The impacts of a global pandemic on the efficacy and stability of contemporary wildlife conservation: South Africa as a case study.

Conservationists speculated on potential benefits to wildlife of lockdown restrictions because of the COVID-19 pandemic but voiced concern that restrictions impeded nature conservation. We assessed the effects of lockdown restrictions on biodiversity conservation in South Africa, a biodiverse country with economic inequality and reliance on wildlife resources. We solicited expert opinion using the IUCN's Threats Classification Scheme to structure a questionnaire and illustrated responses with individual case studies from government parastatal and non-governmental conservation organisations. The most highly reported threats were biological resource use, residential/commercial developments, invasive species, and human intrusions. The trends reported by 90 survey respondents were supported by case studies using environmental compliance data from parastatal conservation organisations. Lack of tourism revenue and funding were cited as hindrances to conservation. Mechanisms to prevent environmental degradation in the face of global emergencies must be implemented and 'ring-fenced' to ensure conservation is not a casualty during future global crises.

Kinase Mutations and Imatinib Response in Patients With Metastatic Gastrointestinal Stromal Tumor.

PURPOSE: Most gastrointestinal stromal tumors (GISTs) express constitutively activated mutant isoforms of KIT or kinase platelet-derived growth factor receptor alpha (PDGFRA) that are potential therapeutic targets for imatinib mesylate. The relationship between mutations in these kinases and clinical response to imatinib was examined in a group of patients with advanced GIST. PATIENTS AND METHODS: GISTs from 127 patients enrolled onto a phase II clinical study of imatinib were examined for mutations of KIT or PDGFRA. Mutation types were correlated with clinical outcome. RESULTS: Activating mutations of KIT or PDGFRA were found in 112 (88.2%) and six (4.7%) GISTs, respectively. Most KIT mutations involved exon 9 (n = 23) or exon 11 (n = 85). All KIT mutant isoforms, but only a subset of PDGFRA mutant isoforms, were sensitive to imatinib, in vitro. In patients with GISTs harboring exon 11 KIT mutations, the partial response rate (PR) was 83.5%, whereas patients with tumors containing an exon 9 KIT mutation or no detectable mutation of KIT or PDGFRA had PR rates of 47.8% (P = .0006) and 0.0% (P < .0001), respectively. Patients whose tumors contained exon 11 KIT mutations had a longer event-free and overall survival than those whose tumors expressed either exon 9 KIT mutations or had no detectable kinase mutation. CONCLUSION: Activating mutations of KIT or PDGFRA are found in the vast majority of GISTs, and the mutational status of these oncoproteins is predictive of clinical response to imatinib. PDGFRA mutations can explain response and sensitivity to imatinib in some GISTs lacking KIT mutations.

MMP-1 expression has an independent prognostic value in breast cancer.

BACKGROUND: Breast cancer consists of a variety of tumours, which differ by their morphological features, molecular characteristics and outcome. Well-known prognostic factors, e.g. tumour grade and size, Ki-67, hormone receptor status, HER2 expression, lymph node status and patient age have been traditionally related to prognosis. Although the conventional prognostic markers are reliable in general, better markers to predict the outcome of an individual tumour are needed. Matrix metalloproteinase-1 (MMP-1) expression has been reported to inversely correlate with survival in advanced cancers. In breast cancer MMP-1 is often upregulated, especially in basal-type breast tumours. The purpose of this retrospective study was to analyse MMP-1 expression in breast cancer cells and in cancer associated stromal cells and to correlate the results with traditional prognostic factors including p53 and bcl-2, as well as to patient survival in breast cancer subtypes. METHODS: Immunohistochemical analysis of MMP-1, ER, PR, Ki-67, HER2, bcl-2, p53 and CK5/6 expression was performed on 125 breast cancers. Statistical analyses were carried out using Kruskal-Wallis and Mann-Whitney -tests. In pairwise comparison Bonferroni-adjustment was applied. Correlations were calculated using Spearman rank-order correlation coefficients. Kaplan-Meier survival analyses were carried out to compare breast cancer-specific survival curves. Factors significantly associated with disease-specific survival in univariate models were included in multivariate stepwise. RESULTS: Positive correlations were found between tumour grade and MMP-1 expression in tumour cells and in stromal cells. P53 positivity significantly correlated with MMP-1 expression in tumour cells, whereas HER2 expression correlated with MMP-1 both in tumour cells and stromal cells. MMP-1 expression in stromal cells showed a significant association with luminal A and luminal B, HER2 overexpressing and triple-negative breast cancer subtypes. CONCLUSIONS: The most important finding of this study was the independent prognostic value of MMP-1 as well as Ki-67 and bcl-2 expression in tumour cells. Our study showed also that both tumoural and stromal MMP-1 expression is associated with breast tumour progression and poor prognosis. A significant difference of MMP-1 expression by cancer associated stromal cells in luminal A, luminal B and triple-negative breast cancer classes was also demonstrated.

