RNA-protein crosslinking to AMP residues at internal positions in RNA with a new photocrosslinking ATP analog.

A new photocrosslinking purine analog was synthesized and evaluated as a transcription substrate for Escherichia coli RNA polymerase. This analog, 8-[(4-azidophenacyl)thio]adenosine 5'-triphosphate (8-APAS-ATP) contains an aryl azide photocrosslinking group that is attached to the ATP base via a sulfur-linked arm on the 8 position of the purine ring. This position is not involved in the normal Watson-Crick base pairing needed for specific hybridization. Although 8-APAS-ATP could not replace ATP as a substrate for transcription initiation, once stable elongation complexes were formed, 8-APAS-AMP could be site-specifically incorporated into the RNA, and this transcript could be further elongated, placing the photoreactive analog at internal positions in the RNA. Irradiation of transcription elongation complexes in which the RNA contained the analog exclusively at the 3' end of an RNA 22mer, or a 23mer with the analog 1 nt from the 3' end, produced RNA crosslinks to the RNA polymerase subunits that form the RNA 3' end binding site (beta, beta'). Both 8-APAS-AMP and the related 8-azido-AMP were subjected to conformational modeling as nucleoside monophosphates and in DNA-RNA hybrids. Surprisingly, the lowest energy conformation for 8-APAS-AMP was found to be syn, while that of 8-azido-AMP was anti, suggesting that the conformational properties and transcription substrate properties of 8-azido-ATP should be re-evaluated. Although the azide and linker together are larger in 8-APAS-ATP than in 8-N(3)-ATP, the flexibility of the linker itself allows this analog to adopt several different energetically favorable conformations, making it a good substrate for the RNA polymerase.

Influence of nitric oxide on the intracellular reduced glutathione pool: different cellular capacities and strategies to encounter nitric oxide-mediated stress.

Different cell types exhibit huge differences towards the cytotoxic action of NO. In search for an explanation, we used subtoxic concentrations of the NO-donors S-nitrosocysteine (SNOC) for short-term challenge and of (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1- ium-1,2-diolate (DETA/NO) for longer periods of exposure, respectively, and subtoxic concentrations of the oxidant H2O2 to determine the impact on intracellular reduced glutathione (GSH) concentrations. We find that GSH concentrations are always decreased, but that different cell types show different responses. Incubation of the relatively NO-sensitive murine lymphocytes with both NO-donors, but not with H2O2, resulted in a nearly complete loss of intracellular GSH. Short-term NO-treatment of P815 mastocytoma cells, also sensitive to NO-mediated cell death, decreased GSH to a similar extent only if either glutathione reductase (GSHR) activity or y-glutamylcysteine synthetase (gammaGCS) activity were inhibited concomitantly by specific inhibitors. Long-term NO-treatment of P815 cells, however, resulted in a significant decrease of GSH that could be further enhanced by inhibiting gammaGCS activity. In contrast, neither short-term nor long-term NO-exposure nor H2O2-treatment affected intracellular GSH levels of L929 fibroblasts, which were previously shown to be extremely resistant towards NO, whereas concomitant gammaGCS inhibition, but not GSHR inhibition, completely decreased GSH concentrations. These results show that different cell types use different pathways trying to maintain glutathione concentrations to cope with nitrosative stress, and the overall capability to maintain a critical amount of GSH correlates with susceptibility to NO-induced cell death.

Nitric oxide mediates intracytoplasmic and intranuclear zinc release.

We previously described that NO. leads to destruction of ZnS clusters and release of Zn2+ from various proteins including zinc finger transcription factors. To assess the relevance in living cells, we investigated, whether exogenous NO. leads to an increase of cytoplasmic and nuclear free Zn2+. L929 cells, mouse splenocytes, or rat aorta endothelial cells were labeled with Zinquin-E, a Zn2+-specific fluorophore, and were treated with two different spontaneous NO donors, S-nitrosocysteine or DETA/NO. Both NO donors strongly increased the Zn2+-dependent fluorescence in the cellular cytosol and also in nuclei as compared to controls. NO-dependent Zn2+ release in splenocytes was quantitated by flow cytometry. These results show for the first time, that nitrosative stress mediates intracellular and intranuclear Zn2+ release which may be relevant in altering gene expression patterns.

Enhanced green fluorescent protein fusion proteins of herpes simplex virus type 1 thymidine kinase and cytochrome P450 4B1: applications for prodrug-activating gene therapy.

