Hyperbaric storage at room temperature of watermelon juice as an alternative to refrigeration

Mestrado em Bioquímica, ramo Bioquímica AlimentarO armazenamento hiperbárico (AH) é um método de conservação alimentar que se baseia no uso da pressão como principal fator para retardar a ação microbiana e de degradação de qualidade de forma a aumentar o tempo de vida útil dos alimentos. O objetivo deste estudo centra-se na avaliação do AH à temperatura ambiente de sumo de melancia utilizando diferentes condições de pressão (0,1-100 MPa) a temperatura ambiente (≈21 °C) durante 10 dias de armazenamento. Esta avaliação foi realizada recorrendo a análises físico-químicas microbiológicas e bioquímicas. Tanto no sumo armazenado a 0,1 MPa e ≈21 °C como refrigerado a carga microbiológica aumentou para valores superiores a 6 Log UFC/mL para todos os microrganismos. Nas amostras a 50 MPa houve um aumento nos mesofilos aeróbios totais, Enterobacteriaceae e psicrófilos aeróbios totais e uma diminuição nos bolores e leveduras em ≈ 2 log UFC/mL. O AH a pressões de 75 e 100 MPa reduziu a carga microbiológica para valores ≤ 1 Log UFC/mL após 10 dias. A maioria dos parâmetros físicos-químicos manteve-se constante no AH a 50 e 100 MPa.). A 100 MPa obteve-se a maior concentração de licopeno em comparação com as outras condições de armazenamento. A atividade enzimática da peroxidase (POD) (40,63 ± 1,98) e da pectina metilesterase (PME) (43,65 ± 1,52) foi inferior no AH a 50 e 100 MPa enquanto a atividade enzimática da polifenoloxidase (PPO) foi superior nestas amostras (49,15 ± 6,32 e 54,20 ± 1,58 respetivamente). Foi realizado também um ensaio de AH utilizando sumo de melancia pasteurizado previamente por alta pressão a 600 MPa à ≈21 °C durante 6 minutos e armazenado a 50 MPa (à temperatura ambiente) durante 75 dias. Neste ensaio verificou-se que a carga microbiana manteve-se constante com valores ≤ 1 Log UFC/mL durante os 75 dias de armazenamento. Os parâmetros físico-químicos mantiveram-se constantes ao longo dos 75 dias de armazenamento em todas as condições de armazenamento. No caso da atividade enzimática após 75 dias de armazenamento a atividade da PPO, POD e PME nas amostras a 50 MPa foi reduzida em 26, 50 e 45% respetivamente verificando-se uma redução similar no caso das amostras refrigeradas (23, 45 e 40%, respetivamente). Por outro lado nas amostras armazenadas a 0,1 MPa observou-se uma redução superior de 45, 66 e 58% respetivamente logo após 15 dias de armazenamento. Com base nos resultados obtidos este método pode ser considerado como uma possível alternativa à refrigeração para a conservação de sumo de melancia.Hyperbaric Storage (HS) is a method of food preservation that relies on the use of pressure as the main factor to retard microbial and quality degradation in order to increase the food shelf life. The objective of this study is based on the HS at room temperature of watermelon juice using different pressure conditions (0.1-100 MPa) at room temperature (≈ 21 °C) for 10 days of storage. Microbiological physicochemical and biochemical analyses were carried out. In juice stored at 0.1 MPa and ≈21 °C and under refrigeration the microbiological load increased to values ≥ 6 Log CFU/mL for all microorganisms. In samples at 50 MPa there was an increase in total aerobic mesophiles, Enterobacteriaceae and total aerobic psychrophiles, and a decrease in yeasts and molds of about 2 log UFC. The HS at pressures of 75 and 100 MPa caused a reduction of the microbiological load to values ≤ 1 Log CFU/mL after 10 days. Most of the physicochemical parameters kept constant in the HS at 50 and 100 Mpa. At 100 MPa higher lycopeneconcentration was obtained which increased compared to the other storage conditions. The activities of peroxidase (POD) and pectin methylesterase (PME) were lower in HS at 50 and 100 MPa. While the enzymatic activity of polyphenoloxidases (PPO) was higher in these samples. A different HS assay was also performed using high pressure pasteurized watermelon juiceat 600 MPa at ≈21 ° C for 6 minutes and stored at 50 MPa (room temperature) for 75 days. In this assay it was verified that the microbial load remained constant with values ≤1 log CFU/mL during the 75 days of storage. The physicochemical parameters remained constant throughout the 75 days of storage in all storage. In the case of enzymatic activity after 75 days of storage the PPO, POD and PME activityes were reduced in the 50 MPa samples by 26, 50 and 45% respectively being similar to refrigerated samples (23, 45 and 40%, respectively). On the other hand samples stored at 0.1 MPa it was observed a higher reduction of about 45, 66 and 58% after 15 days Based on the obtained results this method can be considered as a possible alternative to refrigeration for the conservation of watermelon juice

