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Chemotherapy-refractive multiple myeloma (MM) is serious and life-threatening, and better treatments are urgently needed. BCMA is a prominent marker on the cell surface of MM cells, rendering it an accepted target for antibody therapy. Considering that MM is a liquid tumor and immunotherapy has enjoyed success against leukemia, we devise an approach designed to enhance NK cell activity against MM. Ordinarily, NK cells function to naturally survey the body and eliminate malignant cells. Our platform approach is designed to enhance NK function. A tri-specific immune-engaging TriKE is manufactured, consisting of a camelid nanobody VHH antibody fragment recognizing CD16 expressed on NK cells and an scFv antibody fragment specifically recognizing BCMA. These two fragments are crosslinked by the human cytokine interleukin-15 (IL-15) known to have prominent activating effects on NK cells. The molecule, when tested by flow cytometry, shows activation of NK cells in their numbers and activity. Additionally, the molecule demonstrates anti-cancer effects in an in vivo xenograft model of human MM. We believe that the drug will have the capability of enhancing NK cells at the site of the immune synapse, i.e., the effector:target cell interface, and this will promote cancer remissions.

Simple Summary HER2 is a marker known to be over-expressed on breast cancer, rendering it one of the most useful solid tumor targets for antibody-based therapies. Despite expression on ovarian cancer, results targeting HER2 in this setting have been disappointing, thus requiring more aggressive approaches. Natural killer (NK) cells are known as principal mediators of cancer cell killing, but cancer cells find ways to deter them. We devised a tri-specific biological drug containing antibody fragments that simultaneously binds NK cells and cancer cells and at the same time delivers a natural cytokine signal that triggers robust NK cell expansion. In vitro studies show the drug augments NK cell killing of a number of HER2-positive human cell lines, while enhancing NK cell activation and proliferation. Studies in mice engrafted with human ovarian cancer showed the drug has anti-tumor efficacy, clearly demonstrating its ability to bolster NK cells in their ability to contain tumor cell growth. Abstract Clinical studies validated antibodies directed against HER2, trastuzumab, and pertuzumab, as useful methodology to target breast cancer cases where HER2 is expressed. The hope was that HER2 targeting using these antibodies in ovarian cancer patients would prove useful as well, but clinical studies have shown lackluster results in this setting, indicating a need for a more comprehensive approach. Immunotherapy approaches stimulating the innate immune system show great promise, although enhancing natural killer (NK) function is not an established mainstream immunotherapy. This study focused on a new nanobody platform technology in which the bispecific antibody was altered to incorporate a cytokine. Herein we describe bioengineered CAM1615HER2 consisting of a camelid VHH antibody fragment recognizing CD16 and a single chain variable fragment (scFv) recognizing HER2 cross-linked by the human interleukin-15 (IL-15) cytokine. This tri-specific killer engager (TriKETM) showed in vitro prowess in its ability to kill ovarian cancer human cell lines. In addition, we demonstrated its efficacy in inducing potent anti-cancer effects in an in vivo xenograft model of human ovarian cancer engrafting both cancer cells and human NK cells. While previous approaches with trastuzumab and pertuzumab faltered in ovarian cancer, the hope is incorporating targeting and cytokine priming within the same molecule will enhance efficacy in this setting.

Select subsets of immune effector cells have the greatest propensity to mediate antitumor responses. However, procuring these subsets is challenging, and cell-based immunotherapy is hampered by limited effector-cell persistence and lack of on-demand availability. To address these limitations, we generated a triple-gene-edited induced pluripotent stem cell (iPSC). The clonal iPSC line was engineered to express a high affinity, non-cleavable version of the Fc receptor CD16a and a membrane-bound interleukin (IL)-15/IL-15R fusion protein. The third edit was a knockout of the ecto-enzyme CD38, which hydrolyzes NAD+. Natural killer (NK) cells derived from these uniformly engineered iPSCs, termed iADAPT, displayed metabolic features and gene expression profiles mirroring those of cytomegalovirus-induced adaptive NK cells. iADAPT NK cells persisted in vivo in the absence of exogenous cytokine and elicited superior antitumor activity. Our findings suggest that unique subsets of the immune system can be modeled through iPSC technology for effective treatment of patients with advanced cancer. acceptedVersion

