James McNulty

Pancratistatin: A natural anti-cancer compound that targets mitochondria specifically in cancer cells to induce apoptosis

The major hurdle in the fight against cancer is the non-specific nature of current treatments. The search for specific drugs that are non-cytotoxic to normal cells and can effectively target cancer cells has lead some researchers to investigate the potential anti-cancer activity of natural compounds. Some natural compounds, such as Taxol, have been shown to posses some anti-cancer potential. Pancratistatin (PST) is a natural compound that was isolated from the spider lily Pancratium littorale and shown to exhibit antineoplastic activity. The specificity of PST to cancer cells and the mechanism of PST’s action remain unknown. This study provides a detailed look at the effect of PST treatment on cancerous and normal cells. Our results indicate that PST induced apoptosis selectively in cancer cells and that the mitochondria may be the site of action of PST in cancer cells. A biochemical target available specifically in cancer cells may lead to the development of new and more effective cancer fighting agents.

The ultrasound promoted Knoevenagel condensation of aromatic aldehydes

Abstract Application of ultrasound has been found to greatly assist the Knoevenagel aldol condensation reaction of activated methylenes with aromatic aldehydes under mild conditions. The outcome of the ultrasound-promoted reaction depends upon the electronic nature of the armoatic aldehyde, the solvent employed and the addition of acids, bases or ammonium salts.

Pancratistatin causes early activation of caspase-3 and the flipping of phosphatidyl serine followed by rapid apoptosis specifically in human lymphoma cells.

Recently a major research effort has been focused on the development of anticancer drugs by targeting the components of a biochemical pathway to induce apoptosis in cancerous cells. Some of the natural products (e.g. paclitaxel) have been proven to be useful in inducing apoptosis in cancer cells with limited specificity. Pancratistatin, a natural product isolated and characterized over a decade ago, has been shown to be cytostatic and antineoplastic. We investigated the specificity and biochemical mechanism of action of pancratistatin. Pancratistatin seemed to show more specificity than VP-16 or paclitaxel as an efficient inducer of apoptosis in human lymphoma (Jurkat) cells, with minimal effect on normal nucleated blood cells. Caspase-3 activation and exposure of phosphatidyl serine on the outer leaflet of the plasma membrane were earlier events than the generation of ROS and DNA fragmentation observed following pancratistatin treatment. This indicates a possible involvement of caspase-3 and plasma membrane proteins in the induction phase of apoptosis. Our results indicate that pancratistatin does not cause DNA double-strand breaks or DNA damage prior to the execution phase of apoptosis in cancer cells. Parallel experimentation with VP-16, a currently used medication for cancer treatment, indicated that VP-16 causes substantial DNA damage in normal non-cancerous blood cells, while pancratistatin does not cause any DNA double-strand breaks or DNA damage in non-cancerous cells. Taken together, our finding that pancratistatin induces apoptosis in cancer cells using non-genomic targets, and more importantly does not seem to have any affect non-cancerous cells, presents a significant platform to develop non-toxic anticancer therapies.

The First Well‐Defined Silver(I)‐Complex‐Catalyzed Cycloaddition of Azides onto Terminal Alkynes at Room Temperature

The copper(I)-catalyzed Huisgen dipolar cycloaddition of terminal alkynes 1 with azides 2 to yield 1,4and 1,4,5-substituted 1,2,3-triazoles 3 (azide–alkyne cycloaddition; AAC reaction) has been transformed in recent years, since its description as the prototype “click” reaction, to become a general process with applications in diverse areas ranging from functional materials to biological chemistry. The generally accepted mechanism for the reaction, outlined in Scheme 1, involves a stepwise process initiated through generation of a copper(I) acetylide complex I, which complexes to the azide 2, undergoes the cycloaddition, generating a metalated triazole III, which is then protonated to yield the product 3, regenerating the catalyst.

Structure-activity studies on the lycorine pharmacophore: a potent inducer of apoptosis in human leukemia cells.

Abstract The direct chemoselective differential functionalization of the ring-C hydroxyl groups present in the Amaryllidaceae alkaloid lycorine is described allowing for selective manipulation of the 1,2-hydroxyl groups. A mini-library comprised of synthetic and natural lycorane alkaloids was prepared and their apoptosis-inducing activity investigated in human leukemia (Jurkat) cells. Further insights into the nature of this interesting apoptosis-inducing pharmacophore are described, including the requirement of both free hydroxyl groups in ring-C.

Microwave‐Assisted, Aqueous Wittig Reactions: Organic‐Solvent‐ and Protecting‐Group‐Free Chemoselective Synthesis of Functionalized Alkenes

