Evaluation of systemic targeting of RET oncogene-based MTC with tumor-selective peptide-tagged Ad vectors in clinical mouse models.

Significant advantage of targeted antitumoral treatment consists in the possibility to restrict maximum therapeutic efficacy to the malignant cell population by reducing toxicity in healthy tissues. Using different clinical models for aggressive medullary thyroid carcinoma (MTC), we have recently identified peptide ligands that bind highly selective to tumor cells. By linking the most convincing SRESPHP peptide to an adenoviral (Ad) vector expressing the MTC-related oncogene inhibitor RETΔTK, gene transfer was specifically directed to neoplastic tissue after systemic virus administration. We show that peptide-mediated delivery of RETΔTK significantly enhanced apoptosis, resulting in a strong inhibition of orthotopic and xenograft tumor growth. Conversely, tumors treated with controls expanded their initial size without notable cell death. According to the therapeutic effect, strong virus accumulation was found exclusively in thyroid carcinomas. Strikingly, application of native tropism depleted viral vector linked to tumor-selective peptide was accompanied by a substantial reduction of Ad binding to the liver. Of note, single systemic injection of a low dose (10e8 pfu/mouse) of MTC-specific Ad.RETΔTK induced regression of multiple tumors at different sites in all treated animals. In sum, our results open up the possibility for an efficient cancer cell-specific therapy of primary MTC, their migrating populations and potentially metastases.

Transcriptome analysis in mouse tumors induced by Ret-MEN2/FMTC mutations reveals subtype-specific role in survival and interference with immune surveillance.

Activating mutations in the Ret proto-oncogene are responsible for occurrence of multiple endocrine neoplasia (MEN) type 2A and 2B, and familial medullary thyroid carcinoma (FMTC). A striking genotype-phenotype correlation between the mutated RET codon and clinical manifestation implies that tumorigenesis is conditioned by the type of mutation. We investigated gene expression profiles between and within distinct MEN2 subtypes through whole-genome microarray analysis in tumors induced by NIH-3T3 cells transformed with defined RET-MEN2A (C609Y, C634R), MEN2B, (A883F, M918T), and FMTC (Y791F) mutations. Expression profiling identified a statistically significant modification of 1494 genes, 628 down- and 866 upregulated in MEN2B compared with MEN2A/FMTC tumors. By contrast, no obvious alterations were observed among individual MEN2B and MEN2A type mutations, or between MEN2A and FMTC. Functional clustering of differential genes revealed RET-MEN2B specific upregulation of genes associated with novel growth and survival pathways. Intriguingly, RET-MEN2A/FMTC-specific tumors were characterized by a considerable number of genes involved in the host antitumor immune response via stimulation of natural killer/T-cell proliferation, migration, and cytotoxicity, which were completely absent in RET-MEN2B related cancers. QPCR on tumors versus cultured NIH-RET cell lines demonstrated that they are largely attributed to the host innate immune system, whereas expression of CX3CL1 involved in leukocyte recruitment is exclusively RET-MEN2A/FMTC tumor cell dependent. In correlation, massive inflammatory infiltrates were apparent only in tumors carrying MEN type 2A/FMTC mutations, suggesting that RET-MEN2B receptors specifically counteract immune infiltration by preventing chemokine expression, which may contribute to the different clinical outcome of both subtypes.

[Involvement of CNS in leukaemia and lymphomas--CSF meningeosis and immunocytochemical phenotyping]. / ZNS-Beteiligung bei Leukämien und Lymphomen--Meningeosis im Liquor cerebrospinalis und Möglichkeiten der immunzytochemischen Zellphänotypisierung.

The detection of tumour cells in the cerebrospinal fluid (CSF) of leukaemia and lymphoma patients is a challenge for CSF cytological investigation. Nevertheless, it is generally possible to confirm the involvement of the central nervous system (CNS) by conventional microscopic detection of tumour cells. Immunocytochemical staining techniques are a valuable complement of conventional cytology. Immunocytochemical cell phenotyping is indicated for identification of atypical cells in patients with suspected primary CNS lymphoma or malignant haematological disease.

