Laminin alpha 5 regulates mammary gland remodeling through luminal cell differentiation and Wnt4-mediated epithelial crosstalk

Epithelial attachment to the basement membrane (BM) is essential for mammary gland development, yet the exact roles of specific BM components remain unclear. Here, we show that Laminin alpha 5 (Lama5) expression specifically in the luminal epithelial cells is necessary for normal mammary gland growth during puberty, and for alveologenesis during pregnancy. Lama5 loss in the keratin 8-expressing cells results in reduced frequency and differentiation of hormone receptor expressing (HR+) luminal cells. Consequently, Wnt4-mediated crosstalk between HR+ luminal cells and basal epithelial cells is compromised during gland remodeling, and results in defective epithelial growth. The effects of Lama5 deletion on gland growth and branching can be rescued by Wnt4 supplementation in the in vitro model of branching morphogenesis. Our results reveal a surprising role for BM-protein expression in the luminal mammary epithelial cells, and highlight the function of Lama5 in mammary gland remodeling and luminal differentiation.Peer reviewe

Punasoluvalmisteiden lämpötilaseuranta

Opinnäytetyön aiheena oli punasoluvalmisteiden lämpötilaseuranta. Työssä tutustutaan vereen, verensiirtoihin, punasoluvalmisteisiin sekä punasoluvalmisteiden lämpötilaseurantaan säilytyksen ja kuljetuksen aikana. Työ toteutettiin yhteistyössä HUSLABin ja Suomen Punaisen Ristin Veripalvelun kanssa. Työn tavoitteena oli laatia HUSLAB-ohjeistus Helsingin kolmelle kotisairaalalle verensiirtovalmisteiden tilaamista ja käsittelyä varten. Ennen ohjeistuksen laatimista tutustuttiin HUSLABin Meilahden verikeskuksen toimintaan ja seurattiin fyysisesti mukana verikeskuksesta kotisairaaloihin lähteviä verivalmistekuljetuksia. Kuljetusten havainnoinnin pohjalta HUSLABille raportoitiin valmisteiden kuljetusten ja käsittelyn senhetkinen tila sekä kehitystarpeet uutta ohjeistusta ajatellen. Veripalvelussa tehtiin tutkimuksia, joissa punasoluvalmisteita altistettiin erilaisille ulkoisille lämpötiloille ja seurattiin niiden lämpenemistä ja viilenemistä. Veripalvelussa tehdyissä tutkimuksissa testattiin myös pakkausmenetelmien toimivuutta helle- ja pakkasolosuhteissa. Työn teoriaosuus toteutettiin kirjallisuuskatsauksena ja käytännön osuus kokeina Veripalvelussa sekä prosessinkehityksenä HUSLABille. Teoriaosuudessa tutustuttiin punasoluvalmisteille kansainvälisesti määriteltyihin säilytysvaatimuksiin, kuljetuksenaikaisiin pakkausmenetelmiin sekä erilaisiin käytössä oleviin lämpötilaseurantateknologioihin. Teoriatietoja voitiin hyödyntää fyysistä seurantaa arvioitaessa. Käytännön tutkimusosuuden perusteella HUSLABin nykyiset kuljetusmenetelmät ovat riittä-vät turvaamaan verivalmisteiden oikeat kuljetuslämpötilat. Verivalmisteiden käsittely kotisai-raaloissa vaihteli. Osassa kotisairaaloista oli tarvetta tarkistaa käsittelytapoja. Opinnäytetyön tuloksena laadittu uusi ohjeistus korostaa oikeita toimintatapoja ja yhtenäistää valmisteiden käsittelymenetelmiä kotisairaaloissa.The subject of this thesis was temperature monitoring of red blood cell products. The thesis focuses on blood, blood transfusions, red blood cell products and their temperature monitoring during storage and transport. The thesis was carried out in cooperation with HUSLAB and The Finnish Red Cross Blood Services. The objective of the thesis was to compose HUSLAB instructions for the ordering and handling of blood transfusion products for three home hospitals in Helsinki. The operation of HUSLAB blood center and blood transports to home hospitals were followed before the composition of the instructions. Based on the observations, a report was made for HUSLAB about the state of blood product transports and handling. The report also included development needs to take into account when composing the new instructions. At the premises of Red Cross Blood Services, red blood cell products were predisposed to different external temperatures and their heating up and cooling down was monitored. The research also evaluated how well the blood product packaging endured heat and frost. The theoretical part of this thesis was carried out as a literature research and the practical part as tests in the Blood Services and as a process development for HUSLAB. In the theoretical part, internationally defined storage requirements, packaging procedures and different temperature monitoring technologies were studied. The procedures witnessed during the observation could then be evaluated based on the theory. The findings show that the HUSLAB transport methods of blood products are adequate to guarantee safe transport temperatures for blood. The handling of blood products in the home hospitals varied. In some of the home hospitals, there was a need for reviewing the handling process. The new HUSLAB instructions composed based on the results of this thesis highlight the proper procedures and unify the handling process in the home hospitals

