Trastuzumab deruxtecan versus trastuzumab emtansine in HER2-positive metastatic breast cancer: long-term survival analysis of the DESTINY-Breast03 trial.

Trastuzumab deruxtecan (T-DXd) demonstrated significantly improved efficacy over trastuzumab emtansine (T-DM1) in DESTINY-Breast03 (median follow-up, 28 months). We report updated efficacy and safety analyses, including secondary and exploratory efficacy endpoints (median follow-up, 41 months) of DESTINY-Breast03. Patients with advanced HER2-positive metastatic breast cancer previously treated with taxane and trastuzumab were randomized to T-DXd (5.4 mg per kg (261 patients)) or T-DM1 (3.6 mg per kg (263 patients)). The primary endpoint was progression-free survival (PFS) by blinded independent central review and was previously reported. The key secondary endpoint was overall survival (OS). Other secondary endpoints included objective response rate, duration of response and PFS (all by investigator assessment) and safety. At data cutoff, 20 November 2023, median PFS by investigator assessment was 29.0 versus 7.2 months (hazard ratio (HR), 0.30; 95% confidence interval (CI), 0.24-0.38), the 36-month PFS rate was 45.7% versus 12.4% and median OS was 52.6 versus 42.7 months (HR, 0.73; 95% CI, 0.56-0.94) with T-DXd versus T-DM1, respectively. Treatment-emergent adverse events were consistent with the previous analyses. No new instances of grade ≥3 interstitial lung disease or pneumonitis occurred (all grade rate, 16.7% (T-DXd) versus 3.4% (T-DM1)). With longer follow-up, T-DXd continued to demonstrate superior efficacy over T-DM1 with a manageable safety profile. ClinicalTrials.gov registration: NCT03529110 .

Developmental stages and episode-specific regulatory genes in andromonoecious melon flower development.

BACKGROUND AND AIMS: Given the lack of specific studies on floral development in melon (Cucumis melo L.), we carried out an extensive study involving morphological and transcriptomic analyses to characterize floral development in this species. METHODS: Using an andromonoecious line, we analysed the development of floral buds in male and hermaphrodite flowers with both light microscopy and scanning electron microscopy. Based on flower lengths, we established a correlation between the developmental stages and four main episodes of floral development and conducted an extensive RNA sequencing analysis of these episodes. KEY RESULTS: We identified 12 stages of floral development, from the appearance of the floral meristems to anthesis. The main structural differences between male and hermaphrodite flowers appeared between stages 6 and 7; later stages of development leading to the formation of organs and structures in both types of flowers were also described. We analysed the gene expression patterns of the four episodes in flower development to find the genes that were specific to each given episode. Among others, we identified genes that defined the passage from one episode to the next according to the ABCDE model of floral development. CONCLUSIONS: This work combines a detailed morphological analysis and a comprehensive transcriptomic study to enable characterization of the structural and molecular mechanisms that determine the floral development of an andromonoecious genotype in melon. Taken together, our results provide a first insight into gene regulation networks in melon floral development that are crucial for flowering and pollen formation, highlighting potential targets for genetic manipulation to improve crop yield of melon in the future.

Editing melon eIF4E associates with virus resistance and male sterility.

The cap-binding protein eIF4E, through its interaction with eIF4G, constitutes the core of the eIF4F complex, which plays a key role in the circularization of mRNAs and their subsequent cap-dependent translation. In addition to its fundamental role in mRNA translation initiation, other functions have been described or suggested for eIF4E, including acting as a proviral factor and participating in sexual development. We used CRISPR/Cas9 genome editing to generate melon eif4e knockout mutant lines. Editing worked efficiently in melon, as we obtained transformed plants with a single-nucleotide deletion in homozygosis in the first eIF4E exon already in a T0 generation. Edited and non-transgenic plants of a segregating F2 generation were inoculated with Moroccan watermelon mosaic virus (MWMV); homozygous mutant plants showed virus resistance, while heterozygous and non-mutant plants were infected, in agreement with our previous results with plants silenced in eIF4E. Interestingly, all homozygous edited plants of the T0 and F2 generations showed a male sterility phenotype, while crossing with wild-type plants restored fertility, displaying a perfect correlation between the segregation of the male sterility phenotype and the segregation of the eif4e mutation. Morphological comparative analysis of melon male flowers along consecutive developmental stages showed postmeiotic abnormal development for both microsporocytes and tapetum, with clear differences in the timing of tapetum degradation in the mutant versus wild-type. An RNA-Seq analysis identified critical genes in pollen development that were down-regulated in flowers of eif4e/eif4e plants, and suggested that eIF4E-specific mRNA translation initiation is a limiting factor for male gametes formation in melon.

