Monitoring of stable isotope composition of precipitation reveals thunderstorm dynamics.

The summer of 2019 is particularly well known for the famous heatwaves that swept across the European continent, with its associated drought and record-breaking air temperatures. This was followed by powerful thunderstorms, characterised by hail and heavy rain that damaged the crops on a regional scale. Here, we investigated one of the largest storm cells, lasting more than 6 h, which struck southwestern Romania. High-temporal resolution sampling of storm precipitation was performed for stable isotope measurements, rainfall and air temperature, to follow the storm dynamics. Hydrogen and oxygen isotope measurements show an abrupt decreasing temporal trend followed by superimposed V-shaped patterns interpreted as reflecting moisture replenishment by successive rain bands. To model the stable isotope values of precipitation in relation to the general trend of decreasing air temperatures, we applied a numerical Rayleigh condensation model for a non-constant α isotopic fractionation factor between liquid water and water vapour. The storm is powered by four consecutive moisture fronts, each following a Rayleigh distribution. About 40 % of the water vapour condenses during the sampled storm due to adiabatic expansion and cooling, which lowers saturation. Condensation ceases when cooling and absolute humidity can no longer sustain the dew point, stopping the rain. The timing of the event, occurring late at night and early in the morning, its duration of over 6 h as well as its synoptic scale may indicate a mesoscale convective complex.

Ursus spelaeus (Rosenmüller, 1794) during the MIS 3: new evidence from the Cioclovina Uscata Cave and radiocarbon age overview for the Carpathians.

Ursus spelaeus, the Late Pleistocene a cave bear is known from numerous accumulations found in the fossil sector of caves situated in the Carpathian and Apuseni Mountains. In this study, we present new radiocarbon data along a profile of the Cioclovina Uscata Cave, which is situated in the South Carpathians. The data suggest that, during the entire Marine Isotope Stage 3 (MIS 3) interval, the cave was serving as a shelter for U. spelaeus, with the oldest dated bone indicating an age of > 47,710 and the youngest one, an age of 31,820 ± 400 years cal BP. Histogram plots of 110 radiocarbon data from different caves of the Carpathian and Apuseni Mountains as Cioclovina Uscata, Peștera (Cave) cu Oase, Peștera Muierii, or Peștera Urșilor, respectively, show a maximum expansion of the cave bear population between 50,000 and 40,000, a decline between 40,000 and 35,000 and a partial recovery from 35,000-30,000 years cal BP. Radiocarbon data of Homo sapiens remains, younger than 35,000 years cal BP, support the fact that H. sapiens accessed the same caves where the cave bear persisted to hibernate. Besides general cool conditions and restricted food sources, the presence of H. sapiens constituted an additional stress factor driving the cave bear to extinction.

Multi-element stable isotope geochemistry and arsenic speciation of hydrothermal vent fauna (Alviniconcha sp., Ifremeria nautilei and Eochionelasmus ohtai manusensis), Manus Basin, Papua New Guinea.

Deep-sea hydrothermal vent communities, revealing patterns of niche partitioning, live in a limited area characterised by sharp physico-chemical gradients. In this study, we investigated carbon, sulfur, nitrogen stable isotopes as well as arsenic (As) speciations and concentrations for two snails (Alviniconcha sp. and Ifremeria nautilei) and a crustacean, (Eochionelasmus ohtai manusensis), occupying distinct niches in the hydrothermal vent field of the Vienna Woods, Manus Basin, Western Pacific. δ13C values of Alviniconcha sp. (foot), I. nautilei (foot and chitin) and E. o. manusensis (soft tissue) are similar, from -28 to -33‰ (V-PDB). The δ15N values of Alviniconcha sp. (foot and chitin), I. nautilei (foot and chitin) and E. o. manusensis (soft tissue) range from 8.4 to 10.6‰. The δ34S values of Alviniconcha sp. (foot and chitin), I. nautilei (foot) and E. o. manusensis (soft tissue) range from 5.9 to 11.1‰. Using stable isotopes, for the first time, we inferred a Calvin-Benson (RuBisCo) metabolic pathway for Alviniconcha sp. along with the presence of γ-Proteobacteria symbionts for the Vienna Woods communities. For I. nautilei, a feeding pattern is proposed with γ-Proteobacteria symbiosis and a Calvin-Benson-Bassham diet with mixotrophic feeding. E. ohtai manusensis is filtering bacteria with a CBB feeding strategy, with δ15N values indicating possible higher position in the trophic chain. Arsenic concentrations in the dry tissue of Alviniconcha (foot), I. nautilei (foot) and E. o. manusensis (soft tissue) are high, from 4134 to 8478 µg/g, with inorganic As concentrations of 607, 492 and 104 µg/g, respectively and dimethyl arsenic (DMA) concentrations of 11.12, 0.25 and 11.2 µg/g, respectively. Snails occurring in a vent proximal position have higher As concentration than barnacles, a pattern not observed for S concentrations. Arsenosugars were not put in evidence indicating that the available organic material for the vent organisms are not surface derived.

