Oral paclitaxel chemotherapy for brain tumors: ideal combination treatment of paclitaxel and P-glycoprotein inhibitor

Oral chemotherapy has many advantages over parenteral chemotherapeutics administration. To use the advantages of the oral chemotherapy and maximize anti-tumor effects of the chemotherapeutic agent, we designed HM30181A (a P-glycoprotein inhibitor) and a paclitaxel oral co-administration chemotherapeutic method. HM30181A is used to aid paclitaxel absorption from gut lumen into blood and to inhibit paclitaxel exclusion out of the brain tumor mass by endothelial cells, which inhibits paclitaxel access to tumor cells in the brain parenchyma. We applied HM30181A and paclitaxel oral co-administration methods to the treatment of tumors in the brain using the K1735 melanoma brain metastasis animal model and the U-87 MG glioblastoma animal model. Administrations were performed twice per week for 28 days and the therapeutic effect was examined using tumor volume change. We observed that 32 mg/kg HM30181A and 16 mg/kg of paclitaxel (dose ratio 2:1) oral co-administration showed significant therapeutic effects in both animal models, but when the doses or dose ratio was changed, the effects could not be observed. Therefore, adjustments of doses and dose ratio of the agents seems to be essential in realizing oral HM30181A and paclitaxel treatment in brain tumors. These results suggest that if the doses and dose ratio can be successfully adjusted, the oral co-administration of HM30181A and paclitaxel can be used to treat tumors in the brain.     

P-glycoprotein expression in brain tumors

Summary Overexpression of P-glycoprotein (P-gp) in cancer cells can result in resistance to several chemotherapy agents (multidrug resistance) including doxorubicin and vincristine. The drugs to which resistance develops also penetrate the blood brain barrier poorly. P-gp expression in brain capillary endothelial cells suggests that P-gp may restrict drug entry into brain tumors and thus be another mechanism of drug resistance. To seek evidence for either of these roles in the drug resistance of brain tumors, we examined the location of expression of P-gp in 49 brain tumors, using an anti-P-gp mouse monoclonal antibody and immunohistochemistry. P-gp expression was observed in tumor cells of two glioblastomas and a meningeal sarcoma but not in low-grade primary or metastatic tumors. In low-grade primary tumors, P-gp was present in all vascular endothelial cells. In the vascular endothelial cells of anaplastic primary brain tumors and brain metastases, P-gp expression was heterogeneous or absent. These findings are consistent with a role for P-gp in the resistance of some brain tumors to chemotherapy agents.     

HM30181A,a potent P-glycoprotein inhibitor,potentiates the absorption and in vivo antitumor efficacy of paclitaxel in an orthotopic brain tumor model

Objective:Delivery of chemotherapeutic drugs to the brain has remained a major obstacle in the treatment of glioma, owing to the presence of the blood-brain barrier and the activity of P-gp, which pumps its substrate back into the systemic circulation. The aim of the present study was to develop an intravenous formulation of HM30181A (HM) to inhibit P-gp in the brain to effectively deliver paclitaxel (PTX) for the treatment of malignant glioma.Methods:Two formulations of solubilized HM were designed on the basis of different solid dispersion strategies: i) spray-drying [polyvinlypyrrolidone (PVP)-HM] and ii) solvent evaporation [HP-β-cyclodextrin (cyclodextrin)-HM]. The P-gp inhibition of these 2 formulations was assessed on the basis of rhodamine 123 uptake in cancer cells. Blood and brain pharmacokinetic parameters were also determined, and the antitumor effect of cyclodextrin-HM with PTX was evaluated in an orthotopic glioma xenograft mouse model.Results:Although both PVP-HM and cyclodextrin-HM formulations showed promising P-gp inhibition activity in vitro, cyclodextrin-HM had a higher maximum tolerated dose in mice than did PVP-HM. Pharmacokinetic study of cyclodextrin-HM revealed a plasma concentration plateau at 20 mg/kg, and the mice began to lose weight at doses above this level. Cyclodextrin-HM (10 mg/kg) administered with PTX at 10 mg/kg showed optimal antitumor activity in a mouse model, according to both tumor volume measurement and survival time (P < 0.05).Conclusions:In a mouse orthotopic brain tumor model, the intravenous co-administration of cyclodextrin-HM with PTX showed potent antitumor effects and therefore may have potential for glioma therapy in humans.     

