The inducible caspase-9 suicide gene system as a 'safety switch' to limit on-target, off-tumor toxicities of chimeric antigen receptor T cells

Published online: 28 October 2014.Immune modulation has become a central element in many cancer treatments, and T cells genetically engineered to express chimeric antigen receptors (CAR) may provide a new approach to cancer immunotherapy. Autologous CAR T cells that have been re-directed toward tumor-associated antigens (TAA) have shown promising results in phase 1 clinical trials, with some patients undergoing complete tumor regression. However, this T-cell therapy must carefully balance effective T-cell activation, to ensure antitumor activity, with the potential for uncontrolled activation that may produce immunopathology. An inducible Caspase 9 (iCasp9) "safety switch" offers a solution that allows for the removal of inappropriately activated CAR T cells. The induction of iCasp9 depends on the administration of the small molecule dimerizer drug AP1903 and dimerization results in rapid induction of apoptosis in transduced cells, preferentially killing activated cells expressing high levels of transgene. The iCasp9 gene has been incorporated into vectors for use in preclinical studies and demonstrates effective and reliable suicide gene activity in phase 1 clinical trials. A third-generation CAR incorporating iCasp9 re-directs T cells toward the GD2 TAA. GD2 is over-expressed in melanoma and other malignancies of neural crest origin and the safety and activity of these GD2-iCAR T cells will be investigated in CARPETS and other actively recruiting phase 1 trials.Tessa Gargett and Michael P. Brow

Higher order contributions to the effective action of N=2 super Yang-Mills

We apply heat kernel techniques in N=1 superspace to compute the one-loop effective action to order $F^5$ for chiral superfields coupled to a non-Abelian super Yang-Mills background. The results, when combined with those of hep-th/0210146, yield the one-loop effective action to order $F^5$ for any N=2 super Yang-Mills theory coupled to matter hypermultiplets.Comment: 23 pages, references adde

Cave deposits as a sedimentary trap for the Marine Isotope Stage 3 environmental record: The case study of Pod Hradem, Czech Republic

Pod Hradem Cave, located in the Moravian Karst, Czech Republic, offers an excellent opportunity for environmental reconstructions of Marine Isotope Stage 3 (MIS 3) in Central Europe due to its detailed sedimentary record dated 50,000 to 28,000 cal BP. Identifying the natural environments of the Middle to Upper Palaeolithic (MUP) transition is necessary to understand the settlement strategies and related behaviour of both Neanderthals and Anatomically Modern Humans, both of whom may have occupied the region at the same time. A multidisciplinary excavation was carried out between 2011 and 2016. Detailed analyses of the sediments, vertebrate microfauna, pollen and charcoal revealed minor but observable fluctuations in climate, with little change in the surrounding vegetation. The Pod Hradem palaeoenvironmental dataset is complex, but generally reflects a predominantly glacial climate with a range of vegetation types and habitats during the Late Pleistocene, followed by the warmer and more humid Holocene. The MUP transition as recorded in Pod Hradem Cave was a glacial environment interrupted by two relatively warmer periods. Central Europe experienced extreme climate fluctuations during MIS3, as recorded from different sedimentary archives, but it seems that the Pod Hradem Cave environment may have acted as a buffer zone, ameliorating those extremes, and providing a suitable refuge for both bears seeking winter hibernation dens and occasionally visiting humans.Thisproject was funded from the SoMoPro programme. Research leading tothese results has received a financial contribution from the EuropeanCommunity within the Seventh Framework Programme (FP/2007–2013) under Grant Agreement No. 229603. The research was alsoco-financed by the South Moravian Region and the Department ofAnthropology & Department of Geological Sciences (departmentalfunding - Masaryk University) and the internal programme of theInstitute of Geology CAS in Prague No. RVO 67985831

Cytolytic DNA vaccine encoding lytic perforin augments the maturation of- and antigen presentation by- dendritic cells in a time-dependent manner

The use of cost-effective vaccines capable of inducing robust CD8+ T cell immunity will contribute significantly towards the elimination of persistent viral infections and cancers worldwide. We have previously reported that a cytolytic DNA vaccine encoding an immunogen and a truncated mouse perforin (PRF) protein significantly augments anti-viral T cell (including CD8+ T cell) immunity. Thus, the current study investigated whether this vaccine enhances activation of dendritic cells (DCs) resulting in greater priming of CD8+ T cell immunity. In vitro data showed that transfection of HEK293T cells with the cytolytic DNA resulted in the release of lactate dehydrogenase, indicative of necrotic/lytic cell death. In vitro exposure of this lytic cell debris to purified DCs from naïve C57BL/6 mice resulted in maturation of DCs as determined by up-regulation of CD80/CD86. Using activation/proliferation of adoptively transferred OT-I CD8+ T cells to measure antigen presentation by DCs in vivo, it was determined that cytolytic DNA immunisation resulted in a time-dependent increase in the proliferation of OT-I CD8+ T cells compared to canonical DNA immunisation. Overall, the data suggest that the cytolytic DNA vaccine increases the activity of DCs which has important implications for the design of DNA vaccines to improve their translational prospects.Danushka K. Wijesundara, Wenbo Yu, Ben J. C. Quah, Preethi Eldi, John D. Hayball, Kerrilyn R. Diener, Ilia Voskoboinik, Eric J. Gowans, and Branka Grubor-Bau