Simple Summary B7-H3 costimulatory ligand has sparked tremendous interest as a target for antibody-directed therapy of solid tumors. This is attributed to high expression of B7-H3 on the surface of tumor and low expression on normal tissues. Natural killer cells are known as key effectors of the innate immune system against cancer and are being explored clinically in a number of immunotherapy settings. Here, we describe a unique platform technology incorporating a cytokine as a bispecific antibody cross-linker to create a tri-specific NK cell engager, or TriKE, directed against the antigen B7-H3. The data shows this immunotherapeutic construct promotes specific natural killer cell expansion. It also induces antibody-dependent cellular cytotoxicity due to simultaneous recognition of both natural killer cells and B7-H3-expressing cancer cells. Finally, using a xenogeneic mouse model, the data shows that the TriKE induces better natural killer cell-mediated control of ovarian cancer. Taken together, the findings presented here indicate that a B7-H3-targeted TriKE has the potential to enhance natural killer cell immunotherapy in solid tumor settings and supports further development. Abstract We improved the bispecific antibody platform that primarily engages natural killer (NK) cells to kill cancer cells through antibody-dependent cellular cytotoxicity (ADCC) by adding IL-15 as a crosslinker that expands and self-sustains the effector NK cell population. The overall goal was to target B7-H3, an established marker predominantly expressed on cancer cells and minimally expressed on normal cells, and prove that it could target cancer cells in vitro and inhibit tumor growth in vivo. The tri-specific killer engager (TriKETM) was assembled by DNA shuffling and ligation using DNA encoding a camelid anti-CD16 antibody fragment, a wild-type IL-15 moiety, and an anti-B7-H3 scFv (clone 376.96). The expressed and purified cam1615B7H3 protein was tested for in vitro NK cell activity against a variety of tumors and in vivo against a tagged human MA-148 ovarian cancer cell line grafted in NSG mice. cam1615B7H3 showed specific NK cell expansion, high killing activity across a range of B7-H3+ carcinomas, and the ability to mediate growth inhibition of aggressive ovarian cancer in vivo. cam1615B7H3 TriKE improves NK cell function, expansion, targeted cytotoxicity against various types of B7-H3-positive human cancer cell lines, and delivers an anti-cancer effect in vivo in a solid tumor setting.

