The oncofetal protein, 5T4, is a suitable target for antibody-guided anti-cancer chemotherapy with calicheamicin.

The oncofetal protein, 5T4, is a tumor-associated protein displayed on the cell membrane of various carcinomas. This molecule is a promising target for anti-tumor vaccine development and for targeted therapy with staphylococcus exotoxin. The potential use of 5T4 as a target for antibody-guided chemotherapy has not been demonstrated. We report oncolytic efficacy and selectivity in vitro and in vivo with immuno-conjugates of calicheamicin (CM) and the anti-5T4 antibody, H8. CM is a potent cytotoxic drug that causes double strand breaks in DNA. Conjugates of CM and H8 were constructed with acid-labile as well as acid-stabile linkers. In vitro, when applied to monolayers of 5T4(+) cells, CM-conjugates targeting 5T4 were consistently more toxic than either free drug or a non-binding control CM-conjugate. This difference was less pronounced on 5T4-deficient cells. In vivo, four 5T4-positive subcutaneous tumor models were treated with conjugates. Efficacy was demonstrated by reduction of tumor growth relative to controls treated with drug vehicle. To evidence selectivity, the efficacy of the anti-5T4 conjugates was compared to the efficacy of H8, a mixture of H8 and calicheamicin, calicheamicin alone or calicheamicin conjugated to the anti-CD33 antibody, hP67.6. In addition, the efficacy and selectivity of an acid-labile conjugate of H8 was evaluated in an orthotopic model for 5T4(+) lung cancer. Increased survival following treatment was used as a parameter of efficacy. Calicheamicin conjugates of H8 were effective and selective in all the examined tumor models. Differences in efficacy between the acid-labile and acid-stabile conjugates depended on the investigated tumor model.

Evaluation of cDNA libraries from different developmental stages of Schistosoma mansoni for production of expressed sequence tags (ESTs).

A comparative study of the gene expression profile in different developmental stages of Schistosoma mansoni has been initiated based on the expressed sequence tag (EST) approach. A total of 1401 ESTs were generated from seven different cDNA libraries constructed from four distinct stages of the parasite life cycle. The libraries were first evaluated for their quality for a large-scale cDNA sequencing program. Most of them were shown to have less than 20% useless clones and more than 50% new genes. The redundancy of each library was also analyzed, showing that one adult worm cDNA library was composed of a small number of highly frequent genes. When comparing ESTs from distinct libraries, we could detect that most genes were present only in a single library, but others were expressed in more than one developmental stage and may represent housekeeping genes in the parasite. When considering only once the genes present in more than one library, a total of 466 unique genes were obtained, corresponding to 427 new S. mansoni genes. From the total of unique genes, 20.2% were identified based on homology with genes from other organisms, 8.3% matched S. mansoni characterized genes and 71.5% represent unknown genes.

Minilibraries constructed from cDNA generated by arbitrarily primed RT-PCR: an alternative to normalized libraries for the generation of ESTs from nanogram quantities of mRNA.

The generation of expressed sequenced tags (ESTs) depends on the arbitrary selection of individual cDNA clones from libraries. The efficiency of this process reflects the clonal structure of the library used and can be significantly increased using size selected, directional, normalized cDNA libraries. This strategy, however, is not readily applicable when mRNA is limiting, as is the case in the study of complex microorganisms such as parasites, fetal tissues or tumor biopsies. We show here that the construction and systematic sequencing of minilibraries of cDNAs produced by arbitrarily primed PCR provides an alternative means of efficiently generating ESTs in situations where only nanogram quantities of RNA are available. This methodology greatly compensates for unequal message abundance, avoids the need for complex library construction, is equally applicable to the analysis of abundant or rare biological material and is ideally suited to multicenter programmes.

Crossed cerebellar and uncrossed basal ganglia and thalamic diaschisis in Alzheimer's disease.

We detected crossed cerebellar as well as uncrossed basal ganglia and thalamic diaschisis in Alzheimer's disease by positron emission tomography (PET) using 18F-fluorodeoxyglucose. We studied a series of 26 consecutive, clinically diagnosed Alzheimer cases, including 6 proven by later autopsy, and compared them with 9 age-matched controls. We calculated asymmetry indices (AIs) of cerebral metabolic rate for matched left-right regions of interest (ROIs) and determined the extent of diaschisis by correlative analyses. For the Alzheimer group, we found cerebellar AIs correlated negatively, and thalamic AIs positively, with those of the cerebral hemisphere and frontal, temporal, parietal, and angular cortices, while basal ganglia AIs correlated positively with frontal cortical AIs. The only significant correlation of AIs for normal subjects was between the thalamus and cerebral hemisphere. These data indicate that PET is a sensitive technique for detecting diaschisis.