Impact of different hyperbaric storage conditions on microbial, physicochemical and enzymatic parameters of watermelon juice

Hyperbaric storage (HS) of raw watermelon juice, up to 10days at 50, 75, and 100MPa at variable/uncontrolled room temperature (18-23°C, RT) was studied and compared with storage at atmospheric pressure (AP) under refrigeration (4°C, RF) and RT, being evaluated microbiological (endogenous and inoculated), physicochemical parameters, and enzymatic activities. Ten days of storage at 50MPa resulted in a microbial growth evolution similar to RF, while at 75/100MPa were observed microbial load reductions on endogenous and inoculated microorganisms (Escherichia coli and Listeria innocua, whose counts were reduced to below the detection limit of 1.00 log CFU/mL), resulting in a shelf-life extension compared to RF. The physicochemical parameters remained stable at 75MPa when compared to the initial raw juice, except for browning degree that increased 1.72-fold, whilst at 100MPa were observed higher colour variations, attributed to a lycopene content decrease (25%), as well as reductions on peroxidase residual activity (16.8%) after 10days, while both polyphenol oxidase and pectin methylesterase residual activities were similar to RF. These outcomes hint HS as a reliable alternative to RF as a new food preservation methodology, allowing energy savings and shelf-life extension of food products. This is the first paper studying the effect of HS on inoculated microorganisms and on a broad number of physicochemical parameters and on endogenous enzymatic activities, for a preservation length surpassing the shelf-life by RF

Two fish in a pod. Mislabelling on board threatens sustainability in mixed fisheries

Accuracy in reporting captures is a key element to achieve fisheries sustainability. However, identification of the catches might be a challenge when two or more species are morphologically similar and caught jointly, like the mixed fisheries of black hakes in East Atlantic African waters. Black hakes (Merluccius senegalensis and M. polli) are tough to differentiate without previous training due to their high morphological resemblance. The two species are managed as a single stock, although the biological differences between them suggest the need of a separate management. In this study, a total of 806 black hakes were visually identified by fishers on deck of fishing vessels operating in Mauritania and Senegal waters, then assigned to a species by sequencing 450bp of the Mitochondrial Control Region. Comparing the results with visual identification we found 31.4% of the total catch were incorrectly labelled on board by the fishermen. The accuracy of the fishers' identification depended on the depth of capture and on fish size, larger individuals caught from deeper waters being more correctly assigned to M. polli. Mislabelling biased to M. polli suggests that M. senegalensis, already catalogued as endangered, is being underreported, which could endanger the conservation of this species and threaten the sustainability of black hake fisheries. Our results highlight the need for separate evaluation of the stocks in mixed fisheries for morphologically similar fish. Thus, monitoring through DNA barcoding in the very first step of the seafood chain surveys would improve accurate species delimitation and reduce its impact on the correct assessment of the stocks.info:eu-repo/semantics/publishedVersio

Immune microenvironment characterisation and dynamics during anti-HER2-based neoadjuvant treatment in HER2-positive breast cancer

