDR1 P-glycoprotein ABC-transporter.

Cyclic depsipeptide cyclo-[D-Hmp(1)-L-MeVal(2)-L-Phe(3)-L-MePhe(4)-L-Pro(5)-L-aIle+ ++(6)-L-MeVal(7)-L-Leu(8)-L-betaHOMeVal(9)], the antifungal antibiotic aureobasidin A (AbA), was reported to interfere with ATP-binding cassette (ABC) transporters in yeast and mammalian cells, particularly the MDR1 P-glycoprotein (Pgp), a transmembrane phospholipid flippase or "hydrophobic vacuum cleaner" that mediates multidrug resistance (MDR) of cancer cells. In a standardized assay that measures Pgp function by the Pgp-mediated efflux of the calcein-AM Pgp substrate and uses human lymphoblastoid MDR-CEM (VBL(100)) cells as highly resistant Pgp-expressing cells and the cyclic undecapeptide cyclosporin A (CsA) as a reference MDR-reversing agent (IC(50) of 3.4 microM), AbA was found to be a more active Pgp inhibitor (IC(50) of 2.

The abamectin derivative ivermectin is a potent P-glycoprotein inhibitor.

Among the compounds endowed with the capacity to reverse the P-glycoprotein (Pgp)-mediated multidrug resistance of cancer cells, a powerful agent was found to be the cyclosporin D derivative SDZ PSC 833. After in vivo treatment with SDZ PSC 833, mice showed a decreased tolerability to cyclosporin A (CsA), but also to ivermectin, a widely used polycyclic lactone pesticide of Streptomyces avermitilis origin. The sequels were suggestive of CsA- or ivermectin-induced central nervous system dysfunction; they were interpreted as caused by the neutralization of the Pgp at the blood-brain barrier level, implying that CsA and ivermectin were Pgp substrates.

Ranking of P-glycoprotein substrates and inhibitors by a calcein-AM fluorometry screening assay.

In order to compare the capacities of a variety of compounds to interfere with P-glycoprotein (Pgp) function, a novel assay was set up to work on a large screening scale. The model assay measures the capacity of parental sensitive (Par) and multidrug-resistant (MDR) cells to efflux a small fixed amount of acetoxymethyl calcein (calcein-AM) after their pretreatment with concentration ranges of known Pgp modulators. This microplate cytometry-based assay was performed with two different pairs of cell lines, the human lymphocytic leukemia CEM cells and the murine monocytic leukemia P388 cells.

Decreased biotolerability for ivermectin and cyclosporin A in mice exposed to potent P-glycoprotein inhibitors.

SDZ PSC 833 or SDZ 280-446 are strong blockers of the function of class I mdr gene-encoded P-glycoprotein molecules, which were developed for the reversal of multi-drug-resistance of tumor cells. When treated with such drugs, normal mice may display hypersensitivity to cyclosporin A and ivermectin. The recorded signs of acute toxicity are compatible with alterations of the murine central nervous system functions and with earlier data suggesting that P-glycoprotein expressed at the murine blood-brain barrier might be involved in the exclusion of cyclosporin A or ivermectin from brain tissue.

Decreased uptake of cyclosporin A by P-glycoprotein (Pgp) expressing CEM leukemic cells and restoration of normal retention by Pgp blockers.

The P-glycoprotein (Pgp) molecules which are expressed on multidrug resistant (MDR) tumor cells can efflux a variety of cytostatics. In both normal and tumoral epitheliums, Pgp molecules are selectively expressed on the apical surface of the epithelial cells. Such a distribution seems to be responsible for the transcellular transport of Pgp substrates, including cyclosporin A (CsA), from the basal to the apical side.

Lymphotoxicity and myelotoxicity of doxorubicin and SDZ PSC 833 combined chemotherapies for normal mice.