To monitor therapeutic transgene expression, we developed fusion genes of enhanced green fluorescent protein (EGFP) with two different prodrug-activating enzyme genes: herpes simplex virus type 1 thymidine kinase (HSV-tk) and rabbit cytochrome P450 4B1 (cyp4b1). Expression of the resulting fusion proteins, TK-EGFP and 4B1-EGFP, rendered transduced human and rodent glioma cells sensitive to cytotoxic treatment with the corresponding prodrugs ganciclovir and 4-ipomeanol. Ganciclovir and 4-ipomeanol sensitivity was comparable with that achieved with the native HSV-TK and CYP4B1 proteins. As shown by fluorescence microscopy, TK-EGFP was expressed predominantly intranuclearly, whereas 4B1-EGFP was detectable in the cytoplasm, thereby displaying the orthotopic subcellular distribution of the corresponding native enzymes. The fluorescence intensity correlated well with the corresponding prodrug sensitivity, as shown by fluorescence-activated cell sorter analysis. EGFP expression was also used for the selection of stably HSV-tk-transduced cells by flow cytometric cell sorting. Resulting cell populations showed a homogeneity of fluorescence intensity similar to single-cell clones after antibiotic selection. In conclusion, tk-egfp and 4b1-egfp fusion genes are valuable tools for monitoring prodrug-activating gene therapy in living cells. EGFP fusion genes/proteins provide a simple and reproducible means for the detection, selection, and characterization of cells expressing enzyme genes for prodrug activation.

Behavioral effects of psychomotor stimulant infusions into amygdaloid nuclei.

The role of amygdaloid nuclei in locomotion, stereotypy, and conditioned place preference (CPP) produced by psychomotor stimulants was examined. Five 2-day conditioning trials were conducted over 10 consecutive days. Rats received bilateral intracranial infusions of saline, cocaine (25-100 micrograms/side), or amphetamine (0.31-20 micrograms/side) into the ventricles (ICV), basolateral amygdala (BlA), or central amygdala (CeA) and were confined to a compartment. On alternating days, rats received sham infusions and were confined to a different compartment. Locomotion was measured daily, stereotypy was measured on trials 1 and 5, and CPP was measured 24 h after conditioning. ICV infusions of cocaine or amphetamine produced locomotion, rearing, and CPP. Intra-BlA and intra-CeA infusions of the highest dose of cocaine produced locomotion. In contrast, intra-CeA infusions of amphetamine potently produced locomotion and CPP. Intra-BlA infusions of amphetamine, however, did not produce any behavioral changes. These results suggest that the CeA, but not the BlA, is involved in initiating reward and locomotion produced by amphetamine.

Potential tumor or organ-imaging agents. 30. Radioiodinated phospholipid ethers.

A radioiodinated analogue of a naturally occurring alkyl lysophospholipid (ALP) was synthesized for evaluation as a potential tumor-localizing imaging agent. rac-1-[12-(m-Iodophenyl)dodecyl]-2-methylglycero-3-phosphocholine (ET-12IP-OMe, 14) was radiolabeled with iodine-125 via an isotope-exchange procedure. Tissue distribution studies with [125I]ET-12IP-OMe in tumor-bearing rats revealed an immediate tumor uptake of radioactivity. Although radioactivity was also present in nontarget tissues at this time, clearance of tracer from the tumor was much slower and thus provided a suitable tumor to nontarget tissue ratio at 24 h. As a result of this selective accumulation, it was possible to clearly delineate the tumor with gamma-camera scintigraphy.

In vitro evaluation of phosphocholine and quaternary ammonium containing lipids as novel anti-HIV agents.

A series of synthetic lipids containing a two- or three-carbon backbone substituted with a thio, oxy, or amidoalkyl functionality and either a phosphocholine or quaternary ammonium moiety was evaluated as potential anti-HIV-1 agents. Several analogues were identified as possessing activity with the most promising compound being rac-3-octadecanamido-2-ethoxypropylphosphocholine (8). Compound 8 exhibited an IC50 for the inhibition of plaque formation of 0.16 microM which was 84-fold lower than the IC50 value determined for CEM-SS cell growth inhibition. Initial mechanistic studies have indicated that these compounds, unlike AZT, are not reverse transcriptase (RT) inhibitors, but instead appear to inhibit a late step in HIV replication involving virus assembly and infectious virus production. Since these lipids are acting via a different mechanism, they represent an alternative approach to the chemotherapeutic treatment of AIDS as well as candidates for combination therapy with AZT.

