Gilberto Schwartsmann

Natural products in anticancer therapy.

Many pharmaceutical agents have been discovered by screening natural products from plants, animals, marine organisms and microorganisms. Vincristine, irinotecan, etoposide and paclitaxel are examples of plant-derived compounds that are being employed in cancer treatment, and dactinomycin, bleomycin and doxorubicin are anticancer agents derived from microbial sources. Citarabine is an example of an anticancer agent originating from a marine source. Other agents originating from marine sources are bryostatin-1, aplidine, dolastatin 10 and ET-743, which have recently entered phase I and II clinical trials.

Marine organisms as a source of new anticancer agents

Various active anticancer agents are derived from plants and terrestrial microorganisms. The isolation of C-nucleosides from the Caribbean sponge, Cryptotheca crypta, four decades ago, provided the basis for the synthesis of cytarabine, the first marine-derived anticancer agent to be developed for clinical use. Cytarabine is currently used in the routine treatment of patients with leukaemia and lymphoma. Gemcitabine, one of its fluorinated derivatives, has also been approved for use in patients with pancreatic, breast, bladder, and non-small-cell lung cancer. Over the past decade, several new experimental anticancer agents derived from marine sources have entered preclinical and clinical trials. This field has expanded significantly as a result of improvements in the technology of deep-sea collection, extraction, and large-scale production through aquaculture and synthesis. In this paper, examples of marine-derived experimental agents that are currently undergoing preclinical and early clinical evaluation are briefly discussed. A summary of the available information on the results of phase I and II trials of agents such as aplidine, ecteinascidin-734 (ET-734), dolastatin 10 and bryostatin 1 is also presented.

Extracellular nucleotides and nucleosides induce proliferation and increase nucleoside transport in human glioma cell lines

Extracellular purines (adenosine triphosphate (ATP), adenosine 5′-diphosphate (ADP) and adenosine) and pyrimidines (uridine 5′-triphosphate (UTP) and UDP) are important signaling molecules that mediate diverse biological effects via P1 and P2 purinergic receptors. The human glioma cell lines U87 MG, U251 MG and U138 MG were treated with purines and pyrimidines for 24 or 48h and proliferation was measured by [3H]-thymidine incorporation, flow cytometry and cell counting. The studies showed that extracellular nucleotides and nucleosides induce proliferation of the studied glioma cells. Incorporation of [3H]-thymidine followed the order of ATP ≅ guanosine ≅ inosine ≅ adenosine > UTP > ADP while ATPγS and 2MeSATP had no effect. The effect of ATP was partially inhibited by suramin and by reactive blue 2 (RB2). Co-treatment with the following antagonists of P1 purinoreceptors DPCPX, CPT or 8PT did not block the effect of adenosine while a specific antagonist of the A3 receptor, MRS1220, totally blocked the effect of adenosine. ATP and adenosine also increased the overall uptake of [3H]-thymidine into the cell, producing a positive effect on the [3H]-thymidine incorporation measurements. These data indicate that the uptake of thymidine and proliferation of gliomas can be induced by purines and pyrimidines via both P1 and P2 purinoceptors.

Anticancer, antichemotactic and antimicrobial activities of marine sponges collected off the coast of Santa Catarina, southern Brazil

Abstract This study reports the in vitro screening of 10 marine sponges (Porifera) collected from the coastline of Santa Catarina, southern Brazil, in the search for novel pharmaceuticals. Organic and aqueous extracts were tested for anticancer, antibacterial, antifungal and antichemotactic activities. Eight of the ten species tested demonstrated activity in one or more of the bioassays. Organic extracts of Polymastia janeirensis Boury-Esnauls, 1973, Haliclona aff tubifera George and Wilson, 1919, Mycale arcuiris Lerner and Hajdu, 2002 and Raspailia ( syringella ) sp. each demonstrated cytotoxicity at 100 μg/ml in an in vitro screening assay against the HT29 colorectal tumour cell line. Further analysis against three human tumour cell lines (HT29, U373 and NCI-H460) demonstrated IC 50 concentrations ranging from 25 to 50 μg/ml. Aqueous extracts of six species P. janeirensis , M. arcuiris , Raspailia ( syringella ) sp., Guitarra sp., Tedania ignis Duchassaing and Michelotti, 1864 and Pseudaxinella reticulata Ridley and Dendy, 1886 each significantly ( p ≤0.05) retarded the migration of polymorphonuclear (PMN) leukocytes in a chemotactic assay. In antibacterial assays, only H. aff tubifera (four of five bacterial strains) and Axinella corrugata George and Wilson, 1919 (one of five bacterial strains) demonstrated activity. None of the 10 species demonstrated measurable antifungal activity. These extracts are currently undergoing further analysis to identify the active constituents.

Intrahippocampal infusion of the bombesin/gastrin-releasing peptide antagonist RC-3095 impairs inhibitory avoidance retention

Bombesin (BN)-like peptides regulate cell proliferation and cancer growth as well as neuroendocrine and neural functions. We evaluated the effects of the BN/gastrin-releasing peptide (GRP) antagonist RC-3095 on memory formation. Male Wistar rats were given a bilateral infusion of saline or RC-3095 (0.2, 1.0 or 5.0 microg) into the dorsal hippocampus either immediately or 2 h after training in an inhibitory avoidance (IA) task. Retention test trials were carried out 1.5 h (short-term memory) and 24 h (long-term memory) after training. RC-3095 impaired both short- and long-term retention only when given immediately after training. The results suggest that the hippocampal BN/GRP receptor system modulates IA memory formation.