In the mouse, the P-glycoprotein-directed chemosensitizer SDZ PSC 833 could both restore a therapeutic window for doxorubicin against multidrug-resistant tumors, by inhibiting P-glycoprotein function, and increase the anti-cancer drug efficacy against drug-sensitive tumors, by increasing doxorubicin bioavailability. Since the success of such combined chemotherapy treatments might have been limited by the myelotoxicity of doxorubicin and the P-glycoprotein expression on some blood cells, their lymphotoxicity and myelotoxicity was studied on normal B6D2F1 mice, and whenever possible, the persistence of blood cell alterations was also searched for in scid recipients of lymphohaematopoietic grafts from the donor mice. Analyzed parameters were blood, lymphoid and myeloid cell numbers, proliferative responses to T- and B-cell mitogens, and serum immunoglobulin levels.

Development of grafted gld cells in athymic and euthymic recipients.

Mice homozygous for the gld (generalized lymphoproliferative disease) and lpr (lymphoproliferation) mutations display similar autoimmune and lymphoproliferative diseases. Both result from defective apoptosis, the targets of the lpr and gld mutations being the genes for, respectively, an apoptosis-signalling receptor [the Fas antigen receptor (FasR)] and its counter-receptor [the Fas ligand (FasL)]. Though this definitely causes the development and accumulation of large numbers of unusual Thy-1+ B220+ cells in peripheral lymphoid organs, details on how this actually occurs are still lacking.

Interactions of B6 wild and B6 gld cells engrafted within athymic nude beige recipients.

The murine gld mutation is targetted to the gene coding for the ligand of the Fas receptor for apoptosis. Gld mice display a lymphoproliferative and autoimmune syndrome that can be transferred in both irradiated euthymic wild and athymic beige (nubg) recipients. In order to test whether a supply of normal wild cells could correct the development of the gld syndrome, nubg mice were grafted with mixtures of gld and wild spleen cells from congenic donors which differed for the allotypes of the T-cell Thy1 membrane glycoprotein and/or of the B-cell Ig heavy chain.

Serological abnormalities induced by engraftment of viable motheaten hematopoietic cells in nude beige recipients.

C57BL/6J (B6) mice homozygous for the viable motheaten (mev) mutation are short-lived and display severe immunodeficiency, autoimmunity and inflammatory disease. B6 mice doubly homozygous for the nude (nu) and beige (bg) mutations (nubg mice) are also short-lived and immunodeficient. Nevertheless, grafts of mev lympho-hematopoietic cells increased life expectancy of nubg recipients.

Influence of the lpr environment on the lymph node cell phenotypes in C57BL/6 nubg and nulpr chimeras.

Mice homozygous for the lpr gene show a marked lymphoproliferative syndrome. Most T cells which accumulate in their lymphoid organs belong to a fairly unusual subpopulation. Although being CD44+ T cells expressing neither CD4 nor CD8, they are CD3 T-cell receptor (TCR) alpha beta positive and express both Thy-1 and B220, the B-cell form of the CD45 marker.

Modulation of human P-glycoprotein epitope expression by temperature and/or resistance-modulating agents.

Three monoclonal antibodies (mAb), MRK16, MM4.17 and MC57, directed against distinct epitopes on the external domain of human P-glycoprotein (Pgp), were used to follow its expression on multidrug resistant (MDR)-cells. The linear MM4.

The Yaa gene-dependent B-cell deficiency worsens the generalized lymphadenopathy and autoimmunity of C57BL/6-gld male mice.

The BXSB mice are unique among murine models for systemic lupus erythematosus in that males are much more severely affected than females. The BXSB male disease is associated with a Y-chromosome-linked gene, which is an autoimmunity accelerator gene (Yaa). The Yaa mutation affects the B-cell subset, which becomes hyper-responsive to T-cell signals.

Myeloid and lymphoid cell alterations in normal mice exposed to chemotherapy with doxorubicin and/or the multidrug-resistance reversing agent SDZ PSC 833.

The cyclosporin SDZ PSC 833 (PSC) is a potent in vivo chemosensitizer for tumor cells with P-glycoprotein(Pgp)-dependent multidrug resistance (MDR). However, Pgp expression also occurs in CD8+ T cells, NK cells, macrophages and stem cells. In order to find whether PSC might display specific myelotoxicity or potentiate the toxicity of anti-cancer drugs, healthy mice were exposed to single doxorubicin (DOX) and combined (DOX + PSC) chemotherapy protocols known to be near or above the borderline of toxicity for tumor-bearing mice.