Randomized study of paclitaxel and tamoxifen deposition into human brain tumors: implications for the treatment of metastatic brain tumors.

PURPOSE: Drug resistance in brain tumors is partially mediated by the blood-brain barrier of which a key component is P-glycoprotein, which is highly expressed in cerebral capillaries. Tamoxifen is a nontoxic inhibitor of P-glycoprotein. This trial assessed, in primary and metastatic brain tumors, the differential deposition of paclitaxel and whether tamoxifen could increase paclitaxel deposition. EXPERIMENTAL DESIGN: Patients for surgical resection of their primary or metastatic brain tumors were prospectively randomized to prior paclitaxel alone (175 mg/m(2)/i.v.) or tamoxifen for 5 days followed by paclitaxel. Central and peripheral tumor, surrounding normal brain and plasma, were analyzed for paclitaxel and tamoxifen. RESULTS: Twenty-seven patients completed the study. Based on a multivariate linear regression model, no significant differences in paclitaxel concentrations between the two study arms were found after adjusting for treatment group (tamoxifen versus control). However, in analysis for tumor type, metastatic brain tumors had higher paclitaxel concentrations in the tumor center (1.93-fold, P = 0.10) and in the tumor periphery (2.46-fold, P = 0.039) compared with primary brain tumors. Pharmacokinetic analyses showed comparable paclitaxel areas under the serum concentration between treatment arms. CONCLUSIONS: Paclitaxel deposition was not increased with this tamoxifen schedule as the low plasma concentrations were likely secondary to concurrent use of P-450-inducing medications. However, the statistically higher paclitaxel deposition in the periphery of metastatic brain tumors provides functional evidence corroborating reports of decreased P-glycoprotein expression in metastatic versus primary brain tumors. This suggests that metastatic brain tumors may respond to paclitaxel if it has proven clinical efficacy for the primary tumor's histopathology.     

The influence of the P-glycoprotein inhibitor zosuquidar trihydrochloride (LY335979) on the brain penetration of paclitaxel in mice

We determined the effect of zosuquidar·3HCl, an inhibitor of P-gp, on the penetration of the anticancer drug paclitaxel into the brain. Zosuquidar·3HCl was administered orally at 25 and 80 mg/kg 1 h before i.v. paclitaxel and i.v. at 20 mg/kg 10 min and 1 h before paclitaxel. The concentrations of paclitaxel in plasma and tissues and of zosuquidar·3HCl in plasma were quantified by high-performance liquid chromatography. The results revealed 3.5-fold and 5-fold higher paclitaxel levels in the brain of wild-type mice treated orally with 25 and 80 mg/kg zosuquidar·3HCl, respectively. However, complete inhibition as in P-gp knockout mice (11-fold increase) was not achieved. Zosuquidar·3HCl also increased the paclitaxel concentrations in plasma and tissues to levels similar to those observed in P-gp knockout mice, suggesting selective P-gp inhibition of zosuquidar·3HCl. When zosuquidar·3HCl was administered i.v. 10 min before paclitaxel, the paclitaxel levels in the brain of wild-type mice increased by 5.6-fold, whereas the increase was only 2.1-fold when zosuquidar·3HCl was administered 1 h before paclitaxel. This suggests that the inhibition of P-gp at the blood-brain barrier by zosuquidar·3HCl is rapidly reversible and that the concentrations of zosuquidar·3HCl in the plasma have already declined to levels insufficient to inhibit P-gp at the blood-brain barrier. In conclusion, zosuquidar·3HCl is only moderately active as an inhibitor of P-gp at the blood-brain barrier.     

Penetration of intra-arterially administered vincristine in experimental brain tumor