A thin layer of phytoplankton observed in the Philippine Sea with a synthetic moored array of autonomous gliders

Author Posting. © American Geophysical Union, 2009. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Journal of Geophysical Research 114 (2009): C10020, doi:10.1029/2009JC005317.A synthetic moored array composed of five buoyancy-propelled autonomous underwater gliders was used to characterize mesoscale variability and phytoplankton distribution in a 100 km × 100 km domain in the Philippine Sea east of Luzon Strait for 10 days in May 2004. The study area, located east of the Kuroshio near the subtropical front, is dominated by strong internal tides, by energetic westward-propagating mesoscale eddies with azimuthal velocities exceeding 50 cm/s, and by a deep (130 m) maximum in chlorophyll fluorescence. Each glider in the array was instructed to maintain geographic position while repeatedly profiling to 200-m depth. Good station-keeping performance enabled the resulting series of vertical profiles to be interpreted in the same manner as a physically moored chain of instruments. Although organized primarily as a demonstration of glider capabilities, this field exercise provides a unique data set for examining biological-physical interactions in the open ocean. Here we report on the evolution of a thin layer of phytoplankton observed near the deep chlorophyll maximum. Coincident observations of fine structure in temperature and salinity suggest that the thinning process of this layer was driven primarily by physical forcing, most probably vertical shear associated with energetic diurnal internal waves, as opposed to a biological mechanism, such as convergent swimming, grazing, or spatial variation in growth rate.The Office of Naval Research provided support for fieldwork and analysis through grants N-00014-00-1-0256 and N-00014-05-1-0367

Endometrial and Menstrual Blood Mesenchymal Stem/Stromal Cells: Biological Properties and Clinical Application

A highly proliferative mesenchymal stem/stromal cell (MSC) population was recently discovered in the dynamic, cyclically regenerating human endometrium as clonogenic stromal cells that fulfilled the International Society for Cellular Therapy (ISCT) criteria. Specific surface markers enriching for clonogenic endometrial MSC (eMSC), CD140b and CD146 co-expression, and the single marker SUSD2, showed their perivascular identity in the endometrium, including the layer which sheds during menstruation. Indeed, cells with MSC properties have been identified in menstrual fluid and commonly termed menstrual blood stem/stromal cells (MenSC). MenSC are generally retrieved from menstrual fluid as plastic adherent cells, similar to bone marrow MSC (bmMSC). While eMSC and MenSC share several biological features with bmMSC, they also show some differences in immunophenotype, proliferation and differentiation capacities. Here we review the phenotype and functions of eMSC and MenSC, with a focus on recent studies. Similar to other MSC, eMSC and MenSC exert immunomodulatory and anti-inflammatory impacts on key cells of the innate and adaptive immune system. These include macrophages, T cells and NK cells, both in vitro and in small and large animal models. These properties suggest eMSC and MenSC as additional sources of MSC for cell therapies in regenerative medicine as well as immune-mediated disorders and inflammatory diseases. Their easy acquisition via an office-based biopsy or collected from menstrual effluent makes eMSC and MenSC attractive sources of MSC for clinical applications. In preparation for clinical translation, a serum-free culture protocol was established for eMSC which includes a small molecule TGFβ receptor inhibitor that prevents spontaneous differentiation, apoptosis, senescence, maintains the clonogenic SUSD2+ population and enhances their potency, suggesting potential for cell-therapies and regenerative medicine. However, standardization of MenSC isolation protocols and culture conditions are major issues requiring further research to maximize their potential for clinical application. Future research will also address crucial safety aspects of eMSC and MenSC to ensure these protocols produce cell products free from tumorigenicity and toxicity. Although a wealth of data on the biological properties of eMSC and MenSC has recently been published, it will be important to address their mechanism of action in preclinical models of human disease. © Copyright © 2020 Bozorgmehr, Gurung, Darzi, Nikoo, Kazemnejad, Zarnani and Gargett

Multidecadal variations in the early Holocene outflow of Red Sea Water into the Arabian Sea

