Clinical trials using chemopreventive vitamin D analogs in breast cancer

This article comprehensively reviews the clinical trials and considers the future directions of the use of vitamin D and its analogs in the treatment or chemoprevention of breast cancer. Chemopreventive treatment strategies strive to delay the onset of certain cancers, prevent the progression of malignant disease after diagnosis, or delay the advent of recurrence after curative treatment. We first summarize the epidemiological evidence that led to the hypothesis that vitamin D may have an anti-cancer activity. Vitamin D shows great potential as a therapy for breast cancer; however, its use in clinical trials has been hindered by the induction of hypercalcemia at a concentration required to suppress cancer cell proliferation. This has led to the development of less calcemic analogs of vitamin D. We review the clinical trials with breast cancer patients using vitamin D analogs.     

Pharmacokinetics of the chemopreventive agent oltipraz and of its metabolite M3 in human subjects after a single oral dose.

The dithiolethione oltipraz (OPZ) has activity as a chemopreventive agent in animal models and is in early clinical trials. OPZ undergoes metabolism by molecular rearrangement to yield a pyrrolopyrazine derivative, M3, which we have previously shown to be inactive in the induction of detoxication genes. M3 is metabolized further: at least 10 possible conjugates have been described in three species. We developed a new high-performance liquid chromatography method to simultaneously measure plasma concentrations of OPZ and of M3. This method was applied to serial plasma samples in a Phase I clinical trial, in which OPZ was administered at single doses varying from 125 to 1000 mg/m2. OPZ and M3 concentration-time profiles were highly variable among individuals, and the occurrence of secondary concentration peaks suggested substantial enterohepatic cycling. Absorption was rapid, and the mean time to peak was 2.2 h. Maximum plasma concentration values were proportional to the dose. Harmonic mean half-lives at these doses ranged from 9.3-22.7 h. There were indications of dose-dependent pharmacokinetic properties because apparent clearance and volume of distribution at steady state increased with dose, although these changes were not statistically significant as a result of high interpatient variability. Accordingly, there were less than proportional increases in the OPZ and M3 area under the curve and maximum plasma values. Interpretation of OPZ and M3 disposition is confounded by the unknown bioavailability factor; however, the most likely inferences are that bioavailability of OPZ decreases with increasing dose and that metabolism to M3 is saturable.     

The chemopreventive efficacy of inhaled oltipraz particulates in the B[a]P-induced A/J mouse lung adenoma model

This study explored the efficacy of oltipraz, a dithiolthione to prevent lung cancer by delivering it directly to the lung as inhaled particulates to obtain maximum efficacy with no toxicity. Two exposure regimens were used to compare the efficacies of early (Regimen-A) versus late (Regimen-B) intervention in prevention of lung tumorigenesis in A/J mice. Female A/J mice were exposed to 10, 30 and 100 mg/m(3) exposure concentrations of oltipraz for 1.0 h a day for 5 days per week for 4 weeks in Regimen A. During the second and third week, mice received totally 6 mg of B[a]P via gavage and after 16 weeks, they were killed for tumor counting and pathology. In Regimen B, mice were treated first with B[a]P and, after a gap of 4 weeks, exposed to oltipraz at 100 mg/m(3) for 16 additional weeks. At 22 weeks, animals were killed and necropsied for tumor scoring. The spontaneous tumors were few in untreated A/J mice (0.7 tumors/lung), whereas there was an average of 16.5 tumors per lung in the B[a]P group (20-fold induction). Evaluation of lung tumor multiplicity following exposure to oltipraz showed that oltipraz inhibited the tumor development in a dose-dependent manner (10-100 mg/m(3)) with inhibition ranging from 37 to 53% in Regimen A and 51% in Regimen B, when compared with the B[a]P group. Analysis of the tumor incidence showed that 81.5% of the animals had 10 or more tumors in the B[a]P group, whereas, in oltipraz exposure groups, there was a significant decrease in Regimen A (24-36%) and in Regimen B (42%). The data from this study show that oltipraz is an effective agent for lung cancer prevention, when it is delivered directly to the target tissue as aerosolized particulates.     