Efficacy and safety of imatinib mesylate in advanced gastrointestinal stromal tumors.

BACKGROUND: Constitutive activation of KIT receptor tyrosine kinase is critical in the pathogenesis of gastrointestinal stromal tumors. Imatinib mesylate, a selective tyrosine kinase inhibitor, has been shown in preclinical models and preliminary clinical studies to have activity against such tumors. METHODS: We conducted an open-label, randomized, multicenter trial to evaluate the activity of imatinib in patients with advanced gastrointestinal stromal tumor. We assessed antitumor response and the safety and tolerability of the drug. Pharmacokinetics were assessed in a subgroup of patients. RESULTS: A total of 147 patients were randomly assigned to receive 400 mg or 600 mg of imatinib daily. Overall, 79 patients (53.7 percent) had a partial response, 41 patients (27.9 percent) had stable disease, and for technical reasons, response could not be evaluated in 7 patients (4.8 percent). No patient had a complete response to the treatment. The median duration of response had not been reached after a median follow-up of 24 weeks after the onset of response. Early resistance to imatinib was noted in 20 patients (13.6 percent). Therapy was well tolerated, although mild-to-moderate edema, diarrhea, and fatigue were common. Gastrointestinal or intraabdominal hemorrhage occurred in approximately 5 percent of patients. There were no significant differences in toxic effects or response between the two doses. Imatinib was well absorbed, with pharmacokinetics similar to those reported in patients with chronic myeloid leukemia. CONCLUSIONS: Imatinib induced a sustained objective response in more than half of patients with an advanced unresectable or metastatic gastrointestinal stromal tumor. Inhibition of the KIT signal-transduction pathway is a promising treatment for advanced gastrointestinal stromal tumors, which resist conventional chemotherapy.

Mechanisms of glutamate receptor induced proliferation of astrocytes.

Astrocytes express mainly metabotropic glutamate receptor 3 and metabotropic glutamate receptor 5 receptor subtypes, which show opposing effects on cellular proliferation upon activation. In this study, we investigated the mechanisms by which activation of these receptors modulates astrocyte proliferation. Activation of metabotropic glutamate receptor 5 with (S)-3,5-dihydroxyphenylglycine increased phospholipase D activity in astrocytes as well as astrocyte proliferation. The 3,5-dihydroxyphenylglycine-induced proliferation was inhibited in the presence of the metabotropic glutamate receptor 5 antagonist (2-methyl-6-(phenylethynyl)pyridine), the protein kinase C inhibitor GF109203X, brefeldin A and 1-butanol. Activation of metabotropic glutamate receptor 3 with (2'S,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)glycine-IV (DCG-IV) inhibited astrocyte proliferation without affecting metabotropic glutamate receptor 5-mediated phospholipase D activity. Metabotropic glutamate receptor 3 activation, however, only partially inhibited metabotropic glutamate receptor 5-mediated proliferation. In conclusion, metabotropic glutamate receptor 5 stimulates astrocyte proliferation via a protein kinase C-phospholipase D-phosphatidic acid-dependent pathway, whereas metabotropic glutamate receptor 3-mediated inhibition of astrocyte proliferation does not involve phospholipase D, and is independent of metabotropic glutamate receptor 5-mediated effects.

