Distinct mechanisms of bisphosphonate action between osteoblasts and breast cancer cells: identity of a potent new bisphosphonate analogue

While the effects of bisphosphonates on bone-resorbing osteoclasts have been well documented, the effects of bisphosphonates on other cell types are not as well studied. Recently, we reported that bisphosphonates have direct effects on bone-forming human fetal osteoblast cells (hFOB) [1]. In this report, the role of the mevalonate pathway in the actions of bisphosphonates on hFOB, and MDA-MB-231 human breast cancer cells was examined. These studies included a novel bisphosphonate analog, the anhydride formed between arabinocytidine 5 phosphate and etidronate (Ara-CBP). Ara-CBP was the most potent inhibitor of hFOB and MDA-MB-231 cell proliferation, and stimulator of hFOB cell mineralization compared to etidronate, the anhydride formed between AMP and etidronate (ABP), pamidronate, and zoledronate. Inhibition of hFOB cell proliferation by Ara-CBP and zoledronate was partially reversed by mevalonate pathway intermediates, and stimulation of hFOB cell mineralization was completely reversed by mevalonate pathway intermediates. These results suggest that zoledronate and Ara-CBP act, at least in part, via inhibition of the mevalonate pathway in hFOB cells. In contrast, none of the mevalonate pathway intermediates reversed the inhibition of MDA-MB-231 cell proliferation by the bisphosphonates, or the effects of pamidronate on hFOB cells. As a positive control, the effects of mevastatin on hFOB and MDA-MB-231 cells were completely reversed by mevalonate. In summary, these data suggest that zoledronate and Ara-CBP induce human osteoblast differentiation via inhibition of the mevalonate pathway. In contrast, the inhibition of MDA-MB-231 cell proliferation by the bisphosphonates appears to be through mechanisms other than inhibition of the mevalonate pathway.     

Inhibitors of the mevalonate pathway as potential therapeutic agents in multiple myeloma

Clinical studies have suggested that bisphosphonates may prolong the survival of sub-sets of myeloma patients. Newer nitrogen containing bisphosphonates such as zoledronate act, at least in part, by inhibiting farnesyl diphosphate synthase and subsequent protein prenylation, furthermore, limited data suggests that zoledronate exerts a direct anti-tumour effect against human myeloma cell lines. We therefore investigated the anti-myeloma potential of zoledronate in comparison to, and in combination with, two other inhibitors of the mevalonate pathway: the HMGCoA reductase inhibitor fluvastatin and the farnesyl transferase inhibitor SCH66336. We found that fluvastatin was able to inhibit the proliferation of myeloma cells more effectively than zoledronate or SCH66336 and that combinations of zoledronate and fluvastatin, but not zoledronate and SCH66336 acted synergistically. Our data indicated that the anti-proliferative effect of mevalonate pathway inhibitors is mediated principally via prevention of geranylgeranylation and is the result of both cell cycle arrest and apoptosis induction. Microarray and quantitative real-time PCR analyses further demonstrated that genes related to apoptosis, cell cycle control, and the mevalonate pathway were particularly affected by zoledronate and fluvastatin, and that some of these genetic effects were synergistic. We conclude that the mechanisms of geranylgeranylation inhibition mediated anti-myeloma effects warrant further evaluation and may provide novel targets for future therapeutic development.     

Bisphosphonates in cancer therapy

Bisphosphonates inhibit osteoclast-mediated bone resorption in metastatic bone disease. A wealth of preclinical data have begun to shed light on the complex mechanisms by which bisphosphonates inhibit bone resorption and interfere with the formation and growth of bone metastases. Nitrogen-containing bisphosphonates inhibit the mevalonate pathway, which results in the inhibition of osteoclast function and the induction of apoptosis in osteoclasts and tumor cells alike. There is now extensive evidence that bisphosphonates have cytostatic activity against tumor cell lines and inhibit tumor cell adhesion and invasion of the extracellular matrix. These data are supported by a growing body of evidence from animal models demonstrating that bisphosphonates can reduce skeletal tumor burden. However, it remains unclear whether this reduction reflects a direct antitumor effect or an indirect effect via osteoclast inhibition and alteration of the bone microenvironment. Further preclinical studies are needed to elucidate these biochemical mechanisms fully; ultimately, well-controlled clinical trials will be required to investigate whether the antitumor potential of bisphosphonates translates into a significant clinical benefit for patients with cancer.     