Positron emission tomography and histopathology in Pick's disease.

PET using 18F-2-fluoro-2-deoxyglucose was carried out on a case of Pick's disease established by necropsy. A sharply decreased cortical metabolic rate for glucose was obtained in specific gyri, especially in the frontal lobes, where there was extensive gliosis and neuronal loss. More moderate decreases were found in areas with numerous Pick bodies and inflated neurons but less gliosis. The PET pattern was sufficiently distinctive to suggest that it might be possible to distinguish Pick's from Alzheimer's disease premortem.

Comparison of PET, MRI, and CT with pathology in a proven case of Alzheimer's disease.

Positron emission tomography (PET) with fluorodeoxyglucose (FDG), magnetic resonance imaging (MRI), and CT were carried out in a patient with Alzheimer's disease 16 months before he died. At autopsy, the gross appearance of the brain correlated with MRI and CT, which showed some regional atrophy. These were much less revealing than PET, which correlated with microscopic findings of neuronal loss and proliferation of glia. In areas of moderately impaired local cerebral metabolic rate of glucose, as revealed by reduced FDG uptake, there was some gliosis, primarily around the numerous senile plaques. In areas of severe metabolic impairment, there was a profound loss of neurons, extensive gliosis, and a diminished appearance of plaques. PET-FDG is a better measure of the severity of Alzheimer's disease than MRI or CT, because it reflects the degree of neuronal pathology.

Positron emission tomography in the early diagnosis of Huntington's disease.

We studied 10 patients with early Huntington's disease and 7 normal age-matched controls with positron emission tomography (PET) using fluorodeoxyglucose. Subjects had little or no caudate nucleus atrophy and had not received any medications. The results demonstrated that hypometabolism of glucose preceded tissue loss. Furthermore, patients with minimal neurologic or psychiatric symptoms and no obvious CT changes may be differentiated from normal persons with high accuracy by PET. PET is helpful in the early diagnosis of Huntington's disease irrespective of the mode of presentation. PET may also be useful for preclinical detection and may supplement information from DNA studies.

PET studies of cerebral glucose metabolism in idiopathic torticollis.

Regional cerebral glucose metabolism was studied in 16 patients with idiopathic torticollis, using positron emission tomography. Analysis of subcortical regions revealed no consistent focal abnormality of cerebral metabolic rate for glucose, but there was a bilateral breakdown of the normal relationships between the thalamus and basal ganglia. The findings suggest disruption of the pallidothalamic projections in this focal dystonia and may imply a disturbance of GABA.

Regional cerebral glucose metabolism in identical twins.

The present study reports on the degree of similarity in regional cerebral glucose metabolism in seven sets of female identical twins. All scans were done with subjects at rest. Glucose metabolism was measured with positron emission tomography (PET). Significant intraclass correlations were found for the orbital and prefrontal cortex as well as the basal ganglia (caudate/putamen). It is suggested that these correlations reflect the functional aspects of these areas, namely attention and posturing movements.

Multiple line source array for SPECT transmission scans: simulation, phantom and patient studies.

UNLABELLED: Accurate attenuation and scatter corrections in quantitative SPECT studies require attenuation maps of the density distribution in the scanned object. These can be obtained from simultaneous emission/transmission scans. METHODS: A new method has been developed using a multiple line source array (MLA) for transmission scans, and its performance has been investigated using computer simulations and experimental data. The activity in the central lines of the MLA was higher than at the edges of the system, so that more transmission photons would be directed toward the thicker parts of the human body. A series of transmission-only and simultaneous emission/transmission studies were performed for different phantom configurations and human subjects. Attenuation maps were generated and used in reconstruction of attenuation-corrected emission images. RESULTS: The mu coefficients for attenuation maps obtained using the MLA system and simulated and experimental data display no artifacts and are qualitatively and quantitatively correct. For phantoms, the agreement between the measured and the true value of mu for water was found to be better than 4%. The attenuation-corrected emission images for the phantom studies demonstrate that the activity in the heart can be accurately reconstructed. A significant qualitative improvement was also obtained when the attenuation correction was used on patient data. CONCLUSION: Our results indicate that the MLA transmission source can be used in simultaneous transmission/emission imaging to generate accurate attenuation maps. These maps allow for performing an object-specific, attenuation correction of the emission images.

Characterization of dose distribution in radiation therapy plans.