Role of chemopreventive agents in cancer therapy

Tumorigenesis or carcinogenesis is a multi-step process that is induced primarily by carcinogens leading to the development of cancer. Extensive research in the last few years has revealed that regular consumption of certain fruits and vegetables can reduce the risk of acquiring specific cancers. Phytochemicals derived from such fruits and vegetables, referred to as chemopreventive agents include genistein, resveratrol, diallyl sulfide, S-allyl cysteine, allicin, lycopene, capsaicin, curcumin, 6-gingerol, ellagic acid, ursolic acid, silymarin, anethol, catechins and eugenol. Because these agents have been shown to suppress cancer cell proliferation, inhibit growth factor signaling pathways, induce apoptosis, inhibit NF-kappaB, AP-1 and JAK-STAT activation pathways, inhibit angiogenesis, suppress the expression of anti-apoptotic proteins, inhibit cyclooxygenase-2, they may have untapped therapeutic value. These chemopreventive agents also have very recently been found to reverse chemoresistance and radioresistance in patients undergoing cancer treatment. Thus, these chemopreventive agents have potential to be used as adjuncts to current cancer therapies.     

Nigerian foodstuffs with prostate cancer chemopreventive polyphenols

Dietary polyphenols are antioxidants that can scavenge biological free radicals, and chemoprevent diseases with biological oxidation as their main etiological factor. In this paper, we review our laboratory data vis-?-vis available literature on prostate cancer chemopreventive substances in Nigerian foodstuffs. Dacryodes edulis fruit, Moringa oleifera and Syzygium aromaticum contained prostate active polyphenols like ellagic acid, gallate, methylgallate, catechol, kaempferol quercetin and their derivatives. Also Canarium schweinfurthii Engl oil contained ten phenolic compounds and lignans, namely; catechol, p-hydroxybenzaldehyde, dihydroxyphenylacetic acid, tyrosol, p-hydroxybenzoic acid, dihydroxybenzoic acid, vanillic acid, phloretic acid, pinoresinol, secoisolariciresinol. In addition, tomatoes (Lycopersicon esculentum Mill) which contains the powerful antioxidant and anti-prostate cancer agent, lycopene; cabbage (Brassica oleracea) containing indole-3-carbinol; citrus fruits containing pectin; Soursop (Annona muricata) containing annonaceous acetogenins; soya beans (Glycine max) containing isoflavones; chilli pepper (Capsicum annuum) containing capsaicin, and green tea (Camellia sinensis) containing (-) epigallocatechin gallate (EGCG), (-) epicatechin, (-) epicatechin-3-gallate and (-) epigallocatechin -3-gallate which are widely reported to posses prostate cancer chemopreventive compounds are also grown in Nigeria and other African countries. Thus, the high incidence of prostate cancer among males of African extraction can be dramatically reduced, and the age of onset drastically increased, if the population at risk consumes the right kinds of foods in the right proportion, beginning early in life, especially as prostate cancer has a latency period of about 50 years.     

Beer constituents as potential cancer chemopreventive agents

Beer is a complex alcoholic beverage made from barley (malt), hop, water and yeast. Phenolic constituents of beer are derived from malt (70-80%) and hop (20-30%). Structural classes include simple phenols, benzoic- and cinnamic acid derivatives, coumarins, catechins, di-, tri- and oligomeric proanthocyanidins, (prenylated) chalcones and flavonoids as well as alpha- and iso-alpha-acids derived from hop. Compounds belonging to different structural classes have distinct profiles of biological activity in in vitro test systems, and in combination might lead to enhanced effects. Scientific evidence has accumulated over the past 10 years pointing to the cancer preventive potential of selected hop-derived beer constituents, i.e., prenylflavonoids including xanthohumol and isoxanthohumol, and hop bitter acids. Chemopreventive activities observed with these compounds relevant to inhibition of carcinogenesis at the initiation, promotion and progression phases, as well as results from in vivo studies on metabolism, bioavailability and efficacy are summarised in this review.     

Using chemopreventive agents to enhance the efficacy of cancer therapy

Emerging evidence suggests that cancer preventative agents might be combined with chemotherapy or radiotherapy for the more effective treatment of cancer. Recent studies suggest that genistein and other dietary compounds that prevent cancer may enhance the efficacy of cancer therapeutics by modifying the activity of key cell proliferation and survival pathways, such as those controlled by Akt, nuclear factor-kappaB, and cyclooxygenase-2. In this article, we summarize the findings of recent investigations of chemopreventive agents in combination with cancer treatment regimens.     