Vincristine is an integral part of the "PCV" regimen that is commonly administered to treat primary brain tumors. The efficacy of vincristine as a single agent in these tumors has been poorly studied. This study was designed to determine whether vincristine enters normal rat brain or an intracranially or subcutaneously implanted glioma and to assess the presence of the efflux pump P-glycoprotein (P-gp) on tumor and vascular endothelial cells. The 9L rat gliosarcoma was implanted intracranially and subcutaneously in three Fischer 344 rats. On day 7, [3H]vincristine (50 microCi, 4.8 microg) was injected into the carotid artery, and the animals were euthanized 10 or 20 min later. Quantitative autoradiography revealed that vincristine levels in the liver were 6- to 11-fold greater than in the i.c. tumor, and 15- to 37-fold greater than in normal brain, the reverse of the expected pattern with intraarterial delivery. Vincristine levels in the s.c. tumor were 2-fold higher than levels in the i.c. tumor. P-gp was detected with JSB1 antibody in vascular endothelium of both normal brain and the i.c. tumor, but not in the tumor cells in either location, or in endothelial cells in the s.c. tumor. These results demonstrate that vincristine has negligible penetration of normal rat brain or i.c. 9L glioma despite intra-arterial delivery and the presence of blood-brain barrier dysfunction as demonstrated by Evan's blue. Furthermore, this study suggests that P-gp-mediated efflux from endothelium may explain these findings. The lack of penetration of vincristine into brain tumor and the paucity of single-agent activity studies suggest that vincristine should not be used in the treatment of primary brain tumors.     

Expression and functional activity of the ABC-transporter proteins P-glycoprotein and multidrug-resistance protein 1 in human brain tumor cells and astrocytes

The poor prognosis of glioma patients is partly based on the minor success obtained from chemotherapeutic treatments. Resistance mechanisms at the tumor cell level may be, in addition to the blood–brain barrier, involved in the intrinsic chemo-insensitivity of brain tumors. We investigated the expression of the drug-transporter proteins P-glycoprotein (P-gp) and multidrug-resistance protein 1 (MRP1) in cell lines (N=24) and primary cell cultures (N=36) from neuroectodermal tumors, as well as in brain tumor extracts (N=18) and normal human astrocytes (N=1). We found that a considerable expression of P-gp was relatively rare in glioma cells, in contrast to MRP1, which was constitutively overexpressed in cells derived from astrocytomas as well as glioblastomas. Also, normal astrocytes cultured in vitro expressed high amounts of MRP1 but no detectable P-gp. Meningioma cells frequently co-expressed P-gp and MRP1, while, most of the neuroblastoma cell lines express higher P-gp but lower MRP1 levels as compared to the other tumor types. Both, a drug-exporting and a chemoprotective function of P-gp as well as MRP1 could be demonstrated in selected tumor cells by a significant upregulation of cellular ³H-daunomycin accumulation and daunomycin cytotoxicity via administration of transporter antagonists. Summing up, our data suggest that P-gp contributes to cellular resistance merely in a small subgroup of gliomas, but frequently in neuroblastomas and meningiomas. In contrast, MRP1 is demonstrated to play a constitutive role in the intrinsic chemoresistance of gliomas and their normal cell counterpart.     

Therapy of hematogenous melanoma brain metastases with endostatin.

PURPOSE: Cerebral metastases represent the most common type of brain tumors. This study investigated the effects of endogenous endostatin on hematogenous cerebral melanoma metastases. EXPERIMENTAL DESIGN: Murine K1735 melanoma cells were transfected with the mouse endostatin cDNA. Experimental tumors were induced either by s.c. injection, intracerebral implantation, or via injection into the internal carotid artery to simulate hematogenous metastatic spread. The effects of endostatin expression on tumor incidence, growth pattern, and vascularity were analyzed. RESULTS: In vitro secretion of endostatin by 2.5 x 10(5) cells within 24 hours was 0.12 +/- 0.03 ng, 4.35 +/- 0.4, and 1.18 +/- 0.7 ng/mL for wild type and two endostatin-transfected K1735 clones termed K1735-endo/2 and K1735-endo/8, respectively. Tumor inhibition in vivo correlated with endogenous endostatin production. Within 25 days, growth of s.c. K1735-endo/2 tumors was <20% compared with wild-type controls. Following intracerebral implantation the average survival time of mice was 27.8 +/- 2.6 versus 13.3 +/- 3.7 days in the K1735-endo/2 versus the wild-type group, respectively. Intracarotid injection of 1 x 10(5) wild-type cells killed the mice within 24 +/- 1.8 days. In contrast, endostatin expression prevented macroscopic metastatic tumor growth in 11 of 12 mice, although viable microscopic tumor pockets were detectable in all animals. CONCLUSION: Endostatin inhibits tumor progression of multiple cerebral metastases in vivo. Hematogenous micrometastases are more efficiently suppressed than tumors resulting from high focal cell numbers which may be due to a higher angiogenic signaling exerted by massive cell deposits. Endostatin may prevent solid tumor growth more effectively by inhibition of early angiogenesis.     