[The role of blood-brain barrier in the pathogenesis of Alzheimer dementia--implications for immunological therapies for plaque dissolution]. / Die Funktion der Blut-Hirn-Schranke für die Pathogenese der Alzheimer-Demenz--Implikationen für immunologische Therapien zur Plaqueauflösung.

Alzheimer's disease (AD) is a devastating neurodegenerative disorder affecting more than 27 million people worldwide and leading to severe social-economic problems. One characteristic hallmark of AD--the amyloid plaques--are still being discussed to be one important triggering factor. However, current animal and autopsy studies refer to soluble and highly toxic A block oligomers as the deadly agent for the neurons. Current therapies mainly rely on the abatement of symptoms without antagonizing the etiology of the disease. Potential new approaches address reduced production, increased degradation and/or evacuation of toxic A block peptides from the brain. Among others one important group of target-proteins are the ABC transporters of the blood-brain barrier which contribute importantly to the detoxification of the brain. Changes of specific transport functions evoke important alterations for the known pathogenesis and future therapies of AD, especially approaches that target plaque dissolution and plaque reduction.

French maritime pine bark treatment decelerates plaque development and improves spatial memory in Alzheimer's disease mice.

BACKGROUND: Plant extracts are increasingly investigated as potential drugs against Alzheimer's disease (AD) and dementia in general. Pycnogenol is an extract from the bark of the French maritime pine (Pinus pinaster Aiton subsp. atlantica) with known anti-oxidative and neuroprotective effects. HYPOTHESIS/PURPOSE: Pycnogenol is thought to improve cognitive functions in elderly. We wanted to investigate and quantify these effects in a model system of cerebral ß-amyloidosis/AD. STUDY DESIGN/METHODS: This study experimentally assessed the effects of Pycnogenol on AD-related pathology in a ß-amyloidosis mouse model. APP-transgenic mice and controls were treated orally in a pre-onset and post-onset treatment paradigm. The effects of Pycnogenol were characterized by analysing ß-amyloid (Aß) plaques, number of neurons, glia coverage, myelination pattern, and cortical coverage with axons using immunohistochemistry. Aß levels were quantified using ELISA and gene expression levels of APP-processing enzymes ADAM10, BACE1 and IDE protein levels were determined by Western blot. Behavioural changes in circadian rhythm were monitored and spatial memory / cognition was assessed using a water maze test. RESULTS: Pycnogenol significantly decreased the number of plaques in both treatment paradigms but did not alter levels of soluble Aß or the gene expression of APP-processing enzymes. The morphological analyses revealed no changes in the number of neurons, astrocytes, microglia, the myelination pattern, or the morphology of axons. Behavioural testing revealed an improvement of the spatial memory in the pre-onset treatment paradigm only. CONCLUSION: Our results suggest to evaluate clinically a potential use of Pycnogenol in the prevention or in early stages of mild cognitive impairment (MCI) and AD.

Gene structure, expression in Escherichia coli and biochemical properties of the NAD+ -dependent glyceraldehyde-3-phosphate dehydrogenase from Pinus sylvestris chloroplasts.

Photosynthetic eukaryotes typically possess two distinct glyceraldehyde-3-phosphate dehydrogenases, an NAD+ -specific enzyme in the cytosol (GapC: EC 1.2.1.12) and an NADP+ -dependent enzyme in the chloroplast (GapAB: EC 1.2.1.13). The gymnosperm Pinus sylvestris is an exception in that it is known to express a gene encoding a transit peptide-bearing GapC-like subunit that is imported into chloroplasts (GapCp), but the enzymatic properties of this novel GAPDH have not been described from any source. We have expressed the mature GapCp unit from Pinus in Escherichia coli and have characterized the active enzyme. GapCp has a specific activity of 89 units per milligram and is strictly NAD+ -dependent, showing no detectable activity with NADP+. Values of the apparent Km for NAD+ and glyceraldehyde-3-phosphate were determined as 62 and 344 microM, respectively. The Pinus GapCpl gene possesses 12 introns, two in the region encoding the transit peptide and ten in the region encoding the mature subunit, all of which are found at positions strictly conserved across genes for higher plant GapC. A cDNA encoding a homologue of GapCp was isolated from the heterosporous fern Marsilea quadrifolia, indicating that NAD+ -dependent chloroplast GAPDH also occurs in other higher plants.

