Tumor regression with regional distribution of the targeted toxin TF-CRM107 in patients with malignant brain tumors.

We investigated regional therapy of recurrent malignant brain tumors with transferrin-CRM107, a conjugate of human transferrin (Tf) and a genetic mutant of diphtheria toxin (CRM107) that lacks native toxin binding. Physiological barriers to delivering proteins to tumor and surrounding infiltrated brain were circumvented with high-flow interstitial microinfusion. At least a 50% reduction in tumor volume on magnetic resonance imaging (MRI) occurred in 9 of 15 patients who could be evaluated (60%), including two complete responses. Peritumoral toxicity developed 1-4 weeks after treatment in three of three patients at 1.0 microg/ml, but in zero of nine patients treated at lower concentrations. No symptomatic systemic toxicity occurred. Regional perfusion with Tf-CRM107 produces tumor responses without systemic toxicity in patients with malignant brain tumors refractory to conventional therapy. Direct interstitial infusion can be used successfully to distribute a large protein in the tumor and infiltrated brain surrounding the tumor.

Therapy of malignant brain tumors by intratumoral implantation of retroviral vector-producing cells.

Intratumoral implantation of murine cells modified to produce retroviral vectors containing the herpes simplex virus-thymidine kinase (HSV-TK) gene induces regression of experimental brain tumors in rodents after ganciclovir (GCV) administration. We evaluated this approach in 15 patients with progressive growth of recurrent malignant brain tumors. Antitumor activity was detected in five of the smaller tumors (1.4 +/- 0.5 ml). In situ hybridization for HSV-TK demonstrated survival of vector-producing cells (VPCs) at 7 days but indicated limited gene transfer to tumors, suggesting that indirect, "bystander," mechanisms provide local antitumor activity in human tumors. However, the response of only very small tumors in which a high density of vector-producing cells had been placed suggests that techniques to improve delivery and distribution of the therapeutic gene will need to be developed if clinical utility is to be achieved with this approach.

In vivo gene transfer with retroviral vector-producer cells for treatment of experimental brain tumors.

Direct in situ introduction of exogenous genes into proliferating tumors could provide an effective therapeutic approach for treatment of localized tumors. Rats with a cerebral glioma were given an intratumoral stereotaxic injection of murine fibroblasts that were producing a retroviral vector in which the herpes simplex thymidine kinase (HS-tk) gene had been inserted. After 5 days during which the HS-tk retroviral vectors that were produced in situ transduced the neighboring proliferating glioma cells, the rats were treated with the anti-herpes drug ganciclovir. Gliomas in the ganciclovir- and vector-treated rats regressed completely both macroscopically and microscopically. This technique exploits what was previously considered to be a disadvantage of retroviral vectors--that is, their inability to transfer genes into nondividing cells. Instead, this feature of retroviruses is used to target gene delivery to dividing tumor cells and to spare nondividing neural tissue.

Lymphoid cell-glioma cell interaction enhances cell coat production by human gliomas: novel suppressor mechanism.

Certain human glioma lines produce mucopolysaccharide coats that impair the generation of cytolytic lymphocytes in response to these lines in vitro. Coat production is substantially enhanced by the interaction of glioma cells with a macromolecular factor released by human peripheral blood mononuclear cells in culture. This interaction thus constitutes an unusual mechanism by which inflammatory cells may nonspecifically suppress the cellular immune response to at least one class of solid tumors in humans.

Correlation of end-tidal CO2 with transcranial Doppler flow velocity is decreased during chemoregulation in delayed cerebral vasospasm after subarachnoid haemorrhage--results of a pilot study.

