Search

Unlabelled: Increasing the delivery of therapeutic drugs to the brain improves outcome for patients with brain tumors. Osmotic opening of the blood-brain barrier (BBB) can markedly increase drug delivery, but achieving consistent, good quality BBB disruption (BBBD) is essential. We evaluated four experiments compared with our standard isoflurane/O2 protocol to improve the quality and consistency of BBBD and drug delivery to brain tumor and normal brain in a rat model. Success of BBBD was assessed qualitatively with the large molecular weight marker Evans blue albumin and quantitatively by measuring delivery of the low molecular weight marker [3H]-methotrexate. With isoflurane/O2 anesthesia, the effects of two BBBD drugs of different osmolalities were evaluated at two different infusion rates and infusion durations. Arabinose was superior to saline (P = 0.006) in obtaining consistent Evans blue staining in 16 of 24 animals, and it significantly increased [3H]-methotrexate delivery compared with saline in the tumor (0.388 +/- 0.03 vs 0.135 +/-0.04; P = 0.0001), brain around the tumor (0.269 +/- 0.03 vs 0.035 +/- 0.03; P = 0.0001), brain distant to the tumor (0.445 +/- 0.05 vs 0.034 +/- 0.07; P = 0.001), and opposite hemisphere (0.024 +/- 0.00 vs 0.016 +/- 0.00; P = 0.0452). Forty seconds was better than 30 s (P = 0.0372) for drug delivery to the tumor. Under isoflurane/O2 anesthesia (n = 30), maintaining hypocarbia was better than hypercarbia (P = 0.025) for attaining good BBBD. A propofol/ N2O regimen was compared with the isoflurane/O2 regimen, altering blood pressure, heart rate, and PaCO2 as covariates (n = 48). Propofol/N2O was superior to isoflurane/O2 by both qualitative and quantitative measures (P < 0.0001). Neurotoxicity and neuropathology with the propofol/N2O regimen was evaluated, and none was found. These data support the use of propofol/N2O along with maintaining hypocarbia to optimize BBBD in animals with tumors., Implications: Propofol/N2O anesthesia may be better than isoflurane/O2 for optimizing osmotic blood-brain barrier disruption for delivery of chemotherapeutic drugs to brain tumor and normal brain.

Background and Purpose: Osmotic disruption of the blood-brain barrier (BBB) provides a method for transvascular delivery of therapeutic agents to the brain. The apparent global delivery of viral-sized iron oxide particles to the rat brain after BBB opening as seen on MR images was compared with the cellular and subcellular location and distribution of the particles., Methods: Two dextran-coated superparamagnetic monocrystalline iron oxide nanoparticle contrast agents, MION and Feridex, were administered intraarterially in rats at 10 mg Fe/kg immediately after osmotic opening of the BBB with hyperosmolar mannitol. After 2 to 24 hours, iron distribution in the brain was evaluated first with MR imaging then by histochemical analysis and electron microscopy to assess perivascular and intracellular distribution., Results: After BBB opening, MR images showed enhancement throughout the disrupted hemisphere for both Feridex and MION. Feridex histochemical staining was found in capillaries of the disrupted hemisphere. Electron microscopy showed that the Feridex particles passed the capillary endothelial cells but did not cross beyond the basement membrane. In contrast, after MION delivery, iron histochemistry was detected within cell bodies in the disrupted hemisphere, and the electron-dense MION core was detected intracellularly and extracellularly in the neuropil., Conclusion: MR images showing homogeneous delivery to the brain at the macroscopic level did not indicate delivery at the microscopic level. These data support the presence of a physiological barrier at the basal lamina, analogous to the podocyte in the kidney, distal to the anatomic (tight junction) BBB, which may limit the distribution of some proteins and viral particles after transvascular delivery to the brain.

Objective: To compare transient blood-brain barrier disruption (BBBD) by hypertonic mannitol with pharmacological modification of the blood-tumor barrier by the vasoactive peptide bradykinin for delivery of small and large agents to nude rat intracerebral xenografts., Methods: Female nude rats (n = 104) with 6-day intracerebral human small cell lung carcinoma tumors were treated using BBBD (n = 24), intracarotid bradykinin (n = 38), or saline (controls, n = 32) administered intra-arterially. During or immediately after infusion, the rats were given radiolabeled agent (methotrexate or dextran 70; Dupont NEN, Boston, MA). The rats were killed 10 minutes later, and samples of tumor and brain regions were obtained for scintillation counting. Twenty-two additional rats were examined using magnetic resonance imaging after administering one of two contrast agents (gadoteridol or iron oxide nanoparticles) or saline (controls) in conjunction with BBBD or bradykinin., Results: After BBBD, the delivery of both small (methotrexate) and large (dextran 70) radiolabeled tracers was increased 2- to 6-fold in the tumor and 3- to 20-fold in surrounding brain, as compared with saline controls. After bradykinin treatment, there was minimal change in delivery of methotrexate or dextran 70 to tumor and brain around tumor, with the greatest increase less than 60% over controls. Magnetic resonance imaging demonstrated increased delivery of both small and large contrast agents to the treated hemisphere after BBBD. In comparison, no increased tumor enhancement could be detected after bradykinin treatment., Conclusion: BBBD resulted in global delivery of a variety of agents in a wide range of sizes. In this human brain tumor xenograft model, bradykinin was not effective at increasing delivery to the tumor of any agent tested.