Retrospective study of neoadjuvant versus adjuvant radiochemotherapy in locally advanced noninflammatory breast cancer : survival advantage in cT2 category by neoadjuvant radiochemotherapy.

PURPOSE: This retrospective study compares patients treated between 1991 and 1998 with neoadjuvant radiotherapy +/- chemotherapy (RCT) or adjuvant RCT for locally advanced noninflammatory breast cancers (LABC) in terms of pathologic complete response (pCR), 10-year relapse-free (RFS), and overall survival (OS). PATIENTS AND METHODS: Preoperative RCT in 315 and adjuvant RCT in 329 cases consisted in 50 Gy (5 x 2 Gy/week) to the breast and the supra-/infraclavicular lymph nodes. 101 neoadjuvant patients received - in case of breast conservation - a 10-Gy interstitial boost with (192)Ir afterloading before and 214 neoadjuvant patients a preoperative electron boost after external-beam radiotherapy. In the neoadjuvant RCT group, chemotherapy was applied prior to radiotherapy in 192 patients, and simultaneously in 113; ten had no chemotherapy. In the adjuvant RCT group, chemotherapy was applied to 44 patients before surgery and to 166 after surgery; 119 had no chemotherapy. RESULTS: Breast conservation became possible in 50.8% after neoadjuvant RCT for LABC with a pCR rate at surgery of 29.2%. A complete nodal remission (pN0) after RCT was observed in 56% (89/159) of the cN+ (clinically node-positive) neoadjuvant patients. There were trends in favor of preoperative RCT for RFS and OS (hazard ratio [HR] = 0.85; p = 0.09 for RFS; HR = 0.8130; p = 0.1037 for OS). For patients with cT2 tumors the RFS and OS were statistically significantly better (HR = 0.5090; p = 0.0130 for RFS; HR = 0.4390; p = 0.0026 for OS) after neoadjuvant compared to adjuvant RCT. CONCLUSION: Neoadjuvant RCT achieved a pCR rate of 29.2% and a statistically significantly better RFS and OS in patients with cT2-category breast cancer.

Correction to: The role of passive immunization in the age of SARS-CoV-2: an update.

An amendment to this paper has been published and can be accessed via the original article.

The role of passive immunization in the age of SARS-CoV-2: an update.

The rapid spread of the corona virus pandemic is an existential problem for many people in numerous countries. So far, there is no effective vaccine protection or proven therapy available against the SARS-CoV-2 virus. In this review, we describe the role of passive immunization in times of the corona virus. Passive immunization could be a bridging technology to improve the immune defense of critically ill patients until better approaches with effective medications are available.

Gene expression of circulating tumour cells in breast cancer patients.

