Frameless stereotactic radiosurgery for the treatment of primary intracranial tumours in dogs.

Stereotactic radiosurgery (SRS) is a procedure that delivers a single large radiation dose to a well-defined target. Here, we describe a frameless SRS technique suitable for intracranial targets in canines. Medical records of dogs diagnosed with a primary intracranial tumour by imaging or histopathology that underwent SRS were retrospectively reviewed. Frameless SRS was used successfully to treat tumours in 51 dogs with a variety of head sizes and shapes. Tumours diagnosed included 38 meningiomas, 4 pituitary tumours, 4 trigeminal nerve tumours, 3 gliomas, 1 histiocytic sarcoma and 1 choroid plexus tumour. Median survival time was 399 days for all tumours and for dogs with meningiomas; cause-specific survival was 493 days for both cohorts. Acute grade III central nervous system toxicity (altered mentation) occurred in two dogs. Frameless SRS resulted in survival times comparable to conventional radiation therapy, but with fewer acute adverse effects and only a single anaesthetic episode required for therapy.

Episodic cerebellar dysfunction in a bichon frise: a canine case of episodic ataxia?

A four-year-old neutered male bichon frise was presented for the evaluation of chronic, progressive yet episodic neurological dysfunction that was predominantly cerebellar in nature. Diagnostic testing including haematology, serum chemistry, magnetic resonance imaging, cerebrospinal fluid analysis and urine organic acid screening was normal. Trial therapies with phenobarbital, prednisone and acetazolamide were unsuccessful. Treatment with 4-aminopyridine led to complete resolution of the signs.

Radiosurgery using a stereotactic headframe system for irradiation of brain tumors in dogs.

Radiation therapy of brain tumors in dogs typically involves administration of multiple fractions over several weeks. Fractionation is used to minimize damage to normal tissue. Radiosurgery uses multiple non-coplanar stereotactically focused beams of radiation in a series of arcs to deliver a single dose to the target with extreme accuracy. The large number of beams facilitates a high degree of conformation between the treatment area and the target tumor and allows for a steep dose gradient; the use of nonintersecting arcs minimizes exposure of normal tissue. Computed tomography with a stereotactic localizer secured to the skull allows generation of a 3-dimensional image of the target and provides accurate spatial coordinates for computerized treatment planning and delivery. Three dogs were treated with radiosurgery, using 1,000 to 1,500 cGy. A linear accelerator mounted on a rotating gantry was used to generate and deliver the radiation. Two dogs with meningiomas survived 227 and 56 weeks after radiosurgery. A dog with an oligodendroglioma survived 66 weeks. No complications were observed following the use of this technique.

Design of MKC-442 (emivirine) analogues with improved activity against drug-resistant HIV mutants.

Two analogues of the nonnucleoside inhibitor of HIV-1 RT, MKC-442 (emivirine), containing different C6 substituents have been designed to be less susceptible to the commonly found drug-resistance mutation of Tyr181Cys. Compound TNK-6123 had a C6 thiocyclohexyl group designed to have more flexibility in adapting to the mutated drug-binding site. GCA-186 had additional 3',5'-dimethyl substituents aimed at forming close contacts with the conserved residue Trp229. Both compounds showed approximately 30-fold greater inhibitory effect than MKC-442 to the Tyr181Cys mutant virus as well as to the clinically important Lys103Asn virus. X-ray crystallographic structure determination of complexes with HIV-1 RT confirmed the predicted binding modes. These strategies might be used to improve the resilience of other NNRTI series against common drug-resistance mutations.

Crystallographic analysis of the binding modes of thiazoloisoindolinone non-nucleoside inhibitors to HIV-1 reverse transcriptase and comparison with modeling studies.

We have determined the crystal structures of thiazoloisoindolinone non-nucleoside inhibitors in complex with HIV-1 reverse transcriptase to high-resolution limits of 2.7 A (BM +21.1326) and 2. 52 A (BM +50.0934). We find that the binding modes of this series of inhibitors closely resemble that of "two-ring" non-nucleoside reverse transcriptase inhibitors. The structures allow rationalization of stereochemical requirements, structure-activity data, and drug resistance data. Comparisons with our previous structures suggest modifications to the inhibitors that might improve resilience to drug-resistant mutant forms of reverse transcriptase. Comparison with earlier modeling studies reveals that the predicted overlap of thiazoloisoindolinones with TIBO was largely correct, while that with nevirapine was significantly different.