Despite their recognised role in HER2-positive (HER2+) breast cancer (BC), the composition, localisation and functional orientation of immune cells within tumour microenvironment, as well as its dynamics during anti-HER2 treatment, is largely unknown. We here investigate changes in tumour-immune contexture, as assessed by stromal tumour-infiltrating lymphocytes (sTILs) and by multiplexed spatial cellular phenotyping, during treatment with lapatinib-trastuzumab in HER2+ BC patients (PAMELA trial). Moreover, we evaluate the relationship of tumour-immune contexture with hormone receptor status, intrinsic subtype and immune-related gene expression. sTIL levels increase after 2 weeks of HER2 blockade in HR-negative disease and HER2-enriched subtype. This is linked to a concomitant increase in cell density of all four immune subpopulations (CD3+, CD4+, CD8+, Foxp3+). Moreover, immune contexture analysis showed that immune cells spatially interacting with tumour cells have the strongest association with response to anti-HER2 treatment. Subsequently, sTILs consistently decrease at the surgery in patients achieving pathologic complete response, whereas most residual tumours at surgery remain inflamed, possibly reflecting a progressive loss of function of T cells. Understanding the features of the resulting tumour immunosuppressive microenvironment has crucial implications for the design of new strategies to de-escalate or escalate systemic therapy in early-stage HER2+ BC

The remarkably low affinity of CD4/peptide-major histocompatibility complex class II protein interactions

The αβ T-cell co-receptor CD4 enhances immune responses more than one million-fold in some assays, and yet the affinity of CD4 for its ligand, peptide-major histocompatibility class II (pMHC II) on antigen-presenting cells, is so weak that it was previously unquantifiable. Here, we report that a soluble form of CD4 failed to bind detectably to pMHC II in surface plasmon resonance-based assays, establishing a new upper limit for the solution affinity at 2.5 mM. However, when presented multivalently on magnetic beads, soluble CD4 bound pMHC II-expressing B cells, confirming that it is active and allowing mapping of the native co-receptor binding site on pMHC II. Whereas binding was undetectable in solution, the affinity of the CD4/pMHC II interaction could be measured in two dimensions (2D) using CD4- and adhesion molecule-functionalized, supported lipid bilayers, yielding a 2D dissociation constant, Kd, of ~5000 molecules/μm2. This value is 2-3 orders of magnitude higher than previously measured 2D Kd values for interacting leukocyte surface proteins. Calculations indicated, however, that CD4/pMHC II binding would increase rates of T-cell receptor (TCR) complex phosphorylation by three-fold via the recruitment of Lck, with only a small, 2-20% increase in the effective affinity of the TCR for pMHC II. The affinity of CD4/pMHC II therefore appears to be set at a value that increases T-cell sensitivity by enhancing phosphorylation, without compromising ligand discrimination.This work was supported by the Wellcome Trust and the UK Medical Research Council. PJ was supported by grants from the Swedish Research Council (number: 623-2014- 6387 and 621-2014-3907). OD is supported by a Sir Henry Dale Fellowship jointly funded by the Wellcome Trust and the Royal Society (Grant Number: 098363)

SOLTI-1805 TOT-HER3 Study Concept: A Window-of-Opportunity Trial of Patritumab Deruxtecan, a HER3 Directed Antibody Drug Conjugate, in Patients With Early Breast Cancer

Background: Preclinical data support a key role for the human epidermal growth factor receptor 3 (HER3) pathway in hormone receptor (HR)-positive breast cancer. Recently, new HER3 directed antibody drug conjugates have shown activity in breast cancer. Given the need to better understand the molecular biology, tumor microenvironment, and mechanisms of drug resistance in breast cancer, we designed this window-of-opportunity study with the HER3 directed antibody drug conjugate patritumab deruxtecan (HER3-DXd; U3-1402). Trial Design: Based on these data, a prospective, multicenter, single-arm, window-of-opportunity study was designed to evaluate the biological effect of patritumab deruxtecan in the treatment of naïve patients with HR-positive/HER2-negative early breast cancer whose primary tumors are ≥1 cm by ultrasound evaluation. Patients will be enrolled in four cohorts according to the mRNA-based ERBB3 expression by central assessment. The primary endpoint is a CelTIL score after one single dose. A translational research plan is also included to provide biological information and to evaluate secondary and exploratory objectives of the study