Development of in vivo models for studies of brain metastasis

To develop an in vivo model for studies of brain metastasis, varying doses of K-1735 tumor cells with differing metastatic potentials were injected into the carotid arteries of anesthetized mice. Direct intracerebral administration of cells produced tumors in the brain parenchyma, and all tumor cells produced melanotic lesions at these sites. Studies with radioactively-labelled tumor cells confirmed that tumor cells reached the meninges and extracerebral organs. Distant metastases were found in the lungs and hearts of mice which received K-1735 cells into the carotid artery, but not in animals which received tumorigenic non-metastatic cells. Regardless of the type of cells injected, no growths were found in the meninges. The high degree of reproducibility recommends this model for studies designed to investigate the biology and therapy of cancer metastasis to the brain.     

Halofuginone inhibits angiogenesis and growth in implanted metastatic rat brain tumor model--an MRI study

Tumor growth and metastasis depend on angiogenesis; therefore, efforts are made to develop specific angiogenic inhibitors. Halofuginone (HF) is a potent inhibitor of collagen type alpha1(I). In solid tumor models, HF has a potent antitumor and antiangiogenic effect in vivo, but its effect on brain tumors has not yet been evaluated. By employing magnetic resonance imaging (MRI), we monitored the effect of HF on tumor progression and vascularization by utilizing an implanted malignant fibrous histiocytoma metastatic rat brain tumor model. Here we demonstrate that treatment with HF effectively and dose-dependently reduced tumor growth and angiogenesis. On day 13, HF-treated tumors were fivefold smaller than control (P < .001). Treatment with HF significantly prolonged survival of treated animals (142%; P = .001). In HF-treated rats, tumor vascularization was inhibited by 30% on day 13 and by 37% on day 19 (P < .05). Additionally, HF treatment inhibited vessel maturation (P = .03). Finally, in HF-treated rats, we noticed the appearance of a few clusters of satellite tumors, which were distinct from the primary tumor and usually contained vessel cores. This phenomenon was relatively moderate when compared to previous reports of other antiangiogenic agents used to treat brain tumors. We therefore conclude that HF is effective for treatment of metastatic brain tumors.     

Phase I/II Study of Weekly Oraxol for the Second‐Line Treatment of Patients With Metastatic or Recurrent Gastric Cancer

Lessons Learned

• Oraxol, a novel oral formulation of paclitaxel, displayed modest efficacy as second-line chemotherapy for gastric cancer.
• Considering its favorable toxicity profiles, further studies are warranted in various solid tumors including gastric cancer.

Background.

Oraxol consists of paclitaxel and HM30181A, a P-glycoprotein inhibitor, to increase the oral bioavailability of paclitaxel. This phase I/II study (HM-OXL-201) was conducted to determine the maximum tolerated dose (MTD) and recommended phase II dose (RP2D) of Oraxol. In addition, we investigated the efficacy and safety of Oraxol as second-line chemotherapy for metastatic or recurrent gastric cancer (GC).

Methods.

In the phase I component, paclitaxel was orally administered at escalating doses (90, 120, or 150 mg/m² per day) with a fixed dose (15 mg/day) of HM30181A. Oraxol was administrated 6 times per cycle (days 1, 2, 8, 9, 15, and 16) every 4 weeks. In the phase II component, the efficacy and safety of Oraxol were evaluated.

Results.

In the phase I component, the MTD could not be determined. Based on toxicity and pharmacokinetic data, the RP2D of oral paclitaxel was determined to be 150 mg/m². In the phase II component, 4 of 43 patients (9.3%) achieved partial responses. Median progression-free survival and overall survival were 2.6 and 10.7 months, respectively. Toxicity profiles were favorable, and the most common drug-related adverse events (grade ≥3) were neutropenia and diarrhea.

Conclusion.

Oraxol exhibited modest efficacy and favorable toxicity profiles as second-line chemotherapy for GC.     

