The treatment of moderate to severe psoriasis with a new anti-CD11a monoclonal antibody.

BACKGROUND: Anti-CD11a (hu1124) is a humanized monoclonal antibody directed against the CD11a subunit of LFA-1. This study investigated whether treatment with anti-CD11a antibody provides clinical benefit to patients with moderate to severe plaque psoriasis. METHODS: This was a double-blind, placebo-controlled, phase II, multicenter study. In total, 145 patients with minimum Psoriasis Area and Severity Index scores of 12 and affected body surface area of 10% or more were sequentially enrolled into low-dose (0.1 mg/kg, n = 22) or high-dose (0.3 mg/kg, n = 75) groups. Within groups, patients were randomized to treatment or placebo (n = 48) in a 2:1 ratio. Drug was administered intravenously at weekly intervals for 8 weeks. RESULTS: The percentage of subjects achieving more than 50% improvement in physician's global assessment at day 56 (1 week after final dose) was 15% and 48% for placebo and 0.3 mg/kg of drug, respectively (P =.002). A physician's global assessment of excellent (>75% improvement) was greater in the 0.3 mg/kg group versus placebo (25% vs 2%, P =.0003). Average Psoriasis Area and Severity Index scores at day 56 were 13.9 +/- 7.5 (placebo) and 10.9 +/- 8.4 (0.3 mg/kg) (P <.0001). Epidermal thickness was reduced in the 0.3 mg/kg group compared with the placebo group (37% vs 19%, P =.004). Treatment was well tolerated; mild to moderate flu-like complaints were the most common adverse events. White blood cell counts and lymphocyte counts transiently increased. Depletion of circulating lymphocytes did not occur. CONCLUSIONS: Anti-CD11a antibody administered intravenously in 8 weekly doses of 0.3 mg/kg was well tolerated and induced clinical and histologic improvements in psoriasis.

Fluorescence recovery after photobleaching reveals that LPS rapidly transfers from CD14 to hsp70 and hsp90 on the cell membrane.

Although CD14 has been implicated in the immune recognition of bacterial lipopolysaccharide (LPS) from Gram-negative bacteria and also peptidoglycan (PGN) and lipoteichoic acid (LTA) from the outer cell wall of Gram-positive bacteria, accumulating evidence has suggested the possible existence of other functional receptor(s). In this study, we have used fluorescence recovery after photobleaching (FRAP) in order to get the first dynamic picture of the innate recognition of bacteria. We have found that the diffusion coefficient of CD14 remains unaffected after LPS ligation and that the diffusion coefficients of FITC-LPS and FITC-LTA bound to cells differ from that of CD14. Furthermore, FITC-LPS/LTA rapidly become immobile when bound to cells, suggesting that FITC-LPS/LTA must briefly associate with CD14 in the initial attachment process and rapidly move on to an immobile receptor or to a complex of receptors. Further FRAP experiments revealed that heat shock protein 70 (hsp70) and hsp90 are immobile in cell membranes, and antibodies against them were found to block the transfer of LPS to the immobile receptor and to inhibit interleukin 6 production upon LPS stimulation. These experiments indicated that LPS transfers from CD14 to hsp70 and hsp90, which may be part of an LPS/LTA multimeric receptor complex. Thus, hsps are implicated as mediators of the innate activation by bacteria.

Finite element model of antibody penetration in a prevascular tumor nodule embedded in normal tissue.

We have developed a pharmacokinetic model for monoclonal antibodies (mAb) to aid in investigating protocols for targeting small primary tumors or sites of metastatic disease. The model describes the uptake of systemically-administered antibody by a prevascular spherical tumor nodule embedded in normal tissue. The model incorporates plasma kinetics, transcapillary transport, interstitial diffusion, binding reactions, and lymphatic clearance. Antigen internalization can easily be incorporated. Simulations obtained from a three-dimensional finite element analysis are used to assess errors in predictions from earlier models in which the influence of the normal tissue was collapsed into a boundary condition at the tumor surface. The model employing a Dirichlet boundary condition substantially overpredicted the mean total tumor mAb concentration at all times. Although the model with a concentration-dependent flux (composite) boundary condition underpredicted mAb concentration, the discrepancy with finite element results is only notable at early times. Sensitivity analyses were performed on mAb dose and on the coefficients for mAb diffusion in the tissue regions, since reported antibody diffusivity values have varied over 30-fold. The results of the study suggest that mAb diffusivity and mAb binding site density in tumors should have major influences on optimizing doses and scheduling of mAb administration in tumor targeting protocols.

