A spectroscopic and molecular dynamics study on the aggregation process of a long-acting lipidated therapeutic peptide: the case of semaglutide

The aggregation properties of semaglutide, a lipidated peptide drug agonist of the Glucagon-like peptide 1 receptor recently approved for the treatment of type 2 diabetes, have been investigated by spectroscopic techniques (UV-Vis absorption, steady-state and time-resolved fluorescence, and electronic circular dichroism) and molecular dynamics simulations. We show that in the micromolar concentration region, in aqueous solution, semaglutide is present as monomeric and dimeric species, with a characteristic monomer-to-dimer transition occurring at around 20 μM. The lipid chain stabilizes a globular morphology of the monomer and dimer species, giving rise to a locally well-defined polar outer surface where the lipid and peptide portions are packed to each other. At very long times, these peptide clusters nucleate the growth of larger aggregates characterized by blue luminescence and a β-sheet arrangement of the peptide chains. The understanding of the oligomerization and aggregation potential of peptide candidates is key for the development of long acting and stable drugs

a propensity score analysis exploring the impact of the addition of adjuvant chemotherapy act to hormone therapy aht in a multi center series of resected luminal early stage pure invasive lobular breast carcinoma ilc

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Die Bedeutung von Rhamnolipiden in der Pathogenese epithelialer Pseudomonas aeruginosa Infektionen

Pseudomonas aeruginosa (P. aeruginosa) ist ein ubiquitär vorkommendes, gramnegatives, monotrich begeißeltes, aerob lebendes Stäbchenbakterium. Das opportunistisches Pathogen befällt Menschen, Tiere, Insekten, Nematoden und Pflanzen und ist unter anderem aufgrund seiner Toleranz gegenüber einer Vielzahl von Desinfektionsmitteln und Antibiotika einer der wichtigsten Verursacher von nosokomialen Infektionen. Die Ergebnisse dieser Arbeit geben einen Hinweis darauf, dass P. aeruginosa das angeborene Immunsystem mit Hilfe von Rhamnolipid, einem vom Bakterium selbst produzierten Surfactant, umgeht. In der vorliegenden Arbeit konnte nachgewiesen werden, dass die Flagellin-induzierte hBD2-Expression in humanen Keratinozyten durch bakterielle Überstände von P. aeruginosa, die in der stationären Phase generiert wurden, supprimiert wird. Der supprimierende Faktor konnte in Versuchen als hitzestabil und durch Säure ausfällbar identifiziert werden. Außerdem blieb er bei Behandlung mit Proteinase K unbeeinflusst, es handelt sich deshalb nicht um ein Protein. Weitere Untersuchungen konnten den Faktor als Rhamnolipid identifizieren. Eine Kostimulation von Keratinozyten mit aufgereinigtem Rhamnolipid in Mengen unterhalb der zytotoxischen Konzentration zusammen mit Flagellin zeigte ebenfalls eine Supprimierung der induzierten hBD2-Expression. Ähnliche Effekte konnten bei Kostimulation der Keratinozyten mit Flagellin und BAPTA-AM, einem intrazellulären Calciumchelator, erreicht werden. Neben Flagellin als Induktor ließ sich auch die PMA-induzierte hBD2-Expression supprimieren. Dies lässt vermuten, dass Rhamnolipide mit Calcium-abhängigen Signalkaskaden, wie der PKC-Signalkaskade, interferieren. Die Versuche dieser Arbeit zeigen damit einen Weg, über den es P. aeruginosa möglich ist, sich auf der Haut zu etablieren, ohne seinen Hauptpathogenitätsfaktor, das Flagellin, verstecken zu müssen

tumor infiltrating lymphocytes tils as an independent prognostic factor for early her2 breast cancer patients treated with adjuvant chemotherapy and trastuzumab in the randomized shorther trial

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Immune characterization of breast cancer metastases: prognostic implications.