Use of cancer chemopreventive phytochemicals as antineoplastic agents

A lot of information has been gathered on cellular mechanisms by which chemopreventive phytochemicals, such as curcumin (a spice in curry) or epigallocatechin gallate (extracted from tea), interfere with carcinogenesis. A comparison of this knowledge with what we know about molecularly targeted chemotherapeutic agents suggests that it might be worthwhile to investigate the usefulness of such phytochemicals in the treatment of established malignant diseases. Phytochemicals use a plethora of antisurvival mechanisms, boost the host's anti-inflammatory defence, and sensitise malignant cells to cytotoxic agents. The restricted systemic availability of agents such as curcumin and epigallocatechin gallate, needs to be taken into account if they are to be developed as cochemotherapeutic drugs.     

Retinoids as chemopreventive agents for breast cancer.

Although several retinoids have been evaluated for prevention of mammary carcinogenesis in rats and mice, retinyl acetate (RA) and N-(4-hydroxyphenyl)retinamide (4-HPR) proved most effective. In rats, dietary administration of the retinoids reduced the incidence and number, and increased the latency of N-methyl-N-nitrosourea (MNU)-induced mammary cancers. 4-HPR reduced the number of hyperplastic alveolar nodules (HAN) in MTV- mice and the number of tumors in MTV+ mice. Other studies indicate that the synergistic effect of retinoid administration and hormonal deprivation is more efficacious in prevention of MNU-induced mammary cancer than either modality alone. Furthermore, retinoids alone and the combination of retinoid and tamoxifen inhibit the appearance of mammary cancers following the surgical removal of the first cancer as well as inhibit the growth of established cancers. Again, the combined modality was the most effective. Retinoids also exert an antiproliferative effect upon the mammary epithelium in vivo, which is represented morphologically by a bare duct system with little branching, end buds, and few, if any, alveoli. In organ culture, retinoids inhibit mammary end bud differentiation and proliferation induced by insulin and prolactin or carcinogens.     

Inhibition of angiogenesis by the cancer chemopreventive agent conjugated linoleic acid

Dietary conjugated linoleic acid (CLA) has been shown previously to inhibit rat mammary carcinogenesis. In addition to direct effects on mammary epithelial cells,including decreased proliferation and induction of apoptosis, CLA may exert its effects indirectly by inhibiting the differentiation of mammary stromal cells to an endothelial cell type. Specifically, CLA was found to decrease the ability of mammary stromal cells to form complex anastomosing microcapillary networks in vitro on Engelbreth-Holm-Swarm-derived reconstituted basement membrane. This suggested that CLA might inhibit angiogenesis in vivo. To test this possibility, CD2/F(1) mice were placed on synthetic diets containing 0, 1, or 2% CLA for 6 weeks, before angiogenic challenge by s.c. injection with an angiogenic gel substrate (Matrigel pellet assay). After 7 days, the pellets from animals fed the control diet were infiltrated by abundant branching networks of blood vessels with patent lumen-containing RBCs. In contrast, pellets from the CLA-fed animals contained fewer infiltrating cells, which formed limited branching cellular networks, the majority of which had collapsed lumen and no RBCs. Both levels of dietary CLA showed similar effects, with the number of RBC-containing vessels per 20x field decreased to a third of that seen in control. Dietary CLA decreased serum levels of vascular endothelial growth factor (VEGF) and whole mammary gland levels of VEGF and its receptor Flk-1. Both cis-9, trans-11 and trans-10, cis-12 CLA isomers were effective in inhibiting angiogenesis in vitro in a dose-dependent fashion. The ability of CLA to inhibit angiogenesis may contribute to its efficacy as a chemopreventive agent.     