Irradiation of rat brain reduces P-glycoprotein expression and function

The blood-brain barrier (BBB) hampers delivery of several drugs including chemotherapeutics to the brain. The drug efflux pump P-glycoprotein (P-gp), expressed on brain capillary endothelial cells, is part of the BBB. P-gp expression on capillary endothelium decreases 5 days after brain irradiation, which may reduce P-gp function and increase brain levels of P-gp substrates. To elucidate whether radiation therapy reduces P-gp expression and function in the brain, right hemispheres of rats were irradiated with single doses of 2-25 Gy followed by 10 mg kg(-1) of the P-gp substrate cyclosporine A (CsA) intravenously (i.v.), with once 15 Gy followed by CsA (10, 15 or 20 mg kg(-1)), or with fractionated irradiation (4 x 5 Gy) followed by CsA (10 mg kg(-1)) 5 days later. Additionally, four groups of three rats received 25 Gy once and were killed 10, 15, 20 or 25 days later. The brains were removed and P-gp detected immunohistochemically. P-gp function was assessed by [(11)C]carvedilol uptake using quantitative autoradiography. Irradiation increased [(11)C]carvedilol uptake dose-dependently, to a maximum of 20% above non irradiated hemisphere. CsA increased [(11)C]carvedilol uptake dose-dependently in both hemispheres, but more (P<0.001) in the irradiated hemisphere. Fractionated irradiation resulted in a lost P-gp expression 10 days after start irradiation, which coincided with increased [(11)C]carvedilol uptake. P-gp expression decreased between day 15 and 20 after single dose irradiation, and increased again thereafter. Rat brain irradiation results in a temporary decreased P-gp function.     

Active specific immunotherapy against occult brain metastasis

We determined whether lyophilized High Five (H5) insect cells engineered to produce IFN-beta (H5BVIFN-beta) could induce systemic immunity against occult brain metastases. C3H/HeN mice were injected s.c. with syngeneic UV-2237M fibrosarcoma or K-1735M2 melanoma cells. Intralesional injection of 2 x 10(6) lyophilized H5BVIFN-beta cells produced complete regression of the s.c. tumors. Six weeks later, UV-2237M fibrosarcoma cells or K-1735M2 melanoma cells were injected into the internal carotid artery of naive or treated mice. UV-2237M brain metastases developed in naive mice or mice cured of K-1735M2 tumors but not in mice cured of UV-2237M tumors. Similarly, K-1735M2 brain metastases developed in naive mice or mice cured of UV-2237M fibrosarcomas but not in mice cured of K-1735M2 melanoma. In the next set of studies, mice were injected s.c. with UV-2237M fibrosarcoma cells. On day 7, UV-2237M fibrosarcoma cells or K-1735M2 cells were implanted into the internal carotid artery, and on day 10, the s.c. tumors were injected with lyophilized H5BVIFN-beta. Both the s.c. tumors and the occult brain metastases produced from carotid injections were eradicated in a tumor-specific manner. The regression of the brain metastases was abrogated by depletion of CD4(+) or CD8(+) cells from immunized mice. These results demonstrate that specific systemic immunity can be induced by lyophilized H5BVIFN-beta and that the resultant immune response can eliminate established brain metastasis.     

Vascular gene expression patterns are conserved in primary and metastatic brain tumors

Malignant primary glial and secondary metastatic brain tumors represent distinct pathological entities. Nevertheless, both tumor types induce profound angiogenic responses in the host brain microvasculature that promote tumor growth. We hypothesized that primary and metastatic tumors induce similar microvascular changes that could function as conserved angiogenesis based therapeutic targets. We previously isolated glioma endothelial marker genes (GEMs) that were selectively upregulated in the microvasculature of proliferating glioblastomas. We sought to determine whether these genes were similarly induced in the microvasculature of metastatic brain tumors. RT-PCR and quantitative RT-PCR were used to screen expression levels of 20 candidate GEMs in primary and metastatic clinical brain tumor specimens. Differentially regulated GEMs were further evaluated by immunohistochemistry or in situ hybridization to localize gene expression using clinical tissue microarrays. Thirteen GEMs were upregulated to a similar degree in both primary and metastatic brain tumors. Most of these genes localize to the cell surface (CXCR7, PV1) or extracellular matrix (COL1A1, COL3A1, COL4A1, COL6A2, MMP14, PXDN) and were selectively expressed by the microvasculature. The shared expression profile between primary and metastatic brain tumors suggests that the molecular pathways driving the angiogenic response are conserved, despite differences in the tumor cells themselves. Anti-angiogenic therapies currently in development for primary brain tumors may prove beneficial for brain metastases and vice versa.     