HOPE (SOLTI-1903) breast cancer study: real-world, patient-centric, clinical practice study to assess the impact of genomic data on next treatment decision-choice in patients with locally advanced or metastatic breast cancer

Background Metastatic breast cancer (mBC) causes nearly all BC-related deaths. Next-generation sequencing (NGS) technologies allow for the application of personalized medicine using targeted therapies that could improve patients' outcomes. However, NGS is not routinely used in the clinical practice and its cost induces access-inequity among patients. We hypothesized that promoting active patient participation in the management of their disease offering access to NGS testing and to the subsequent medical interpretation and recommendations provided by a multidisciplinary molecular advisory board (MAB) could contribute to progressively overcome this challenge. We designed HOPE (SOLTI-1903) breast cancer trial, a study where patients voluntarily lead their inclusion through a digital tool (DT). The main objectives of HOPE study are to empower mBC patients, gather real-world data on the use of molecular information in the management of mBC and to generate evidence to assess the clinical utility for healthcare systems.Trial design After self-registration through the DT, the study team validates eligibility criteria and assists patients with mBC in the subsequent steps. Patients get access to the information sheet and sign the informed consent form through an advanced digital signature. Afterwards, they provide the most recent (preferably) metastatic archival tumor sample for DNA-sequencing and a blood sample obtained at the time of disease progression for ctDNA analysis. Paired results are reviewed by the MAB, considering patient's medical history. The MAB provides a further interpretation of molecular results and potential treatment recommendations, including ongoing clinical trials and further (germline) genetic testing. Participants self-document their treatment and disease evolution for the next 2 years. Patients are encouraged to involve their physicians in the study. HOPE also includes a patient empowerment program with educational workshops and videos about mBC and precision medicine in oncology. The primary endpoint of the study was to describe the feasibility of a patient-centric precision oncology program in mBC patients when a comprehensive genomic profile is available to decide on a subsequent line of treatment

Patritumab deruxtecan in HER2-negative breast cancer: part B results of the window-of-opportunity SOLTI-1805 TOT-HER3 trial and biological determinants of early response

Patritumab deruxtecan (HER3-DXd) exhibits promising efficacy in breast cancer, with its activity not directly correlated to baseline ERBB3/HER3 levels. This research investigates the genetic factors affecting HER3-DXd's response in women with early-stage hormone receptor-positive and HER2-negative (HR+/HER2-) breast cancer. In the SOLTI-1805 TOT-HER3 trial, a single HER3-DXd dose was administered to 98 patients across two parts: 78 patients received 6.4 mg/kg (Part A), and 44 received a lower 5.6 mg/kg dose (Part B). The CelTIL score, measuring tumor cellularity and infiltrating lymphocytes from baseline to day 21, was used to assess drug activity. Part A demonstrated increased CelTIL score after one dose of HER3-DXd. Here we report CelTIL score and safety for Part B. In addition, the exploratory analyses of part A involve a comprehensive study of gene expression, somatic mutations, copy-number segments, and DNA-based subtypes, while Part B focuses on validating gene expression. RNA analyses show significant correlations between CelTIL responses, high proliferation genes (e.g., CCNE1, MKI67), and low expression of luminal genes (e.g., NAT1, SLC39A6). DNA findings indicate that CelTIL response is significantly associated with TP53 mutations, proliferation, non-luminal signatures, and a distinct DNA-based subtype (DNADX cluster-3). Critically, low HER2DX ERBB2 mRNA, correlates with increased HER3-DXd activity, which is validated through in vivo patient-derived xenograft models. This study proposes chemosensitivity determinants, DNA-based subtype classification, and low ERBB2 expression as potential markers for HER3-DXd activity in HER2-negative breast cancer. Patritumab deruxtecan (HER3-DXd) is a promising therapy for breast cancer, targeting HER3. Here, the authors analyse the genomic factors that affect the response to HER3-DXd in patients with early-stage HER2-negative breast cancer as part of the SOLTI-1805 TOT-HER3 clinical trial and report outcomes for Part B of the trial using lower HER3-DXd dose in patients with HER2-negative breast cancer