Reverse transcriptase and protease inhibitors mutational viral load in HIV infected pregnant women with transmitted drug resistance in Argentina.

OBJECTIVE: Argentina has reported high levels of transmitted drug resistance (TDR), in HIV-infected pregnant women by population sequencing. We aimed to describe, in patients with TDR, the percentage of quasispecies harboring resistance mutations (RAMs) and mutational load (ML). METHODS: Retrospective study in a cohort of 40 naïve HIV-infected pregnant women, whose pretreatment samples had been genotyped by TRUGENE (period 2008-2014). Samples were re-sequenced with Ultra-deep Sequencing and ML was calculated considering baseline HIV-1 RNA load multiplied by the frequency of quasispecies harboring RAMs. RESULTS: TDR for NNRTIs, NRTIs and PIs was 17.5% (n=7 patients), 10% (n=4), 12.5% (n=5) respectively. Predominant NNRTI RAMs were K103N (n=4; 10%) and G190A/E/S (n=3; 7.5%). For NNRTIs, 78% of RAMs were present in >93.5% of viral population and ML was >1000 copies/mL (c/mL) for 89%, with a median (IQR) of 8330 c/ml (7738-29796). The following NRTI RAMs were described (per patient: % of quasispecies, ML): T215I (99.7%, 11014 c/ml); D67G (1.28%, 502 c/mL); M41L (79.8%, 88578 c/mL) and M184I (1.02%, 173 c/mL). Most frequent PI-RAMs were I85V, M46I, I50V and L90M (n=2, 5% each). For PIs, quasispecies with RAMs were <2.3% of viral population and ML was <350 c/mL for 77.8% of them. CONCLUSIONS: NNRTI-RAMs are predominant within the viral population, usually exceeding the threshold of 1000 c/mL, indicating potential higher risk of perinatal transmission. Conversely, PI mutations appear mostly as minority variants, with potential lower risk of transmission. Among NRTI, quasispecies harboring RAMs and ML values were variable.

Reducing herbicide use and leaching in agronomically performant maize-based cropping systems: An 8-year study.

Irrigated maize-based Cropping Systems (CS) are questioned because of the high risk of herbicide transfer to water. An 8-year systemic experiment was conducted to i) compute a multi-performance comparison between a Conventional Maize Monoculture (MMConv) and four CS that aimed to reduce irrigation and herbicide leaching: MMLI, a low-input MM using cover crop and Integrated Weed Management (IWM) techniques; MMStill, a Strip-tillage MM using cover crop; MMCT, a Conservation Tillage MM with cover crop; Maize-MSW, an IWM Maize rotated with Soybean and Wheat and ii) determine the main drivers and evaluate the influence of CS on herbicide leaching in maize. Drainage water was collected through 1-m depth lysimeter plates and analysed for 6 herbicide molecules and 1 degradation metabolite. MMLI yielded 10.7 t ha-1 close to MMConv (11.5 t ha-1) despite a lower herbicide use (-57%) and irrigation (-21%). MMLI and Maize-MSW had less drainage events compared to MMConv. MMCT and MMStill both yielded less (respectively 7.6 t ha-1 and 6.2 t ha-1) while their herbicide use increased (both +24%). Mean annual herbicide losses were 0.5 ± 1.0 g ha-1 for MMLI, 0.7 ± 1.2 g ha-1 for Maize-MSW, 1.3 ± 2.1 g ha-1 for MMStill, 2.0 ± 4.8 g ha-1 MMConv and 3.0 ± 9.6 g ha-1 for MMCT. Herbicide leaching remained variable but was consistently and mostly influenced by drainage volume. According to the CS, only 1.5 to 6.0 drainage events were responsible for 90% of the herbicide losses. High leaching peaks were identified for mesotrione and glyphosate and may indicate that preferential flows occurred, especially under MMCT. Quantity applied had limited influence on herbicide leaching. To reduce the herbicide leaching risk, CS must concomitantly manage water quality and quantity through a combination of agroecological practices, as in MMLI, a CS able to reach other technical objectives. Present study recommends assessing CS through a diversity of performance indicators.