Probe calibration in transient microdialysis in vivo.

PURPOSE: We examine the theoretical basis for calibrating microdialysis probes in vivo for pharmacokinetic experiments in which the extracellular analyte concentrations vary in time. METHODS: A software package, MICRODIAL. was used to simulate microdialysis for illustrative transient situations with linear concentration dependence. RESULTS: For a constant distant extracellular analyte concentration. the calibration factor (extraction fraction, Ed) exhibits a mass transfer transient associated with the development of spatial concentration profiles within the tissue and the probe. Processes clearing the analyte from the extracellular fluid (ECF) strongly influence the rapidity of approach to steady-state and affect the magnitude of the steady-state calibration factor, Ess(d). For situations in which the distant ECF concentration varies in time as a result of exchange with the plasma compartment, different time profiles of the distant ECF and plasma concentrations yield different transient E(d). For the linear, transient cases examined, the area-under-the-curve (AUC 0-infinity) time integral of the distant ECF concentration was found to be proportional to the outflow dialysate concentration-time integral with Ess(d) being the proportionality constant. CONCLUSIONS: The options for calibrating microdialysis probes in solid tissues appear limited under non-steady state conditions; however, AUC integrals for linear systems may be determined by continuous microdialysis sampling and steady-state probe calibration approaches.

A CD14-independent LPS receptor cluster.

Bacterial lipopolysaccharide (LPS), the major structural component of the outer wall of Gram-negative bacteria, is a potent initiator of an inflammatory response and serves as an indicator of bacterial infection. Although CD14 has been identified as the main LPS receptor, accumulating evidence has suggested the possible existence of other functional receptor(s). In this study, using affinity chromatography, peptide mass fingerprinting and fluorescence resonance energy transfer, we have identified four new proteins that form an activation cluster after LPS ligation and are involved in LPS signal transduction. Here we present evidence that implicates heat shock proteins 70 and 90, chemokine receptor 4 and growth differentiation factor 5 as the main mediators of activation by bacterial lipopolysaccharide.

Interactions of bacterial lipopolysaccharide and peptidoglycan with a 70 kDa and an 80 kDa protein on the cell surface of CD14+ and CD14- cells.

Bacterial cell wall components, lipopolysaccharide (LPS), lipoteichoic acid (LTA), and peptidoglycan (PGN) are known to stimulate cells of the immune, inflammatory and vascular systems contributing to septic shock. CD14 has been identified as the main LPS receptor, a process that is accelerated by the serum protein LPS-binding protein (LBP). CD14 has also been found to bind LTA and PGN from the cell wall of gram positive bacteria. Recently, toll-like receptor proteins TLR-2 and TLR-4 have been shown to be required for LPS and LTA-induced intracellular signalling. Although CD14 functions as either a glycosylphosphatidylinositol (GPI)-anchored molecule that does not transverse the cell membrane or as a soluble serum protein, the mechanisms by which the CD14-LPS/LTA complex interacts with the TLRs remains to be elucidated. We have looked directly for cell surface protein(s) that bind LPS or LTA in a CD14-dependent manner. Using biochemical approaches we have identified two proteins of molecular weight 70 kDa (LAP-1) and 80 kDa (LAP-2) that can be precipitated from both CD14(+) and CD14(-) cells with LPS- or LTA-specific antibodies. Binding of LPS and LTA to LAP-1 and -2 required serum. While soluble CD14 (sCD14) was sufficient to allow precipitation of these two proteins from CD14(-) cells, serum could not be replaced by purified sCD14 and/or LBP when mCD14-expressing cells were used.

Differences in uptake and metabolism of retinoic acid between estrogen receptor-positive and -negative human breast cancer cells.