Novel aspects of mevalonate pathway inhibitors as antitumor agents

The mevalonate pathway for cholesterol biosynthesis and protein prenylation has been implicated in various aspects of tumor development and progression. Certain classes of drugs, such as statins and bisphosphonates, inhibit mevalonate metabolism and therefore have also been tested as antitumor agents. This concept is strongly supported by the recent finding that mutant p53, which is present in more than half of all human cancers, can significantly upregulate mevalonate metabolism and protein prenylation in carcinoma cells. The first evidence that mevalonate pathway inhibitors may have the potential to reverse the malignant phenotype has already been obtained. Moreover, recently discovered immunomodulatory properties of statins and bisphosphonates may also contribute to their known anticancer effects. Drug-induced inhibition of protein prenylation may induce sequential cellular stress responses, including the unfolded protein response and autophagy, that eventually translate into inflammasome-dependent and caspase-1-mediated activation of innate immunity. This review focuses on these novel capabilities of mevalonate pathway inhibitors to beneficially affect tumor biology and contribute to tumor immune surveillance.     

Antitumor activity of the intratumoral injection of fowlpox vectors expressing a triad of costimulatory molecules and granulocyte/macrophage colony stimulating factor in mesothelioma

Deficiency in costimulatory molecule expression has been implicated in the ability of tumors to escape immune effectors. The activity of the intratumoral administration of recombinant fowlpox vectors expressing a triad of costimulatory molecules (rF-TRICOM) was evaluated in the asbestos-induced AB12 and AC29 mouse models of mesothelioma. Mesothelioma cell infected with rF-TRICOM expressed high levels of the costimulatory molecules. Prolongation of survival was observed in mice receiving rF-TRICOM in AB12 and AC29 intraperitoneal models. Complete tumor regressions were observed in mice receiving intratumoral rF-TRICOM in the AB12 subcutaneous tumor model. Tumor regressions were associated with the development of serum IgG reactivities to mesothelioma-associated determinants and specific systemic cytolytic activity, and responding mice were capable of rejecting tumors upon re-challenge. Antitumor activity was also observed in mice with established AB12 tumor vaccinated with irradiated rF-TRICOM-infected AB12 cells. The antitumor activity of intratumoral rF-TRICOM was superior to that of the intratumoral injection of a fowlpox vector expressing granulocyte-macrophage colony stimulating factor (rF-GM-CSF). AB12 and AC29 tumors were found to produce GM-CSF and to have substantial macrophage infiltration. Production of GM-CSF decreased in vivo in tumors injected with rF-TRICOM. rF-TRICOM and wild-type fowlpox inhibited the growth of AB12 and AC29 cells in vitro; less inhibition was observed with rF-GM-CSF. These results indicate that the intratumoral injection of rF-TRICOM has significant activity in mouse models of mesothelioma and can elicit a systemic antitumor immune response. The results also suggest potential limitations to the intratumoral administration of cytokines, such as GM-CSF, in mesothelioma.     

How Do Bisphosphonates Inhibit Bone Metastasis In Vivo?

Bisphosphonates are potent inhibitors of osteoclast-mediated bone resorption and have demonstrated clinical utility in the treatment of patients with osteolytic bone metastases. They also exhibit direct antitumor activity in vitro and can reduce skeletal tumor burden and inhibit the formation of bone metastases in vivo. However, whether such effects are caused by a direct action of bisphosphonates on tumor cells or indirectly through inhibition of bone resorption remains unclear. To address this question, we used here a structural analog of the bisphosphonate risedronate, NE-58051, which has a bone mineral affinity similar to that of risedronate, but a 3000-fold lower bone antiresorptive activity. In vitro, risedronate and NE-58051 inhibited proliferation of breast cancer and melanoma cell lines. In vivo, risedronate and NE-58051 did not inhibit the growth of subcutaneous B02 breast tumor xenografts or the formation of B16F10 melanoma lung metastasis. In contrast to NE-58051, risedronate did inhibit B02 breast cancer bone metastasis formation by reducing both bone destruction and skeletal tumor burden, indicating that the antitumor effect of bisphosphonates is achieved mainly through inhibition of osteoclast-mediated bone resorption.     