BACKGROUND: Tumor-infiltrating lymphocytes (TILs) evaluated in primary breast cancer (BC) convey prognostic information. Limited data in the metastatic setting are available. METHODS: Secondary lesions from 94 BC patients, 43 triple-negative (TN) and 51 HER2-positive, were evaluated for TILs and expression of CD8, FOXP3, and PD-L1 by immunohistochemistry. RESULTS: TILs levels on metastasis were generally low (median 5%) and did not differ between TN and HER2+ tumors. Younger patients showed significantly lower TILs (p\u2009=\u20090.002). In HER2+ patients, TILs were higher in lung metastases as compared to other sites (p\u2009=\u20090.038). TILs composition was different across metastatic sites: skin metastases presented higher FOXP3 (p =\u20090.002) and lower CD8/FOXP3 ratio (p\u2009=\u20090.032). Patients treated for metastatic BC prior to biopsy had lower CD8 (overall: p\u2009=\u20090.005, HER2+: p\u2009=\u20090.011, TN: p\u2009=\u20090.075). In TN patients, median overall survival (OS) was 11.8 and 62.9 months for patients with low and high TILs, respectively (HR 0.29, 95%CI 0.11-0.76, log-rank p\u2009=\u20090.008). CD8/FOXP3 ratio was also prognostic in TN patients (median OS 8.0, 13.2, and 54.0 months in 1st, 2nd and 3th tertile, log-rank p\u2009=\u20090.019). Both TILs and CD8/FOXP3 ratio were independent factors at multivariate analysis. Counterintuitively, in HER2+ BC, low TILs tumors showed better prognosis (median OS 53.7 vs 39.9 months in TILs low and TILs high, not statistically significant). CONCLUSIONS: Our findings indicate the relevance of TILs as prognostic biomarker for TNBC even in the advanced setting and provide novel hypothesis-generating data on potential sources of immune heterogeneity of metastatic BC

Association of tumor-infiltrating lymphocytes with distant disease-free survival in the ShortHER randomized adjuvant trial for patients with early HER2+ breast cancer.

BACKGROUND: There is the need to identify new prognostic markers to refine risk stratification for HER2-positive early breast cancer patients. The aim of this study was to evaluate the association of tumor-infiltrating lymphocytes (TILs) with distant disease-free survival (DDFS) in patients with HER2-positive early breast cancer enrolled in the ShortHER adjuvant trial which compared 9 weeks versus 1-year trastuzumab in addition to chemotherapy, and to test the interaction between TILs and treatment arm. PATIENTS AND METHODS: Stromal TILs were assessed for 866 cases on centralized hematoxylin and eosin-stained tumor slides. The association of TILs as 10% increments with DDFS was assessed with Cox models. Kaplan-Meier curves were estimated for patients with TILs\u2009 6520% and TILs\u2009<20%. Median follow-up was 6.1\u2009years. RESULTS: Median TILs was 5% (Q1-Q3 1%-15%). Increased TILs were independently associated with better DDFS in multivariable model [hazard ratio (HR) 0.73, 95% confidence interval (CI) 0.59-0.89, P\u2009=\u20090.006, for each 10% TILs increment]. Five years DDFS rates were 91.1% for patients with TILs\u2009<20% and 95.7% for patients with TILs\u2009 6520% (P\u2009=\u20090.025). The association between 10% TILs increments and DDFS was significant for patients randomized to 9\u2009weeks of trastuzumab (HR 0.60, 95% CI 0.41-0.88) but not for patients treated with 1\u2009year of trastuzumab (HR 0.89, 95% CI 0.71-1.12; test for interaction P\u2009=\u20090.088). For patients with TILs\u2009<20%, the HR for the comparison between the short versus the long arm was 1.75 (95% CI 1.09-2.80, P=0.021); whereas, for patients with TILs\u2009 6520% the HR for the comparison of short versus long arm was 0.23 (95% CI 0.05-1.09, P\u2009=\u20090.064), resulting in a significant interaction (P\u2009=\u20090.015). CONCLUSIONS: TILs are an independent prognostic factor for HER2-positive early breast cancer patients treated with adjuvant chemotherapy and trastuzumab and may refine the ability to identify patients at low risk of relapse eligible for de-escalated adjuvant therapy