PURPOSE: Our previous work had shown that retinoic acid (RA) inhibits cell growth and induces apoptosis in estrogen receptor-positive (ER-positive) MCF-7 and T-47D human breast carcinoma cells, but not in ER-negative human breast carcinoma cells MB-231 and MB-453. The purpose of this work was to determine whether these differences might be due to differences in uptake and metabolism of the drug between ER-positive and ER-negative cells. METHODS: We measured RA uptake in cultured human breast cancer cells and determined its metabolism by high-pressure liquid chromatographic analysis. RESULTS: The two ER-positive cell lines reached maximum RA uptake at about 2 h, followed by a sharp decline, so that most RA had disappeared from the cells and from the medium by 24 h and was found as oxidation products in the culture medium. In contrast, the two ER-negative cell lines showed a pattern of lower accumulation without the sharp increase and subsequent steep decline, so that by 24 h there was more RA in these cells and their culture medium than in the RA-responsive ER-positive cells, even though at 2 h the ER-negative cells had taken up less RA than the ER-positive cells. Kinetic analysis of the uptake of RA in MCF-7 cells was consistent with rapid movement across the cell membranes and the actual rate determined by diffusion of albumin-bound retinoid to the cells. CONCLUSIONS: This study is the first to demonstrate profound differences in RA accumulation and confirms previous results on different rates of RA metabolism between ER-positive and ER-negative human breast cancer cells. The findings reported here, therefore, may introduce additional elements to be considered in the design of new drugs for cancer chemoprevention and therapy.

Considerations in developing successful, population-based molecular screening and prevention of lung cancer.

The current mortality rate for lung cancer exceeds 85%, as it has for the last 3 decades. This statistic reflects the utility of the major diagnostic tool that has been used during this period to diagnose lung cancer: the chest X-ray. The overwhelming majority of new cases of lung cancer that are detected with chest X-rays involve individuals who already have regional or distant metastatic disease. Because the systemic treatment of this disease has not improved greatly, patients with metastatic disease rarely are cured. This article reviews the issues involved with the development of sputum-based cellular diagnostics for early stage lung cancer. The biomarker, heterogeneous nuclear ribonucleoprotein A2/B1, is the lead marker for this approach. It has been used in several studies in independent cohorts that have suggested that its overexpression in bronchial epithelial cells is associated highly with the development of lung cancer. This marker is detectable 1 year or more prior to the detection of lung cancer by chest X-ray. Finding this early airway-confined phase of lung cancer may allow for the evolution of new management approaches for very early stage lung cancer. Research activities, such aerosolized chemoprevention, are discussed.

Anti-neutrophil cytoplasmic autoantibodies (ANCA) to bactericidal/permeability-increasing (BPI) protein recognize the carboxyl terminal domain.

OBJECTIVES: to identify the region of bactericidal/permeability-increasing protein (BPI) recognized by anti-BPI ANCA. METHODS: sera from 140 patients with a variety of clinical diagnoses (20 systemic vasculitis, 12 cystic fibrosis, 22 bronchiectasis/chronic obstructive airways disease, three diabetes mellitus, 13 chronic renal failure, 12 primary sclerosing cholangitis, eight ulcerative colitis, three Crohn's disease, seven cancer, and 40 other or unknown diagnoses) known to be reactive against native (nBPI), were screened by solid phase enzyme linked immunosorbent assay (ELISA) against a panel of recombinant fusion proteins; holo BPI (rBPI), recombinant lipopolysaccharide binding protein (rLBP), an N-terminal fragment of rBPI (rBPI21 ) and 'fusion' proteins containing the C- or N-terminal ends of BPI spliced with N-or C-ends of LBP, respectively. RESULTS: a strong correlation was seen between the degree of reactivity to rBPI and the BPI C-terminal fusion protein, r=0.69, P < 0.001, as well as between nBPI and rBPI protein, r=0.55, P < 0.001, but not between nBPI and the N-terminal region of BPI (rBPI21), or proteins containing only the N-terminal fragment. Binding to proteins containing the BPI C-terminus was confirmed to be specific by fluid phase inhibition ELISA and Western blot analyses. CONCLUSIONS: together these data suggest that circulating autoantibodies to BPI from patients with different diseases recognize the C-terminal region of BPI.

A pilot phase I trial of continuous hyperthermic peritoneal perfusion with high-dose carboplatin as primary treatment of patients with small-volume residual ovarian cancer.