Activated microglia do not mediate the early deposition of Abeta in carriers of the apolipoprotein Eepsilon4 allele.

Activated microglia are a prominent component of the senile plaques in end-stage Alzheimer's disease, but whether microglia contribute to the initiation of the lesions remains unknown. In a previous postmortem study of non-demented elderly cases, we found that amyloidogenesis is advanced by at least 10 years in carriers of the apoEepsilon4 allele. To determine whether microglia are involved in the initial stages of beta-amyloid pathogenesis and whether apoE genotype influences microglial activation, we quantified HLA-DR-immunoreactive microglia in the medial temporal lobe of 229 non-demented humans of various APOE genotypes who had died between 50 and 91 years of age. Our results show that the number of HLA-DR-immunoreactive microglia increases with advancing age in both the gray matter and the white matter. In contrast to amyloid plaques and neurofibrillary tangles, there is no significant correlation between apoE genotype and density of microglia, although apoEepsilon4 homozygotes tended to have more microglia than did other apoE groups. In sections double-immunostained for Abeta and activated microglia, activated microglia were associated with dense-cored plaques but not with diffuse plaques, suggesting that microglial activation is a relatively late event in the genesis of beta-amyloid. Activation of microglia thus appears not to be the initial impetus for Abeta-deposition in the elderly.

Expression of multidrug transporters in dysembryoplastic neuroepithelial tumors causing intractable epilepsy.

OBJECTIVE: Dysembryoplastic neuroepithelial tumors (DNT) are relatively benign brain lesions that often cause medically intractable epilepsy. There is mounting evidence that multidrug transporters such as P-glycoprotein (P-gp) or multidrug resistance-associated proteins (MRP) play an important role in the development of resistance to antiepileptic drugs (AED). MATERIAL AND METHODS: In the present study, we examined the expression of several multidrug transporters in 14 cases of DNT. The peritumoral brain tissue as well as 9 cases of arteriovenous malformations (AVM) served as controls. P-gp, MRP2, MRP5 and breast cancer resistance protein (BCRP) expression was evaluated qualitatively and quantitatively using immunohistochemistry. RESULTS: All transporters were overexpressed quantitatively in DNT, but each revealed a different labeling pattern. P-gp and BCRP were predominantly located in the endothelium of brain vessels. MRP5 was detected primarily in endothelial cells, but notably also in neurons. The expression of P-gp, MRP2 and MRP5 was low in AVM, whereas BCRP demonstrated strong staining. Examination of MDR1 gene polymorphisms revealed no correlation with P-gp expression whereas the MRP2 exon 10 G1249A polymorphism was associated with different MRP2 labelling. CONCLUSIONS: Our results show that multidrug transporters are overexpressed in DNT. This finding supports the view that several of these transport proteins may play an important role in the mechanisms of drug resistance in epileptic brain tissue.

[Patient-related rejection of nasal IPPV therapy. Patients, reasons, follow-up]. / Patientenbedingte Ablehnung der nasalen IPPV-Therapie. Patienten, Gründe, Verlauf.

PATIENTS: We retrospectively analysed the course of 33 patients who had rejected nasal IPPV-therapy (1988 to February 1996). RESULTS: The death-rate was higher (48%) compared to nasal IPPV patients in the same time (18%). The patients were divided in 3 main diagnostic groups (COPD, restrictive thoracic wall, neuromuscle disease). We observed the highest death-rate in COPD patients (66%) and the lowest death-rate in the group with scoliosis or chest wall disease (23%). This is the same result tendencially as in patients with nasal IPPV (mortality-rate COPD 66%, restrictive chest wall 6%).

[Noninvasive intermittent ventilation. A prospective data collection in patients with hypoventilation syndrome]. / Nichtinvasive intermittierende Beatmung. Eine prospektive Datenerhebung bei Patienten mit Hypoventilationssyndrom.