BACKGROUND: Cerebrovascular responses to variations in blood pressure and CO2 are attenuated during delayed vasospasm after subarachnoid hemorrhage (SAH). Transcranial Doppler sonography (TCD) is routinely used to assess the presence of vasospasm, but cerebral blood flow velocities (CBF-V) measured by TCD do not necessarily reflect cerebral blood flow (CBF) or the severity of vasospasm. We hypothesized that the correlation of end-tidal pCO2 levels with CBF-V and CBF is equally decreased in subjects with cerebral vasospasm during variations in pCO2. METHODS: Four cynomolgus monkeys were assigned to the vasospasm group and eight animals to the control group. The animals in the vasospasm group underwent placement of an autologous subarachnoid blood clot and vasospasm was confirmed by angiography on day 7. In both groups, CBF and CBF-V were measured simultaneously while end-tidal pCO2 was altered. CBF was measured using a thermal probe placed on the cortical surface and CBF-V was measured using a commercial TCD device. RESULTS: Pearson's correlation coefficient between CBF-V values and pCO2 levels in the control group was strong (r = 0.94, p < 0.001) while it was moderate in the vasospasm group (r = 0.54, p = 0.04). The correlation of CBF values with pCO2 in healthy controls was equally strong (r = 0.87, p = 0.005), while there was no correlation in the vasospasm group (r = -0.09, p = 0.83). CONCLUSION: In this pilot study, correlations of CBF-V with pCO2 values during chemoregulation testing were lower in animals with vasospasm than in healthy ones. This correlation coefficient based on modifications in pCO2 may potentially facilitate the non-invasive assessment of vasospasm.

Expression of the vascular permeability factor/vascular endothelial growth factor gene in central nervous system neoplasms.

Expression of the vascular permeability factor/vascular endothelial growth factor (VEGPF) gene was investigated in human central nervous system (CNS) neoplasms and normal brain. Adsorption of capillary permeability activity from human glioblastoma multiforme (GBM) cell conditioned medium and GBM cyst fluids by anti-VEGPF antibodies demonstrated that VEGPF is secreted by GBM cells and is present in sufficient quantities in vivo to induce vascular permeability. Cloning and sequencing of polymerase chain reaction-amplified GBM and normal brain cDNA demonstrated three forms of the VEGPF coding region (567, 495, and 363 nucleotides), corresponding to mature polypeptides of 189, 165, and 121 amino acids, respectively. VEGPF mRNA levels in CNS tumors vs. normal brain were investigated by the RNase protection assay. Significant elevation of VEGPF gene expression was observed in 81% (22/27) of the highly vascular and edema-associated CNS neoplasms (6/8 GBM, 8/8 capillary hemangioblastomas, 6/7 meningiomas, and 2/4 cerebral metastases). In contrast, only 13% (2/15) of those CNS tumors that are not commonly associated with significant neovascularity or cerebral edema (2/10 pituitary adenomas and 0/5 nonastrocytic gliomas) had significantly increased levels of VEGPF mRNA. The relative abundance of the forms of VEGPF mRNA was consistent in tumor and normal brain: VEGPF495 > VEGPF363 > VEGPF567. In situ hybridization confirmed the presence of VEGPF mRNA in tumor cells and its increased abundance in capillary hemangioblastomas. Our results suggest a significant role for VEGPF in the development of CNS tumor neovascularity and peritumoral edema.

Mechanism of dexamethasone suppression of brain tumor-associated vascular permeability in rats. Involvement of the glucocorticoid receptor and vascular permeability factor.

Brain tumor-associated cerebral edema arises because tumor capillaries lack normal blood-brain barrier function; vascular permeability factor (VPF, also known as vascular endothelial growth factor, VEGF) is a likely mediator of this phenomenon. Clinically, dexamethasone reduces brain tumor-associated vascular permeability through poorly understood mechanisms. Our goals were to determine if suppression of permeability by dexamethasone might involve inhibition of VPF action or expression, and if dexamethasone effects in this setting are mediated by the glucocorticoid receptor (GR). In two rat models of permeability (peripheral vascular permeability induced by intradermal injection of 9L glioma cell-conditioned medium or purified VPF, and intracerebral vascular permeability induced by implanted 9L glioma), dexamethasone suppressed permeability in a dose-dependent manner. Since 80% of the permeability-inducing activity in 9L-conditioned medium was removed by anti-VPF antibodies, we examined dexamethasone effects of VPF expression in 9L cells. Dexamethasone inhibited FCS- and PDGF-dependent induction of VPF expression. At all levels (intradermal, intracranial, and cell culture), dexamethasone effects were reversed by the GR antagonist mifepristone (RU486). Dexamethasone may decrease brain tumor-associated vascular permeability by two GR-dependent mechanisms: reduction of the response of the vasculature to tumor-derived permeability factors (including VPF), and reduction of VPF expression by tumor cells.

Continuous administration of synthetic ovine corticotropin-releasing factor in man. Physiological and pathophysiological implications.