PURPOSE: Because intraperitoneal (i.p.) therapy may provide a therapeutic advantage and because hyperthermia enhances carboplatin (CBDCA) cytotoxicity, we evaluated the feasibility, toxicity, and pharmacokinetics of CBDCA given via continuous hyperthermic peritoneal perfusion (CHPP) in patients with small-volume residual ovarian cancer. PATIENTS AND METHODS: Six patients underwent optimal cytoreductive procedures (residual disease < or =5 mm) as initial treatment of stages II and III epithelial ovarian adenocarcinoma. All patients received a 90-min CHPP at a CBDCA dose of 800-1200 mg/m2, with the perfusate being recirculated rapidly from a reservoir through a heat exchanger, resulting in i.p. temperatures of 41-43 degrees C. Plasma, perfusate, and urine samples were collected and platinum was quantified by flameless atomic absorption spectrophotometry. RESULTS: At no time did any patient's core temperature exceed 40 degrees C. Peak perfusate platinum concentrations were 8- to 15-fold higher than peak ultrafilterable plasma concentrations. The permeability-area product was extremely high and variable (14-90 ml/min), resulting in a regional advantage of 1.9-5.3. The percentage of the dose absorbed ranged widely from 27% to 77%. Dose-limiting hematologic toxicity was observed at a dose of 1200 mg/m2 and this was associated with a CBDCA AUC in plasma of 11 mg min ml(-1). CONCLUSION: CHPP with CBDCA was safely given to three patients at a dose of 800 mg/m2, and dose-limiting hematologic toxicities observed at 1200 mg/m2, correlated with the plasma CBDCA exposure established when lower doses of CBDCA are given systemically. The pharmacokinetic data are consistent with the expected effect of vigorous mixing on the exposed peritoneal surface area. Variable drug absorption and clearance make the prediction of systemic exposure highly uncertain. These findings may have important implications for novel therapies given i.p.

Population pharmacokinetics and pharmacodynamics of the anti-CD11a antibody hu1124 in human subjects with psoriasis.

The pharmacokinetics of hu1124, a human anti-CD11a antibody, were investigated in human subjects with psoriasis. CD11a is a subunit of LFA-1, a cell surface molecule involved in T cell mediated immune responses. Subjects received a single dose of 0.03, 0.1, 0.3, 0.6, 1, 2, 3, or 10 mg/kg of hu1124 intravenously over 1-3 hr. Blood samples were collected at selected times from 60 min to 72 days after administration. Plasma samples were assayed for hu1124 by ELISA, and pharmacokinetic analyses were performed on the drug plasma concentrations. As the dose of hu1124 was increased, the clearance decreased from 322 ml/day per kg at 0.1 mg/kg to 6.6 ml/day per kg at 10 mg/kg of hu1124. The plasma hu1124 concentration-time profile suggested that the clearance of hu1124 was saturable above 10 micrograms/ml. In addition, treatment with hu1124 caused a rapid reduction in the level of CD11a expression on CD3-positive lymphocytes (T cells) to about 25% of pretreatment levels. Regardless of the hu1124 dose administered, cell surface CD11a remained at this reduced level as long as hu1124 was detectable (> 0.025 microgram/ml) in the plasma. When hu1124 levels fell below 3 micrograms/ml, the drug was rapidly cleared from the circulation and expression of CD11a returned to normal within 7-10 days thereafter. In vitro, half-maximal binding of hu1124 to lymphocytes was achieved at about 0.1 microgram/ml and saturation required more than 10 micrograms/ml. One of the receptor-mediated pharmacokinetic/pharmacodynamic models which was developed describes the dynamic interaction of hu1124 binding to CD11a, resulting in the removal of hu1124 from the circulation and reduction of cell surface CD11a. The model accounts for the continually changing number of CD11a molecules available for removing hu1124 from the circulation based on prior exposure of cells expressing CD11a to hu1124. In addition, the model also accounts for saturation of CD11a molecules by hu1124 at drug concentrations of approximately 10 micrograms/ml, thereby reducing the clearance rate of hu1124 with increasing dose.

Tissue-level transport mechanisms of intraperitoneally-administered monoclonal antibodies.