Virus-Infected Melon Plants Emit Volatiles that Induce Gene Deregulation in Neighboring Healthy Plants.

It is well described that viral infections stimulate the emission of plant volatiles able to recruit viral vectors thereby promoting virus spread. In contrast, much less is known on the effects that emitted volatiles may have on the metabolism of healthy neighboring plants, which are potential targets for new infections through vector transmission. Watermelon mosaic virus (WMV) (genus Potyvirus, family Potyviridae) is an aphid-transmitted virus endemic in cucurbit crops worldwide. We have compared gene expression profiles of WMV-infected melon plants with those of healthy or healthy-but-cohabited-with-infected plants. Pathogenesis-related (PR) and small heat shock protein encoding genes were deregulated in cohabited plants, and PR deregulation depended on the distance to the infected plant. The signaling was short distance in the experimental conditions used, and cohabiting had a moderate effect on the plant susceptibility to WMV. Static headspace experiments showed that benzaldehyde and γ-butyrolactone were significantly over-emitted by WMV-infected plants. Altogether, our data suggest that perception of a volatile signal encoded by WMV-infected tissues triggers a response to prepare healthy tissues or/and healthy neighboring plants for the incoming infections.

Endoscopic Endonasal Approach for Clival Chordoma with Subarachnoid Invasion: A Two-Dimensional Step-by-Step Operative Video.

Chordomas are slow-growing, low-grade, locally invasive, and locally aggressive tumors. They peak at 40-60 years of age, with a male preponderance (2:1). Belonging to the sarcoma family and thought to develop from the notochord remnant, they are most commonly found in the midline, with half located at the sacrum and about one third at the skull base.1 Their treatment mainly consists of surgical excision, followed by radiation therapy.2 The endoscopic endonasal approach provides direct access to the clival chordomas with no need for brain retraction or manipulation of neurovascular structures.3-5 Herein we present a step-by-step resection technique of a clival chordoma invading the subarachnoid space and touching the brainstem and vertebrobasilar vessels in a 46-year-old man with headaches, with a prior failed attempt of resection at an outside institution, resulting in a biopsy only in the palate. An endoscopic endonasal transclival approach was performed and gross total removal was achieved (Video 1). The patient had an uneventful recovery with no deficits, and he was then sent to proton beam therapy.

Effects of the Aphanizomenon flos-aquae Extract (Klamin®) on a Neurodegeneration Cellular Model.

Cyanobacteria have been recognized as a source of bioactive molecules to be employed in nutraceuticals, pharmaceuticals, and functional foods. An extract of Aphanizomenon flos-aquae (AFA), commercialized as Klamin®, was subjected to chemical analysis to determine its compounds. The AFA extract Klamin® resulted to be nontoxic, also at high doses, when administered onto LAN5 neuronal cells. Its scavenging properties against ROS generation were evaluated by using DCFH-DA assay, and its mitochondrial protective role was determined by JC-1 and MitoSOX assays. Klamin® exerts a protective role against beta amyloid- (Aß-) induced toxicity and against oxidative stress. Anti-inflammatory properties were demonstrated by NFßB nuclear localization and activation of IL-6 and IL-1ß inflammatory cytokines through ELISA. Finally, by using thioflavin T (ThT) and fluorimetric measures, we found that Klamin® interferes with Aß aggregation kinetics, supporting the formation of smaller and nontoxic structures compared to toxic Aß aggregates alone. Altogether, these data indicate that the AFA extract may play a protective role against mechanisms leading to neurodegeneration.

Seminal plasma affects the survival rate and motility pattern of raw llama spermatozoa.

