The Marine Natural Product Furospinulosin 1 Induces Apoptosis in MDA-MB-231 Triple Negative Breast Cancer Cell Spheroids, But Not in Cells Grown Traditionally with Longer Treatment.

Cancer cells grown in spheroid conditions interact with each other and the extracellular matrix, providing a better representation of the in vivo environment than two-dimensional cultures and are a more clinically relevant model. A discrete screening of genetically diverse marine samples in the spheroid assay led to the identification of a novel activity for the known compound furospinulosin 1. This compound shows activity against MDA-MB-231 triple negative breast cancer cells grown as spheroids and treated for 24 or 48 h. No cytotoxicity was seen in traditional two-dimensional adherent cultures treated for a longer time (72 h). A reverse phase protein array (RPPA) confirmed the limited activity of the compound in cells grown traditionally and revealed changes in protein expression when cells are grown as spheroids that are associated with better clinical prognosis. Analysis of the RPPA data through the Broad institute's connectivity map suggested the hypothesis that furospinulosin 1 functions as an MEK inhibitor. Analysis of the RPPA data through STRING supports the apoptosis observed. The selectivity exhibited by furospinulosin 1 for triple negative breast cancer cells only when grown as spheroids makes it an interesting compound with strong therapeutic potential that merits further study.

Discovery of Survivin Inhibitors Part 1: Screening the Harbor Branch Pure Compound Library.

Survivin is a 16.5 KDa protein whose functions include promoting cellular mitosis, angiogenesis, and senescence as well as inhibiting apoptosis. Higher survivin expression is found in cancer tissues than normal tissues, and this expression correlates with disease progression and aggressiveness. Survivin has been validated as a clinical target for cancer. Small molecules are important antagonists of survivin levels in cancer cells. A structurally diverse library of genetically encoded small molecules (natural products) derived from marine plants, invertebrates, and microbes was screened for their ability to reduce expression levels of survivin in the DLD-1 colon adenocarcinoma and the A549 nonsmall cell lung carcinoma cell lines. This led to the identification of this novel activity for the known compounds eryloside E, ilicicolin H, tanzawaic acid A, and p-hydroxyphenopyrrozin. Both eryloside E and ilicicolin H showed the ability to reduce survivin expression in the low micromolar range against both cell lines.

Analogues of the Potent Antitumor Compound Leiodermatolide from a Deep-Water Sponge of the Genus Leiodermatium.

Two new analogues of the potent antitumor compound leiodermatolide, which we call leiodermatolides B and C, have been isolated from specimens of a deep-water sponge of the genus Leiodermatium collected off Florida. The compounds were purified using standard chromatographic methods, and the structures defined through interpretation of the HRMS and 1D and 2D NMR data. Leiodermatolide B (2) lacks the C-21 hydroxy group found in leiodermatolide and has equal potency as the parent compound, providing a simpler analogue for possible clinical development. It inhibits the proliferation of the AsPC-1 human pancreatic adenocarcinoma cell line with an IC50 of 43 nM. Leiodermatolide C (3) has a modified macrolide ring and is over 85-fold less potent with an IC50 of 3.7 µM against the same cell line. These compounds add to the knowledge of the pharmacophore of this class of potent antitumor agents.

Inhibition of IL-8 secretion on BxPC-3 and MIA PaCa-2 cells and induction of cytotoxicity in pancreatic cancer cells with marine natural products.