Kinase mutations and imatinib response in patients with metastatic gastrointestinal stromal tumor.

PURPOSE: Most gastrointestinal stromal tumors (GISTs) express constitutively activated mutant isoforms of KIT or kinase platelet-derived growth factor receptor alpha (PDGFRA) that are potential therapeutic targets for imatinib mesylate. The relationship between mutations in these kinases and clinical response to imatinib was examined in a group of patients with advanced GIST. PATIENTS AND METHODS: GISTs from 127 patients enrolled onto a phase II clinical study of imatinib were examined for mutations of KIT or PDGFRA. Mutation types were correlated with clinical outcome. RESULTS: Activating mutations of KIT or PDGFRA were found in 112 (88.2%) and six (4.7%) GISTs, respectively. Most KIT mutations involved exon 9 (n = 23) or exon 11 (n = 85). All KIT mutant isoforms, but only a subset of PDGFRA mutant isoforms, were sensitive to imatinib, in vitro. In patients with GISTs harboring exon 11 KIT mutations, the partial response rate (PR) was 83.5%, whereas patients with tumors containing an exon 9 KIT mutation or no detectable mutation of KIT or PDGFRA had PR rates of 47.8% (P =.0006) and 0.0% (P <.0001), respectively. Patients whose tumors contained exon 11 KIT mutations had a longer event-free and overall survival than those whose tumors expressed either exon 9 KIT mutations or had no detectable kinase mutation. CONCLUSION: Activating mutations of KIT or PDGFRA are found in the vast majority of GISTs, and the mutational status of these oncoproteins is predictive of clinical response to imatinib. PDGFRA mutations can explain response and sensitivity to imatinib in some GISTs lacking KIT mutations.

Synthesis and pharmacology of willardiine derivatives acting as antagonists of kainate receptors.

The natural product willardiine (8) is an AMPA receptor agonist while 5-iodowillardiine (10) is a selective kainate receptor agonist. In an attempt to produce antagonists of kainate and AMPA receptors analogues of willardiine with substituents at the N3 position of the uracil ring were synthesized. The N3-4-carboxybenzyl substituted analogue (38c) was found to be equipotent at AMPA and GLUK5-containing kainate receptors in the neonatal rat spinal cord. The N3-2-carboxybenzyl substituted analogue (38a) proved to be a potent and selective GLUK5 subunit containing kainate receptor antagonist when tested on native rat and human recombinant AMPA and kainate receptor subtypes. The GLUK5 kainate receptor antagonist activity was found to reside in the S enantiomer (44a) whereas the R enantiomer (44b) was almost inactive. 5-Iodo substitution of the uracil ring of 44a gave 45, which was found to have enhanced potency and selectivity for GLUK5.

Increased proliferation activity measured by immunoreactive Ki67 is associated with survival improvement in rectal/recto sigmoid cancer.

AIM: To assess the expression of Ki67 as prognosticator in rectal/recto sigmoid cancer. METHODS: Samples from 146 patients with rectal and recto sigmoid cancer were studied for expression of Ki67 and its prognostic significance in comparison with clinico-pathological predictors of survival. Formalin-fixed, paraffin-embedded tissues from 6 (4.1%) patients with T1, 26 (17.8%) with T2, 94 (64.4%) with T3, and 20 (13.7%) with T4 tumors were studied. Ki67 expression was determined immunohistochemically. Samples were divided according to mean value into high (>40%) and low (< or =40%) expression. Areas of extensive proliferation (>50%) were defined as "hot spot" areas. RESULTS: Hot spot areas were present in samples regardless of histopathological grade. Lower TNM and Dukes stage and higher expression of Ki67 and presence of Ki67 hot spot areas in histopathological samples were associated with better survival, whereas no association was observed with histopathological grade (P = 0.78). In Cox multivariate regression analysis, significant prognostic factors were Dukes stage (P<0.001), presence of lymph node metastases (P = 0.015), age (P = 0.035) and presence of Ki67 hot spot areas (P = 0.044). CONCLUSION: Proliferative activity as measured by Ki67 in rectal cancer is associated with survival improvement compared with patients with low Ki67. Areas of prognostically significant increased proliferation were found independently of histopathological tumor grade.