The objectives of this study were to evaluate the effect over time of different percentages of seminal plasma (SP) on llama sperm characteristics in raw semen and correlate the techniques routinely used to evaluate sperm viability and acrosome status with the Fluorescein Isothiocyanate -Arachis hypogea agglutinin/Propidium Iodide (FITC-PNA/PI). Eighteen ejaculates, obtained from 6 male llamas using electroejaculation, were incubated in 0.1% collagenase in HEPES-TALP (HT), centrifuged and resuspended with SP and HT: 0, 10, 50 and 100% SP. Samples were incubated (37 °C) until evaluation at 0; 1.5 and 3 h. Split plot and factorial designs were used to analyze sperm motility, viability, membrane function and acrosome status and Spearman's test was used for correlation. At 0 h, samples with 100% SP showed oscillatory motility; whereas in samples with 0 and 10% SP, progressive motility was predominant. Viability, membrane function and total motility decreased significantly at 3 h of incubation in samples with 100% SP. Sperm with intact acrosomes were fewer in 0% SP media at all times. FITC-PNA/PI correlated with 6-Carboxyfluorescein Diacetate and Propidium Iodide (CFDA/PI) and with Coomassie Blue (CB) stains (r = 0.8; p = 0.0 and r = 0.5; p = 0.0 respectively). CONCLUSIONS: the motility pattern of llama sperm is influenced by the concentration of SP. The use of SP as the only medium is not able to maintain sperm motility, viability and membrane function for 3 h. A certain percentage of SP is necessary in the medium to avoid spontaneous acrosome reactions. The correlations observed could help to shorten evaluation times and reduce costs in sperm laboratories.

Morphology, morphometry and chromatin distribution in llama sperm nuclei.

The objectives of this study were as follows: (i) to describe and evaluate the frequencies of different morphologies of llama sperm nuclei, (ii) to determine morphometric values of nuclear parameters, (iii) to describe and estimate the frequencies of different classes of chromatin distribution and (iv) to measure haploid DNA content and analyse its nuclear distribution. The study was performed using ejaculates collected from seven males, and sperm nuclei were stained with the Feulgen reaction. Normal morphology ranged from 78.36% to 93.92%, and abnormalities included short, small, large, pyriform, narrow, micro and round nuclei. Important differences in nuclei considered normal were found between some males. The following average values were obtained for each sperm nuclear morphometric parameter analysed: area 11.64 µm2 , perimeter 13.16 µm, length 5.12 µm, width 2.81 µm, ellipticity 1.85 and form 0.83. Differences between males were significant for all the parameters (p < .01). Light microscope observations and cytophotometric determinations allowed discriminating between three classes of chromatin distribution: homogeneous, diffuse and showing a clear band. Significant differences between males were found for the frequencies of the three classes (p < .01). Cluster analysis methods were used to estimate the resemblance between males according to the characteristics of their sperm nuclei. A great intermale variability was found for morphological, morphometric and chromatin distribution data. These parameters would have low dependence between them.

Bioactive compounds from extra virgin olive oils: Correlation between phenolic content and oxidative stress cell protection.

When compared with other edible vegetable oils, the extra virgin olive oil (EVOO) exhibits excellent nutritional properties due to the presence of biophenolic compounds. Although they constitute only a very small amount of the unsaponifiable fraction of EVOO, biophenols strongly contribute to the sensorial properties of this precious food conferring it, for example, the bitter or pungent taste. Furthermore, it has been found that biophenols possess beneficial effects against many human pathologies such as oxidative stress, inflammation, cardiovascular diseases, cancer and aging-related illness. In the present work, the biophenolic content of 51 Italian and Spanish EVOOs was qualitatively and quantitatively identified and their antioxidant ability analyzed by oxygen radical absorbance capacity (ORAC) assay. Results indicated that the maximum relationship can be found if the ORAC value is correlated with the concentration of the large family composed by ligstroside and oleuropein derivatives together with their degradation products, hydroxytyrosol and tyrosol. Then, selected biophenolic extracts were tested in NIH-3T3 cell line to verify their ability in the recovery of the oxidative stress revealed by DCFH-DA assay. Results were linearly correlated with the concentration of ligstroside aglycone (aldehyde and hydroxyl form).

Bone marrow stem cell applied in the canine veterinary clinics / Células tronco de medula óssea utilizadas na clínica veterinária canina

Cell therapy represents an old therapeutic practice initiated with the transfusion of whole blood in different clinical situations. There is now a breakthrough in the study of multipotent stem cell therapy because of its functionality in regeneration of tissues, which promotes attention of the scientific community. Bone marrow is one of the main sources of multipotent stem cells, composed by hematopoietic stem cells responsible for the renewal of the cellular components of the blood, and mesenchymal stem cells that aid in the regeneration of tissues. These cells have a strong potential for the treatment of several diseases, due their main characteristics such as high plasticity, capacity for self-renewal and immunomodulation. Although, there are many studies that show good results with the use of cell therapy as a form of treatment for several diseases, some studies still show inconclusive or unsatisfactory results. Therefore, the objective of this study was to review the application of bone marrow stem cells in the canine model since improvements on the knowledge of the technique are necessary to enable its applicability with safety and efficacy.(AU)