Evaluation of the cancer chemopreventive efficacy of rice bran in genetic mouse models of breast, prostate and intestinal carcinogenesis

Brown rice is a staple dietary constituent in Asia, whereas rice consumed in the Western world is generally white, obtained from brown rice by removal of the bran. We tested the hypothesis that rice bran interferes with development of tumours in TAg, TRansgenic Adenocarcinoma of the Mouse Prostate (TRAMP) or Apc(Min) mice, genetic models of mammary, prostate and intestinal carcinogenesis, respectively. Mice received rice bran (30%) in AIN-93G diet throughout their post-weaning lifespan. In TAg and TRAMP mice, rice bran did not affect carcinoma development. In TRAMP or wild-type C57Bl6/J mice, dietary rice bran increased kidney weight by 18 and 20%, respectively. Consumption of rice bran reduced numbers of intestinal adenomas in Apc(Min) mice by 51% (P<0.01), compared to mice on control diet. In parallel, dietary rice bran decreased intestinal haemorrhage in these mice, as reflected by increased haematocrit. At 10% in the diet, rice bran did not significantly retard Apc(Min) adenoma development. Likewise, low-fibre rice bran (30% in the diet) did not affect intestinal carcinogenesis, suggesting that the fibrous constituents of the bran mediate chemopreventive efficacy. The results suggest that rice bran might be beneficially evaluated as a putative chemopreventive intervention in humans with intestinal polyps.     

The chemopreventive agent oltipraz possesses potent antiangiogenic activity in vitro, ex vivo, and in vivo and inhibits tumor xenograft growth.

Angiogenesis plays a pivotal role in tumor growth and represents a key target for chemopreventive intervention. On the basis of the structural features and lack of target organ specificity of the synthetic dithiolethione oltipraz, inhibition of angiogenesis was assessed as a potential mechanism for its broad-based chemopreventive activity. The effects of oltipraz on the development and maturation of a vascular network was determined in vitro using two-dimensional capillary tube formation assays with human umbilical vein endothelial cells plated on Matrigel and ex vivo using primary rat aortic ring explant cultures in three-dimensional collagen gels, respectively. The antiangiogenic and antitumor efficacy of oltipraz administration in vivo in nude mice was evaluated by determining its effects on neovascularization in s.c. Matrigel implants seeded with vascular endothelial growth factor and basic fibroblast growth factor-stimulated porcine aortic endothelial cells and on tumor growth and angiogenesis in SVR murine angiosarcoma xenografts implanted s.c. A dose-dependent reduction (0.4-100 microM) in microvessel formation was observed in both human and rodent bioassays after oltipraz exposure, with inhibition approaching 100% in the rat aortic ring assay at the highest concentration (P < 0.01). Similarly, oltipraz (40 microM) inhibited complete capillary tube formation by human umbilical vein endothelial cells by 62% (P < 0.05) relative to control cultures. p.o. administration of oltipraz (250 mg/kg/day for 6 days) to nude mice implanted with porcine aortic endothelial cell-Matrigel plugs resulted in a 42% reduction in neovascularization (P < 0.05) relative to vehicle-treated control mice. Administration of the same dose of oltipraz to athymic mice bearing established s.c. SVR angiosarcoma xenografts for 10 days resulted in a significant inhibition of tumor growth as early as day 4 of dosing (P < 0.005), with a maximum inhibition of tumor growth (81%, P < 0.001) relative vehicle-treated mice by day 10. The observed efficacy of oltipraz in this model is comparable with that of SU 5416 and TNP-470, known antiangiogenic agents currently under clinical development. Plasma levels of oltipraz at the termination of in vivo efficacy studies were 66.4 +/- 7 microM as determined by reversed phase high-performance liquid chromatography, a concentration range associated with significant antiangiogenic activity of oltipraz in vitro and ex vivo. These data suggest that the chemopreventive agent oltipraz may be effective in the treatment of advanced stage cancers and metastases, in part, because of its antiangiogenic activity in vivo.     

Chronic dosing of oltipraz in people at increased risk for colorectal cancer.