Crystal structures of HIV-1 reverse transcriptase in complex with carboxanilide derivatives.

The carboxanilides are nonnucleoside inhibitors (NNIs) of HIV-1 reverse transcriptase (RT), of potential clinical importance. The compounds differ in potency and in their retention of potency in the face of drug resistance mutations. Whereas UC-84, the prototype compound, only weakly inhibits many RTs bearing single point resistance mutations, inhibition by UC-781 is little affected. It has been proposed that UC-38 and UC-781 may form quaternary complexes with RT at a site other than the known binding pocket of other NNIs. X-ray crystal structures of four HIV-1 RT-carboxanilide complexes (UC-10, UC-38, UC-84, and UC-781) reported here reveal that all four inhibitors bind in the usual NNI site, forming binary 1:1 complexes with RT in the absence of substrates with the amide/thioamide bond in cis conformations. For all four complexes the anilide rings of the inhibitors overlap aromatic rings of many other NNIs bound to RT. In contrast, the second rings of UC-10, UC-84, and UC-781 do not bind in equivalent positions to those of other "two-ring" NNIs such as alpha-APA or HEPT derivatives. The binding modes most closely resemble that of the structurally dissimilar NNI, Cl-TIBO, with a common hydrogen bond between each carboxanilide NH- group and the main-chain carbonyl oxygen of Lys101. The binding modes differ slightly between the UC-10/UC-781 and UC-38/UC-84 pairs of compounds, apparently related to the shorter isopropylmethanoyl substituents of the anilide rings of UC-38/UC-84, which draws these rings closer to residues Tyr181 and Tyr188. This in turn explains the differences in the effect of mutated residues on the binding of these compounds.

3'-Azido-3'-deoxythymidine drug resistance mutations in HIV-1 reverse transcriptase can induce long range conformational changes.

HIV reverse transcriptase (RT) is one of the main targets for the action of anti-AIDS drugs. Many of these drugs [e.g., 3'-azido-3'-deoxythymidine (AZT) and 2',3'-dideoxyinosine (ddI)] are analogues of the nucleoside substrates used by the HIV RT. One of the main problems in anti-HIV therapy is the selection of a mutant virus with reduced drug sensitivity. Drug resistance in HIV is generated for nucleoside analogue inhibitors by mutations in HIV RT. However, most of these mutations are situated some distance from the polymerase active site, giving rise to questions concerning the mechanism of resistance. To understand the possible structural bases for this, the crystal structures of AZT- and ddI-resistant RTs have been determined. For the ddI-resistant RT with a mutation at residue 74, no significant conformational changes were observed for the p66 subunit. In contrast, for the AZT-resistant RT (RTMC) bearing four mutations, two of these (at 215 and 219) give rise to a conformational change that propagates to the active site aspartate residues. Thus, these drug resistance mutations produce an effect at the RT polymerase site mediated simply by the protein. It is likely that such long-range effects could represent a common mechanism for generating drug resistance in other systems.

Adult-onset motor neuron disease in three cats.

Motor neuron diseases of domestic animals have rarely been described. Three cats with adult-onset, chronic, progressive generalized muscle weakness characterized initially by trembling on exertion and later by extreme difficulty in walking, cervical ventroflexion, dysphagia, and marked muscle atrophy were elevated. Spinal reflexes were evident early but were nondetectable as the disease progressed. Electromyography revealed fibrillation potentials, with nerve conduction velocities within the reference range. Histologic examination of muscle specimens revealed denervation. Marked neuron loss and gliosis were detected in the ventral horns of the spinal cord, with atrophy of ventral nerve rootlets. Less dramatic neuron loss was seen in brain stem motor nuclei. Electron microscopic examination of the ventral horns disclosed hypertrophied astrocytes, with densely arrayed intermediate filaments, swollen axons with large filamentous accumulations, and many macrophages with lipofuscin-like inclusions. Clinical and pathologic findings were consistent with a progressive neurodegenerative disease affecting spinal and some bulbar motor nuclei.