Comparative genotoxic effect of vincristine, vinblastine, and vinorelbine in somatic cells of Drosophila melanogaster.

In this study, the vinca alkaloids vincristine (VCR), vinblastine (VBL) and vinorelbine (VNR) were investigated for genotoxicity in the wing Somatic Mutation and Recombination Test (SMART) of Drosophila. Our in vivo experiments demonstrated that all drugs assessed induced genetic toxicity, causing increments in the incidence of mutational events, as well as in somatic recombination. Another point to be considered is the fact that VNR was able to induce, respectively, approximately 13.0 and 1.7 times more mutant clones per millimolar exposure unit as their analogues VCR and VBL. The replacement of a CH(3) attached to vindoline group in VBL by a CHO in VCR seems to be responsible for the approximately seven times higher potency of the former. In contrast, the structural modifications on VNRs catharantine group could be related to its higher genotoxic potency, as well as its similar mutagenic and recombinagenic action.

Protein Kinase C-Mediated in vitro Invasion of Human Glioma Cells through Extracellular-Signal-Regulated Kinase and Ornithine Decarboxylase

We investigated the involvement of protein kinase C (PKC) in the in vitro invasiveness of the A-172, U-87 and U-373 human glioma cell lines, as well as the role of ornithine decarboxylase (ODC) and/or extracellular-signal-regulated kinase (ERK) in the actions of PKC. Thus, cells were treated under serum-free conditions with the PKC activator phorbol 12-myristate 13-acetate (PMA), or with the PKC inhibitors bisindolylmaleimide I (GF 109203X) or calphostin C in the absence or presence of the ODC inhibitor D,L-α-difluoromethylornithine (DFMO), and/or the mitogen-activated protein kinase/extracellular-signal-regulated kinase inhibitor 2′-amino-3′-methoxyflavone (PD 098059). Subsequently, cells were assessed for membrane-type 1 matrix metalloproteinase (MT1-MMP) mRNA contents, 72-kD latent, and 59/62-kD activated matrix metalloproteinase 2 (MMP-2) in conditioned media, as well as invasiveness. For these purposes, we used Northern blot analysis, gelatine zymography, and an in vitro filter invasion assay, respectively. Data were related to those found with untreated cells. PKC activity was 2- to 3-fold stimulated by PMA (100 nM for 30 min), and about 2-fold inhibited by calphostin C (40 nM for 2 h) or GF 109203X (5 µM for 20 min). This was accompanied by a similar increase or decrease, respectively, in MT1-MMP mRNA expression, 59/62-kD MMP-2 activity, and in vitro invasion. Inhibition of ODC activity (about 2-fold by 24 h DFMO 5 mM), ERK activation (almost completely by 20 min PD 098059 50 µM), or both these enzymes simultaneously led to a reduction by about half in levels of MT1-MMP mRNA, 59/62-kD MMP-2 activity, and invasion in untreated as well as PMA-stimulated cells. The use of these compounds did not significantly alter the inhibitory effects of GF 109203X or calphostin C. Modulation of PKC and/or ERK activity resulted in corresponding changes in ERK and/or ODC activities, but interference with ODC affected neither ERK nor PKC. Our data suggest a regulatory role for PKC, in co-operation with ERK and ODC, in glioma cell invasion, by modulation of MT1-MMP mRNA expression and MMP-2 activation.

Irinotecan and oxaliplatin: an overview of the novel chemotherapeutic options for the treatment of advanced colorectal cancer.

Colorectal cancer is one of the most frequent malignancies in humans and an important cause of cancer death. Metastatic colorectal cancer remains incurable with available systemic therapeutic options. The most active cytotoxic drug against this malignancy, the antimetabolite 5-fluorouracil, was developed more than forty years ago, and as a single agent produces responses in only 10 to 15% of patients which in general last less than one year. Efforts to ameliorate these poor results resulted in the 5-fluorouracil/leucovorin combination, which enhances response rates about two-fold, without, however, significantly improving survival rates. The recent emergence of a handful of new 5-fluorouracil analogues and folate antagonists, as well as the topoisomerase I inhibitor irinotecan, and the third-generation platinum compound oxaliplatin, is likely to alter this gloomy scenario. These agents are at least as effective as 5-fluorouracil in patients with advanced colorectal carcinoma, both untreated and previously treated with 5-fluorouracil-based regimens. This has led to the approval of irinotecan as second-line treatment for 5-fluorouracil-refractory disease, while the use of oxaliplatin has been suggested for patients having a defective 5-fluorouracil catabolism. Recently, FDA approved the combination of irinotecan with 5-fluorouracil and leucovorin for first-line treatment of advanced colon cancer. Based on the synergistic preclinical antitumor effects of some of these agents, their meaningful single-agent activity, distinct mechanisms of cytotoxicity and resistance, and only partially overlapping toxicity profiles, effective combination regimens are now being developed, which are likely to lead to a new, more hopeful era for patients suffering from advanced colorectal carcinoma.

Modulation of Oxidative Stress in Response to Gamma-radiation in Human Glioma Cell Lines

Radiation therapy is routinely used in the management of primary central nervous system malignancies. However, the efficacy of this therapeutic modality is limited by the occurrence of resistance. In the present study, we investigated whether modulation of oxidative stress might underlie glioma cell radioresistance. Superoxide dismutase activity in irradiated M059J cells was two-fold higher than that in untreated controls, but did not significantly change in U-87 and U-138 cells. This is accompanied by an increase in reactive oxygen species content and decreases in cells viability. Pharmacological or genetic modulation of oxidative stress could be associated with an enhancement in the susceptibility of tumor cells to radiation therapy.