Propriétés antitumorales du bisphosphonate zolédronate et implications thérapeutiques potentielles en clinique

Zoledronate, just as other bisphosphonates, inhibit osteoclast mediated bone resorption. This is the reason why they are used in the treatment of bone metastasis, in order to block osteolysis. Zoledronate and some other bisphosphonates (clodronate, pamidronate, ibandronate, alendronate, risédronate, minodronate) also exhibit antitumor properties in vitro. They act directly on tumor cells by blocking tumor cell adhesion, invasion and proliferation, and by inducing tumor cell apoptosis. However, their high bone mineral affinity decreases their bioavailability to a significant extent and, thus, should weaken their in vivo antitumor potential. Despite of this, several studies (most of them being performed with zoledronate) show that bisphosphonates have an in vivo antitumor activity. This review focuses on zoledronate and on results obtained in several experimental models showing that this bisphosphonate interferes with the growth of tumors and metastases which are thriving in tissues others than the skeletal tissue. The significance of these findings is discussed in the light of several ongoing clinical trials which examine the benefits of using zoledronate and other bisphosphonates in the adjuvant treatment of cancers at an early stage of the disease.     

Bisphosphonates: biological response modifiers in breast cancer

Bone recurrence constitutes one third of initial sites of relapse and one half of distant sites of relapse at 10 years from diagnosis of breast cancer. Bone pain, fracture (including vertebral fracture resulting from increased bone resorption following chemotherapy-induced menopause), and hypercalcemia are components of skeletal morbidity. The pathophysiology of malignant osteopathy occurs because of the secretion of substances (such as parathyroid hormone-related peptide), by the malignant cell, which stimulate osteoclast function; this in turn feeds further growth, which causes a vicious cycle. Interruption of this cycle by bisphosphonates may inhibit the growth of malignant cells. Bisphosphonates are drugs that inhibit bone turnover by decreasing bone resorption. Side effects of bisphosphonates include upper gastrointestinal symptoms (in oral nitrogen-containing bisphosphonates) and diarrhea (in oral non-nitrogen-containing bisphosphonates) and an acute phase-like reaction with intravenous (I.V.) pamidronate. Bisphosphonates have different molecular mechanisms of action: Nitrogen-containing bisphosphonates (eg, pamidronate and alendronate) inhibit the mevalonate-signaling pathway while the non-nitrogen-containing drugs (eg, clodronate) incorporate into adenosine triphosphate analogues. There is in vitro evidence that these drugs also possess anticancer properties. In hypercalcemia patients, treatment with pamidronate and zoledronate produce prompt and efficient normocalcemia. Intravenous pamidronate and zoledronate, oral clodronate, and ibandronate reduce skeletal complications in patients with bone metastases; I.V. pamidronate and clodronate are useful for bone pain relief. Three adjuvant bisphosphonate trials are discussed herein: 2 small open-label studies giving conflicting results and a large placebo-controlled trial of oral clodronate. This latter trial shows a reduction in the incidence of skeletal metastases (while the patients are on therapy) and an improved survival at 5 years.     

Lowering bone mineral affinity of bisphosphonates as a therapeutic strategy to optimize skeletal tumor growth inhibition in vivo

Bisphosphonates bind avidly to bone mineral and are potent inhibitors of osteoclast-mediated bone destruction. They also exhibit antitumor activity in vitro. Here, we used a mouse model of human breast cancer bone metastasis to examine the effects of risedronate and NE-10790, a phosphonocarboxylate analogue of the bisphosphonate risedronate, on osteolysis and tumor growth. Osteolysis was measured by radiography and histomorphometry. Tumor burden was measured by fluorescence imaging and histomorphometry. NE-10790 had a 70-fold lower bone mineral affinity compared with risedronate. It was 7-fold and 8,800-fold less potent than risedronate at reducing, respectively, breast cancer cell viability in vitro and bone loss in ovariectomized animals. We next showed that risedronate given at a low dosage in animals bearing human B02-GFP breast tumors reduced osteolysis by inhibiting bone resorption, whereas therapy with higher doses also inhibited skeletal tumor burden. Conversely, therapy with NE-10790 substantially reduced skeletal tumor growth at a dosage that did not inhibit osteolysis, a higher dosage being able to also reduce bone destruction. The in vivo antitumor activity of NE-10790 was restricted to bone because it did not inhibit the growth of subcutaneous B02-GFP tumor xenografts nor the formation of B16-F10 melanoma lung metastases. Moreover, NE-10790, in combination with risedronate, reduced both osteolysis and skeletal tumor burden, whereas NE-10790 or risedronate alone only decreased either tumor burden or osteolysis, respectively. In conclusion, our study shows that decreasing the bone mineral affinity of bisphosphonates is an effective therapeutic strategy to inhibit skeletal tumor growth in vivo.     