Allosteric inhibitors against HIV-1 reverse transcriptase: design and synthesis of MKC-442 analogues having an omega-functionalized acyclic structure.

Based on X-ray crystallographic analysis of MKC-442/human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) complex, analogues in which the N1-substituent is replaced with omega-functionalized alkyl groups were designed to improve the affinity for the enzyme. Synthesis of these compounds was carried out starting from MKC-442 by a sequence of reactions (N3-protection, removal of N1-ethoxymethyl group, alkylation, and N3-deprotection). The compounds were evaluated for anti-HIV activity. Structure-activity relationships are discussed in terms of the possible interaction with the enzyme.

Unique features in the structure of the complex between HIV-1 reverse transcriptase and the bis(heteroaryl)piperazine (BHAP) U-90152 explain resistance mutations for this nonnucleoside inhibitor.

The viral reverse transcriptase (RT) provides an attractive target in the search for anti-HIV therapies. The nonnucleoside inhibitors (NNIs) are a diverse set of compounds (usually HIV-1 specific) that function by distorting the polymerase active site upon binding in a nearby pocket. Despite being potent and of generally low toxicity, their clinical use has been limited by rapid selection for resistant viral populations. The 2.65-A resolution structure of the complex between HIV-1 RT and the bis(heteroaryl)piperazine (BHAP) NNI, 1-(5-methanesulfonamido-1H-indol-2-yl-carbonyl)-4- [3-(1-methyl-ethylamino) pyridinyl] piperazine (U-90152), reveals the inhibitor conformation and bound water molecules. The bulky U-90152 molecule occupies the same pocket as other NNIs, but the complex is stabilized quite differently, in particular by hydrogen bonding to the main chain of Lys-103 and extensive hydrophobic contacts with Pro-236. These interactions rationalize observed resistance mutations, notably Pro-236-Leu, which occurs characteristically for BHAPs. When bound, part of U-90152 protrudes into the solvent creating a channel between Pro-236 and the polypeptide segments 225-226 and 105-106, giving the first clear evidence of the entry mode for NNIs. The structure allows prediction of binding modes for related inhibitors [(altrylamino)piperidine-BHAPs] and suggests changes to U-90152, such as the addition of a 6 amino group to the pyridine ring, which may make binding more resilient to mutations in the RT. The observation of novel hydrogen bonding to the protein main chain may provide lessons for the improvement of quite different inhibitors.

Head trauma.

No definitive, clinically proven drug therapy currently exists for reversal or amelioration of the injurious cellular events initiated by head trauma. The patient's physiologic status should therefore be monitored and managed as best as possible bearing in mind all the factors that can aggravate intracranial abnormalities. All treatment options have positive and negative aspects that should be considered and balanced on an individual patient basis. A major concern in the management of head-injured patients is the avoidance or treatment of intracranial hypertension. This may be achieved largely through medical means and accurate efforts can only be made if intracranial pressure and the variables that influence it are monitored directly. A definite place exists for surgery in the management of head-injured patients, though such a decision is best guided by objective evaluation of the intracranial vault.

Acute, flaccid quadriplegia in three cats with suspected coral snake envenomation.

Three cats were evaluated for acute, ascending, flaccid quadriplegia; depression; and reduced nociception. Complete or partial neuromuscular junction blockade was found on nerve stimulation studies during electromyographic examinations. Two of the cases had wounds on the chin or paw compatible with coral snake bites. Although a coral snake was found in only one case, coral snake envenomation was suspected because potential for exposure, clinical signs, and electrodiagnostic findings were similar to dogs reported with this condition and to cats with tiger snake envenomation. Only one case received coral snake antivenin. All cases recovered within seven-to-10 days.

Complexes of HIV-1 reverse transcriptase with inhibitors of the HEPT series reveal conformational changes relevant to the design of potent non-nucleoside inhibitors.

Crystal structures of HIV-1 reverse transcriptase (RT) complexed with a range of chemically diverse non-nucleoside inhibitors (NNIs) have shown a single pocket in which the inhibitors bind and details of the inhibitor-protein interactions. To delineate the structural requirements for an effective inhibitor, we have determined the structures of three closely related NNIs which vary widely in their potencies. Crystal structures of HIV-1 RT complexed with two very potent inhibitors, MKC-442 and TNK-651, at 2.55 angstroms resolution complement our previous analysis of the complex with the less effective inhibitor, HEPT. These structures reveal conformational changes which correlate with changes in potency. We suggest that a major determinant of increased potency in the analogues of HEPT is an improved interaction between residue Tyr181 in the protein and the 6-benzyl ring of the inhibitors which stabilizes the structure of the complex. This arises through a conformational switching of the protein structure triggered by the steric bulk of the 5-substituent of the inhibitor pyrimidine ring.