As a method of considering only significant radiation doses to different tissues, the ICRU Report 50 recommends taking the dose given to a significant tissue volume (minimum diameter greater then 15 mm) instead of choosing a single, potentially insignificant, voxel value. In order to find this significant volume, we have adapted an emission imaging analysis method to radiation therapy planning. The resulting method finds and characterizes the dose distribution in the volumes of interest in a way that includes spatial arrangement. The data can be used to signal significant hot or cold volumes in the dose plan and to score the plans based on significant dose to the tissues.

The Theory of Three-Dimensional Image Reconstruction for PET.

A review is made of selected recent publications on three-dimensional image reconstruction for PET. A new algorithm is proposed which is designed to fully utilize all emitted gamma rays which can be detected in a truncated spherical detector. By such full utilization of emitted rays the new algorithm should produce images of better statistical accuracy than could be produced by previously known algorithms.

Use of transputers in a 3-D positron emission tomograph.

The use of a VME-bus-based transputer network as a parallel processing engine for positron volume imaging (PVI) is discussed. The authors find that the speedups of parallel networks depend on two major factors, the ratio of computation to communication for a task and the size of the task, and give a simple model to explore the limits on speedups. Through actual implementation it is shown that real-time PVI data acquisition can be achieved with about 20 transputer nodes, and it is estimated that three-dimensional (3-D) image reconstruction can be achieved within 10 min using 200 nodes. Larger images and a larger number of histograms can readily be accommodated using the same parallel algorithms, as the model presented places no limits on the size of the images. The versatility and scalability of transputers makes them very suitable for use in PVI tomographs in that the same transputers can be used for speeding up data acquisition, image reconstruction, and display.

Full data utilization in PVI using the 3D radon transform.

An algorithm is described for three-dimensional (3D) image reconstruction in positron volume imaging (PVI) using the inversion of the 3D radon transform (RT) for a truncated cylindrical detector geometry. A discrete version of the 3D RT is formed in a 3D histogram from the event-line coordinates of the detected events. The histogram entries represent the plane integrals of the activity in the field of view. Conventional inversion of the x-ray transform (XT) excludes oblique events in non-iterative reconstruction methods. by employing a spatially varying angular acceptance of events into the histogrammed plane integrals of the 3D RT, it is possible to include most of the detected oblique events in the reconstruction of the image using the standard 3D RT inversion formula. the oblique events are added to the histogram with a partial weight compared to those in complete (XT) projections. This single-pass reconstruction image has better statistical noise properties than images formed by RT inversion from complete XT projections, but only for some detector geometries is it significantly better. Monte Carlo simulations were used to study the statistical noise in images reconstructed using the new algorithm. The inherent difference in the axial versus the transaxial statistical noise in images reconstructed from truncated detectors is noted and is found to increase by including oblique events with this new algorithm.

Performance of the dynamic single photon emission computed tomography (dSPECT) method for decreasing or increasing activity changes.

Radionuclide imaging is now widely used whenever functional information is required. We present a new approach to dynamic SPECT imaging (dSPECT method) that uses a single slow rotation of a conventional camera and allows us to reconstruct a series of 3D images corresponding to the radiotracer distribution in the body at various times. Using simulations of various camera configurations and acquisition protocols, we have shown that this method is able to reconstruct washout half-lives with an accuracy greater than 90% when used with triple-head SPECT cameras. Accuracy decreases when using fewer camera heads, but dual-head geometries still give an accuracy greater than 80% for short and 90% for long half-lives and about 50-75% for single-head systems. Dynamic phantom experiments have yielded similar results. Presence of attenuation and background activity does not affect the accuracy of the dSPECT reconstructions. In all situations investigated satisfactory dynamic images were produced. A preliminary normal volunteer study measuring renal function was performed. The reconstructed dynamic images may be presented as a three-dimensional movie showing movement of the tracer through the kidneys and the measurement of the regional renal function can be performed. The time-activity curves determined from this dSPECT data are very similar to those obtained from dynamic planar scans.

Design of an efficient position sensitive gamma ray detector for nuclear medicine.

The shortcomings of conventional scintillation cameras are analysed theoretically with a view towards improving performance at gamma ray energies above 140 keV. A camera design is proposed which incorporates several new features to obtain good spatial resolution from thicker crystals of sodium iodide. Computer simulations show that in addition to having good efficiency and spatial resolution, the new design allows parallax error correction and (possibly) Compton scattering correction at gamma energies up to 511 keV.

Cortical glutaminase, beta-glucuronidase and glucose utilization in Alzheimer's disease.