The bisphosphonate incadronate inhibits intraperitoneal dissemination in an in vivo pancreatic cancer model

Pancreatic cancer is characterized by intraperitoneal dissemination and often by large volumes of ascites. Aminobisphosphonates exhibit potent antitumor effects and are currently being tested against human solid tumors. Several aminobisphosphonates inhibit cancer cell migration by preventing the activation of Rho through inhibition of the mevalonate pathway. We evaluated the ability of an aminobisphosphonate, incadronate, to inhibit the growth of disseminated pancreatic cancer in vivo. We established an in vivo pancreatic cancer model with i.p. carcinomatosis in nude mice. Incadronate administration started from the day of tumor inoculation, and reduced tumor burden and ascites accumulation. Further, we evaluated the effect of incadronate on the inhibition of pancreatic cancer cell proliferation, migration and invasion in vitro. Incadronate induced growth inhibition and apoptotic death of pancreatic cancer cells. It also inhibited migration presumably by preventing the activation of Rho by lysophosphatidic acid. Thus, the in vivo antitumor effect may result from the inhibition of cancer cell proliferation and migration. The potent effects of incadronate in reducing tumor burden and ascites suggest that it will be of value in regimens for the treatment of pancreatic cancer.     

双膦酸盐抗血管形成机制研究进展

双膦酸盐具有直接抗肿瘤和抗血管形成作用,具体机制仍未肯定.现有研究认为双膦酸盐通过干扰甲羟戊酸信号通路的关键激酶,使小G蛋白异戊酸化受阻等机制抑制内皮祖细胞分化和小管样形成,阻止血管发生;抑制血管内皮细胞的增殖、黏附和迁移能力,干扰血管形成;调节肿瘤血管形成细胞因子分泌,抑制血管形成相关基因等功能发挥其抗血管形成作用.     

Bisphosphonate treatment inhibits the growth of prostate cancer cells

The presence of skeletal metastases in patients suffering from cancer leads to a variety of clinical complications. Bisphosphonates are a class of drugs with a potent bone resorption inhibition activity that have found increasing utility in treating and managing patients with metastatic bone disease. Several clinical trials have demonstrated that bisphosphonates have clinical value in the treatment and management of skeletal metastases derived from advanced prostate cancer. Currently, the mechanism(s) through which bisphosphonates exert their activity is only beginning to be understood. We have studied the effects of bisphosphonate treatment on the growth of prostate cancer cell lines in vitro. Treatment of PC3, DU145, and LNCaP cells with pamidronate or zoledronate significantly reduced the growth of all three cell lines. Using flow cytometry, pamidronate treatment (100 microM) was shown to induce significant amounts of cell death in all three cell lines studied. In contrast, treatment with zoledronate (100 microM) did not induce cell death, instead exerting dramatic effects on cell proliferation, as evidenced by a major increase in cells present in the G0-G1 and S phase. Although both drugs reduced prostate cancer cell growth in the presence of serum, zoledronate was more potent under these conditions, disrupting growth at doses as low as 25 microM in the presence of 5% fetal bovine serum. These results raise the intriguing possibility that the observed clinical utility of bisphosphonates in managing skeletal metastases may in part derive from direct inhibition of prostate cancer cell growth in the bone microenvironment.     