Molecular structures and conformational studies of triarylcyclopropyl and related nonsteroidal antiestrogens.

Molecular structures and conformational characteristics of a series of 1,1-dichloro-2,2,3-triarylcyclopropanes (DTACs), which were reported previously to be distinctly antiestrogenic and inhibitors of the estrogen-receptor-positive MCF-7 human breast cancer cells in culture, are reported. In addition, structural and conformational features of the DTACs were compared to the first-known nonsteroidal antiestrogen, MER25, and the clinically useful antiestrogen Tamoxifen. The molecular structures of four DTAC compounds were determined by X-ray diffraction. Crystallographic structures show that the DTAC molecules have nearly the same relative conformation for the three aryl rings which is designated as a "nonpropeller" conformation in contrast to the observed "propeller" conformation for the three rings in all known triarylethylenes. Systematic conformational searches were performed to find the conformational preferences of DTACs, MER25, and Tamoxifen using idealized model compounds built from their respective crystal structure. Energy-minimization and conformational-search studies demonstrated that all DTAC molecules have a common, single global minimum energy conformer for their central core containing the dichlorotriarylcyclopropyl system, which is similar to that found in their crystal structures. Conformational search of MER25 showed that the molecule can assume a number of low-energy conformers of which two, one anti (A1) and one gauche (G1A), have about the same energy. The anti conformation is similar to the one observed in its crystal structure and resembles the estrogenic E-isomer of Tamoxifen, while the lowest energy gauche conformer of MER25 resembles more closely the antiestrogenic Z-isomer of Tamoxifen. NMR spectroscopic analysis of MER25 showed that the molecule exists predominantly in the anti conformation in solution. A comparative review of the structural features and bioactivities of Tamoxifen, DTACs, and MER25 provides a possible explanation for their low estrogen receptor binding affinity which is common to these compounds together with their antiestrogenic activity.

Synthesis and biological evaluation of a series of 1,1-dichloro-2,2,3-triarylcyclopropanes as pure antiestrogens.

A series of 1,1-dichloro-2,2,3-triarylcyclopropanes (DTACs) was synthesized and evaluated as pure antiestrogens. Addition of 4-methoxy- or 4-(benzyloxy)phenyl Grignard reagents to p-methoxy, p-benzyloxy, or unsubstituted deoxybenzoins, followed by dehydration of the resulting carbinols produced a mixture of E and Z olefins, which were reacted with dichlorocarbene to give O-protected DTACs. The E and Z isomers were separated by fractional crystallization and the central or geminal phenyl ring was deprotected to provide phenolic DTACs. Alkylation with (N,N-dimethylamino)ethyl chloride yielded basic cyclopropanes. Two chlorodiarylindenes were isolated as thermolysis products of the DTACs, and one was converted to a phenol by hydrogenolysis. All DTACs and indenes were competitive inhibitors of [3H]estradiol binding in the immature rat uterine cytosol receptor assay, with relative binding affinities of 0.1-3.6% of estradiol. None of the new compounds were estrogenic in the 3-day immature mouse uterotrophic assay at doses up to 750 micrograms. In the 3-day immature mouse antiuterotrophic assay, five DTACs with either a methoxy (5a), benzyloxy (4d, 5c), or (dimethylamino)ethoxy (7a, 7b) central ring side chain produced significant decreases in uterine weight at doses up to 750 micrograms. One compound, (Z)-1,1-dichloro-2-[4-[2-(dimethylamino)ethoxy]-phenyl]-2-(4- methoxyphenyl)-3-phenylcyclopropane (7b), elicited a dose-dependent decrease in vivo comparable to MER 25. These same five compounds, as well as the lead compound Analog II, were active in vitro against the estrogen-dependent MCF-7 human breast tumor cell line in a dose-dependent fashion.

Synthesis of phosphocholine and quaternary amine ether lipids and evaluation of in vitro antineoplastic activity.