Analysis of mGluR1a constitutive internalization using a pulse-chase enzyme-linked immuno-sorbant assay (ELISA).

The surface expression of G protein-coupled receptors is regulated by internalization. For many receptors, a constitutive level of internalization in the absence of agonist has been reported. The constitutive internalization of metabotropic glutamate receptor 1a (mGluR1a) has been described, but in general little attention has been dedicated to this important aspect of receptor regulation. Here we describe a pulse-chase ELISA method that allows the investigation of mGluR1a constitutive internalization. When investigated by pulse-chase ELISA, the constitutive internalization of mGluR1a was inhibited by dominant negative mutant constructs of arrestin-2 or Eps-15. This observation, besides indicating the arrestin- and clathrin-dependence of mGluR1a constitutive internalization, also confirmed the physiological relevance of the method described in this article. Confocal microscopy experiments to study receptor localization further validated the pulse-chase labelling procedure. The application of the pulse-chase ELISA to mGluR1b, revealed that this splice variant undergoes marginal constitutive internalization. Two COOH-terminal deletion mutants of mGluR1a, DMI (Arg847stop) and DMII (Arg868stop), were also tested for constitutive internalization. Interestingly, only DMII underwent significant constitutive internalization, suggesting that the region between Arg847 and Arg868 might play a regulatory role in mGluR1a trafficking. Taken together, the pulse-chase ELISA appears to be an efficient tool to analyze the constitutive internalization of different mGluR1 splice variants.

Clinical presentation of gastrointestinal stromal tumors and treatment of operable disease.

Gastrointestinal stromal tumors (GISTs) are generally found in the stomach or small intestine and less commonly in the colon, rectum or an intra-abdominal site. The patients symptoms on presentation are most commonly gastrointestinal bleeding. Surgery remains the standard treatment for nonmetastatic GISTs, but rates of disease recurrence are significant--5% in primary disease and 90% in locally advanced disease. Five-year survival following surgical resection varies between 35% and 65% on the basis of several published studies. Clinical knowledge of the prognosis of patients with GISTs remains rather limited--small tumor size, low-grade mitotic index and stomach location are factors associated with a more favorable prognosis.

On the unreliability of published DNA sequences.

• Here, the reliability of published fungal nucleic acid sequences is tested by the critical re-evaluation of 206 named sequences obtained from public-access databases. • Sequences from the ribosomal RNA (rRNA) gene cluster were examined as these are commonly used to establish fungal phylogeny and evolution, and are also increasingly employed in the identification of fungi from nonculture based studies. • Fifty-one rRNA internal transcribed spacer (ITS) sequences were obtained for species of Amanita, 55 ITS sequences were obtained for species of Phoma and 100 rRNA small subunit sequences were obtained from representative genera of the order Helotiales. In each case, the fungal group was selected partly on the basis of sequences deposited by three or more laboratories in order to avoid sample bias. The results suggest that up to 20% of the sequences available for each group may be unreliable, and this proportion is supported by additional informal observations.

Quality of life following laparoscopic Nissen fundoplication: assessing short-term and long-term outcomes.