BACKGROUND: The diagnostic tools to predict the prognosis in patients suffering from breast cancer (BC) need further improvements. New technological achievements like the gene profiling of circulating tumour cells (CTC) could help identify new prognostic markers in the clinical setting. Furthermore, gene expression patterns of CTC might provide important informations on the mechanisms of tumour cell metastasation. MATERIALS AND METHODS: We performed realtime-PCR and multiplex-PCR analyses following immunomagnetic separation of CTC. Peripheral blood (PB) samples of 63 patients with breast cancer of various stages were analyzed and compared to a control group of 14 healthy individuals. After reverse-transcription, we performed multiplex PCR using primers for the genes ga733.3, muc-1 and c-erbB2. Mammaglobin1, spdef and c-erbB2 were analyzed applying realtime-PCR. RESULTS: ga733.2 overexpression was found in 12.7% of breast cancer cases, muc-1 in 15.9%, mgb1 in 9.1% and spdef in 12.1%. In this study, c-erbB2 did not show any significant correlation to BC, possibly due to a highly ambient expression. Besides single gene analyses, gene profiles were additionally evaluated. Highly significant correlations to BC were found in single gene analyses of ga733.2 and muc-1 and in gene profile analyses of ga733.3*muc-1 and GA7 ga733.3*muc-1*mgb1*spdef. CONCLUSION: Our study reveals that the single genes ga733.3, muc-1 and the gene profiles ga733.3*muc-1 and ga733.3*3muc-1*mgb1*spdef can serve as markers for the detection of CTC in BC. The multigene analyses found highly positive levels in BC patients. Our study indicates that not single gene analyses but subtle patterns of multiple genes lead to rising accuracy and low loss of specificity in detection of breast cancer cases.

Complete Clinical Remission of Stage IV Triple-Negative Breast Cancer Lung Metastasis Administering Low-Dose Immune Checkpoint Blockade in Combination With Hyperthermia and Interleukin-2.

The prognosis of triple-negative breast cancer with metastases after chemotherapy remains dismal. We report the case of a 50-year-old female with first disease recurrence at the axillary lymph node and, later on, bilateral pulmonary metastases with severe shortness of breath. The patient received low-dose immune checkpoint blockade (concurrent nivolumab and ipilimumab) weekly over 3 weeks with regional hyperthermia 3 times a week, followed by systemic fever-range hyperthermia induced by interleukin-2 for 5 days. She went into complete remission of her pulmonary metastases with transient WHO I-II diarrhea and skin rash. The patient remained alive for 27 months after the start of treatment, with recurrence of metastases as a sternal mass, and up to 3 cm pleural metastases. This exceptional response should instigate further research efforts with this protocol, which consists only of approved drugs and treatments.

Expression analysis of pancreatic cancer cell lines reveals association of enhanced gene transcription and genomic amplifications at the 8q22.1 and 8q24.22 loci.

Despite tremendous effort and progress in the diagnostics of pancreatic cancer with respect to imaging techniques and molecular genetics, only very few patients can be cured by surgery leading to a 5-year survival rate of only 3%. Especially the lack of chemotherapeutical options in this entity requires a better understanding of the molecular mechanisms leading to pancreatic carcinoma growth and progression in order to develop novel treatment regimens. To identify signaling pathways that are critical for this tumor entity, we compared six well-established pancreatic cancer cell lines (Capan-1, Capan-2, HUP-T3, HUP-T4, KCL-MOH, PaTu-8903) with colon cancer cell lines and tumor cell lines of non-epithelial origin by expression profiling. For this purpose we employed Human Genome Focus Arrays representing about 8500 well annotated human genes. We identified 353 genes with significantly high expression in the group of pancreatic carcinomas. Based on Gene Ontology annotations these genes are especially involved in Rho protein signal transduction, proteasome activator activity, cell motility, apoptotic program, and cell-cell adhesion processes indicating these pathways to be interesting candidates for the design of targeted therapies. Most pancreatic carcinomas are characterized by mutations in the TP53 and the KRAS genes and the absence of microsatellite instability, which could also be confirmed for our panel of pancreatic carcinoma cell lines. Looking for individual differences within this group that may be responsible for more or less aggressive behavior, we identified genomic amplifications at the 8q22.1 and the 8q24.22 loci to be associated with enhanced gene transcription. Because we have previously shown that gains of genomic material from the long arm of chromosome 8 have an adverse effect on the outcome of pancreatic carcinoma patients, we conclude that functional analysis of amplified genes at 8q22 and/or 8q24 may lead to an improved understanding of pancreatic carcinoma progression.

STAT5 phosphorylation in malignant melanoma is important for survival and is mediated through SRC and JAK1 kinases.