The in vitro antineoplastic activity of many phosphorus-containing (e.g., phosphocholines) and non-phosphorus-containing (e.g., quaternary ammonium salts) ether lipids has been evaluated in the HL-60 promyelocytic cell line. These compounds are analogues of ET-18-OMe (1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine). Structural modification of 1-(alkylamido)-, -(alkylthio)-, and -(alkyloxy)propyl backbones has provided further insight into the structure-activity relationships of these lipids. In this study, a long saturated C-1 chain and a three-carbon backbone with a single short C-2 substituent were preferred. At the positively charged nitrogen of phosphocholines, fewer than three substituents caused a significant loss of activity, and substituents larger than methyl decreased activity slightly. In the nonphosphorus compounds, many nitrogen heterocycles and also a sulfonium moiety were incorporated without changing the degree of activity; however, a thiazolium group decreased activity. The most active compound, 29 [N-[3-(hexadecyloxy)-2-methoxypropyl]-3-(hydroxymethyl)pyridinium bromide], was approximately twice as active as the reference standard, ET-18-OMe, in a trypan blue dye exclusion assay.

Synthesis of quaternary amine ether lipids and evaluation of neoplastic cell growth inhibitory properties.

Novel quaternary amine ether lipids have been synthesized and tested for inhibition of neoplastic cell proliferation with the HL-60 promyelocytic leukemia cell line. These compounds contain a positively charged quaternary amine functional group attached either directly to the glycerol backbone or at the end of an alkoxy chain. The biological testing has identified several analogues with activity equivalent to or greater than that exhibited by the reference compound in this assay, ET-18-OMe (1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine). Among the most active analogues are compounds 11, [N,N,N-triethyl-3-(hexadecyloxy)-2-ethoxy-1-propylammonium bromide] and 22 [N-[4-[3-(hexadecyloxy)-2-ethoxypropoxy]-1-butyl]pyridinium bromide], which are approximately 3 times as active as the reference standard.

Synthesis and biological activity of novel quaternary ammonium derivatives of alkylglycerols as potent inhibitors of protein kinase C.

Alkylglycerols such as rac-1-O-octadecyl-2-O-methylglycerophosphochocholine (Et-18-OMe) have shown an inhibitory effect on the metastasis and growth of various cancer cell lines. Alkyl phospholipids have been shown to accumulate at the surface in several cell lines, the selectivity of which is still not clearly understood. A consequence of this action may lead to the inhibition of cell membrane related protein kinase C (PKC). The goal of this research was to develop ether lipid inhibitors of PKC to augment antineoplastic activity. This led to the synthesis and in vitro testing of a series of novel quaternary ammonium derivatives of alkylglycerols. The biological testing of these analogues on PKC stimulated with rac-1-O-oleoyl-2-O-acetylglycerol showed several analogues with inhibition comparable to that of Et-18-OMe.

Synthesis and evaluation of novel ether lipid nucleoside conjugates for anti-HIV-1 activity.

Combinations of an amidoalkylphosphocholine, 8, and AZT have been found to cause an apparent synergistic action in suppressing infectious HIV-1 replication. In addition, amidoalkyl, oxyalkyl, and thioalkyl ether lipids have been chemically linked to anti-HIV-1 nucleosides (AZT and DDI) through phosphate and phosphonate linkages. These conjugates have shown promising in vitro anti-HIV-1 activity. Also, the conjugates have a 5-10-fold reduction in cell cytotoxicity compared to AZT alone. The most active compound, an amidoalkyl ether lipid-AZT conjugates, 4A, was found to have a differential selectivity of 1793 in a syncytial plaque assay. In comparison, AZT alone has a value of 1281.

Tumor visualization with a radioiodinated phospholipid ether.

The known ability of phospholipid ethers to accumulate in certain tumors prompted the synthesis and evaluation of a radioiodinated phospholipid ether analog as a potential tumor imaging agent. Tissue distribution studies with [125I]-rac-1-0-[12-(m-iodophenyl)dodecyl-2-0-methylglycero-3- phosphocholine in rats bearing the Walker 256 carcinosarcoma showed the tumor to contain the highest concentration of radioactivity at 24 hr (15% of the dose) and a tumor-to-blood ratio of 13. Scintigraphic images taken at 24 hr compared favorably with those obtained with (67Ga)-citrate. In contrast with the latter, however, the phospholipid ether showed little propensity to accumulate in an inflammatory lesion in the rat. Tumor visualization was also accomplished in a rabbit bearing the V x 2 adenocarcinoma. We conclude that phospholipid ethers may represent a new class of carrier molecules for the transport of radionuclides to tumors.

Measurement of vascular volume in experimental rat tumors by 19F magnetic resonance imaging.