The continuous 24-h infusion of a maximally stimulating dose (1 micrograms/kg per h) of ovine corticotropin-releasing factor (CRF) in man caused a modest elevation of plasma cortisol (17.2 +/- 1.4 micrograms/dl) and urinary-free cortisol (173 +/- 43 micrograms/24 h) concentrations, which was far less than that seen with a maximally stimulating dose of ACTH (50.4 +/- 2.2 micrograms/dl and 1,200 +/- 94 micrograms/24 h, respectively). The circadian rhythms of plasma ACTH and cortisol were preserved during CRF administration. An intravenous bolus injection of 1 microgram/kg of ovine CRF given to normal volunteers under basal conditions resulted in elevated plasma ACTH and cortisol peak levels (28 +/- 6 pg/ml and 15.0 +/- 1.0 micrograms/dl, respectively). However, no plasma ACTH and cortisol responses were observed when an identical CRF stimulation test was given at the end of the continuous infusion. These findings suggest that the stimulatory activity of exogenous CRF on the ACTH-secreting cells of the pituitary gland is restrained by the negative feedback of cortisol. The persistent circadian rhythm of ACTH, despite a constant level of plasma CRF during the infusion, suggests that the circadian variation in the activity of the hypothalamic-pituitary-adrenal axis cannot be explained solely by circadian periodicity of the endogenous CRF stimulus.

Neural substrates of cue reactivity and craving in gambling disorder.

Cue reactivity is an established procedure in addictions research for examining the subjective experience and neural basis of craving. This experiment sought to quantify cue-related brain responses in gambling disorder using personally tailored cues in conjunction with subjective craving, as well as a comparison with appetitive non-gambling stimuli. Participants with gambling disorder (n=19) attending treatment and 19 controls viewed personally tailored blocks of gambling-related cues, as well as neutral cues and highly appetitive (food) images during a functional magnetic resonance imaging (fMRI) scan performed ~2-3 h after a usual meal. fMRI analysis examined cue-related brain activity, cue-related changes in connectivity and associations with block-by-block craving ratings. Craving ratings in the participants with gambling disorder increased following gambling cues compared with non-gambling cues. fMRI analysis revealed group differences in left insula and anterior cingulate cortex, with the gambling disorder group showing greater reactivity to the gambling cues, but no differences to the food cues. In participants with gambling disorder, craving to gamble correlated positively with gambling cue-related activity in the bilateral insula and ventral striatum, and negatively with functional connectivity between the ventral striatum and the medial prefrontal cortex. Gambling cues, but not food cues, elicit increased brain responses in reward-related circuitry in individuals with gambling disorder (compared with controls), providing support for the incentive sensitization theory of addiction. Activity in the insula co-varied with craving intensity, and may be a target for interventions.

A comparison of immunometric and radioimmunoassay measurement of ACTH for the differential diagnosis of Cushing's syndrome.

OBJECTIVE: Measurement of plasma ACTH levels by radioimmunoassay (RIA) is used to identify adrenal causes (AA) of Cushing's syndrome (CS) and to distinguish ectopic CS (EAS) from Cushing's disease (CD) using CRH stimulation testing and inferior petrosal sinus sampling (IPSS). We wished to determine whether diagnostic criteria developed with RIA would also be applicable for immunoradiometric (IRMA) or immunochemiluminescent (ICMA) assays. SUBJECTS AND METHODS: ACTH was measured by RIA, immunoradiometric and/or immunochemiluminescent assay on samples obtained during three types of diagnostic testing in a tertiary referral setting: a) basally (63 CD, 5 AA, 2 EAS and 37 non-CS patients); b) in 44 CD patients following CRH; c) in 6 ectopic CS and 17 CD patients during IPSS. The primary outcome was comparison of diagnostic utility. RESULTS: a) IRMA results, while lower, correlated highly with RIA (r=0.9, p<0.0001) and had similar sensitivity (100 vs 80%) and specificity (89 vs 94%) for the diagnosis of AA (p=0.3); b) the sensitivity for CD of CRH testing using IRMA was similar to that of RIA (85 vs 83%, p=1.0); c) during IPSS, IRMA had similar sensitivity (100%) and specificity (100 vs 83%) compared with ICMA or RIA (p=1.0). CONCLUSIONS: ACTH immunometric assays correlate closely to RIA and offer similar diagnostic utility.

Morphology of interleukin-2-stimulated human peripheral blood mononuclear effector cells killing glioma-derived tumor cells in vitro.