The Yaa mutation induces the development of autoimmunity in mice heterozygous for the gld (generalized lymphadenopathy disease) mutation.

Mice homozygous for either the generalized lymphoproliferative disease (gld) or the lymphoproliferation (lpr) nonallelic mutations develop similar syndromes combining systemic autoimmunity and lymphoproliferative disease. Though essentially recessive, the lpr and gld mutations may be expressed in the heterozygous state: [lpr/+] mice displayed a mild "lpr-like" autoimmunity, and the [lprcg/+ gld/+] mice developed a "gld-like" disorder, showing interactions of the gld gene product with the nonallelic lprcg product. The Y-chromosome-linked autoimmune accelerator (Yaa) mutation being an autoimmunity accelerator, its association with an heterozygous gld gene might also bring a gld-like syndrome.

Derivatives of a novel cyclopeptolide. 2. Synthesis, activity against multidrug resistance in CHO and KB cells in vitro, and structure-activity relationships.

A series of derivatives of the novel cyclopeptolide 1 was prepared, and their ability to chemosensitize multi drug resistant CHO and KB cells in vitro was evaluated. In contrast to the parent compound, several of the derivatives were found to be highly active. In particular, conversion of the R-lactic acid residue of 1 into its S-isomer via lactone ring cleavage and recyclization with inversion resulted in a marked enhancement of activity.

SDZ 280-125: a cyclopeptolide endowed with an in vitro cyclosporin A-like profile of activity for the reversion of the P-glycoprotein-mediated multidrug resistance of tumor cells.

Tumor cells whose multidrug resistance is caused by the P-glycoprotein (Pgp) mediated anti-cancer drug (ACD) efflux can be chemosensitized by cyclosporins, whose derivatives were found to display a whole range of resistance-modulating activities. Similarly, derivatives of the non-immunosuppressive natural fungus cyclic peptolide SDZ 90-215 were recently shown to display a broad range of chemosensitizing activities. With highly resistant cells expressing high levels of Pgp, one such compound (SDZ 280-125) was shown here to restore both a normal sensitivity to the growth-inhibitory effects of ACD and a normal retention of an anthracycline antibiotic.

Extent and persistence of P-glycoprotein inhibition in multidrug-resistant P388 cells after exposure to resistance-modifying agents.

The low daunomycin (DAU) retention in P388 cells displaying P-glycoprotein (Pgp)-mediated multidrug resistance (MDR) can be increased by the presence of various resistance-modifying agents (RMAs). Taking the DAU retention restoration as an indicator of Pgp function inhibition and using a few RMAs, including SDZ PSC 833, SDZ 280-446, cyclosporin A (CsA) and verapamil, we compared different conditions of MDR cell exposure to the RMA. The 'co + post-RMA' treatments (RMA present during both DAU uptake and efflux phases) generally led to higher DAU retention levels than the 'co-RMA' treatments (RMA present during the DAU uptake phase only).

Atypical multi-drug resistance (MDR): low sensitivity of a P-glycoprotein-expressing human T lymphoblastoid MDR cell line to classical P-glycoprotein-directed resistance-modulating agents.

Verapamil, cyclosporin A (CsA), the cyclosporin derivative SDZ PSC 833 and the novel cyclopeptolide SDZ 280-446 were tested for their capacity to chemosensitize a P-glycoprotein (Pgp)-expressing multi-drug resistant (MDR) variant of the CEM human T lymphoblastoid cell subline (CCRF ACTD 400+). That MDR-CEM cell subline had been previously selected for MDR by actinomycin D and displayed a very high resistance phenotype: 3700-fold for actinomycin D, 3900-fold for vincristine, 1200-fold for taxol, 1000-fold for daunomycin (DAU) and 400-fold for colchicine. Interestingly, these MDR-CEM cells displayed little chemosensitization by resistance-modulating agents (RMA) which presumably work by inhibiting Pgp function.