RATIONALE AND OBJECTIVES: In-vivo assessment of tumor vascular volume may provide useful information for treatment planning or the assessment of treatment effectiveness. The goals of our study were to measure percent vascular volume in two experimental tumor sublines using 19F magnetic resonance imaging (MRI) and to assess changes in tumor blood volume with growth. METHODS: An emulsion of perfluorotributylamine (FTBA) was used as a vascular contrast agent for 19F MRI: The amount of emulsion in the tumor vasculature was measured by 19F MRI and used to calculate percent vascular volume. A separate ex-vivo study of vascular volume was conducted using the dye Hoechst 33342. A total of five rats were studied by MRI and 14 by the ex-vivo method. RESULTS: The ranges of percent vascular volume values measured in the imaging and Hoechst dye studies were 2% to 9% and 1.25 to 7%, respectively. A trend toward decreasing percent vascular volume with increasing tumor volume was evident in one tumor subline. CONCLUSIONS: The quantitative 19F MRI technique was effective for measuring percent vascular volume and changes in vascular volume with growth.

Fluorinated blood substitute retention in the rat measured by fluorine-19 magnetic resonance imaging.

RATIONALE AND OBJECTIVES: Emulsions of perfluorocarbons (PFCs) have been tested as blood substitutes. However, evidence exists that there is long-term retention of some PFCs by the organs of the reticuloendothelial system (RES). The authors investigate organ retention of the blood substitute component, perfluorotripropylamine (FTPA), using fluorine-19 (19F) magnetic resonance imaging (MRI). METHODS: Various dosages of an emulsion of FTPA were administered to five rats. At intervals up to 86 weeks after infusion, 19F MRI was used to measure the amount of FTPA in liver and spleen. The data were fit to both linear and exponential elimination models, and organ retention half-lives were calculated. RESULTS: The exponential half-lives for combined liver and spleen FTPA ranged from 110 to 190 days. Linear half-lives ranged from 175 to 300 days. CONCLUSIONS: FTPA retained by the liver and spleen may be quantified by 19F MRI: The half-lives that were measured are longer than those reported previously for FTPA.

Somatosensory evoked potentials as a measure of experimental cerebral ischemia.

Somatosensory evoked potentials (SEP's) reflect the integrity of the central neuronal pathway, and as such may be used to assess function that remains during a variety of cerebral insults. To evaluate the natural history and utility of SEP's during experimental cerebral ischemia and infarction, SEP's were measured in 17 adult cats at 24 hours and 1 hour prior to right middle cerebral artery (MCA) occlusion, and again immediately afterward and at either 6 hours (five cats) or 24 hours (six cats) post-occlusion. Before occlusion of the right MCA, the SEP's were identical in the right and left hemispheres. After occlusion, there was a significant slowing of the interpeak latency of the first positive peak (P1) in the right hemisphere (3.53 +/- 0.6 msec before compared to 3.99 +/- 0.6 msec after occlusion, p less than 0.001). Maximal slowing in right hemisphere P1 latency was seen in those animals in which the stroke extended into the thalamus (4.38 +/- 0.1 msec). This was significantly slower than left hemisphere values (3.92 +/- 0.32 msec, p less than 0.01). The ipsilateral cortical components of the SEP's, the second positive peak (P2), and the major negative deflection (MN) were slowed in all cats immediately after right MCA occlusion compared to preocclusion measurements (p less than 0.001). Severe infarcts in the mid-suprasylvian and posterior ectosylvian gyri (including the somatosensory cortex) resulted in a greater slowing of the latency of MN compared to less severe infarcts in that region (20.6 +/- 3.9 msec versus 16.4 +/- 1.1 msec, p less than 0.05). There was a precipitous decrease in the amplitude or voltage of the ipsilateral P2-MN complex immediately after occlusion (5.32 +/- 0.4 microV before compared to 0.98 +/- 0.3 microV after occlusion, p less than 0.001). Therefore, the central latencies and cortical amplitudes of the SEP's are sensitive experimental tools as indicators of the onset and extent of a cerebral ischemic insult.

Indomethacin-mediated improvement following middle cerebral artery occlusion in cats. Effects of anesthesia.