Malignant potential of cells isolated from lymph node or brain metastases of melanoma patients and implications for prognosis

We studied the correlation between the formation of brain metastasis and the malignant growth potential of seven human melanoma cell lines, isolated from lymph node metastases (A375-SM, TXM-1, DM-4) or from brain metastases (TXM-13, TXM-18, TXM-34, TXM-40), and the potential of three variants of the mouse K-1735 melanoma. Growth rates in different concentrations of fetal bovine serum and colony-forming efficiency in semisolid agarose were measured, and the tumorigenicity and metastatic ability were determined in nude mice (for the human melanoma cell lines) or in C3H/HeN mice (for the K-1735 variants). The ability to form brain metastasis was tested by injection of cells into the carotid artery. A high colony-forming efficiency in agarose, especially at concentrations of agarose greater than 0.6%, corresponded with high tumor take rates, rapid tumor growth rates, and metastatic colonization of the lungs of the recipient mice. For the human melanomas, the lymph node metastasis-derived cells were more tumorigenic and metastatic than the brain metastasis-derived cells. In the K-1735 mouse melanoma, the tumorigenic and metastatic behavior of the cells after i.v. and s.c. injection corresponded with growth in agarose cultures. However, for growth in the brain after intracarotid injection, the different melanoma cell lines showed similar frequencies of tumor take, regardless of tumorigenicity in other sites of the recipient mice, although mice given injections of brain metastasis-derived cells survived longer than mice given injections of lymph node metastasis (human melanoma) or lung metastasis (K-1735 M-2)-derived cell lines. The results from the human and mouse melanoma cell lines show that the brain metastasis-derived cell lines were not more malignant than the lymph node or lung metastasis-derived cells. These data imply that the production of brain metastasis is not always the final stage of a metastatic cascade.     

Serpin inhibitors of urokinase and thrombin in normal rat brain and the 9L brain tumor: evidence for elevated expression of protease nexin I-like inhibitor and a novel sodium dodecyl sulfate-activated tumor antithrombin

Increasing attention is being paid to alterations of the hemostatic balance in tumors, in general, and brain tumors, in particular. Apparently divergent results, showing excess fibrinolysis (i.e., increased plasminogen activator activity) or its inhibition (i.e., increased inhibitor activity), have been reported. The 9L rat brain tumor is a gliosarcoma and a model used to study treatment paradigms for human gliomas. To study the roles of fibrin and fibrinolysis in this brain tumor model, we used these features to investigate the nature of the plasminogen activator (PA) and thrombin inhibitors in normal rat brain and in the 9L rat brain tumor, growing both in vitro and in vivo in rat brain. The results indicate that cells cultured from the tumor in vitro express PA inhibitory activity which is both of the protease nexin I and PA inhibitor 1 types. However, the serpin PA inhibitory activity in extracts of both the normal brain and tumor is of the protease nexin I/PA inhibitor 3 type. This activity is higher in the tumor than in the surrounding "normal" tissue. In addition, we present evidence for a novel thrombin inhibitor which (a) is present only in the tumor growing in rat brain and undetectable either in the normal brain tissue or in vitro, (b) is in a latent, but sodium dodecyl sulfate-activatable, state, and (c) does not bind urokinase. In current studies, investigators are exploring the roles of these molecules and the target serine proteases they inhibit in the pathogenesis of gliomas.     

Vascularization of melanoma by mobilization and remodeling of preexisting latent vessels to patency

Tumors must manipulate the host vasculature to provide a blood supply adequate for their proliferation. Although tumors may arise as avascular masses, there is increasing evidence that some tumors begin to proliferate by first co-opting preexisting host blood vessels. By fluorescent vascular imaging, we provide evidence that the vasculature in orthotopically implanted melanoma arises from a preexisting red cell-deficient vascular network that remodels to patency to accommodate the requirements of the expanding tumor mass. Topical application of vascular endothelial growth factor to vascular beds generated immediate and robust vascular transitions that were morphologically similar to tumor-induced transitions. N(phi)-nitro-L-arginine, a nitric oxide inhibitor, significantly inhibited the growth of a syngeneic K1735M2 melanoma by reducing blood supply to the tumor by a mechanism independent of endothelial cell proliferation. These findings suggest that tumor-induced remodeling of red cell-deficient vessels to patency contributes to tumor vascularization and growth.     