This is the first morphological study of interleukin-2-stimulated human peripheral blood mononuclear (PBM) cells resulting in lymphokine-activated killer (LAK) cell activity against human glioma-derived tumor cells in vitro, in which high-resolution differential interference video light microscopy, scanning electron microscopy, and transmission electron microscopy were used. A subset of cells within the LAK cell population are the effector cells and have an asymmetric cellular architecture characteristic of cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells. Upon binding to target cells, the LAK effector cell nucleus is positioned away from the target cell, whereas the granules, Golgi apparatus, and microtubules orient toward the target cell. These LAK-glioma cell conjugates form very tight plasma membrane bonds with numerous interdigitations, and vesicles were found in the small extracellular spaces between the cells. This morphology was not observed in unstimulated PBM-glioma cell co-cultures. Glioma-derived cells react to LAK effector cells by blebbing, becoming round, and rapidly detaching from the substrate. The injured glioma-derived cells had a highly condensed cytoplasm and chromatin, lobular nucleus, and severe plasma membrane blebs, which are consistent with an apoptotic rather than an osmotic lysis mechanism of cell death. This study provides morphological evidence that supports a common cytotoxic mechanism for CTLs, NK cells, and LAK effector cells. The cytotoxic mechanism is based on the local exocytosis of vesicles by the effector cell into the small extracellular space between the effector-target cell conjugate. Granules found in CTLs, NK cells, and LAK cells contain a pore-forming protein that inserts holes in the target cell's plasma membrane through which a lethal substance(s) not yet identified is thought to enter the cell.

Pharmacokinetics and toxicology of immunotoxins administered into the subarachnoid space in nonhuman primates and rodents.

Immunotoxins have been suggested as possible therapeutic agents in patients with leptomeningeal carcinomatosis. The pharmacokinetics, stability, and toxicity of immunotoxins injected into the i.t. space were examined in rats and rhesus monkeys. Monoclonal antibodies specific for the human (454A12 and J1) and rat (OX26) transferrin receptors were coupled to recombinant ricin A chain. In monkeys, the maximally tolerated dose of the anti-human transferrin receptor immunotoxin (454A12-rRA) was a dose that yielded a nominal cerebrospinal fluid (CSF) concentration of approximately 1.2 x 10(-7) M. In rats, the 10% lethal dose (LD10) of the anti-human transferrin receptor immunotoxin was a dose yielding a nominal CSF concentration of 8.8 x 10(-7) M whereas the LD10 of the anti-rat transferrin receptor immunotoxin (OX26-rRA) was a dose yielding a nominal CSF concentration of 1.2 x 10(-7) M. Thus, the species-relevant antibody resulted in toxicity at a concentration one-seventh that of the immunotoxin with the irrelevant antibody. A comparison of the area under the concentration curve at the LD10 for rats with the area under the concentration curve at the maximally tolerated dose in monkeys and humans shows that the species-relevant immunotoxin was a better predictor of the toxic dose of the anti-transferrin receptor immunotoxin in humans than the irrelevant immunotoxin. The pharmacokinetics of the 454A12-rRA immunotoxin within the CSF of monkeys showed a biphasic clearance with an early-phase half-life of 1.4 h and a late phase half-life of 10.9 h. The clearance was 4.4 ml/h or approximately twice the estimated clearance due to bulk flow of CSF. Loss by degradation was ruled out because immunoblot analysis showed that the immunotoxin was stable for up to 24 h after administration. Possible losses in addition to sampling include diffusion into brain tissue and transcapillary permeation. The apparent volume of distribution was 10.1 ml or approximately three-fourths the total CSF volume of the monkey. Dose limiting toxicity corresponded with the selective elimination of Purkinje cells in both rats and monkeys and was manifested clinically as ataxia and lack of coordination. The onset of ataxia in monkeys occurred within 5 days and, in the more mild form, was reversible with time. There was evidence of only minimal inflammation within the CSF, and there were no signs of systemic toxicity. Immunotoxins injected into the subarachnoid space may have potential for treatment of leptomeningeal carcinomatosis.

In situ retroviral-mediated gene transfer for the treatment of brain tumors in rats.