Altered signaling pathways are key regulators of cellular functions in tumor cells. Constitutive activation of signal transducer and activator of transcription (STAT)3 and -5 may be involved in tumor formation and progression. We have investigated the role of STAT5 in cutaneous melanoma metastases using various RNA and protein techniques. In melanoma specimens, Stat5b transcripts were upregulated approximately 3.8-fold. In 13 of 21 (62%) human melanoma metastases, STAT5 was phosphorylated in comparison to normal human melanocytes and benign nevi. The STAT5 target gene Bcl-2 was frequently upregulated. The investigation of the underlying mechanism revealed specific STAT5 activation by recombinant human epidermal growth factor (rEGF). rEGF-induced activation of STAT5 occurred in vitro through the non-receptor tyrosine kinases transforming gene (src) of Rous Sarcoma virus and Janus kinase 1. Inhibition of Stat5b expression by small interfering RNA strongly reduced the expression of Bcl-2 and led to decreased cell viability and increased apoptosis in the melanoma cell lines A375 and BLM. Transfection with dominant-negative Stat5b caused enhanced cell death and G1 arrest in A375 cells. Our study identifies phosphorylated STAT5 in melanoma and shows regulation through rEGF; STAT5 may thus act as a survival factor for growth of human melanoma and may represent a potential target for molecular therapy.

Gene expression in acute Stanford type A dissection: a comparative microarray study.

BACKGROUND: We compared gene expression profiles in acutely dissected aorta with those in normal control aorta. MATERIALS AND METHODS: Ascending aorta specimen from patients with an acute Stanford A-dissection were taken during surgery and compared with those from normal ascending aorta from multiorgan donors using the BD Atlas Human1.2 Array I, BD Atlas Human Cardiovascular Array and the Affymetrix HG-U133A GeneChip. For analysis only genes with strong signals of more than 70 percent of the mean signal of all spots on the array were accepted as being expressed. Quantitative real-time polymerase chain reaction (RT-PCR) was used to confirm regulation of expression of a subset of 24 genes known to be involved in aortic structure and function. RESULTS: According to our definition expression profiling of aorta tissue specimens revealed an expression of 19.1% to 23.5% of the genes listed on the arrays. Of those 15.7% to 28.9% were differently expressed in dissected and control aorta specimens. Several genes that encode for extracellular matrix components such as collagen IV alpha2 and -alpha5, collagen VI alpha3, collagen XIV alpha1, collagen XVIII alpha1 and elastin were down-regulated in aortic dissection, whereas levels of matrix metalloproteinases-11, -14 and -19 were increased. Some genes coding for cell to cell adhesion, cell to matrix signaling (e.g., polycystin1 and -2), cytoskeleton, as well as several myofibrillar genes (e.g., alpha-actinin, tropomyosin, gelsolin) were found to be down-regulated. Not surprisingly, some genes associated with chronic inflammation such as interleukin -2, -6 and -8, were up-regulated in dissection. CONCLUSION: Our results demonstrate the complexity of the dissecting process on a molecular level. Genes coding for the integrity and strength of the aortic wall were down-regulated whereas components of inflammatory response were up-regulated. Altered patterns of gene expression indicate a pre-existing structural failure, which is probably a consequence of insufficient remodeling of the aortic wall resulting in further aortic dissection.

Predictors of primary breast cancers responsiveness to preoperative epirubicin/cyclophosphamide-based chemotherapy: translation of microarray data into clinically useful predictive signatures.

BACKGROUND: Our goal was to identify gene signatures predictive of response to preoperative systemic chemotherapy (PST) with epirubicin/cyclophosphamide (EC) in patients with primary breast cancer. METHODS: Needle biopsies were obtained pre-treatment from 83 patients with breast cancer and mRNA was profiled on Affymetrix HG-U133A arrays. Response ranged from pathologically confirmed complete remission (pCR), to partial remission (PR), to stable or progressive disease, "No Change" (NC). A primary analysis was performed in breast tissue samples from 56 patients and 5 normal healthy individuals as a training cohort for predictive marker identification. Gene signatures identifying individuals most likely to respond completely to PST-EC were extracted by combining several statistical methods and filtering criteria. In order to optimize prediction of non responding tumors Student's t-test and Wilcoxon test were also applied. An independent cohort of 27 patients was used to challenge the predictive signatures. A k-Nearest neighbor algorithm as well as two independent linear partial least squares determinant analysis (PLS-DA) models based on the training cohort were selected for classification of the test samples. The average specificity of these predictions was greater than 74% for pCR, 100% for PR and greater than 62% for NC. All three classification models could identify all pCR cases. RESULTS: The differential expression of 59 genes in the training and the test cohort demonstrated capability to predict response to PST-EC treatment. Based on the training cohort a classifier was constructed following a decision tree. First, a transcriptional profile capable to distinguish cancerous from normal tissue was identified. Then, a "favorable outcome signature" (31 genes) and a "poor outcome signature" (26 genes) were extracted from the cancer specific signatures. This stepwise implementation could predict pCR and distinguish between NC and PR in a subsequent set of patients. Both PLS-DA models were implemented to discriminate all three response classes in one step. CONCLUSION: In this study signatures were identified capable to predict clinical outcome in an independent set of primary breast cancer patients undergoing PST-EC.