Characterization of a novel OX40 ligand and CD40 ligand-expressing oncolytic adenovirus used in the PeptiCRAd cancer vaccine platform

Oncolytic viruses (OVs) have been shown to induce anti-cancer immunity and enhance cancer immunotherapies, such as immune checkpoint inhibitor therapies. OV therapies can be further improved by arming OVs with immunostimulatory molecules, including various cytokines or chemokines. Here, we have developed a novel adenovirus encoding two immunostimulatory molecules: cluster of differentiation 40 ligand (CD40L) and tumor necrosis factor receptor superfamily member 4 ligand (OX40L). This novel virus, designated VALOD-102, is designed to activate both innate and adaptive immune responses against tumors. CD40L affects the innate side by licensing antigen-presenting cells to drive CD8(+) T cell responses, and OX40L increases clonal expansion and survival of CD8(+) T cells and formation of a larger pool of memory T cells. VALO-D102 and its murine surrogate VALO-mD901, expressing murine OX40L and CD40L, were used in our previously developed PeptiCRAd cancer vaccine platform. Intratumoral administration of PeptiCRAd significantly increased tumor-specific T cell responses, reduced tumor growth, and induced systemic anti-cancer immunity in two mouse models of melanoma. In addition, PeptiCRAd therapy, in combination with anti-PD-1 immune checkpoint inhibitor therapy, significantly improved tumor growth control as compared to either monotherapy alone.Peer reviewe

Development of mesothelioma-specific oncolytic immunotherapy enabled by immunopeptidomics of murine and human mesothelioma tumors

Abstract Malignant pleural mesothelioma (MPM) is an aggressive tumor with a poor prognosis. As the available therapeutic options show a lack of efficacy, novel therapeutic strategies are urgently needed. Given its T-cell infiltration, we hypothesized that MPM is a suitable target for therapeutic cancer vaccination. To date, research on mesothelioma has focused on the identification of molecular signatures to better classify and characterize the disease, and little is known about therapeutic targets that engage cytotoxic (CD8+) T cells. In this study we investigate the immunopeptidomic antigen-presented landscape of MPM in both murine (AB12 cell line) and human cell lines (H28, MSTO-211H, H2452, and JL1), as well as in patients’ primary tumors. Applying state-of-the-art immuno-affinity purification methodologies, we identify MHC I-restricted peptides presented on the surface of malignant cells. We characterize in vitro the immunogenicity profile of the eluted peptides using T cells from human healthy donors and cancer patients. Furthermore, we use the most promising peptides to formulate an oncolytic virus-based precision immunotherapy (PeptiCRAd) and test its efficacy in a mouse model of mesothelioma in female mice. Overall, we demonstrate that the use of immunopeptidomic analysis in combination with oncolytic immunotherapy represents a feasible and effective strategy to tackle untreatable tumors