Pancreatic cancer presents one of the most negative prognosis of all cancers as it has usually metastasized by the time a patient is diagnosed. The American Cancer Society estimates that 93% of patients will die within 5 years of diagnosis, highlighting the need for new drugs to treat this disease. Interleukin 8 (IL-8) mediates the angiogenesis of tumors arising from Ras mutations, which are present in about 90% of pancreatic adenocarcinomas. Overexpression of IL-8 in pancreatic tumors is believed to promote tumor angiogenesis and to activate survival signaling pathways. A 96-well cell-based enzyme-linked immunosorbent assay was set up to screen the Harbor Branch Oceanographic Institute library of marine natural products to identify those with the ability to inhibit IL-8 production by BxPC-3 pancreatic cancer cells. Over 1000 fractions were screened, resulting in the identification of 10 known marine natural products with this ability. These compounds fall into four classes of compounds including the pyrroloiminoquinone alkaloids secobatzelline A and isobatzelline C; mycalamide A and B, onnamide A, discalamide A, and theopederin K from the mycalamide class of polyketides; the lipopeptide microcolin A; and the cyclic depsipeptides didemnin B and nordidemnin B. In addition, didemnin B, nordidemnin B, and theopederin K induce potent cytotoxicity against four pancreatic cancer cell lines tested. Many of these compounds have been previously reported to inhibit protein synthesis and the decrease in IL-8 production may be nonspecific. Nevertheless, this is a new activity for these compounds and inhibition of IL-8 secretion by pancreatic cancer cells can now be added to the previously reported antiangiogenic activities of the didemnins.

Leiodermatolide, a novel marine natural product, has potent cytotoxic and antimitotic activity against cancer cells, appears to affect microtubule dynamics, and exhibits antitumor activity.

Pancreatic cancer, the fourth leading cause of cancer death in the United States, has a negative prognosis because metastasis occurs before symptoms manifest. Leiodermatolide, a polyketide macrolide with antimitotic activity isolated from a deep water sponge of the genus Leiodermatium, exhibits potent and selective cytotoxicity toward the pancreatic cancer cell lines AsPC-1, PANC-1, BxPC-3, and MIA PaCa-2, and potent cytotoxicity against skin, breast and colon cancer cell lines. Induction of apoptosis by leiodermatolide was confirmed in the AsPC-1, BxPC-3 and MIA PaCa-2 cells. Leiodermatolide induces cell cycle arrest but has no effects on in vitro polymerization or depolymerization of tubulin alone, while it enhances polymerization of tubulin containing microtubule associated proteins (MAPs). Observations through confocal microscopy show that leiodermatolide, at low concentrations, causes minimal effects on polymerization or depolymerization of the microtubule network in interphase cells, but disruption of spindle formation in mitotic cells. At higher concentrations, depolymerization of the microtubule network is observed. Visualization of the growing microtubule in HeLa cells expressing GFP-tagged plus end binding protein EB-1 showed that leiodermatolide stopped the polymerization of tubulin. These results suggest that leiodermatolide may affect tubulin dynamics without directly interacting with tubulin and hint at a unique mechanism of action. In a mouse model of metastatic pancreatic cancer, leiodermatolide exhibited significant tumor reduction when compared to gemcitabine and controls. The antitumor activities of leiodermatolide, as well as the proven utility of antimitotic compounds against cancer, make leiodermatolide an interesting compound with potential chemotherapeutic effects that may merit further research.

Early effects of lasonolide a on pancreatic cancer cells.

Lasonolide A, a novel polyketide-derived macrolide, was previously identified from an extract of the marine sponge Forcepia sp. in an assay for protein kinase C (PKC) inhibitors. Cytotoxicity testing and profiling of lasonolide A in the National Cancer Institute (NCI) 60 cell panel screen revealed that it was potent toward a broad range of cell lines and also suggested a unique mechanism of action. Contrary to expected results, we found lasonolide A to be a strong activator of PKC in Panc-1 pancreatic carcinoma cells. Downstream mitogen-activated protein kinases, ERK 1/2 and p38 were also rapidly phosphorylated in response to lasonolide A, as was Akt. Microscopy studies revealed that lasonolide A induced blebbing and contraction of the cells within minutes of exposure, and the eventual loss of adherence. However, membrane integrity was maintained and the effects were reversible if lasonolide A was washed from the cells after their loss of adherence. Pretreatment of cells with a myosin II inhibitor, blebbistatin, slowed the early onset, but did not prevent the morphological effects of lasonolide A. Cells stained for actin filaments showed some reduction in stress fiber structure after lasonolide A exposure; however, it did not affect the polymerization of purified actin in vitro. Bisindolemaleimide, a PKC inhibitor, and wortmannin, a phosphoinositide 3-kinase; inhibitor, did not reduce lasonolide A-induced contraction or blebbing or the activation of mitogen-activated protein kinases, although Akt phosphorylation was prevented by wortmannin pretreatment. Our results indicate that lasonolide A activates multiple signal transduction pathways and suggest that the origin is upstream of PKC.