Targeted Antivascular Therapy with the Apolipoprotein(a) Kringle V, rhLK8, Inhibits the Growth and Metastasis of Human Prostate Cancer in an Orthotopic Nude Mouse Model

Antivascular therapy has emerged as a rational strategy to improve the treatment of androgen-independent prostate cancer owing to the necessity of establishing a vascular network for the growth and progression of the primary and metastatic tumor. We determined whether recombinant human apolipoprotein(a) kringle V, rhLK8, produces therapeutic efficacy in an orthotopic human prostate cancer animal model. Fifty thousand androgen-independent human prostate cancer cells (PC-3MM2) were injected into the prostate of nude mice. After 3 days, these mice were randomized to receive the vehicle solution (intraperitoneally [i.p.], daily), paclitaxel (8 mg/kg i.p., weekly), rhLK8 (50 mg/kg i.p., daily), or a combination of paclitaxel and rhLK8 for 4 weeks. Treatment with paclitaxel or rhLK8 alone did not show significant therapeutic effects on tumor incidence or on tumor size compared with the control group. The combination of rhLK8 and paclitaxel significantly reduced tumor size and incidence of lymph node metastasis. Significant reduction in microvessel density and cellular proliferation and induction of apoptosis of tumor cells, and tumor-associated endothelial cells, were also achieved. Similarly, PC-3MM2 tumors growing in the tibia showed significant suppression of tumor growth and lymph node metastasis by the combination treatment with rhLK8 and paclitaxel. The integrity of the bone was significantly preserved, and apoptosis of tumor cells and tumor-associated endothelial cells was increased. In conclusion, these results suggest that targeting the tumor microenvironment with the antivascular effect of rhLK8 combined with conventional cytotoxic chemotherapy could be a new and effective approach in the treatment of androgen-independent prostate cancer and their metastases.     

The effect of P-glycoprotein on paclitaxel brain and brain tumor distribution in mice

It may be inferred from the presence of P-glycoprotein (Pgp) in brain capillaries that this drug efflux pump is a factor in limiting the penetration of certain agents into brain tumors. However, by contrast with normal brain capillaries which constitute the blood-brain barrier, brain tumor capillaries are compromised or "leaky," and the extent to which Pgp expression in brain tumor neovasculature retains its capacity to limit drug penetration has not been determined. To address this question, we studied the normal brain and brain tumor distribution of paclitaxel (PAC), a known Pgp substrate, using steady-state PAC dosing regimens in wild-type and Pgp knockout (mdr1a -/- and mdr1b -/-) mice bearing an intracerebral B-16 melanoma. At comparable steady-state PAC plasma concentrations of approximately 5 microg/ml, steady-state PAC brain concentrations in Pgp knockout mice were approximately 3-, 1.8-, and 1.7-fold greater in left brain, right brain, and brain tumor, respectively, than in wild-type mice and statistically different (P < 0.05) in each brain region. Determination of the steady-state brain/plasma concentration ratios or partition coefficients, which take into account any differences in plasma concentrations between each group, indicated a similar pattern as did the absolute brain concentrations. It is concluded that even in the neovasculature of brain tumors, Pgp has the facility to limit drug penetration, although somewhat less so than in normal brain.     

Induction of chemoresistance in HL-60 cells concomitantly causes a resistance to apoptosis and the synthesis of P-glycoprotein.

The appearance of multidrug-resistant (MDR) proteins or the acquisition of a defective apoptotic programme are major drawbacks in the treatment of cancers since both induce a resistance to classical chemotherapy. However, a link between the two mechanisms has not, as yet, been clearly established. In this study, HL-60 cells cultured in the continual presence of a sub-lethal dose of doxorubicin (dox; HL-60/Dox) were used as a model to study acquired chemoresistance. During the induction of chemoresistance, the appearance of a functional P-glycoprotein (P-gp), in addition to the expression of anti-apoptotic Bcl-2, Bcl-XL and pro-apoptotic Bax proteins was assessed. Parental cells which are sensitive to dox, have no P-gp activity and express Bcl-2 and Bax. After 4 weeks of treatment, a functional P-gp was detected in HL-60/Dox cells. In addition, the synthesis of Bcl-2 appeared to be replaced by Bcl-XL while that of Bax remained unchanged. These cells were also resistant to apoptosis induced by both P-gp and non-P-gp substrates. This inability to induce apoptosis could have resulted from the induction of the expression of the inhibitor of apoptosis protein (XIAP). Our data show that acquired chemoresistance could involve a parallel induction of P-gp and an impairment of the apoptotic pathway.