Predictive biological markers for response of invasive breast cancer to anthracycline/cyclophosphamide-based primary (radio-)chemotherapy.

BACKGROUND: The role of biological markers for the prediction of neoadjuvant chemotherapy and radio-chemotherapy may be evaluated using pathological complete response [pCR] in patients with invasive breast cancer. MATERIALS AND METHODS: To investigate this, pre-treatment biopsies from 517 patients with locally advanced breast cancer were analyzed for expression of estrogen receptor [ER], progesterone receptor [PgR], Her-2/neu, epidermal growth factor receptor [EGF-R], p53, Bcl-2 and MIB-1 by immunohistochemistry [IHC], and these data were compared to the pathological response after preoperative epirubicine/cyclophosphamide [EC] chemotherapy (+/- radiotherapy). RESULTS: pCR was more frequent (28.30%, 56/198) in tumors that received radio-chemotherapy compared to chemotherapy alone (11.9%, 38/319, p < 0.0001). Patients with high grading, lower ER, PgR, Bcl-2 or a higher proliferation had a significantly greater benefit from chemotherapy. The overexpressions of Her2/neu or EGF-R were weakly correlated to pCR, while p53 staining did not have any predictive value. Younger patients, with negative PgR and high proliferation index, had the highest benefit from EC therapy (56% pCR). The different multivariate indices of binary regression, PLS-DA and SIMCA, had similar predictive quality and were slightly superior to univariate factors. CONCLUSION: This study emphasizes the value of traditional biological markers and Bcl-2 for use in the individual selection of a primary therapy regimen.

Immediate gene expression changes after the first course of neoadjuvant chemotherapy in patients with primary breast cancer disease.

PURPOSE: Our goal was to identify genes undergoing expressional changes shortly after the beginning of neoadjuvant chemotherapy for primary breast cancer. EXPERIMENTAL DESIGN: The biopsies were taken from patients with primary breast cancer prior to any treatment and 24 hours after the beginning of the neoadjuvant chemotherapy. Expression analyses from matched pair samples representing 25 patients were carried out with Clontech filter arrays. A subcohort of those 25 paired samples were additionally analyzed with the Affymetrix GeneChip platform. All of the transcripts from both platforms were queried for expressional changes. RESULTS: Performing hierarchical cluster analysis, we clustered pre- and posttreatment samples from individual patients more closely to each other than the samples taken from different patients. This reflects the rather low number of transcripts responding directly to the drugs used. Although transcriptional drug response occurring during therapy differed between individual patients, two genes (p21(WAF1/CIP1) and MIC-1) were up-regulated in posttreatment samples. This could be validated by semiquantitative and real-time reverse transcription-PCR. Partial least- discriminant analysis based on approximately 25 genes independently identified by either Clontech or Affymetrix platforms could clearly discriminate pre- and posttreatment samples. However, correlation of certain gene expression levels as well as of differential patterns and clusters as determined by a different platform was not always satisfying. CONCLUSIONS: This study has demonstrated the potential of monitoring posttreatment changes in gene expression as a measure of the pharmacodynamics of drugs. As a clinical laboratory model, it can be useful to identify patients with sensitive and reactive tumors and to help for optimized choice for sequential therapy and obviously improve relapse- free and overall survival.