INTRODUCTION: Noninvasive intermittent ventilation is usually performed in patients with severe ventilatory pump disorder. From 1988 to 3/1995 we treated 163 patients with the aim of home mechanical ventilation (HMV). PATIENTS AND RESULTS: In March 1993 115 of these 163 patients practiced HMV, 22 had already died and 26 had rejected or broken off ventilation therapy. The 115 patients were classified in three main diagnostic groups: Scoliosis or chest wall disease (n = 76), COPD (n = 11) and neuromuscular disease (n = 28). The mean pCO2 at rest of all patients before ventilation therapy was 56 (+/- 12) Torr and fell to 46 (+/- 5) Torr in the course of therapy. The maximum statical inspiration pressure PImax rose from average 3, 8 (+/- 2, 3) to 4, 9 (+/- 2, 0) kPa. There was a probability of surviving two years after onset of ventilation therapy of 85% in the scoliosis group, of 60% in the neuromuscular group and of 30% in the COPD group. CONCLUSIONS: According to results of others home ventilation therapy was very successful in patients with chest wall disease. In some patients with neuromuscular disorder quality of life could be improved and life prolonged. Only half of the COPD patients could be treated successfully, whereas the other half had no benefit from noninvasive ventilation therapy.

[Noninvasive ventilation in acute respiratory insufficiency]. / Nichtinvasive Beatmung bei akuter respiratorischer Insuffizienz.

BACKGROUND AND AIM: Noninvasive positive pressure ventilation (NPPV) via face mask offers in comparison to endotracheal intubation in treating patients with acute respiratory failure (ARF) advantages like allowing swallowing and coughing. We report our experiences and try to verify the indications and the efficacy of NPPV. PATIENTS AND METHODS: In the period of January 1991 until August 1996 109 patients (30 female, 79 male, mean age 61 +/- 12 years) received mechanical ventilation with NPPV representing 25% of all MVs in this term. As baseline capillary blood gases (CBG) were found: pH: 7.30 +/- 0, 10; pCO2: 64 +/- 19; pO2: 60 +/- 19 (all patients received supplemental oxygen). Success of NPPV was determined by an improvement of the baseline CBG. RESULTS: NPPV was successful in 77 (71%) patients. Considering the kind of respiratory insufficiency the patient population was divided into 4 groups 1. acute hypoxemic respiratory failure, 2. acute hypercapnic ventilatory failure, 3. acute decompensation of chronic respiratory insufficiency (CRI) and 4. combined failure. Considering these subgroups we obtained the best results in the group of patients with hypercapnic disturbances. In patients with hypoxemic RF we observed a success of NPPV if the improvement of CBG occurred in the early stage (< or = 12 hours) of NPPV. CONCLUSION: Our data indicate that application of NPPV is an effective and safe alternative to endotracheal intubation in many patients with hypercapnic ventilatory failure. NPPV is also successful in patients with hypoxemic RF with a milder course.

[Long-term breathing via tracheostoma]. / Langzeitheimbeatmung über Tracheostoma.

PATIENTS AND METHODS: From 1988 to 2/1997 we had introduced intermittent positive pressure ventilation (IPPV) in 298 patients. In most cases non-invasive nasal mask ventilation was possible, in 21 patients (7%) a tracheostoma was necessary. These 21 patients were analysed retrospectively due to age, sex, diagnose, ventilation mode, course of illness, home care and costs. RESULTS: We had 13 male and 8 female patients, aged 49 years on average (min. 2, max. 84). 90% had neuromuscular diseases especially muscle dystrophies. Ventilation therapy was performed volume controlled with the cannula unblocked during daytime and blocked at night. Eighteen patients had industrial cannulas (72% Shiley, 28% Rüsch), 3 patients used silver cannulas. Daily ventilation amounted 24 hours in 7 patients, 6 to 14 hours in 14 patients. During the observed time 7 patients remained in stable health situation, in 9 patients the underlying disease was progressive and 5 of them died. IPPV was performed 50.7 months on an average, in living patients 68.8 months, in died 7.6 months. Fifteen patients lived at home, 5 were cared in nursing home, 1 patient stayed in hospital. Outside the hospital the bigger part of costs was paid by sick funds and care funds, the smaller part by social welfare offices. Often costs were divided. Total costs for caring about 24 hours ventilated patient at home amounted up to 21,000 German marks each month.

[Systemic fungal infections in hematologic neoplasms. An autopsy study of 1,053 patients]. / Systemische Pilzinfektionen bei hämatologischen Neoplasien. Eine Autopsiestudie an 1053 Patienten.