Eradication of intraperitoneal and distant tumor by adenovirus-mediated interferon-beta gene therapy is attributable to induction of systemic immunity

Malignant mesothelioma remains an incurable disease for which immune-modulatory therapies, such as exogenous cytokines, have shown some promise. One such cytokine, IFN-beta, has potent antiproliferative and immunostimulatory activity in vitro, but its in vivo use has been limited by toxicity. We thus conducted studies evaluating intracavitary delivery of a replication-deficient adenoviral (Ad) vector encoding for the murine IFN-beta gene (Ad.muIFN-beta) in mouse models of malignant mesothelioma. In contrast to multiple injections of recombinant protein, a single i.p. injection of Ad.muIFN-beta into animals with established tumors elicited remarkable antitumor activity leading to long-term survival in >90% of animals bearing either AB12 or AC29 i.p. mesotheliomas. A control adenovirus vector had minimal antitumor effect in vivo. Significant therapeutic effects were also seen in animals treated with large tumor burdens. Importantly, treatment of i.p. tumor also led to reduction of growth in tumors established at a distant site (flank). A number of experiments suggested that these effects were attributable to an acquired CD8(+) T-cell-mediated response including: (a) the induction of long-lasting antitumor immunity; (b) loss of efficacy of Ad.muIFN-beta in tumor-bearing, immune-deficient (SCID, SCID/beige) mice; (c) detection of high levels of specific antitumor cytolytic activity from unstimulated splenocytes harvested from Ad.muIFN-beta-treated animals that was abolished by CD8(+) T-cell depletion; and (d) abrogation of antitumor effects of Ad.muIFN-beta in tumor-bearing CD8(+) T-cell-depleted animals. These data show that intracavitary IFN-beta gene therapy using an adenoviral vector provides strong CD8(+) T-cell-mediated antitumor effects in murine models of mesothelioma and suggest that this may be a promising strategy for the treatment of localized tumors such as mesothelioma or ovarian cancer in humans.     

Soluble type II transforming growth factor-beta receptor inhibits established murine malignant mesothelioma tumor growth by augmenting host antitumor immunity.

PURPOSE: Transforming growth factor (TGF)-beta blockade has been proposed as an anticancer therapy; however, understanding which tumor patients might benefit most from such therapy is crucial. An ideal target of such inhibitory therapy might be malignant mesothelioma (MM), a highly lethal, treatment-resistant malignancy of mesothelial cells of the pleura and peritoneum that produces large amounts of TGF-beta. The purpose of this study was to explore the possible therapeutic utility of TGF-beta blockade on MM. EXPERIMENTAL DESIGN: To evaluate this hypothesis, we tested the effects of a soluble TGF-beta type II receptor (sTGF-beta R) that specifically inhibits TGF-beta1 and TGF-beta 3 in three different murine MM tumor models, AB12 and AC29 (which produce large amounts of TGF-beta) and AB1 (which does not produce TGF-beta). RESULTS: Tumor growth of both established AB12 and AC29 tumors was inhibited by sTGF-beta R. In contrast, AB1 tumors showed little response to sTGF-beta R. The mechanism of these antitumor effects was evaluated and determined to be primarily dependent on immune-mediated responses because (a) the antitumor effects were markedly diminished in severe combined immunodeficient mice or mice depleted of CD8(+) T cells and (b) CD8(+) T cells isolated from spleens of mice treated with sTGF-beta R showed strong antitumor cytolytic effects, whereas CD8(+) T cells isolated from spleens of tumor-bearing mice treated with of control IgG2a showed no antitumor cytolytic effects. CONCLUSIONS: Our data suggest that TGF-beta blockade of established TGF-beta-secreting MM should be explored as a promising strategy to treat patients with MM and other tumors that produce TGF-beta.     

A novel small-molecule inhibitor of transforming growth factor beta type I receptor kinase (SM16) inhibits murine mesothelioma tumor growth in vivo and prevents tumor recurrence after surgical resection

Malignant mesothelioma is an aggressive and lethal pleural cancer that overexpresses transforming growth factor beta (TGFbeta). We investigated the efficacy of a novel small-molecule TGFbeta type I receptor (ALK5) kinase inhibitor, SM16, in the AB12 syngeneic model of malignant mesothelioma. SM16 inhibited TGFbeta signaling seen as decreased phosphorylated Smad2/3 levels in cultured AB12 cells (IC(50), approximately 200 nmol/L). SM16 penetrated tumor cells in vivo, suppressing tumor phosphorylated Smad2/3 levels for at least 3 h following treatment of tumor-bearing mice with a single i.p. bolus of 20 mg/kg SM16. The growth of established AB12 tumors was significantly inhibited by 5 mg/kg/d SM16 (P < 0.001) delivered via s.c. miniosmotic pumps over 28 days. The efficacy of SM16 was a result of a CD8+ antitumor response because (a) the antitumor effects were markedly diminished in severe combined immunodeficient mice and (b) CD8+ T cells isolated from spleens of mice treated with SM16 showed strong antitumor cytolytic effects whereas CD8+ T cells isolated from spleens of tumor-bearing mice treated with control vehicle showed minimal activity. Treatment of mice bearing large tumors with 5 mg/kg/d SM16 after debulking surgery reduced the extent of tumor recurrence from 80% to <20% (P < 0.05). SM16 was also highly effective in blocking and regressing tumors when given p.o. at doses of 0.45 or 0.65 g/kg in mouse chow. Thus, SM16 shows potent activity against established AB12 malignant mesothelioma tumors using an immune-mediated mechanism and can significantly prevent tumor recurrence after resection of bulky AB12 malignant mesothelioma tumors. These data suggest that ALK5 inhibitors, such as SM16, offer significant potential for the treatment of malignant mesothelioma and possibly other cancers.     