When carboplatin (cis-diammine-1,1-cyclobutane-dicarboxylato-platinum) delivery to brain tumors is optimized with osmotic blood-brain barrier disruption (BBBD), high frequency hearing loss can result. Treatment with sodium thiosulfate (STS) blocked carboplatin cytotoxicity against the LX-1 human small cell lung carcinoma cell line in vitro. STS decreased carboplatin-induced ototoxicity in a guinea pig model, as determined by electrophysiological measurements and analysis of inner ear outer hair cell numbers. Protection was found when STS was administered up to 8 h subsequent to carboplatin but not 24 h after carboplatin. In a rat model of osmotic BBBD, STS was neurotoxic when given immediately after BBBD but not when given 60 min after BBBD, when the barrier is reestablished. Thus, delayed administration of STS may provide a mechanism to reduce the cochlear toxicity caused by BBBD-enhanced carboplatin delivery to the brain.

Delivery of adenovirus, herpes simplex virus (HSV), and paramagnetic monocrystalline iron oxide nanoparticles (MION) to rat brain (n = 64) was assessed after intracerebral inoculation or osmotic disruption of the blood-brain barrier (BBB). After intracerebral inoculation, the area of distribution was 7.93 +/- 0.43 mm2 (n = 9) for MION and 9.17 +/- 1.27 mm2 (n = 9) for replication-defective adenovirus. The replication-compromised HSV RH105 spread to 14.00 +/- 0.87 mm2 (n = 8), but also had a large necrotic center (3.54 +/- 0.47 mm2). No infection was detected when virus was administered intra-arterially without hyperosmotic mannitol. After osmotic BBB disruption, delivery of the viruses and MIONs was detected throughout the disrupted cerebral cortex. Positive staining was found in 4 to 845 cells/100 microns thick coronal brain section (n = 7) after adenovirus administration, and in 13 to 197 cells/section (n = 8) after HSV administration. Cells of glial morphology were more frequently stained after administration of adenovirus, whereas neuronal cells were preferentially stained after delivery of both HSV vectors and MION. In a preliminary test of vector delivery in the feline, MION was detected throughout the white matter tracts after inoculation into normal cat brain. Thus MION may be a tool for use in vivo, to monitor the delivery of virus to the central nervous system. Additionally, BBB disruption may be an effective method to globally deliver recombinant viruses to the CNS.

The delivery of viral vectors to the brain for treatment of intracerebral tumors is most commonly accomplished by stereotaxic inoculation directly into the tumor. However, the small volume of distribution by inoculation may limit the efficacy of viral therapy of large or disseminated tumors. We have investigated mechanisms to increase vector delivery to intracerebral xenografts of human LX-1 small-cell lung carcinoma tumors in the nude rat. The distribution of Escherichia coli lacZ transgene expression from primary viral infection was assessed after delivery of recombinant virus by intratumor inoculation or intracarotid infusion with or without osmotic disruption of the blood-brain barrier (BBB). These studies used replication-compromised herpes simplex virus type 1 (HSV; vector RH105) and replication-defective adenovirus (AdRSVlacZ), which represent two of the most commonly proposed viral vectors for tumor therapy. Transvascular delivery of both viruses to intracerebral tumor was demonstrated when administered intraarterially (i.a.) after osmotic BBB disruption (n = 9 for adenovirus; n = 7 for HSV), while no virus infection was apparent after i.a. administration without BBB modification (n = 8 for adenovirus; n = 4 for HSV). The thymidine kinase-negative HSV vector infected clumps of tumor cells as a result of its ability to replicate selectively in dividing cells. Osmotic BBB disruption in combination with i.a. administration of viral vectors may offer a method of global delivery to treat disseminated brain tumors.