Spinal meningeal sarcoma in a rottweiler puppy.

An 11-week-old rottweiler puppy was presented for evaluation of progressive paraparesis, urinary and faecal incontinence. Radiography and myelography revealed widening of the vertebral canal and remodelling of several lumbar vertebrae associated with an intramedullary spinal mass. Magnetic resonance imaging revealed an extensive, heterogeneous mass involving the lumbosacral spinal cord. Histopathological examination indicated a meningeal sarcoma with very varied cytological architecture.

Abnormal connectivity of the visual pathways in human albinos demonstrated by susceptibility-sensitized MRI.

We studied activation of the human visual cortex (VC) using susceptibility-sensitized MRI at 1.5 Tesla. Three albinos and six healthy controls underwent a series of monocular and binocular photic flash stimulation. Monocular stimulation in albinos caused predominant contralateral activation with a small, well-delineated area in the anterior part of the VC in the ipsilateral hemisphere. This finding was consistent with a chiasmal crossing anomaly in albinism. All controls had symmetric patterns of activation during monocular stimulation. Functional MRI represents a promising method for evaluation of the visual pathways in humans.

Identification of vascular structures as a major source of signal contrast in high resolution 2D and 3D functional activation imaging of the motor cortex at 1.5T: preliminary results.

We have measured the T2* signal response associated with cortical activation due to finger motion at 1.5 Tesla. Both thin slice 2D and 3D images show signal intensity changes which vary from 2% to 32% depending on volunteer, echo time, slice thickness, and in-plane resolution. The largest signal change occurred for the thinnest slices and highest resolution (2 mm3). This is consistent with reducing partial volume effects and a simple difference in phase between the intravascular signal and surrounding parenchyma. No inflow enhancement was seen on the 2D or 3D scans, confirming the nature of the signal difference for this approach was due to local field inhomogeneity effects. Using 3D imaging, multiple effects can be seen simultaneously. With a 3D MRA method, it was possible to locate the vessel that was the source of the T2* behavior; it was in each case a vein on the surface of the cortical parenchyma.

Evaluation of experimental early acute cerebral ischemia before the development of edema: use of dynamic, contrast-enhanced and diffusion-weighted MR scanning.

The ability of dynamic, contrast-enhanced, magnetic susceptibility-weighted scanning to delineate early experimental acute cerebral infarction was compared with that of heavily T2-weighted and diffusion-weighted spin echo scanning. Spontaneously hypertensive rats, which had undergone right middle cerebral artery occlusion, were studied from 15 min to 3 h post ligation on a 1.5-T clinical whole-body imager. In contrast to the diffusion- and T2-weighted spin echo scans, the dynamic, contrast-enhanced technique clearly and consistently delineated the nonperfused regions as early as 15 min post ligation.

Proton NMR imaging of cerebral blood flow using H2(17)O.

Cerebral blood flow was quantitatively mapped by monitoring the cerebral washout of H2(17)O using rapid, single-shot proton NMR imaging. H2(17)O acts as a freely diffusible contrast agent for proton imaging via its scalar-coupled term, enhancing T2 relaxation. Measured values for CBF ranged from 29 to 106 ml/min/100 g over a range of arterial pCO2 between 23 and 81 Torr.

The stability of proton T2 effects of oxygen-17 water in experimental cerebral ischemia.

The gerbil model of unilateral cerebral ischemia has been used to test the temporal and spatial stability of the MRI T2 effects of oxygen-17 water. Following unilateral carotid ligation, symptomatic animals were given a single large intraperitoneal injection of H2(17)O and the distribution and stability of the brain T2 effects were followed with a spin-echo sequence. In contrast to the ischemic areas, the perfused tissue shows a marked and prolonged loss in intensity with little evidence of diffusion of the T2 effect of 17O into the ischemic tissue.