唑来膦酸联合靶向及免疫调节药物的抗肿瘤研究进展

唑来膦酸是第三代含氮双磷酸盐,在临床中广泛用于治疗骨疾病。既往研究发现唑来膦酸在体内外对多种肿瘤细胞均有抗肿瘤作用,同时可以协同增加化疗、内分泌治疗、放疗等抗肿瘤疗效。最近研究发现,唑来膦酸亦可以协同增加靶向及免疫调节药物的抗肿瘤疗效,其抗肿瘤作用机制与抑制甲羟戊酸代谢途径、影响肿瘤信号通路、调节免疫反应、抗血管生成等有关。本文综述了唑来膦酸联合靶向及免疫调节药物在抗肿瘤治疗方面的研究进展。     

Isoprenoid-independent pathway is involved in apoptosis induced by risedronate, a bisphosphonate, in which Bim plays a critical role in breast cancer cell line MCF-7

Bisphosphonates cause apoptosis to various types of cancer cells including breast cancer. Inhibition of the mevalonate pathway was reported to be involved in the apoptosis induced by bisphosphonates, but its precise mechanism has not been unveiled. In the present study, we investigated the molecular mechanism of risedronate, a bisphosphonate, in the apoptosis of the breast cancer cell line MCF-7 in comparison with that of cerivastatin, an HMG CoA reductase inhibitor (statin), since statin has been known to induce apoptosis through an isoprenoid-dependent pathway in these cells. We found that i) risedronate induced MCF-7 cells into apoptosis in a manner similar to cerivastatin with the activation of caspase-9 followed by caspase-6 and -7, that ii) bisphosphonate-induced apoptosis was significantly, but not fully, recovered by the addition of GGOH, an isoprenoid, which completely rescued in case of cerivastatin-induced apoptosis, that iii) risedronate induced G2 arrest with the induction of Bim (BH3-only protein), but that statin induced G1 arrest without it, and that iv) the down-regulation of Bim protein by siRNA significantly attenuated the risedronate-induced apoptosis. These data clearly indicate that both isoprenoid-dependent and -independent pathways might be involved in the apoptosis induced by bisphosphonate, and Bim might be a critical component for the isoprenoid-independent apoptotic pathway.     

Alendronate inhibits lysophosphatidic acid-induced migration of human ovarian cancer cells by attenuating the activation of rho

Alendronate, a nitrogen-containing bisphosphonate, is a potent inhibitor of bone resorption used for the treatment and prevention of osteoporosis. Recent findings suggest that alendronate and other nitrogen-containing bisphosphonates inhibit the mevalonate pathway and thereby inhibit the synthesis of products derived from this metabolite. This, in turn, prevents the prenylation of a number of small GTPases, which regulate cell growth, motility, and invasion. We studied the effect of alendronate on in vitro migration of human ovarian cancer cells. Lysophosphatidic acid (LPA) induced a dose-dependent increase of migration of cancer cells by promoting Rho/Rho-associated kinase signaling. The induction of cancer cell migration by LPA was inhibited by the addition of alendronate in a dose-dependent manner. Treatment of ovarian cancer cells with alendronate resulted in inactivation of Rho, changes of cell morphology, loss of stress fiber formation, and focal adhesion assembly, and the suppression of phosphorylation of myosin light chain and tyrosine phosphorylation of focal adhesion proteins, which are essential processes for cell migration. The effects of alendronate on cancer cells were prevented by the addition of geranylgeranyol, which is the metabolic intermediate of the mevalonate pathway. These results suggest that alendronate inhibits Rho activation by preventing geranylgeranylation, which results in inhibition of LPA-induced migration of human ovarian cancer cells.     