Direct intracerebral injection of recombinant adenoviral vectors within the brain parenchyma or the ventricular system results in a limited volume of distribution of virus, as demonstrated by transgene expression. Global delivery to the central nervous system may increase the use of these vectors but only if the viral vectors can cross the blood-brain barrier and result in transduction of the underlying cells. This short-term study examines whether osmotic disruption with mannitol can result in sufficient opening of the vascular endothelium to allow for passage of replication-defective adenovirus containing the Escherichia coli beta-galactosidase gene (lacZ). Virus was injected into the carotid artery of rats after blood-brain barrier disruption with intracarotid hypertonic mannitol, and the animals were killed and analyzed after 4 days. Histochemical analysis and electron microscopy confirmed expression of the E. coli lacZ gene in the pericapillary astrocytes of the ipsilateral cerebral cortex and deep grey matter. Furthermore, the extent of gene transfer and expression correlated with the degree of barrier opening, as measured by Evans blue staining. Transgene expression was not seen in control animals that received intracarotid saline before recombinant virus injection. These data demonstrate, for the first time, that blood-brain barrier disruption can allow for the delivery of functional viral vectors to the central nervous system.

Delivery of viral particles to the brain is limited by the volume of distribution that can be obtained. Additionally, there is currently no way to non-invasively monitor the distribution of virus following delivery to the central nervous system (CNS). To examine the delivery of virus-sized particles across the blood-brain barrier (BBB), dextran coated, superparamagnetic monocrystalline iron oxide particles, with a hydrodynamic diameter of 20 +/- 4 nm, were delivered to rat brain by direct intracerebral inoculation or by osmotic BBB disruption with hypertonic mannitol. Delivery of these particles was documented by magnetic resonance (MR) imaging and, unexpectedly, neuronal uptake was demonstrated by histochemical staining. Electron microscopy (EM) confirmed iron particle delivery across the capillary basement membrane and localization within CNS parenchymal cells following administration with BBB disruption. This is the first histologic and ultrastructural documentation of the delivery of particles the size of virions across the blood-brain barrier. Additionally, these dextran-coated, iron oxide particles may be useful, in and of themselves, as vectors for diagnostic and/or therapeutic interventions directed at the CNS.

The present studies were undertaken to determine if viral particles can be delivered across the rat blood-brain barrier (BBB). Osmotic BBB modification with intracarotid mannitol (25%) was immediately followed by bolus intracarotid administration of 0.5 ml purified, ultraviolet-inactivated, herpes simplex virus type 1 endogenously labeled with 35S-labeled methionine (2.0 x 10(6) cpm, approximately 5 x 10(8) plaque-forming units/ml). After 60 minutes, intravascular virus was cleared by saline perfusion and the animals were sacrificed. A marked increase (fourfold, p less than or equal to 0.02) in radioactivity was observed in the ipsilateral brain hemisphere when compared to control animals without barrier modification. Administration of intravenous virus immediately after BBB modification displayed no difference in delivery when compared to intracarotid saline-infused controls (without BBB modification) suggesting the importance of a first-pass phenomenon. There were no significant differences in serum concentrations among intracarotid or intravenous groups. These preliminary studies suggest the possibility of delivering viral particles across the BBB with osmotic disruption, which may permit delivery of genetic material in replication-defective viral vectors in the feline model of GM2-gangliosidosis.

Osmotic modification of the blood-brain barrier (BBB) provides an experimental model of vasogenic edema, is totally reversible, and does not cause any structural damage. In the present communication, the effect of corticosteroids on drug delivery to normal rat brain was evaluated in this model. Intraperitoneal dexamethasone was administered at doses ranging from 12 to 48 mg/sq m for 3 days; gentamicin delivery to the brain was then evaluated after either intravenous or intracarotid administration in both control and BBB-modified animals. Only animals receiving the highest dose of dexamethasone and in which the gentamicin was given intravenously demonstrated a statistically significant decrease in drug delivery. The effect of dexamethasone over a wide range of dosages, therefore, exhibited only modest effects on drug delivery to normal brain after osmotic BBB disruption.

The inability of most chemotherapeutic agents to adequately penetrate the blood-brain barrier (BBB), in either normal brain or tumor-infiltrated brain, is a major factor limiting the use of chemotherapy in central nervous system malignancy. This barrier, however, can be opened in a reversible manner by the intra-arterial administration of hyperosmotic agents such as mannitol. It has been suggested that the intravenous administration of dimethyl sulfoxide (DMSO) or 5-fluorouracil (5-FU) can accomplish the same thing in a less invasive manner. We have not been able to confirm these findings. DMSO was administered to 25 rats intravenously at concentrations ranging from 25 to 90% or into the internal carotid artery at a concentration of 30%. The penetration of methotrexate, Evans blue-albumin, and hexosaminidase A was then evaluated at intervals ranging from 1.5 to 3.5 hours after administration. Significant barrier opening was not observed in animals receiving intravenous DMSO. Barrier modification, albeit generally modest, was obtained in animals receiving intracarotid DMSO, but this may have been the result of grand mal seizures, inasmuch as 5 of 6 of these animals had such seizures. Several of the animals receiving i.v. DMSO also had seizures, and all animals developed varying degrees of hematuria. Similarly, 5-FU was administered at a dose of 30 mg/kg i.v. and the permeability of the BBB to either Evans blue-albumin or methotrexate was evaluated. No increased permeability of the BBB to these two markers was observed. In summary, osmotic BBB opening in our hands remains the most consistent and reliable means available to open the BBB in a reversible fashion. Neither intravenous DMSO nor 5-FU seems to increase the delivery of chemotherapy or protein tracer to the central nervous system, and the use of DMSO can result in seizures and hematuria.