Identifying superficial, muscle-invasive, and metastasizing transitional cell carcinoma of the bladder: use of cDNA array analysis of gene expression profiles.

PURPOSE: Expression profiling by DNA microarray technology permits the identification of genes underlying clinical heterogeneity of bladder cancer and which might contribute to disease progression, thereby improving assessment of treatment and prediction of patient outcome. EXPERIMENTAL DESIGN: Invasive (20) and superficial (22) human bladder tumors from 34 patients with known outcome regarding disease recurrence and progression were analyzed by filter-based cDNA arrays (Atlas Human Cancer 1.2; BD Biosciences Clontech) containing 1185 genes. For 9 genes, array data were confirmed using real-time reverse transcription-PCR. Additionally, Atlas array data were validated using Affymetrix GeneChip oligonucleotide arrays with 22,283 human gene fragments and expressed sequence tags sequences in a subset of three superficial and six invasive bladder tumors. RESULTS: A two-way clustering algorithm using different subsets of gene expression data, including a subset of 41 genes validated by the oligonucleotide array (Affymetrix), classified tumor samples according to clinical outcome as superficial, invasive, or metastasizing. Furthermore, (a) a clonal origin of superficial tumors, (b) highly similar gene expression patterns in different areas of invasive tumors, and (c) an invasive-like pattern was observed in bladder mucosas derived from patients with locally advanced disease. Several gene clusters that characterized invasive or superficial tumors were identified. In superficial bladder tumors, increased mRNA levels of genes encoding transcription factors, molecules involved in protein synthesis and metabolism, and some proteins involved into cell cycle progression and differentiation were observed, whereas transcripts for immune, extracellular matrix, adhesion, peritumoral stroma and muscle tissue components, proliferation, and cell cycle controllers were up-regulated in invasive tumors. CONCLUSIONS: Gene expression profiling of human bladder cancers provides insight into the biology of bladder cancer progression and identifies patients with distinct clinical phenotypes.

Non-small lung cancer cells are prime targets for p53 gene transfer mediated by a recombinant adeno-associated virus type-2 vector.

In this study, we elucidated the potential of recombinant adeno-associated virus type-2 (rAAV-2) vectors for lung cancer gene therapy. Cell lines of the three major histological subtypes of non-small cell lung cancer (NSCLC) were highly susceptible for rAAV-2 showing transduction rates between 63.4 and 98.9%. In contrast, cell lines of small cell carcinomas were resistant to rAAV-2 infection. For restoration of p53 function in p53 deficient NSCLC, a rAAV-2 vector was constructed containing wt p53 cDNA. Following transduction with rAAV-p53, cell growth of all NSCLC cell lines was significantly reduced in a dose-dependent manner between 44 and 71.7% in comparison with rAAV-GFP transduced cells. The reduction of tumor cell growth was associated with increased apoptosis. Adding cisplatin to rAAV-p53-infected cells led to a significant growth inhibition between 81 and 91% indicating a synergistic effect between cisplatin and rAAV-p53. Interestingly, the tumor cells surviving cisplatin and rAAV-p53 treatment were inhibited in their ability to form colonies as reflected by a reduction of colony growth between 57 and 90.4%. In conclusion, rAAV-2 vectors exhibit a strong tropism for NSCLC. Successful inhibition of tumor cell growth following transduction with a rAAV-p53 vector underlines the potential role of rAAV-2 in cancer gene therapy.

Gene expression profiling of advanced head and neck squamous cell carcinomas and two squamous cell carcinoma cell lines under radio/chemotherapy using cDNA arrays.

BACKGROUND AND PURPOSE: The response of squamous cell carcinomas of the head and neck (HNSCC) to radio/chemotherapy is accompanied by complex changes in patterns of gene expression. It is highly probable that a better understanding of molecular and genetic changes can help to optimize the treatment of HNSCC. cDNA arrays provide a powerful tool for high-throughput monitoring of gene expression in small clinical specimens. MATERIALS AND METHODS: We used tumour biopsies from four patients with HNSCC which have been taken prior to and during radio/chemotherapy. The patterns of gene expression obtained from clinical samples were compared with gene expression profiles of two squamous cell carcinoma cell lines (FaDU and UD-7A). RESULTS: The experimental data analysis revealed changes in expression levels of several genes during radio/chemotherapy. Despite treatment, independent samples taken from the same cell line or tumour in situ were more similar to each other than either was to other specimens. The data indicate a high gene heterogeneity of HNSCC that is preserved during treatment. CONCLUSIONS: From our preliminary results we conclude that the cDNA array experimental approach can detect differences in gene expression between treated and untreated small tumour biopsies, as well as inter-individual differences in expression profiles between HNSCC tumours. The examination of a greater sample size will be needed to make this preliminary evaluation useful to elucidate the functional significance of individual genes which exhibit altered levels of expression under radiation therapy.