Gene transfer with vectors derived from murine retroviruses is restricted to cells which are proliferating and synthesizing DNA at the time of infection. This suggests that retroviral-mediated gene transfer might permit targeting of gene integration into malignant cells in organs composed mainly of quiescent nonproliferating cells, such as in the brain. Accordingly, selective introduction of genes encoding for susceptibility to otherwise nontoxic drugs ("suicide" genes) into proliferating brain tumors may be used to treat this cancer. We investigated the efficacy and dynamics of in vivo transduction of growing brain tumors with the herpes simplex-thymidine kinase gene followed by administration of the antiviral drug ganciclovir. Ganciclovir is phosphorylated by thymidine kinase to toxic triphosphates that interfere with DNA synthesis, resulting in the preferential death of the transduced tumor cells. Rats inoculated with 4 x 10(4) 9L gliosarcoma cells into the frontal lobe were treated 7 days later with an intratumoral stereotaxic injection of murine fibroblasts (NIH 3T3 cells) that were producing a retroviral vector containing the herpes simplex-thymidine kinase gene. Controls received vector producer and nonproducer NIH 3T3 cell lines containing the Escherichia coli lacZ (beta-galactosidase) gene as well as nonproducer NIH 3T3 cells containing the thymidine kinase gene. The animals were rested for 7 days to allow time for in situ transduction of the proliferating tumor cells with the herpes-thymidine kinase retroviral vector. The animals were then treated with ganciclovir, 15 mg/kg i.p. twice a day for 14 days. Gliomas receiving an injection of 3-5 x 10(6) thymidine kinase producer cells regressed completely in 23 of 30 rats given ganciclovir therapy, while 25 of 26 control rats developed large tumors. Intratumoral injection of a lower concentration of thymidine kinase vector producer cells (1.8 x 10(6)) resulted in a lower frequency of tumor regression (5 of 13 rats). To estimate the efficiency of in vivo gene transfer, 9L brain tumors were given injections of 5 x 10(6) beta-galactosidase vector producer cells. 5-Bromo-4-chloro-3-indolyl-beta-D-galactopyranaside staining revealed maximal staining of beta-galactosidase within the tumor 7-14 days after injection of the vector producer cells. In vivo transduction rates in harvested tumors ranged from 10 to 70%. There was no evidence of transduction of the surrounding normal neural tissue. Occasional blood vessel endothelial cells within or adjacent to the tumor were observed to be 5-bromo-4- chloro-3-indolyl-beta-D-galactopyranaside positive.(ABSTRACT TRUNCATED AT 400 WORDS)

Evidence for the involvement of a potential second tumor suppressor gene on chromosome 17 distinct from p53 in malignant astrocytomas.

Molecular analysis of malignant astrocytomas demonstrated three distinct groups of tumors with chromosome 17p abnormalities, which include (a) deletion of the p53 locus (17p13.1) and mutations in the remaining allele, (b) deletion of the p53 locus but no detectable mutations in the remaining allele, and (c) deletions not including the p53 locus but mutations in one of the alleles. Furthermore, deletion mapping analysis demonstrated allelic loss of genes distal to D17S28/D17S5 markers (17p13.3) in group C tumors. The loss of heterozygosity of genes on chromosome 17 without detectable mutation (group B) or deletion (group C) in the p53 gene implies the presence of a second tumor suppressor gene in the telomeric region of 17p, the homozygous functional inactivation of which may play a role, either alone or in conjunction with p53, in the initiation and/or progression of astrocytic neoplasms.

Vascular protection by chloroquine during brain tumor therapy with Tf-CRM107.

Tf-CRM107 is a conjugate of transferrin and a point mutant of diphtheria toxin that selectively kills cells expressing high levels of the transferrin receptor. Tf-CRM107 has been infused intratumorally into patients with malignant brain tumors. Although approximately half of the patients exhibit tumor responses, patients receiving higher doses of Tf-CRM107 may develop magnetic resonance image (MRI) evidence of toxicity indicative of small vessel thrombosis or petechial hemorrhage. Consistent with these clinical results we found that intracerebral injection of Tf-CRM107 into rats at total doses > or =0.025 microg causes brain damage detectable by MRI and histology. To widen the therapeutic window of Tf-CRM107, we explored ways to prevent this damage to the vasculature. We reasoned that the vasculature may be protected to a greater extent than tumor from Tf-CRM107 infused into brain parenchyma by i.v. injection of reagents with low blood-brain barrier permeability that block the toxicity of Tf-CRM107. Chloroquine, a well-characterized antimalarial drug, blocks the toxicity of diphtheria toxin and Tf-CRM107. Systemic administration of chloroquine blocked the toxicity of Tf-CRM107 infused intracerebrally in rats and changed the maximum tolerated dose of Tf-CRM107 from 0.2 to 0.3 microg. Moreover, chloroquine treatment completely blocked the brain damage detected by MRI caused by intracerebral infusion of 0.05 microg of Tf-CRM107. In nude mice bearing s.c. U251 gliomas, chloroquine treatment had little effect on the antitumor efficacy of Tf-CRM107. Thus, chloroquine treatment may be useful to reduce the toxicity of Tf-CRM107 for normal brain without inhibiting antitumor efficacy and increase the therapeutic window of Tf-CRM107 for brain tumor therapy.