Monoclonal antibodies (MAbs), produced for specific tumor antigens, can be linked with radioisotopes or metabolic toxins and administered intraperitoneally (i.p.) to treat metastatic cancer located on the peritoneum. Despite their specific binding properties, these proteins distribute to the serosal surface of all tissues surrounding the cavity in the same manner as other serum proteins. Recent data have raised a problem of access of the solution containing the MAb to significant portions of the peritoneal surface. If the MAb does arrive at the surface of the tumor, it penetrates via diffusion and convection. The rapidity and depth of penetration of the MAb are very dependent on the binding characteristics of the MAb to the tumor cells. Current data indicate that tumors often have a large interstitial space relative to normal muscle, and this can accelerate both diffusion and convection. However, a highly permeable tumor vasculature in the absence of lymphatic drainage has also been shown to produce interstitial pressure gradients from the center toward the periphery of the tumor, setting up a potential outward flow which may be a significant barrier to the movement of MAbs into the nodule. While theoretical mechanisms of diffusion, convection, and binding are well established, there is still a great need for in vivo data.

Intraventricular immunotoxin therapy for leptomeningeal neoplasia.

OBJECTIVE: The goals of this clinical trial of intraventricular 454A12-rRA therapy were to identify dose-limiting toxicities, to evaluate the pharmacokinetics of single-dose intraventricular 454A12-rRA, and to detect antitumor activity. METHODS: We performed a pilot study of intraventricular therapy with the immunotoxin 454A12-rRA in eight patients with leptomeningeal spread of systemic neoplasia. The immunotoxin 454A12-rRA is a conjugate of a monoclonal antibody against the human transferrin receptor and recombinant ricin A chain, the enzymatically active subunit of the protein toxin ricin. Patients were treated with single doses of 454A12-rRA ranging from 1.2 to 1200 micrograms. RESULTS: The early phase half-life of 454A12-rRA in ventricular cerebrospinal fluid (CSF) averaged 44 +/- 21 minutes, and the late phase half-life averaged 237 +/- 86 minutes. The clearance of the immunotoxin was faster than the clearance of coinjected technetium-99m-diethylenetriamine penta-acetic acid, averaging approximately 2.4-fold greater. No 454A12-rRA degradation was detected by Western blot analysis of ventricular CSF for a period of 24 hours, and bioactivity was retained in CSF paralleling the concentration of immunotoxin. No acute or chronic drug toxicity was identified in patients who received less than or equal to 38 micrograms of 454A12-rRA by intraventricular injection. Doses more than or equal to 120 micrograms caused a CSF inflammatory response that was associated with transient headache, vomiting, and altered mental status. This acute syndrome was responsive to steroids and CSF drainage. No systemic toxicity was detected. In four of the eight patients, a greater than 50% reduction of tumor cell counts in the lumbar CSF occurred within 5 to 7 days after the intraventricular dose of 454A12-rRA; however, no patient had their CSF cleared of tumor, and clinical or magnetic resonance imaging evidence of tumor progression was demonstrated in seven of the eight patients after treatment. CONCLUSION: Tumoricidal concentrations of the immunotoxin 454A12-rRA can be attained safely in the CSF of patients with leptomeningeal tumor spread.

Decay rates of anti-HIV dideoxynucleotides in tissue culture systems: a simple correction for the effect of cell replication.

Measurement of intracellular drug levels in cell culture systems can be of predictive value in establishing rational clinical dosage schedules. Such in vitro measurements carried out with anti-HIV agents of the 2',3'-dideoxynucleoside (ddN) class have shown that many of the pharmacologically active ddNTP metabolites of these agents have relatively long intracellular half-lives and little or no host-cell cytotoxicity. As a consequence, replication of drug-exposed cells continues at an unperturbed rate so that a systematic dilution error occurs in the measurement of ddNTP decay half-times. The aim of this study is to present a simple general formulation for the correction of measured t1/2-values for ddNTPs and for other agents with similar intracellular pharmacokinetic properties. Two factors of practical interest emerge: first, the error is greater for agents with slow intracellular clearance rates than for agents with rapid rates; and second, for cell lines with long doubling times, the measured t1/2-values approach more closely to the true t1/2-values, until with the extreme case (quiescent or "G(o)" cells), the observed and true decay times are identical. The greatest dilution errors are seen with adenodine-based agents such as ddATP and 2'-F-ddATP, while the smallest errors are seen with rapidly cleared agents of the dideoxythymidine class.

Pharmacokinetic problems in peritoneal drug administration: tissue penetration and surface exposure.