The effect of reversible blood-brain barrier modification on the delivery of Adriamycin to the brain was studied in a rodent and canine model. Pharmacokinetic and physiological studies were done in these animals after a wide range of doses of Adriamycin (0.1 to 1.0 mg/kg) were administered into the carotid artery following osmotic barrier modification with mannitol. In the absence of barrier modification, no immunoreactive Adriamycin was detected in the cerebrum; whereas, following barrier modification, up to 4.5 micrograms of drug and/or metabolites per g of brain were found. Optimum tissue levels of Adriamycin and metabolites were achieved following barrier modification when the drug was administered by either bolus or slow continuous (15-min) infusion. Immunoreactive drug was identified in brain for up to 6 hr after administration. Significant functional neurotoxicity occurred at all dose levels, even at 0.1 mg/kg, a level at which Adriamycin concentration in the brain was below the level of detectability. Neuropathological examination revealed the presence of necrosis and hemorrhagic infarcts. Thus, these pharmacological and toxicity studies suggest that Adriamycin (or its metabolites) may produce significant clinical neurotoxicity when even small amounts penetrate the blood-brain barrier.

Antibiotics vary widely in their ability to penetrate the blood-brain barrier. In studies of 70 rats, the permeability of the normal blood-brain barrier to gentamicin was shown to be poor. In experimental brain abscesses, during the cerebritic stage of development, the penetration of intravenous antibiotics was increased compared to normal brain but was very inconsistent. Antibiotic delivery to brain abscess was not significantly altered with the administration of high-dose steroids, but the macrophage and glial response was markedly decreased with high-dose steroid therapy. Reversible osmotic blood-brain barrier modification with mannitol increased the delivery of gentamicin both to brain abscess and to the surrounding brain. It also resulted in more consistent tissue drug levels. The clinical implications of these studies suggest that, because of the inconsistent delivery of gentamicin to brain abscess, the therapeutic efficacy of medical management alone may be quite variable. This mode of therapy could possibly increase the efficacy of medical management of brain abscesses, especially in patients with multiple or surgically inaccessible brain abscesses.

The ability to deliver enzymatically active human hexosaminidase A across the blood-brain barrier and into brain cells of the normal rat was examined. Following osmotic blood-brain barrier modification in the rat, intraarterially administered human hexosaminidase A and B were shown to cross the barrier and enter brain cells. Subcellular fractionation studies demonstrated that most of the human enzyme delivered across the barrier was functionally active and appeared to be inside a subcellular organelle. These studies provide evidence that blood-brain barrier modification permits delivery of functionally active hexosaminidase A into subcellular organelles consistent with that known to be the appropriate site of physiologic activity.

The present studies were undertaken to evaluate the possibility that hexosaminidase A, the enzyme deficient in Tay--Sachs disease, could be effectively delivered to brain. Previous studies from our laboratory have shown that hypertonic mannitol can be used to osmotically produce reversible disruption of the blood--brain barrier in animals (rat and dog) and man without significant neurotoxicity and that such barrier modification significantly increases the delivery of cytoreductive chemotherapy agents to selected areas of brain. By using the rat model of blood--brain barrier modification and radiolabeled enzyme, increased hexosaminidase A delivery to brain has been demonstrated in more than 85 animals. The time of injection of hexosaminidase A after blood--brain barrier disruption is critical for maximum delivery. Rapid (over 30 sec) intra-arterial administration of hexosaminidase A immediately after blood--brain barrier disruption resulted in a marked increase in enzyme delivery to the brain when compared with controls without prior barrier disruption. When the enzyme was administered 15-20 min after barrier disruption, approximately 50% less hexosaminidase A was delivered; when given 60-120 min after barrier modification, the amount delivered was the same as in control animals. This critical time course is very different than that seen in trials of low molecular weight chemotherapeutic agents (methotrexate and adriamycin). These preliminary studies suggest that hexosaminidase A can be delivered to the brain by blood--brain barrier modification and may be indicative of the potential for enzyme replacement in patients who hae Tay--Sachs disease.