Focal cerebral ischemia initiates multiple detrimental effects in the brain. Chief among these are the regional development of ischemic edema, decreased local perfusion, altered neuronal function, and eventual infarction. To determine if pretreatment with the cyclo-oxygenase inhibitor, indomethacin, would result in improvement in these parameters, adult cats were given indomethacin or control solvent (4 mg/kg intraperitoneally twice daily) and were studied for periods up to 24 hours after right middle cerebral artery occlusion. The interaction of anesthetic agents with indomethacin was also examined in separate groups of experimental animals using pentobarbital and ketamine. In cats allowed to recover from pentobarbital anesthesia, indomethacin reduced gray and white matter edema at 6 and 24 hours after occlusion (p less than 0.05). This was noted in densely areas (indomethacin = 84.3%, control = 87.5%), "penumbra" regions (indomethacin = 82.5%, control = 85.3%), and in nonischemic zones (indomethacin = 81.5%, control = 82.3%) at 24 hours. Somatosensory evoked potential amplitude and central latency were also improved in the indomethacin group (p less than 0.05), as was cerebral perfusion (p less than 0.05). In animals anesthetized with continuous ketamine administration, cerebral edema and perfusion as well as evoked potentials were not significantly improved in any region by indomethacin. Regional cerebral blood flow in the group was increased by indomethacin in the nonischemic opposite hemisphere (indomethacin = 64.7 cc/100 gm/min, control = 48.5 cc/100 gm/min, p less than 0.05), but not in the penumbra region of the ischemic hemisphere (indomethacin = 15.0 cc/100 gm/min, control = 18.6 cc/100 gm/min, p less than 0.05), when measured 4 hours after occlusion. This suggested a steal phenomenon. Beneficial effects of indomethacin were evident in the presence of pentobarbital, but not after ketamine anesthesia. This suggests a synergism dependent on decreased arachidonic acid production from pentobarbital-stabilized membranes coupled with diminished production of cyclic endoperoxides from available arachidonate due to inhibition of cyclo-oxygenase with indomethacin.

Effects of verapamil and diltiazem on acute stroke in cats.

To test the effect of verapamil and diltiazem in acute stroke, three groups of mongrel cats of either sex underwent occlusion of the middle cerebral artery (MCA) via a transorbital approach under ketamine anesthesia. The first group served as controls, the second received an intravenous infusion of verapamil (0.1 microgram/kg/min), and the third received an intravenous infusion of diltiazem (0.1 to 1.0 microgram/kg/min). All drug infusions began 2 hours before MCA occlusion and continued for the remainder of the experiment. Before and for up to 24 hours after MCA occlusion, regional cerebral blood flow (rCBF), somatosensory evoked potentials (SSEP's), arterial blood gases, blood pressure, temperature, and hematocrit were measured at least every 2 hours. At the experiment's end, brains were perfused with India ink, removed, sliced, photographed for determination of nonperfused brain area, and weighed, dried, and reweighed for H2O content determination. In these studies, verapamil was associated with worsening of rCBF in ischemic regions and inappropriate increases in rCBF in nonischemic regions, indicating intracerebral steal. Diltiazem increased rCBF in marginally ischemic regions. Changes in SSEP's paralleled blood flow changes, with verapamil decreasing amplitude and conduction velocity while diltiazem slightly improved conduction in the ischemic brain. Verapamil increased the area of nonperfused brain and the content of cerebral H2O. Diltiazem-treated animals had decreased cerebral H2O content, but had a marked increase in the area of nonperfused brain, a finding associated with the high incidence of transtentorial herniation in the diltiazem-treated animals. These findings agree with in vitro studies demonstrating high sensitivity of cerebral blood vessels to calcium channel blockers. These studies further support the notion that calcium channel blockers probably affect several different classes of calcium channels, at different brain sites.

Demonstration of differences in vascular permeability in experimental tumors by use of 19F magnetic resonance imaging.

RATIONALE AND OBJECTIVES: In vivo assessment of tumor vascular permeability may provide useful information for chemotherapy treatment planning or for the assessment of treatment effectiveness. We aimed to assess vascular permeability in two tumor sublines as well as changes in vascular permeability with tumor growth by using 19F magnetic resonance imaging. METHODS: An emulsion of perfluorotributylamine was used as a tumor extravascular contrast agent for 19F MRI. The amount of emulsion that leaked into tumor interstitial space was analyzed qualitatively with imaging. A quantitative study of vascular permeability was done with a separate group of tumors by use of Evans blue dye. RESULTS: One tumor type was more permeable to both perfluorotributylamine emulsion and Evans blue than was the second tumor type. The difference was attributed to a difference in surface area for exchange. In larger tumors of both types, pooling of large amounts of perfluorocarbon occurred and was assumed to be attributable to hemorrhage or blood flow stasis or both. CONCLUSION: 19F MRI is capable of demonstrating the permeability of tumor vessels to macromolecular substances.