BACKGROUND: Mycoses are common complications of haematological neoplasias. For successful antimycotic treatment, a knowledge of preferential underlying disease, frequency, species and site of the mycosis is of importance. PATIENTS AND METHODS: Postmortem material comprising clinical data, autopsy protocols and histological sections obtained between 1976 and 1990 from 1,053 patients with leukaemia and malignant lymphomas following antineoplastic therapy was analysed retrospectively. RESULTS: Autopsy revealed systemic mycoses in 184 patients (17.5%). Between 1976 and 1990, the incidence of fungal infections increased from 12% to 30%, most being found in acute leukaemia (24%). Myeloproliferative syndrome (18%), non-Hodgkin's lymphomas (16%), Hodgkin's disease (10%) and plasmocytoma (2.5%) were less frequently associated with mycoses. With no preference for any particular malignancy in evidence, aspergillosis predominated at histology (85 cases), while candidosis occurred in 75 cases. A combination of two mycoses (aspergillosis and candidosis) (14 patients), zygomycosis (eight patients) and cryptococcosis (two patients) were much less common. While aspergillosis caused mostly pulmonary (81 cases) and cerebral (18 cases) infections, candidosis most frequently affected the GI tract (83 cases). The fungal infection was regarded as the main cause of death in some 76% of the cases. An analysis of bone marrow of patients with mycosis (184 cases) revealed a predominance of hypoplasia (54%) over tumour infiltration (34%) and normal bone marrow (12%). In malignancies with no mycoses (869 cases) in contrast, hypoplasia was significantly less common (19%) than infiltration (59%) or normal bone marrow (22%) (p < 0.001). CONCLUSION: The incidence of mycoses in haematological neoplasias in our post mortem series has continued to increase. Bone marrow hypoplasia in particular predisposes to fungal infection. The lungs are the organs of predilection, and aspergillosis is likely to be the infection presenting.

Spinal cord pathology in chronic experimental Toxoplasma gondii infection.

Infection with the protozoan Toxoplasma (T.) gondii causes chronic infection of the central nervous system and can lead to life-threatening encephalomyelitis in immunocompromised patients. While infection with T. gondii has long time been considered asymptomatic in immunocompetent hosts, this view is challenged by recent reports describing links between seropositivity and behavioral alterations. However, past and current researches are mainly focused on the brain during Toxoplasma encephalitis, neglecting the spinal cord as a key structure conveying brain signals into motion. Therefore, our study aimed to fill the gap and describes the spinal cord pathology in an experimental murine model of toxoplasmosis. In the spinal cord, we found distinct histopathological changes, inflammatory foci and T. gondii cysts similar to the brain. Furthermore, the recruitment of immune cells from the periphery was detected. Moreover, resident microglia as well as recruited monocytes displayed an increased MHC classes I and II expression. Additionally, the expression of pro- and anti-inflammatory cytokines was enhanced in the brain as well as in the spinal cord. In summary, the pathology observed in the spinal cord was similar to the previously described changes in the brain during the infection. This study provides the first detailed description of histopathological and immunological alterations due to experimental T. gondii induced myelitis in mice. Thus, our comparison raises awareness of the importance of the spinal cord in chronic T. gondii infection.

[Gastrointestinal stromal tumor (GIST) of the anterior rectal wall. R0 resection with simultaneous radical retropubic prostatectomy]. / Gastrointestinaler Stromatumor (GIST) der vorderen Rektumwand. R0-reseziert bei simultaner radikaler retropubischer Prostatektomie.

This case report deals with a 79-year-old patient with a gastrointestinal stromal tumor (GIST) of the anterior rectal wall which was unusually located between the rectum and the prostate gland. In addition, this patient suffered from subvesical obstruction accompanied by an elevated PSA level. These circumstances led to our decision to operate on the tumor via simultaneous radical retropubic prostatectomy. In our opinion this resection technique was easier and less traumatic for the patient compared to procedures performed via the abdomen and perineum. This case report demonstrates that in the case of tumors located between the rectum and the prostate gland the differential diagnosis should include not only prostate carcinoma but also rare tumor entities such as GIST.