AIM: To investigate the quality of life following laparoscopic Nissen fundoplication by assessing short-term and long-term outcomes. METHODS: From 1992 to 2005, 249 patients underwent laparoscopic Nissen fundoplication. Short-term outcome data including symptom response, side effects of surgery, endoscopy, and patient's perception of overall success were collected prospectively. Long-term outcomes were investigated retrospectively in patients with a median follow-up of 10 years by assessment of reflux symptoms, side effects of surgery, durability of antireflux surgery, need for additional treatment, patient's perception of success, and quality of life. Antireflux surgery was considered a failure based on the following criteria: moderate to severe heartburn or regurgitation; moderate to severe dysphagia reported in combination with heartburn or regurgitation; regular proton pump inhibitor medication use; endoscopic evidence of erosive esophagitis Savary-Miller grade 1-4; pathological 24-h pH monitoring; or necessity to undergo an additional surgery. The main outcome measures were short- and long-term cure rates and quality of life, with patient satisfaction as a secondary outcome measure. RESULTS: Conversion from laparoscopy to open surgery was necessary in 2.4% of patients. Mortality was zero and the 30-d morbidity was 7.6% (95%CI: 4.7%-11.7%). The median postoperative hospital stay was 2 d [interquartile range (IQR) 2-3 d]. Two hundred and forty-seven patients were interviewed for short-term analysis following endoscopy. Gastroesophageal reflux disease was cured in 98.4% (95%CI: 95.9%-99.6%) of patients three months after surgery. New-onset dysphagia was encountered postoperatively in 13 patients (6.7%); 95% reported that the outcome was better after antireflux surgery than with preoperative medical treatment. One hundred and thirty-nine patients with a median follow-up of 10.2 years (IQR 7.2-11.6 years) were available for a long-term evaluation. Cumulative long-term cure rates were 87.7% (81.0%-92.2%) at 5 years and 72.9% (64.0%-79.9%) at 10 years. Gastrointestinal symptom rating scores and RAND-36 quality of life scores of patients with treatment success were similar to those of the general population but significantly lower in those with failed antireflux surgery. Of the patients available for long-term follow-up, 83% rated their operation a success. CONCLUSION: For the long-term, our results indicate decreasing effectiveness of laparoscopic antireflux surgery, although most of the patients seem to have an overall quality of life similar to that of the general population.

Inverse Laplace Transform of Multidimensional Relaxation Data Without Non-Negativity Constraint.

An algorithm based on Tikhonov regularization in generalized form is described to perform an inverse Laplace transform of multidimensional data without a non-negativity (NN) constraint for spectrum conditioning. Uniform penalty (UP) regularization is used to reduce the requirement for NN, and a further penalty is introduced for zero-crossing (ZC) of the spectrum. This ZC term is weighted with the slope of the curve, which does not prevent negative modes in the spectrum but makes nonphysical undershooting in the vicinity of narrow peaks more expensive. The performance of this algorithm is demonstrated using synthetic data, and the optimization of the free parameters for calculating the regularization matrix is discussed.

Metabotropic glutamate receptor mGlu2 is resistant to homologous agonist-induced desensitization but undergoes protein kinase C-mediated heterologous desensitization.

To investigate the susceptibility of the group II metabotropic glutamate receptor mGlu2 to agonist-induced desensitization, the receptor was stably expressed in Chinese hamster ovary (CHO-mGlu2) or C6 glioma cells (C6-mGlu2). Exposure of CHO-mGlu2 cells to the group II mGlu receptor agonist (2S,1'S,2'S)-2-(carboxycyclopropyl)glycine (LCCG-1; 10 µM) for up to 15 h did not affect the subsequent ability of LCCG-1 to inhibit forskolin-stimulated cAMP accumulation. Similarly, in C6-mGlu2 cells, prolonged exposure to LCCG-1 also did not affect the subsequent ability of LCCG-1 to inhibit cAMP formation. In contrast, exposure of CHO-mGlu2 cells to the protein kinase C activator phorbol myristate acetate (PMA) suppressed the ability of LCCG-1 to inhibit cAMP formation. Using an in vitro model of group II mGlu receptor activity, the hemisected neonatal rat spinal cord preparation, the ability of the selective group II agonist (2R,4R)-4-aminopyrrolidine-2,4-dicarboxylate ((2R,4R)-APDC) to depress the fast component of the dorsal root-evoked ventral root potential (fDR-VRP) did not desensitize when applied for up to 2 h. Together these results indicate that in contrast to most G protein-coupled receptors, the mGlu2 receptor is resistant to agonist-induced homologous desensitization, and that in vitro data suggests that resistance to desensitization is a physiologically relevant property of this mGlu receptor subtype.