Large pyramidal neurons of rat and human neocortex stain immunohistochemically for phosphate-activated glutaminase (PAG). In a limited number of postmortem brains, we find large reductions in cortical PAG activity in Alzheimer's disease (AD). This finding is consistent with histological evidence that pyramidal neurons are affected in AD. The reductions are greater than those found in the same samples in choline acetyltransferase (ChAT) but the possible deleterious effects of coma and similar premortem factors on human PAG activity have yet to be assessed. The activity of beta-glucuronidase, a lysosomal enzyme which occurs in reactive astrocytes, is elevated in the same samples. Positron emission tomography (PET) studies, using 18F-fluorodeoxyglucose (FDG), have demonstrated significant deficiencies in glucose metabolism in the cortex in AD, with the parietal, temporal and some frontal areas being particularly affected. We found in serial scans of 13 AD cases, including one relatively young (44-46 year old) familial case, an exacerbation of the defect over time in most cases. We have found a negative correlation between the regional metabolic rates for glucose (LCMR(s] measured premortem and the beta-glucuronidase activities measured postmortem on a few AD cases that have come to autopsy. The correlations between LCMR(s) and PAG and ChAT activities tend to be positive. The results are consistent with previous suggestions that decreased LCMR(s) in AD reflect local neuronal loss and gliosis.

Cortical metabolism, acetylcholinesterase staining and pathological changes in Alzheimer's disease.

The local cerebral metabolic rate for glucose (LCMRgl) was determined by positron emission tomography (PET) using the 18F-fluorodeoxyglucose method in a series of Alzheimer patients and normal controls. The LCMRgl declined in the cerebral cortex with age, but the decrement was significantly greater in the clinically diagnosed Alzheimer's cases. Comparison of PET and psychological data indicated that, as the disease progressed clinically, the reduction in cortical LCMRgl and the number of cortical regions involved also increased. Variable regions of cortex were involved in the early stages but the temporal, parietal and frontal regions were most typically affected. One case coming to autopsy showed that the severity of the LCMRgl decline paralleled loss of neurons in the cortex and their replacement with astroglia. A case of Pick's disease coming to autopsy had shown a different and highly characteristic pattern of cortical metabolic defect. In this case also a poor metabolic rate was associated with extensive gliosis. Acetylcholinesterase (AChE) staining of the cerebral cortex in elderly normals and Alzheimer's disease cases with a new, highly sensitive method showed that in Alzheimer's disease there was an extensive loss of AChE-positive fibers with senile plaques frequently incorporating AChE-positive fiber debris. AChE staining of the substantia innominata area, where the cells giving rise to these neocortical fibers are presumably located, also showed evidence of degenerating cells and fibers.

18Fluorodeoxyglucose positron emission tomography studies in presumed Alzheimer cases, including 13 serial scans.

Positron emission tomographic (PET) data on local cerebral metabolic rates for glucose (LCMR) are reported for 32 regions of interest (ROI)s in cross-sectional studies on 57 patients with clinically diagnosed Alzheimer's disease (AD) and 20 neurologically normal controls, and in serial studies on 13 of the AD cases, including a familial, young-onset case where the diagnosis has been confirmed at autopsy. Extensive psychological testing was done on all the AD cases. Almost all cortical regions showed a significant decline in LCMR with age in the control subjects. There were the expected cortical metabolic deficits in AD and the serial studies showed a general increase in such deficits over time in 12 of the 13 cases. The regions showing the greatest declines with time in serial studies are the same as those showing the most severe deficiencies in cross-sectional studies. The young-onset case did not show a greater rate of metabolic decline than many of the older cases studied. Results on individual psychological tests tended to correlate with metabolic rates in multiple, rather than single, cortical regions, suggesting intact neuronal networks are required for good performance. The correlations with cortical metabolic activity found were of a sign indicating that the higher the metabolic rates and the better the left:right asymmetry index, the better was the performance.

Cerebral glucose metabolism in Parkinson's disease.

Local cerebral glucose utilization was measured in patients with predominantly unilateral Parkinson's disease using 18F-2-fluoro-deoxyglucose and positron emission tomography. Preliminary results indicate the presence of asymmetric metabolic rates in the inferior basal ganglia. The structure comprising the largest portion of basal ganglia at this level is globus pallidus. These findings are consistent with metabolic studies on animals with unilateral nigrostriatal lesions in which pallidal hypermetabolism on the lesioned side has been demonstrated. Increased pallidal activity is likely secondary to a loss of inhibitory dopaminergic input to the striatum from substantia nigra.