Both theory and clinical studies demonstrate that drug concentrations in the peritoneal cavity can greatly exceed concentrations in the plasma following intraperitoneal administration. This regional advantage has been associated with clinical activity, including surgically documented complete responses in ovarian cancer patients with persistent or recurrent disease following systemic therapy, and has produced a survival advantage in a recent phase III trial. Two pharmacokinetic problems appear to limit the effectiveness of intraperitoneal therapy: poor tumor penetration by the drug and incomplete irrigation of serosal surfaces by the drug-containing solution. We have examined these problems in the context of a very simple, spatially distributed model. If D is the diffusivity of the drug in a tissue adjacent to the peritoneal cavity and k is the rate constant for removal of the drug from the tissue by capillary blood, the model predicts that (for slowly reacting drugs) the characteristic penetration distance is (D/k)1/2 and the apparent permeability of the surface of a peritoneal structure is (Dk)1/2. The permeability-area product used in classical pharmacokinetic calculations for the peritoneal cavity as a whole is the sum of the products of the tissue-specific permeabilities and the relevant superficial surface areas. Since the model is mechanistic, it provides insight into the expected effect of procedures such as pharmacologic manipulation or physical mixing. We observe that large changes in tissue penetration may be difficult to achieve but that we have very little information on the transport characteristics within tumors in this setting or their response to vasoactive drugs. Enhanced mixing is likely to offer significant potential for improved therapy; however, procedures easily applicable to the clinical setting have not been adequately investigated and should be given high priority. Clinical studies indicate that an increase in irrigated area may be achieved in many patients by individualizing the dialysate volume and consideration of patient position.

Biochemical characterization of recombinant fusions of lipopolysaccharide binding protein and bactericidal/permeability-increasing protein. Implications in biological activity.

The physiological response to endotoxin (lipopolysaccharide (LPS)) can be regulated by two closely related LPS-binding proteins, LPS-binding protein (LBP), which potentiates LPS' inflammatory activity via interaction with the monocytic antigen CD14, and bactericidal/permeability-increasing protein (BPI), which neutralizes LPS. Both proteins bind LPS with high affinity sites in their N-terminal domains, whereas interaction between LBP and CD14 is dependent upon the LBP C-terminal domain. We have created fusions of the N- and C-terminal domains from each protein and compared the functional activities and pharmacokinetics of these fusions, the individual N-terminal domains, and the parent proteins. The N-terminal domains of BPI and LBP bound lipid A with their characteristic apparent affinity constants, regardless of the C-terminal fusion partner. In addition, the C-terminal domain of LBP allowed transfer of LPS to CD14 in conjunction with either N-terminal LPS binding domain. Proteins containing a BPI N-terminal domain had greater heparin binding capacities in vitro and were cleared more rapidly from the plasma of whole animals. Taken together, these data better define how closely related proteins such as BPI and LBP can have opposing effects on the body's response to LPS.

Targeted deletion of the lipopolysaccharide (LPS)-binding protein gene leads to profound suppression of LPS responses ex vivo, whereas in vivo responses remain intact.

Gram-negative bacterial lipopolysaccharide (LPS) stimulates phagocytic leukocytes by interacting with the cell surface protein CD14. Cellular responses to LPS are markedly potentiated by the LPS-binding protein (LBP), a lipid-transfer protein that binds LPS aggregates and transfers LPS monomers to CD14. LBP also transfers LPS to lipoproteins, thereby promoting the neutralization of LPS. LBP present in normal plasma has been shown to enhance the LPS responsiveness of cells in vitro. The role of LBP in promoting LPS responsiveness in vivo was tested in LBP-deficient mice produced by gene targeting in embryonic stem cells. Whole blood from LBP-deficient animals was 1,000-fold less responsive to LPS as assessed by the release of tumor necrosis factor (TNF)-alpha. Blood from gene-targeted mice was devoid of immunoreactive LBP, essentially incapable of transferring LPS to CD14 in vitro, and failed to support cellular responses to LPS. These activities were restored by the addition of exogenous recombinant murine LBP to the plasma. Despite these striking in vitro findings, no significant differences in TNF-alpha levels were observed in plasma from wild-type and LBP-deficient mice injected with LPS. These data suggest the presence of an LBP-independent mechanism for responding to LPS. These LBP knockout mice may provide a tool for discovering the nature of the presumed second mechanism for transferring LPS to responsive cells.

Quantification and pharmacokinetics of blood-brain barrier disruption in humans.