Long-term results from a randomized phase II trial of standard- versus higher-dose imatinib mesylate for patients with unresectable or metastatic gastrointestinal stromal tumors expressing KIT.

PURPOSE: The outcome of patients diagnosed with advanced gastrointestinal stromal tumor (GIST) and treated long-term with imatinib mesylate is unknown. A previous report of a randomized phase II trial of imatinib mesylate in patients with incurable GIST detailed high response rates at both the 400 and the 600 mg/d dose levels. We conducted a long-term analysis of patients treated on the trial, including patients followed during an extension phase, to evaluate survival, patterns of failure, and potential prognostic factors, including tumor mutational status. PATIENTS AND METHODS: Patients with advanced GIST were enrolled onto an open-label, multicenter trial and were randomly assigned (1:1) to receive imatinib 400 versus 600 mg/d. Data were prospectively collected on KIT mutational status, total tumor area, and other potential prognostic factors. Patients were followed for a median of 63 months. RESULTS: One hundred forty-seven patients were enrolled: 73 were in arm A (imatinib 400 mg/d), and 74 were in arm B (imatinib 600 mg/d). Response rates, median progression-free survival, and median overall survival were essentially identical on both arms, and median survival was 57 months for all patients. Forty-one patients overall (28%) remained on the drug long-term. Female sex, the presence of an exon 11 mutation, and normal albumin and neutrophil levels were independently associated with better survival. CONCLUSION: Nearly 50% of patients with advanced GIST who were treated with imatinib mesylate survived for more than 5 years, regardless of a 400 or 600 mg/d starting dose.

Prognostic significance of matrix metalloproteinases-1, -2, -7 and -13 and tissue inhibitors of metalloproteinases-1, -2, -3 and -4 in colorectal cancer.

Strong expression of many matrix metalloproteinases (MMPs) has been related to poor survival of colorectal cancer (CRC) patients. The expression of tissue inhibitors of metalloproteinases (TIMPs) has been associated with both a beneficial and a poor outcome and there is thus a need to further clarify the significance of MMPs and TIMPs in CRC. The prognostic significance of 4 MMPs and TIMPs in CRC was evaluated. Formalin-fixed, paraffin-embedded tissue arrayed samples of 351 patients with primary colon or rectal cancer of Dukes' stages A-D were selected for immunohistochemical staining of MMP-1, -2, -7 and -13, and TIMP-1, -2, -3 and -4. High expression of MMP-2 in the malignant epithelium as well as in the surrounding stroma was associated with reduced survival of colon cancer patients. Strong epithelial and stromal cytoplasmic staining of TIMP-3 was associated with a longer survival in rectal cancer patients, and here the interobserver variation for evaluating the degree of staining was lower than for epithelial staining. Strong stromal cytoplasmic staining of TIMP-4 predicted longer survival of rectal cancer patients. Multivariate analysis showed that stromal cytoplasmic TIMP-3 staining was the only marker of independent prognostic value. MMP-2 might be a useful prognostic marker in colon cancer, and TIMP-3 and TIMP-4 in rectal cancer, but the findings associated with stromal staining should be interpreted with some caution. Different biologic behavior or different genetic development may explain the differences between colon and rectal cancers regarding the expression of MMP-2, TIMP-3 and TIMP-4.