Cationic lipid:bacterial DNA complexes elicit adaptive cellular immunity in murine intraperitoneal tumor models

Previous studies with a mycobacterial heat shock protein (hsp-65) have demonstrated some efficacy using cationic liposome-mediated gene transfer in murine i.p. sarcoma models. To further analyze the efficacy of hsp-65 immunotherapy in clinically relevant models of localized cancer, immunocompetent mice bearing i.p. murine mesothelioma were treated with four i.p. doses of a cationic lipid complexed with plasmid DNA (pDNA) containing hsp65, LacZ, or a null plasmid. We observed >90% long-term survival (median survival, 150 days versus approximately 25 days, treated versus saline control, respectively) in a syngeneic, i.p. murine mesothelioma model (AC29). Long-term survivors were observed in all groups treated with lipid complexed with any pDNA. Lipid alone or DNA alone provided no demonstrable survival advantage. In a more aggressive i.p. model of mesothelioma (AB12), we observed >40% long-term survival in groups treated with lipid:pDNA complexes, again irrespective of the transgene. To ask whether these antitumor effects had led to an adaptive immune response against the tumor cell, we rechallenged long-term survivors in both murine models s.c. with the parental tumor cell line. Specific, long-lasting systemic immunity against the tumor was readily demonstrated in both models (AB12 and AC29). Consistent with these results, splenocytes from long-term survivors specifically lysed the parental tumor cell lines. Depleting the CD8+ T-cells from the splenocyte pool eliminated this lytic activity. Lipid:pDNA treatment of athymic, SCID, and SCID/Beige mice bearing a murine i.p. mesothelioma (AC29) resulted in only a slight survival advantage, but there were no long-term survivors. Treatment of immunocompetent mice depleted of specific immune effector cells demonstrated roles for CD8+ and natural killer cells. Although the exact mechanism(s) responsible for these antitumor effects is unclear, the results are consistent with roles for both innate and adaptive immune responses. An initial tumor cell killing stimulated by cationic lipid:pDNA complexes appears to be translated into long-term, systemic immunity against the tumor cell. These results are the first to demonstrate that adaptive immunity against a tumor cell can be induced by the administration of lipid:pDNA complexes. Multiple administrations of cationic lipid complexed with pDNA lacking an expressed transgene could provide a promising generalized immune-mediated modality for treating cancer.     

Bisphosphonates: preclinical review

Bisphosphonates effectively inhibit osteoclast-mediated bone resorption and are integral in the treatment of benign and malignant bone diseases. The evolution of bisphosphonates over the past 30 years has led to the development of nitrogen-containing bisphosphonates (N-BPs), which have a mechanism of action different from that of the nonnitrogen-containing bisphosphonates. Studies conducted over the past decade have elucidated the mechanism of action and pharmacologic properties of the N-BPs. N-BPs exert their effects on osteoclasts and tumor cells by inhibiting a key enzyme in the mevalonate pathway, farnesyl diphosphate synthase, thus preventing protein prenylation and activation of intracellular signaling proteins such as Ras. Recent evidence suggests that N-BPs also induce production of a unique adenosine triphosphate analogue (Apppi) that can directly induce apoptosis. Our increased understanding of the pharmacologic effects of bisphosphonates is shedding light on the mechanisms by which they exert antitumor effects. As a result of their biochemical effects on protein prenylation, N-BPs induce caspase-dependent apoptosis, inhibit matrix metalloproteinase activity, and downregulate alpha(v)beta(3) and alpha(v)beta(5) integrins. In addition, zoledronic acid (Zometa; Novartis Pharmaceuticals Corp.; East Hanover, NJ and Basel, Switzerland) exerts synergistic antitumor activity when combined with other anticancer agents. Zoledronic acid also inhibits tumor cell adhesion to the extracellular matrix and invasion through Matrigel trade mark and has antiangiogenic activity. A growing body of evidence from animal models demonstrates that zoledronic acid and other bisphosphonates can reduce skeletal tumor burden and prevent metastasis to bone. Further studies are needed to fully elucidate these biochemical mechanisms and to determine if the antitumor potential of bisphosphonates translates to the clinical setting.