A terapia celular representa uma antiga prática terapêutica iniciada com a transfusão de sangue total em diferentes situações clínicas. Atualmente há um avanço no estudo da terapia com células-tronco mesenquimais por conta de sua funcionalidade na regeneração de tecidos, o que promove uma crescente atenção do meio científico. A medula óssea é uma das principais fontes de células-tronco multipotentes, no qual se encontram as células-tronco hematopoiéticas, responsável pela renovação dos componentes celulares do sangue, e as células-tronco mesenquimais que auxiliam na regeneração de tecidos. Essas células têm um forte potencial para o tratamento de diversas enfermidades, uma vez que possuem como principais características alta plasticidade, capacidade de auto renovação e imunomodulação. Apesar de haver muitos trabalhos que apresentam bons resultados com a utilização da terapia celular como forma de tratamento para diversas enfermidades, alguns estudos ainda demonstram resultados inconclusivos ou não satisfatórios, por isso, objetivou-se com este trabalho revisar a aplicação das células-tronco derivadas da medula óssea no modelo canino uma vez que é necessário melhorias sobre o conhecimento da técnica para que possibilite a sua aplicabilidade com segurança e eficácia.(AU)

Observed volatilization fluxes of S-metolachlor and benoxacor applied on soil with and without crop residues.

Volatilization may represent a major dissipation pathway for pesticides applied to soils or crops, and these losses may be modified by soil surface conditions or in the presence of plant residues. This paper investigates the effect of surface conditions on volatilization through experimental results. The two experiments consisted of volatilization flux measurements for 3 days after an application of S-metolachlor together with benoxacor: one with two wind tunnels to compare the effect of the presence of crop residues on the soil on volatilization losses and another one at the field scale from bare soil without crop residues. Volatilization fluxes were large immediately after application (between 77 and 223 ng m-2 s-1 for S-metolachlor depending on experimental conditions), decreasing down to a few nanograms per square meter per second on the last day. Volatilization fluxes followed a diurnal cycle driven by environmental conditions. The losses found for both compounds were in accordance with their physicochemical properties. The crop residue on the soil surface modified soil surface conditions-primarily the soil water content essentially, the degradation of S-metolachlor, and the dynamics of volatilization loss.

Comparison of two cooling protocols for llama semen: with and without collagenase and seminal plasma in the medium.

Seminal plasma (SP) of South American Camelids could interfere with the interaction of spermatozoa with the extenders; therefore it becomes necessary to improve semen management using enzymatic treatment. Our objective was to compare two cooling protocols for llama semen. Twelve ejaculates were incubated in 0.1% collagenase and then were divided into two aliquots. One was extended in lactose and egg yolk (LEY) (Protocol A: collagenase and SP present). The other aliquot was centrifuged, and the pellet was resuspended in LEY (Protocol B: collagenase and SP absent). Both samples were maintained at 5°C during 24 hr. Routine and DNA evaluations were carried out in raw and cooled semen. Both cooling protocols maintained sperm viability, membrane function and DNA fragmentation, with Protocol A showing a significantly lowered total and progressive motility (p < .05) and Protocol B showing a significant increase in chromatin decondensation (p < .05). Protocol A avoids centrifugation, reducing processing times and making application in the field simpler. However, as neither protocol showed a significant superiority over the other, studies should be carried out in vivo to evaluate the effect on pregnancy rates of the presence of collagenase and SP in semen samples prior to either cooling or freeze-thawing.

Efficacy of ultrasound-guided fine-needle aspiration performed by surgeons newly trained in thyroid ultrasound.