Isolation, synthesis, and biological activity of aphrocallistin, an adenine-substituted bromotyramine metabolite from the Hexactinellida sponge Aphrocallistes beatrix.

A new adenine-substituted bromotyrosine-derived metabolite designated as aphrocallistin (1) has been isolated from the deep-water Hexactinellida sponge Aphrocallistes beatrix. Its structure was elucidated on the basis of spectral data and confirmed through a convergent, modular total synthetic route that is amenable toward future analogue preparation. Aphrocallistin inhibits the growth of a panel of human tumor cell lines with IC(50) values ranging from 7.5 to >100 microM and has been shown to induce G1 cell cycle arrest in the PANC-1 pancreatic carcinoma cell line. Aphrocallistin has been fully characterized in the NCI cancer cell line panel and has undergone in vitro ADME pharmacological profiling.

Selective cytotoxic activity of the marine-derived batzelline compounds against pancreatic cancer cell lines.

Pancreatic cancer is the fourth leading cause of cancer death in the United States. The prognosis of the disease is very negative, because the cancer will be usually metastasized by the time a patient manifests symptoms. Although combination therapy shows some promise, new drugs to treat the disease are needed. Given our interest in finding new therapies for pancreatic cancer, we sought to determine whether the known cytotoxic activity of the batzellines extended to pancreatic cancer cell lines. The batzellines are pyrroloiminoquinones alkaloids obtained from the deep-water Caribbean sponge Batzella sp (family Esperiopsidae, order Poecilosclerida). We show here that batzellines exhibit selective cytotoxicity towards the pancreatic cancer cell lines AsPC-1, Panc-1, BxPC-3, and MIA PaCa2 compared with the normal African green monkey kidney epithelial cell line Vero. The batzellines cause cytotoxicity by inducing cell cycle arrest that is mediated by their ability to intercalate into DNA and/or inhibit topoisomerase II activity. The cytotoxic abilities of isobatzellines A and C against pancreatic cancer cell lines, their low toxicity against normal cells, and their reported ability to be synthesized makes them interesting compounds with potential chemotherapeutic effects that may merit further research.

Neopeltolide, a macrolide from a lithistid sponge of the family Neopeltidae.

A new marine-derived macrolide designated as neopeltolide (1) has been isolated from a deep-water sponge of the family Neopeltidae. Its structure was elucidated on the basis of spectroscopic data interpretation. Neopeltolide (1) is a potent inhibitor of the in vitro proliferation of the A-549 human lung adenocarcinoma, the NCI-ADR-RES human ovarian sarcoma, and the P388 murine leukemia cell lines, with IC50's of 1.2, 5.1, and 0.56 nM, respectively. Neopeltolide (1) also inhibited the growth of the fungal pathogen Candida albicans with a minimum inhibitory concentration of 0.62 microg/mL.

Lasonolides C-g, five new lasonolide compounds from the sponge Forcepia sp.

Five new marine-derived macrolide compounds, lasonolides C (3), D (4), E (5), F (6), and G (7), have been isolated from the sponge Forcepia sp. along with the parent compound in the series, lasonolide A (1). Their structures were elucidated on the basis of spectral data. Compounds 3-5 inhibit the in vitro proliferation of A-549 human lung adenocarcinoma cells with IC50's of 0.13, 4.5, and 0.31 microM, respectively. Compounds 3-6 inhibit the in vitro proliferation of PANC-1 human pancreatic carcinoma cells with IC50's of 0.38, 4.89, 0.57, and 15.6 microM, respectively. Compound 3 inhibits the in vitro proliferation of the NCI-ADR-RES cell line with an IC50 of 1.12 microM.