The Wittig olefination reaction is regarded as one of the most strategic, widely applicable carbon–carbon doublebond-forming processes available in organic synthesis. The reaction has had an enormous impact on the sophistication of the total synthesis of organic molecules. Some drawbacks of the reaction are the lack of stereocontrol achieved in certain cases, for example, in the synthesis of stilbenes from semistabilised ylides, and the practical issue of phosphane oxide side-product removal. Also, protecting groups are usually required on any acidic protons (OH, NH, etc.) on both the ylide and carbonyl components. Water is a desirable solvent for organic reactions for environmental, economical, safety and chemical processing reasons. It has been used as the reaction medium for Wittig reactions of stabilised ylides to give unsaturated esters. Recently, we reported the first examples of aqueous Wittig reactions of semistabilised ylides derived from trialkylbenzyl and trialkylallyl phosphonium salts. Semistabilised triACHTUNGTRENNUNGethylbenzylidenyl and triethylallylidenyl ylides were shown to be formed chemoselectively in water by using sodium or lithium hydroxide and to react with aromatic, unsaturated, aliphatic and even enolisable aliphatic aldehydes in water, yielding a wide array of olefinic products (Scheme 1). These reactions proceeded with high (E)-olefin selectivity. The triethylphosphane oxide side-product is readily removed from these processes due to its water solubility and, hence, the Wittig reactions of triethylphosphane-derived, semistabilised ylides encapsulate a single solution to two outstanding problems with Wittig olefinations leading to (E)-olefins. This method was applied to the synthesis of valuable transstilbenes, such as resveratrol and trans-3,4,5,4’-tetramethACHTUNGTRENNUNGoxystilbene (DMU-212). High-purity trans-stilbenes are also the central component in light-emitting diodes (LEDs) and organic-based photovoltaic solar cells. In our original work, the phosphonium salts were prepared in the usual fashion, by direct substitution of benzylic or allylic halides with triethylphosphane. Triethylphosphane is a highly odoriferous lachrymator that undergoes rapid oxidation in air and is considered pyrophoric. Allyl and benzyl halides are also known lachrymators and are hydrolytically unstable, generally toxic, alkylating agents. We have now developed a direct alkylation strategy that circumvents these issues, allowing a safe, “off-the-shelf” approach to achieving the above Wittig chemistry. Triethylallyl and triethylbenzyl phosphonium salts are directly available from the reaction of a benzylic or allylic alcohol and air-stable triethylphosphane hydrobromide. We also uncovered a pronounced “microwave effect” in the aqueous olefination reaction, leading to successful Wittig reactions by using weak bases, such as potassium carbonate. The innate reactivity of these ylides in water drew our attention to chemoselectivity issues. We report the unprecedented protecting-group-free, aqueous Wittig reactions of phenols, indoles, pyrroles and ketones, including enolisable substrates. The chemistry employed in the direct synthesis of triethACHTUNGTRENNUNGylallyl and triethylbenzyl phosphonium salts is outlined in Scheme 2. The synthesis of allylic triphenylphosphonium salts from allylic alcohols and acidic Ph3P–HBr was first re[a] Dr. J. McNulty, Dr. P. Das, D. McLeod Department of Chemistry and Chemical Biology McMaster University, 1280 Main Street West Hamilton, Ontario, L8S 4M1 (Canada) Fax: (+1) 905-522-2509 E-mail : jmcnult@mcmaster.ca Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/chem.201000438. Scheme 1. Synthesis of stilbenes, alkenes, and dienes (etc.) by using aqueous Wittig chemistry with semistabilised ylides.

Studies directed towards the refinement of the pancratistatin cytotoxic pharmacophore.

Two deoxy-analogues of the anticancer/antiviral agent pancratistatin containing functionality complementary to the minimum structural pharmacophore were synthesized and subjected to anticancer screening. One of the analogues exhibited selective inhibition of certain tumor cell lines but was significantly less potent than the natural products. The minimum structural pharmacophore has now been refined from eight to three possible structures.

Pancratistatin Selectively Targets Cancer Cell Mitochondria and Reduces Growth of Human Colon Tumor Xenografts

The naturally occurring Amaryllidaceae alkaloid pancratistatin exhibits potent apoptotic activity against a large panel of cancer cells lines and has an insignificant effect on noncancerous cell lines, although with an elusive cellular target. Many current chemotherapeutics induce apoptosis via genotoxic mechanisms and thus have low selectivity. The observed selectivity of pancratistatin for cancer cells promoted us to consider the hypothesis that this alkaloid targets cancer cell mitochondria rather than DNA or its replicative machinery. In this study, we report that pancratistatin decreased mitochondrial membrane potential and induced apoptotic nuclear morphology in p53-mutant (HT-29) and wild-type p53 (HCT116) colorectal carcinoma cell lines, but not in noncancerous colon fibroblast (CCD-18Co) cells. Interestingly, pancratistatin was found to be ineffective against mtDNA-depleted (ρ0) cancer cells. Moreover, pancratistatin induced cell death in a manner independent of Bax and caspase activation, and did not alter β-tubulin polymerization rate nor cause double-stranded DNA breaks. For the first time we report the efficacy of pancratistatin in vivo against human colorectal adenocarcinoma xenografts. Intratumor administration of pancratistatin (3 mg/kg) caused significant reduction in the growth of subcutaneous HT-29 tumors in Nu/Nu mice (n = 6), with no apparent toxicity to the liver or kidneys as indicated by histopathologic analysis and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling. Altogether, this work suggests that pancratistatin may be a novel mitochondria-targeting compound that selectively induces apoptosis in cancer cells and significantly reduces tumor growth. Mol Cancer Ther; 10(1); 57–68. ©2011 AACR.

Synergy of Pancratistatin and Tamoxifen on breast cancer cells in inducing apoptosis by targeting mitochondria.

Pancratistatin (PST), a natural compound obtained from the Hawaiian spider lily, is known to be specific and selective in inducing apoptosis in multiple cancer cell lines while sparing non-cancerous cells and cell lines. Here we report the ability of PST to induce apoptosis specifically in human breast cancer cell lines MCF-7 and Hs-578-T compared to their non cancerous counterparts. In cancer cells PST caused increased levels of reactive oxygen species (ROS), decreased ATP and mitochondrial membrane permeabilization indicating the activation of the mitochondrial pathway of apoptosis. In combination with the anti-estrogen Tamoxifen, PST had a synergic effect. Both compounds caused increased production of ROS when applied to isolated mitochondria from these cancer cell lines supporting the observation that Tamoxifen might work through mechanisms distinct from the canonical estrogen receptor antagonism.

Suzuki cross-coupling reactions of aryl halides in phosphonium salt ionic liquid under mild conditions

The Suzuki cross-coupling of aryl boronic acids with aryl halides, including aryl chlorides, proceeds in the phosphonium salt ionic liquid tetradecyltrihexylphosphonium chloride under mild conditions.