Hyperosmolar blood-brain barrier disruption (HBBBD), produced by infusion of mannitol into the cerebral arteries, has been used in the treatment of brain tumors to increase drug delivery to tumor and adjacent brain. However, the efficacy of HBBBD in brain tumor therapy has been controversial. The goal of this study was to measure changes in vascular permeability after HBBBD in patients with malignant brain tumors. The permeability (K1) of tumor and normal brain blood vessels was measured using rubidium-82 and positron emission tomography before and repeatedly at 8- to 15-minute intervals after HBBBD. Eighteen studies were performed in 13 patients, eight with glioblastoma multiforme and five with anaplastic astrocytoma. The HBBBD increased K1 in all patients. Baseline K1 values were 2.1 +/- 1.4 and 34.1 +/- 22.1 microl/minute/ml (+/- standard deviation) for brain and tumor, respectively. The peak absolute increases in K1 following HBBBD were 20.8 +/- 11.7 and 19.7 +/- 10.7 microl/minute/ml for brain and tumor, corresponding to percentage increases of approximately 1000% in brain and approximately 60% in tumor. The halftimes for return of K1 to near baseline for brain and tumor were 8.1 +/- 3.8 and 4.2 +/- 1.2 minutes, respectively. Simulations of the effects of HBBBD made using a very simple model with intraarterial methotrexate, which is exemplary of drugs with low permeability, indicate that 1) total exposure of the brain and tumor to methotrexate, as measured by the methotrexate concentration-time integral (or area under the curve), would increase with decreasing infusion duration and would be enhanced by 130% to 200% and by 7% to 16%, respectively, compared to intraarterial infusion of methotrexate alone; and 2) exposure time at concentrations above 1 microM, the minimal concentration required for the effects of methotrexate, would not be enhanced in tumor and would be enhanced by only 10% in brain. Hyperosmolar blood-brain barrier disruption transiently increases delivery of water-soluble compounds to normal brain and brain tumors. Most of the enhancement of exposure results from trapping the drug within the blood-brain barrier, an effect of the very transient alteration of the blood-brain barrier by HBBBD. Delivery is most effective when a drug is administered within 5 to 10 minutes after disruption. However, the increased exposure and exposure time that occur with methotrexate, the permeability of which is among the lowest of the agents currently used clinically, are limited and the disproportionate increase in brain exposure, compared to tumor exposure, may alter the therapeutic index of many drugs.

An in vitro flow model to study streaming during pelvic intra-arterial drug infusions.

Regional delivery of suitable drugs by intra-arterial infusion may offer a therapeutic advantage. High concentrations in the tumor are sought with reduced systemic toxicity. Adequate mixing of drug solutions with perfusing blood is essential to provide uniform distribution of drug to tumor-bearing tissue distal to the infusion site. Using a glass model of the iliofemoral and pelvic arteries, we have demonstrated that a streaming phenomenon occurs. Laminar "streamers" of slowly infused drug solution originate at the catheter tip and proceed nonuniformly into distal arterial branches. The intensity of streaming and the pattern of distribution are highly sensitive to catheter tip placement and quite unpredictable. The consequence of regional therapy under streaming conditions is severe maldistribution of drug in the infused tissues with potentially high levels delivered to normal tissues and simultaneous subtherapeutic levels delivered to tumor. Our in vitro model can be used to test appropriate infusion techniques that enhance mixing such as pulsed infusions and novel catheter designs.

Lipopolysaccharide-induced E-selectin expression requires continuous presence of LPS and is inhibited by bactericidal/permeability-increasing protein.

Endothelial cells stimulated by LPS express E-selectin, which plays an important role in mediating neutrophil adhesion during inflammation. E-selectin is induced within 1-2 h, peaks at 4-6 h, and gradually returns to basal level by 24 h. rBPI21, a recombinant N-terminal fragment of human bactericidal/permeability-increasing protein (BPI), inhibited LPS-induced E-selectin expression when added at the same time as, and up to 6 h after, LPS. Delayed administration of rBPI21 also affected LPS-mediated activation of the nuclear factor, NF-kappa B. Two to 4 h following LPS addition to endothelial cells, when NF-kappa B was already activated, addition of rBPI21 resulted in marked reduction of NF-kappa B detectable at 4 or 6 h. These results indicate that endothelial activation requires continuous presence of LPS, and rBPI21 acts to reverse LPS-mediated endothelial activation by interrupting the on-going LPS signal.