Analysis of BRCA1, TP53, and TSG101 germline mutations in German breast and/or ovarian cancer families.

About 5%-10% of breast cancers are considered to be hereditary and associated with germline mutations of specific genes. As yet, the most frequently affected genes identified are BRCA1 and BRCA2, but also other genes such as TP53 are supposed to influence the predisposition toward breast cancer. In the present study, we analyzed patients of 19 German families with early onset breast cancer and/or a family history of breast and/or ovarian cancer for the presence of mutations in BRCA1 and TP53. In addition, we screened for germline mutations in the putative tumor suppressor gene TSG101. For this purpose we used direct sequence analysis of the entire coding regions for all three genes and, in the case of BRCA1, single-strand conformation polymorphism analysis and protein transcription-translation assays. We identified eight previously described polymorphisms and several aberrations in BRCA1: 1 unclassified missense mutation, 3 small protein truncating mutations, 1 novel pseudoexon, and 5 splicing variants. No mutation was detected in TP53. Analysis of TSG101 transcripts revealed an aberrant transcript in two breast cancer patients belonging to the same family, suggesting TSG101 as a predisposing gene in hereditary breast cancer.

rIL-18 triggered gene therapy based on a transduction with the IL-12 plasmid: a new option as immuno-therapy for osteosarcoma?

Gene therapy is a promising new method to treat tumors locally. Immuno-therapy for treatment of osteosarcomas is one option for hopefully improving the survival rate of patients with this tumor. Transduction of OS cells with the pCMV-IL-12neo plasmid induced a significant increase in IFN-gamma expression by mononuclear cells. This is known to induce antitumor effects mediated by the immune system. In combination with an administration of rIL-18, the IFN-gamma increase was multiplied in a dose-dependent manner. This study demonstrated that osteosarcoma cells can be targeted effectively in vitro by plasmids encoding the IL-12 gene. Considering the synergistic pathways it is reasonable to combine a local, gene transfer based on IL-12 with a rIL-18 administration to trigger the potentially promising immuno-effects for adjuvant treatment of osteosarcomas.

Immunotherapy by gene transfer with plasmids encoding IL-12/IL-18 is superior to IL-23/IL-18 gene transfer in a rat osteosarcoma model.

BACKGROUND: Osteosarcomas are primary malignant tumors of bone or soft parts arising from bone-forming mesenchymal cells. Despite dramatic therapeutic advances, namely neo-adjuvant and adjuvant chemotherapy, progress is at a plateau. Cytokine-mediated gene therapy might represent a further advance in the therapy of the osteosarcoma. MATERIALS AND METHODS: We transfected UMR 108 osteosarcoma cells with different plasmids encoding IL-12, IL-23, proIL-18 and ICE (Interleukin-converting enzyme). IFN-gamma induction, which is known to induce antitumor effects mediated by the immune system, and cytotoxic effects of various cytokine combination were investigated. RESULTS: Our results show that local secretion of IL-12 by UMR 108 cells led to an induction of cytotoxic effects mediated by mononuclear cells, which were enhanced by additional administration of recombinant IL-18. In contrast to IL-18, IL-23 showed a moderate increase of IFN-gamma induction when transfected alone and could only slightly increase the IFN-gamma induction mediated by IL-12. IL-18 enhanced IFN-gamma induction when applied alone and was able to increase the IFN-gamma production that was induced by IL-12. CONCLUSION: IL-23 seems to be a less effective immuno-therapeutic for adjuvant treatment of osteosarcomas than IL-12 and IL-18, when taking only IFN-gamma induction into consideration.