We present Holocene stable oxygen isotope data from the deep Arabian Sea off Somalia at a decadal time resolution as a proxy for the history of intermediate/upper deep water. These data show an overall δ18O reduction by 0.5‰ between 10 and ~6.5 kyr B.P. superimposed upon short-term δ18O variations at a decadal-centennial timescale. The amplitude of the decadal variations is 0.3‰ prior, and up to 0.6‰ subsequent, to ~8.1 kyr B.P. We conclude from modeling experiments that the short-term δ18O variations between 10 and ~6.5 kyr B.P. most likely document changes in the evaporation-precipitation balance in the central Red Sea. Changes in water temperature and salinity cause the outflowing Red Sea Water to settle roughly 800 m deeper than today

Spatial variability of mixing in the Southern Ocean

Author Posting. © American Geophysical Union, 2005. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Geophysical Research Letters 32 (2005): L18603, doi:10.1029/2005GL023568.Strain variance from standard hydrographic profiles in the southern hemisphere oceans shows that turbulent mixing is vertically and spatially non-uniform. In the South Atlantic, Indian and South Pacific Oceans, enhanced diffusivities are found over rough topography. Consistent with internal tide generated mixing, the water column diffusivity returns to background levels 500 m to 1000 m off the sea floor. In the Southern Ocean, enhanced diffusivities throughout the entire water column below 1500 m are found in the Antarctic Circumpolar Current over complex topography. Differences in the vertical extent of enhanced diffusivity profiles in the Antarctic Circumpolar Current between the parameterizations based on tidal models and topography and of the present estimate of strain variance imply that elevated vertical diffusivity profiles in the Southern Ocean are due to the interaction between the mean geostrophic current and bottom topography.BMS was supported by the Ocean and Climate Change Institute at the Woods Hole Oceanographic Institution

Bone Marrow-Derived Cells from Male Donors Do Not Contribute to the Endometrial Side Population of the Recipient

Accumulated evidence demonstrates the existence of bone marrow-derived cells origin in the endometria of women undergoing bone marrow transplantation (BMT). In these reports, cells of a bone marrow (BM) origin are able to differentiate into endometrial cells, although their contribution to endometrial regeneration is not yet clear. We have previously demonstrated the functional relevance of side population (SP) cells as the endogenous source of somatic stem cells (SSC) in the human endometrium. The present work aims to understand the presence and contribution of bone marrow-derived cells to the endometrium and the endometrial SP population of women who received BMT from male donors. Five female recipients with spontaneous or induced menstruations were selected and their endometrium was examined for the contribution of XY donor-derived cells using fluorescent in situ hybridization (FISH), telomapping and SP method investigation. We confirm the presence of XY donor-derived cells in the recipient endometrium ranging from 1.7% to 2.62%. We also identify 0.45–0.85% of the donor-derived cells in the epithelial compartment displaying CD9 marker, and 1.0–1.83% of the Vimentin-positive XY donor-derived cells in the stromal compartment. Although the percentage of endometrial SP cells decreased, possibly being due to chemotherapy applied to these patients, they were not formed by XY donor-derived cells, donor BM cells were not associated with the stem cell (SC) niches assessed by telomapping technique, and engraftment percentages were very low with no correlation between time from transplant and engraftment efficiency, suggesting random terminal differentiation. In conclusion, XY donor-derived cells of a BM origin may be considered a limited exogenous source of transdifferentiated endometrial cells rather than a cyclic source of BM donor-derived stem cells

Mixed origin of neovascularization of human endometrial grafts in immunodeficient mouse models

peer reviewedBACKGROUND: In vivo mouse models have been developed to study the physiology of normal and pathologic endometrium. Although angiogenesis is known to play an important role in endometrial physiology and pathology, the origin of neovasculature in xenografts remains controversial. The aim of this study was to assess the origin of the neovasculature of endometrial grafts in different mouse models. METHODS: Human proliferative endometrium (n = 19 women) was grafted s.c. in two immunodeficient mouse strains: nude (n = 8) and severely compromised immunodeficient (SCID; n = 20). Mice were also treated with estradiol, progesterone or levonorgestrel. Fluorescence in-situ hybridization using a centromeric human chromosome X probe, immunohistochemistry (von Willebrand factor and collagen IV) and lectin perfusion were performed to identify the origin of the vessels. RESULTS: More than 90% of vessels within xenografts were of human origin 4 weeks after implantation. Some vessels (9.67 +/- 2.01%) were successively stained by human or mouse specific markers, suggesting the presence of chimeric vessels exhibiting a succession of human and murine portions. No difference in staining was observed between the two strains of mouse or different hormone treatments. Furthermore, erythrocytes were found inside human vessels, confirming their functionality. CONCLUSION: This article shows that human endometrial grafts retain their own vessels, which connect to the murine vasculature coming from the host tissue and become functional