Heterogeneity of subcellular localization of p53 protein in human glioblastomas.

Immunohistochemical analysis of the p53 protein in human glioblastomas with known genetic profiles of p53 mutations and allele losses on chromosome 17p demonstrated a heterogeneous pattern of subcellular compartmentalization of the p53 protein. Tumors with a single wild type copy of the p53 gene but with allelic deletions on chromosome 17p exhibit nuclear and/or cytoplasmic accumulation of p53, whereas tumors with both copies of the wild type gene and no allele losses on chromosome 17 do not accumulate p53. Glioblastomas with one normal and one mutated copy of the p53 gene and allelic deletions on 17p distal to p53, on the other hand, show predominantly cytoplasmic staining, probably originating from the wild type p53 protein. Furthermore, tumors with mutations in the same codon of p53 display quite different intracellular distribution suggesting that, in addition to the genotype of p53, the intracellular microenvironment of a particular tumor is important in determining the subcellular localization of the p53 protein.

In situ cyclopentenyl cytosine infusion for the treatment of experimental brain tumors.

Cyclopentenylcytosine (CPEC; NSC 375575) is a pyrimidine nucleoside analogue that has potent antitumor effects when tested in vitro and also when tested in experimental tumors outside the central nervous system. CPEC exerts its antiproliferative effect through inhibition of CTP synthetase and consequent depletion of CTP and dCTP pools required for cell replication. Due to its poor penetration of the bloodbrain barrier, CPEC has failed to demonstrate therapeutic efficacy in experimental brain tumors after systemic administration. We therefore examined the in vivo activation, distribution, and antitumor effect of CPEC after long-term regional infusion of the drug directly into experimental brain tumors in rats. HPLC analysis of CPEC incubated with homogenized human brain and brain tumor tissue showed minimal degradation of the drug over 24 h. Analysis of rat cerebral 9L gliosarcoma infused with tritium-labeled CPEC demonstrated intratumoral accumulation of the active metabolite CPEC-triphosphate and concomitant depletion of CTP to a much greater extent in tumor tissue than in the adjacent brain. Tumor tissue UTP also decreased, but no significant effects on other ribonucleoside triphosphates were detected. Only trace amounts (< 1%) of CPEC and its metabolites reached peripheral sites, including the liver and kidneys, after intratumoral infusion. Rats treated with continuous intratumoral infusion of CPEC for 4 weeks using s.c. implanted osmotic pumps survived significantly longer than control rats receiving intratumoral saline or i.p. CPEC (P < 0.0001). Long-term intratumoral infusion of CPEC was not associated with any detectable toxicity. Our results support the feasibility of using intratumoral administration of CPEC as a regional therapy for malignant brain tumors.

Reduced systemic drug exposure by combining intraarterial cis-diamminedichloroplatinum(II) with hemodialysis of regional venous drainage.