BACKGROUND: There are a growing number of thyroid ultrasound courses to train endocrinologists, pathologists, and surgeons to perform ultrasound-guided fine-needle aspiration (FNA). However, there are limited data to support the efficacy of ultrasound-guided FNA performed by nonradiologists. METHODS: We compared the efficacy of ultrasound-guided FNA performed by surgeons newly trained in thyroid ultrasound with that of the same technique performed by 1 experienced radiologist. The ratio of nondiagnostic examinations was used to compare the 2 groups. RESULTS: A total of 197 ultrasound-guided FNAs were performed on 172 women (95.53%) and 8 men (4.47%) over a 12-month period. The efficacy of ultrasound-guided FNA did not differ between groups. A nondiagnostic report was found in 23% and 25.7% of the examinations performed by the radiologist and surgeons, respectively. CONCLUSION: The efficacy of ultrasound-guided FNAs performed by our 2 surgeons was similar to that of our radiologist. © 2016 Wiley Periodicals, Inc. Head Neck 39: 439-442, 2017.

Pretreatment HIV-1 drug resistance in Argentina: results from a surveillance study performed according to WHO-proposed new methodology in 2014-15.

BACKGROUND: In Argentina, current national guidelines recommend starting with NNRTI-based regimens. Recently, there have been some local reports regarding concerning levels of NNRTI-transmitted resistance, but surveillance has never been carried out at a national level. OBJECTIVES: To determine the prevalence of HIV drug resistance in people starting ART in Argentina using a WHO-proposed methodology. METHODS: This was a cross-sectional, nationally representative study. Twenty-five antiretroviral-dispensing sites throughout the country were randomly chosen to enrol at least 330 persons starting ART, to generate a point prevalence estimate of resistance-associated mutations (RAMs) with a 5% CI (for the total population and for those without antiretroviral exposure). All consecutive patients older than 18 years starting or restarting ART in the chosen clinics were eligible. Samples were processed with Trugene and analysed using the Stanford algorithm. RESULTS: Between August 2014 and March 2015, we obtained 330 samples from people starting ART. The mean ±â€ŠSD age was 35 ±â€Š11 years, 63.4% were male, 16.6% had prior antiretroviral exposure and the median (IQR) CD4 count was 275 cells/mm3 (106-461). The prevalence of RAMs found was 14% (±4%) for the whole population (3% NRTI-RAMs; 11% NNRTI-RAMs and 2% PI-RAMs) and 13% (±4%) for those without prior antiretroviral exposure (3%, 10% and 2%, respectively). The most common mutation was K103N. CONCLUSIONS: This surveillance study showed concerning levels of HIV drug resistance in Argentina, especially to NNRTIs. Due to this finding, Argentina's Ministry of Health guidelines will change, recommending performing a resistance test for everyone before starting ART. If this is taken up properly, it also might function as a continuing surveillance tool.

Comparison of differents methods of sperm selection of llama raw semen.

The objective of this study was to compare the efficiency of different sperm selection methods applied to the same llama ejaculate. Four treatments were compared: two variants of the swim up technique (with and without seminal plasma), and two different colloids, Androcoll-E-Large and Percoll(®). Using electroejaculation, 21 semen samples were obtained from 7 llama males (n=7, r=3). The ejaculates were incubated in a solution of 0.1% collagenase, to decrease thread formation, and then split into 4 aliquots: one aliquot was layered over a column of Androcoll-E-Large (SLC) and the second over a column of Percoll (45%). The third aliquot was deposited in a tube with culture medium and was incubated at a 45° angle for 30min at 37°C (SU1). The last aliquot was centrifuged to separate the spermatozoa and seminal plasma. The sperm pellet obtained was resuspended, and transferred to a tube with culture medium which was incubated at an angle of 45° for 30min at 37°C (SU2). Both aliquots SLC and P showed higher proportions of progressive motility and plasma membrane functionality (p≤0.05) than raw semen. There were no significant differences (p>0.05) in sperm viability and in normal spermatozoa between raw semen and treatments. Nevertheless, only SLC did not have a significant increase of bent tails. In conclusion SLC centrifugation would be the method of choice for selecting llama spermatozoa.

Correlation and comparison of immunohistochemistry for HER2/neu, using the antibody SP3 and chromogenic in situ hybridization in breast carcinomas samples / Correlação e comparação de imuno-histoquímica para HER2/neu, utilizando o anticorpo SP3 com a hibridização in situ cromógena em amostras de carcinomas mamários