The dithiolethione oltipraz is being developed as a chemopreventive agent for many malignancies, including colorectal cancer, on the basis of its in vivo protective activity against chemically induced tumors in a variety of animal models. This protection has been associated with an enhanced capacity to detoxify reactive carcinogens and, more recently, with increased DNA repair. In a previous single-dose study, elevated detoxification gene expression was observed in the days after oltipraz dosing. Now, in this clinical study, we evaluated the effects of oltipraz when given over a 3-month period. Fourteen individuals with increased risk for colorectal cancer were randomly assigned to one of two oral doses (125 or 250 mg/m2) of oltipraz twice weekly for 12 weeks. Two of seven subjects at the 250 mg/m2 dosage required dose reductions, owing to significant fatigue. The 125 mg/m2 dose level was well tolerated by all patients. Blood or colon tissue (or both) for evaluation of glutathione, glutathione S-transferase, DT-diaphorase activity, and DT-diaphorase mRNA expression were obtained prior to treatment and at weeks 6, 12, and 16. No significant modulation of phase II detoxification enzymes was seen at either dose studied during this period. Phase II trials evaluating a tolerable regimen of oltipraz (as demonstrated in this study) and other possible mechanisms that may be responsible for the protective activity of oltipraz should be pursued.     

Colon cancer chemopreventive drugs modulate integrin-mediated signaling pathways.

Epidemiological studies of colorectal cancer incidence suggest that the development of this disease can be modulated by dietary factors. Among the micronutrients showing significant efficacy in tumor prevention are polyphenolic antioxidants found in fruits and vegetables. Epidemiological studies also indicate that nonsteroidal anti-inflammatory drugs (NSAIDs) decrease the incidence of colorectal cancer. Integrin-mediated cell-matrix contact provides critical signaling that regulates cellular proliferation, migration, and apoptosis. A signaling mediator for this system is focal adhesion kinase (FAK). Thus far, FAK has not been identified as a target for the inhibitory action of any chemopreventive drug in vivo or in vitro. However, the loss of integrin-mediated cell-matrix contact can induce apoptosis (anoikis), and effective chemopreventive agents typically increase the rate of enterocyte apoptosis. Therefore, we asked whether the NSAID, sulindac sulfide, and the phenolic antioxidant, caffeic acid phenethyl ester (CAPE), affected FAK expression or tyrosine phosphorylation in human colon carcinoma cells. We show that subapoptotic doses of both sulindac sulfide and CAPE caused a rearrangement of the actin cytoskeleton and consequently the loss of focal adhesion plaques. These drugs also reduced the tyrosine phosphorylation of FAK and an associated factor, p130Cas. Steady-state levels of these proteins, together with other relevant signaling molecules, remained unchanged after treatments. Finally, we show that both CAPE and sulindac reduced cell invasion, a functional assay for the inhibition of signaling downstream of FAK. These data strongly suggest that chemopreventive drugs can regulate FAK activity. In conclusion, these novel studies add modulation of integrin-mediated signaling to the spectrum of activity of NSAIDs and plant phenolics.     

Camptothecin analogs with enhanced activity against human breast cancer cells. I. Correlation of potency with lipophilicity and persistence in the cleavage complex

The effect of 7-alkyl substitutions on growth inhibition in seven Camptothecin (CPT) ring systems with various groups at the ten position was evaluated in three human breast cancer cell lines that model (1) hormone-sensitive (MCF-7/wt), (2) hormone insensitive (MDA-MB-231), or (3) alkylator-resistant (MCF-7/4-hc) forms of disease. To assess the impact of persistence of cleavage complexes on antiproliferative activity, a post-exposure recovery period in drug-free medium was incorporated into the growth inhibition assay. This modification produced on average a twofold reduction in the growth inhibition endpoint (the IC50), suggesting a greater apoptotic response. The results further revealed a three log range in potency from a mean IC₅₀ of 2 nM (7-butyl-10,11-methylenedioxy-CPT) to 2.5 μM (7-bromomethyl-10-hydryoxy-CPT). Increasing 7-alkyl chain length in six of the ten-substituted CPTs enhanced potency, which was directly correlated with persistence of topoisomerase I-induced DNA cleavage complexes in 10-hydroxy, 10-methoxy, and 10,11-methylenedioxy substituted CPTs. Modeling of the binding mode of 7-butyl-10-amino-CPT revealed a direct hydrogen bond contact for the 10-amino to the side chain of Glu-356 of Core Subdomain I of top1 in addition to known contacts found for other camptothecins. More important, residues 350–356 and 425–431 of Core Subdomain I may provide induced fit stabilization to the lipophilic alkyl moiety at the seven position.