'Values' are sources that carry cultural heritage buildings from the past to the present and to the future. Each cultural heritage building has it’s own unique identity, character and value. Through values, society can understand its past and thus, the desire to preserve cultural heritage within the community arises. However, during conservation and repair interventions, some of these values are disregarded and a decrese of heritage values may occur while attempting to improve these cultural heritage buildings. On the other hand, depending on developing and changing life conditions, cultural heritage buildings may not meet today's needs, so to reactivate these buildings, adaptive reuse projects are realized for heritage buildings in historical areas. In Cyprus, which is very rich in cultural and heritage values, reuse projects have been rapidly increasing. On the other hand, a lot of old and historical buildings are being revitalized in the Historic Walled City of Nicosia with adaptive reuse projects. Although, efforts by the public and investors to reintroduce old heritage buildings, which are rapidly dissappearing to the city and the economy, are seen as a positive development, it should not be ignored that these interventions affect heritage values. In addition, the economic value is emphasized throughout these reuse projects and this causes changes in the authenticity of heritage buildings and changes in values and this causes a detriment to the identity of these buildings. In this study, it is aimed to understand how conservation intervention affects heritage values and the degree of change in their authenticity in the Walled City of Nicosia. To this end, eight cultural heritage buildings that adaptive reuse projects have been applied to in previous decades have been selected and their information was collected through area analysis and survey work in order to evaluate the heritage value and authenticity for each building. These evaluations and analyzes are based on theoretical knowledge derived from reuse works; this study includes the topics of the role of stakeholders, the degree of physical interventions, the appropriateness of the function, the impact of the legal process and the financial resources and their positive and negative effects on heritage values. Keywords: cultural heritage buildings, cultural values, authenticity, adaptive reuse, values-based approach ÖZ: ‘Değerler’ kültürel miras yapılarını geçmişten günümüze ve geleceğe taşıyan kaynaklardır. Her kültürel miras yapısının kendine özgü kimliği, özgün karakteri ve değerleri vardır. Değerler sayesinde toplum geçmişini anlar ve toplum içerisinde kültürel mirası koruma isteği oluşur. Ancak, yapılan koruma müdahaleleri sırasında bazı değerler gözetilmemekte ve kültürel miras yapıları iyileştirilirken miras değerlerinde kayıplar olabilmektedir. Öte yandan, gelişen ve değişen yaşam koşulları ile kültürel miras yapıları günümüz ihtiyaçlarını karşılayamamakta ve miras yapılarının tarihi bölgede tekrar aktif hale gelebilmesi için yeniden işlevlendime projeleri gündeme gelmektedir. Yeniden kullanım projeleri kültürel miras açısından zengin olan Kuzey Kıbrıs’ta zaman içerisinde giderek artmakta ve Tarihi Lefkoşa Suriçi’nde birçok tarihi yapı yeniden işlevlendirilerek hayat bulmaktadır. Yok olmaya yüz tutmuş kültürel miras yapılarının halk ve yatırımcılar tarafından tekarar kente ve ekonomiye kazandırma çabaları olumlu bir gelişme olarak görülse de, yapılan bu müdahalelerin miras değerlerini etkilediği göz ardı edilmemelidir. Ayrıca, ekonomik değerlerin ön plana çıktığı bu dönüşüm projelerinde miras yapılarının özgünlükleri değişmekte ve farklı değerler eklenerek tarihi yapıların kimlikleri zedelenebilmektedir. Bu çalışmada, Lefkoşa Suriçi’nde son on yıl içerisinde gerçekleştirilen koruma müdahalelerinin miras değerlerinin nasıl etkilediğini ve özgünlüklerinin ne derecede değiştiğini ortaya çıkarmak amaçlanmıştır. Bu amaca ulaşmak için, yeniden işlevlendirilmiş kültürel miras yapılarından sekiz tarihi konut yapısı seçilerek yapılan bölge analizleri ile anket çalışmaları yoluyla toplanan bilgiler incelenmiş ve bu yapıların miras değerleri ve özgünlükleri ayrı ayrı değerlendirilmiştir. Teorik bilgilerden elde edilen bir çerçeveye dayandırılan değerlendirmelere ve analizlere bağlı olarak yeniden işlevlendirme çalışmalarında; paydaşların rolü, yapılan fiziksel müdahalelerin dereceleri, verilen fonksiyonun uygunluğu, yasal süreçin etkileri ve finansal kaynaklar ele alınmış, bu etkenlerin miras değerleri üzerindeki olumlu ve olumsuz etkileri ortaya konmuştur. Anahtar Kelimeler: kültürel miras yapıları, kültürel değerler, özgünlük, yeniden işlevlendirme, değer-odaklı yaklaşım Master of Science in Urban Design. Thesis (M.S.)--Eastern Mediterranean University, Faculty of Architecture, Dept. of Architecture, 2019. Co-Supervisor: Assoc. Prof. Dr. Nevter Zafer Cömert and Assoc. Prof. Dr. Ege Uluca Tümer..

Purpose To develop a murine total marrow irradiation (TMI) model in comparison with the total body irradiation (TBI) model. Materials and methods Myeloablative TMI and TBI were administered in mice using a custom jig, and the dosimetric differences between TBI and TMI were evaluated. The early effects of TBI/TMI on bone marrow (BM) and organs were evaluated using histology, FDG-PET, and cytokine production. TMI and TBI with and without cyclophosphamide (Cy) were evaluated for donor cell engraftment and tissue damage early after allogeneic hematopoietic cell transplantation (HCT). Stromal derived factor-1 (SDF-1) expression was evaluated. Results TMI resulted in similar dose exposure to bone and 50% reduction in dose to bystander organs. BM histology was similar between the groups. In the non-HCT model, TMI mice had significantly less acute intestinal and lung injury compared to TBI. In the HCT model, recipients of TMI had significantly less acute intestinal injury and spleen GVHD, but increased early donor cell engraftment and BM:organ SDF-1 ratio compared to TBI recipients. Conclusions The expected BM damage was similar in both models, but the damage to other normal tissues was reduced by TMI. However, BM engraftment was improved in the TMI group compared to TBI, which may be due to enhanced production of SDF-1 in BM relative to other organs after TMI.