ABSTRACT Introduction: Advances in the field of molecular biology have provided the differentiation of molecular subtypes of breast tumors, providing better prognosis and important tools for the treatment of patients with breast cancer. Among these subtypes, the changes in the human epidermal growth factor receptor 2 gene (HER2/neu), increase its copy number and generating HER2 protein amplification. Studies show that patients with breast cancer HER2/neu amplified tend to relapse earlier and have shorter survival time, the monoclonal antibody Trastuzumab is the therapy indicated. The eligibility of patients for therapy is initially made by the immunohistochemistry (IHC) technique, which evaluates the expression level of the HER2 protein. After this evaluation, the cases with equivocal diagnosis (score 2+), are referred to a more accurate technique, the chromogenic in situ hybridization (CISH). Objective: To analyze the sensitivity and specificity of the antibody SP3, and determine their level of agreement with the CISH technique. Material and methods: Retrospective study in the database of the anatomy-pathology laboratory, in CISH tests reports for HER2/neu. Conclusion: The results revealed that clone SP3 showed 100% specificity and 92% sensitivity. IHC reveals variability in its results; however, it is known that the technique is an important tool in the daily routine of laboratories, contributing to the initial screening of patients with breast cancer, which later showed satisfactory results when compared with the CISH technique.

RESUMO Introdução: Avanços no campo da biologia molecular têm proporcionado a diferenciação dos subtipos moleculares das neoplasias mamárias, fornecendo melhor prognóstico e ferramentas importantes para a terapêutica de pacientes com câncer de mama. Entre esses subtipos, as alterações ocorridas no gene receptor tipo 2 do fator de crescimento epidérmico humano (HER2/neu) amplificam o seu número de cópias e geram o aumento da proteína HER2. Estudos mostram que pacientes portadoras de câncer de mama HER2/neu amplificado tendem a ter recaída mais cedo e tempo de sobrevida menor, sendo o anticorpo monoclonal Trastuzumab a terapia indicada. A elegibilidade das pacientes para a terapia é feita inicialmente pela técnica de imuno-histoquímica (IHQ), que avalia o nível de expressão da proteína HER2. Após essa avaliação, os casos que apresentam diagnósticos equívocos (escore 2+) são encaminhados para uma técnica mais precisa, a hibridização cromógena in situ (CISH). Objetivo: Analisar a sensibilidade e a especificidade do anticorpo SP3, além de determinar o seu nível de concordância com a técnica de CISH. Material e métodos: Estudo retrospectivo no banco de dados de um laboratório anatomopatológico, em laudos de exames de CISH para HER2/neu. Conclusão: Os resultados revelaram que o clone SP3 apresentou 100% de especificidade e 92% de sensibilidade. A IHQ revela variabilidade em seus resultados, porém é sabido que a técnica é uma importante ferramenta na rotina diária dos laboratórios, contribuindo na triagem inicial das pacientes portadoras de câncer de mama, que, posteriormente, mostram resultados satisfatórios quando comparados com a técnica de CISH.

Evaluation of the acrosomal status in Lama glama sperm incubated with acrosome reaction inducers.

UNLABELLED: The objectives of this study were to evaluate the effect of different acrosome reaction (AR) inducers on viability and acrosomal status in llama spermatozoa, by using the FITC-PNA/PI technique and evaluate if there is a positive correlation between the FITC-PNA/PI and the Coomassie blue (CB) staining techniques. After incubating twenty ejaculates in 0.1% collagenase the centrifuged pellets were resuspended in TALP-BSA medium. An aliquot was sonicated to remove the acrosomal content (positive control). The rest of the sample was incubated for 3h at 38 °C with 5% CO2 and 100% humidity. TREATMENTS: Three aliquots were further incubated 1h with one of the following AR inducers: calcium ionophore, ionomycin or progesterone. CONTROLS: One without inducers and the other, incubated with dimethyl sulfoxide (vehicle of the inducing agents). Acrosomes were evaluated at time 0 and after 4h incubation. Calcium ionophore was the most potent agent for inducing the AR (67.2 ± 14.4% live+dead AR sperm) (P < 0.05). These samples showed no motility and viability was very low (0-30%). Both ionomycin and progesterone presented significantly higher (P < 0.05) percentages of total AR sperm than the controls, but had similar percentages of dead reacted sperm to the controls. A positive correlation was observed between the intact acrosome FITC-PNA/PI pattern (live+dead sperm) and the acrosome-present CB pattern (r = 0.64; P = 0.000) in all the evaluated samples. CONCLUSIONS: the FITC-PNA/PI technique simultaneously evaluates viability and acrosomal status in llama spermatozoa and calcium ionophore could be used as a control of AR.