During cancer chemotherapy toxicity to normal tissues often limits the tolerable dose. To increase drug delivery to tumor while maintaining tolerable systemic exposure, regional treatments, such as intraarterial drug delivery, have been used. Despite intraarterial delivery, systemic toxicity often remains the dose-limiting sensitivity. If systemic drug exposure could be reduced after intraarterial infusion, the intraarterial dose could be increased, which should increase the therapeutic response. We compared the pharmacokinetic advantage after cisplatin infusion into the internal carotid artery to that obtained after infusing cisplatin into the internal carotid artery during extracorporeal removal of cisplatin from the jugular blood by hemodialysis. Four patients with malignant gliomas received intracarotid cisplatin, 100 mg/m2 over 60 min, every 4 weeks. During one treatment, while cisplatin was infused into the internal carotid artery, the jugular blood was dialyzed extracorporeally at 300 ml/min and returned to the inferior vena cava. Seventy to 96% of the free platinum that entered the dialyzer was removed. By aspirating blood from the jugular vein at 300 ml/min, 30-79% of the ipsilateral carotid blood was collected for extracorporeal circulation. Hemodialysis of the cerebral venous drainage during intracarotid infusion reduced the systemic exposure to cisplatin by 51-61% when compared to the exposure from internal carotid artery infusion without hemodialysis. The pharmacokinetic advantage (brain/body exposure ratio) was increased from 3 to 5/1 during internal carotid artery infusion alone to as much as 15/1 during treatment combining intracarotid infusion with hemodialysis of the jugular blood. Systemic toxicity now limits the dose of cisplatin that can be administered safely. Increased tumor exposure without increased systemic toxicity may be possible with the technique described and greater doses of cisplatin. Assuming no associated local toxicities, the results of the current study indicate that the dose of intracarotid cisplatin can be increased while maintaining tolerable systemic exposure.

Amplification and/or overexpression of platelet-derived growth factor receptors and epidermal growth factor receptor in human glial tumors.

Analysis of genomic organization and expression of platelet-derived growth factor receptors (PDGFR) and epidermal growth factor receptor (EGFR) in human malignant gliomas showed amplification and overexpression of both receptors in distinct subsets of tumors. Amplification of the alpha PDGFR was detected in 4 of 50 glioblastomas (8%). EGFR was amplified in 9 of the 50 tumors (18%). Western blot analysis showed elevated expression of alpha PDGFR and EGFR proteins in 4 (24%) and 3 (18%), respectively, of 17 tumor specimens analyzed. Increased production of alpha PDGFR as well as EGFR proteins was observed in the presence or absence of gene amplification. Three of the 4 tumors with elevated levels of alpha PDGFR also overexpressed the beta PDGFR, which was present as a single copy gene in all 50 tumors analyzed. Our findings suggest that the amplification and/or overexpression either of EGFR or of the alpha PDGFR along with the coordinate overexpression of the beta PDGFR can contribute to the malignant phenotype of distinct subsets of human glioblastoma.

Measurement of thymidine replacement in patients with high grade gliomas, head and neck tumors, and high grade sarcomas after continuous intravenous infusions of 5-iododeoxyuridine.

Based upon the radiation sensitization properties of the halogenated pyrimidines, 5-iododeoxyuridine (IdUrd) and 5-bromodeoxyuridine, long term i.v. infusions of halogenated pyrimidines in conjunction with fractionated radiation therapy have been evaluated in the treatment of a variety of human malignancies. While clinical studies have attempted to measure the halogenated pyrimidine incorporation, few have successfully related tumor response to the incorporation of IdUrd by the tumor. The present study reports the continuous IdUrd labeling index (number of cells labeled) and the IdUrd corrected replacement (percentage of thymidine replacement in the labeled cells of the population) from the tumors of 17 patients who received continuous infusions of IdUrd (1000 mg/m2/24 h). The tumors treated included four high grade gliomas, five head and neck tumors, four high grade sarcomas, and five other tumors of varying types. Less than 25% of the cells in three of four gliomas incorporated IdUrd after 5-7-day IdUrd infusion time. Corrected replacement for the gliomas ranged from 0 to 4%. In contrast, 63-85% of the cells in the head and neck biopsies were labeled with IdUrd after 3-7-day IdUrd infusions suggesting that these large tumors (3-12 cm diameter) have a high fraction of dividing cells. Corrected replacements values for the head and neck tumor patients ranged from 2.9 to 26.3%. The high grade sarcomas also demonstrated a high percentage of IdUrd labeled cells (57-79%) with three patients having corrected replacements of 7.5-14.2%. The continuous labeling and thymidine replacement data for four patients from whom serial biopsies were taken during IdUrd infusion demonstrated both an increasing IdUrd replacement and continuous labeling index with an increasing duration of IdUrd infusion. The clinical response of both the high grade glioma and head and neck tumor patients indicate that the IdUrd replacement and labeling data may provide some important predictive information with regard to the successful use of the halogenated pyrimidines in clinical radiation trials.