Translational Platform Research Articles

Human Serum Albumin-enriched Clopidogrel Bisulfate Nanoparticle Alleviates Cerebral Ischemia-Reperfusion Injury in Rats.

Cerebral ischemia-reperfusion (I/R) injury remains a leading cause of mobility and mortality among patients with ischemic stroke. This study aims to develop a human serum albumin (HSA)-enriched nanoparticle platform for solubilizing clopidogrel bisulfate (CLP) for intravenous administration, and to explore the protective effect of HSA-enriched nanoparticles loaded with CLP (CLP-ANPs) against cerebral I/R injury in transient middle cerebral artery occlusion (MCAO) rat model. CLP-ANPs were synthesized via a modified nanoparticle albumin-bound technology, lyophilized, and then characterized by morphology, particle size, zeta potential, drug loading capacity, encapsulation efficiency, stability and in vitro release kinetics. In vivo pharmacokinetic studies were conducted using Sprague-Dawley (SD) rats. Also, an MCAO rat model was established to explore the therapeutic effect of CLP-ANPs on cerebral I/R injury. CLP-ANPs remained spherical particles with a layer of proteins forming protein corona. Lyophilized CLP-ANPs after dispersion displayed an average size of about 235.6 ± 6.6nm (PDI = 0.16 ± 0.08) with a zeta potential of about - 13.5 ± 1.8mV. CLP-ANPs achieved sustained release for up to 168h in vitro. Next, a single injection of CLP-ANPs dose-dependently reversed the histopathological changes induced by cerebral I/R injury possibly via attenuating apoptosis and reducing oxidative damages in the brain tissues. CLP-ANPs represent a promising and translatable platform system for the management of cerebral I/R injury during ischemic stroke.

MCR toolkit: A GPU-based toolkit for multi-channel reconstruction of preclinical and clinical x-ray CT data.

The advancement of x-ray CT into the domains of photon counting spectral imaging and dynamic cardiac and perfusion imaging has created many new challenges and opportunities for clinicians and researchers. To address challenges such as dose constraints and scanning times while capitalizing on opportunities such as multi-contrast imaging and low-dose coronary angiography, these multi-channel imaging applications require a new generation of CT reconstruction tools. These new tools should exploit the relationships between imaging channels during reconstruction to set new image quality standards while serving as a platform for direct translation between the preclinical and clinical domains. We outline and demonstrate a new Multi-Channel Reconstruction (MCR) Toolkit for GPU-based analytical and iterative reconstruction of preclinical and clinical multi-energy and dynamic x-ray CT data. To promote open science, open-source distribution of the Toolkit will coincide with the release of this publication (GPL v3; gitlab.oit.duke.edu/dpc18/mcr-toolkit-public). The MCR Toolkit source code is implemented in C/C++ and NVIDIA's CUDA GPU programming interface, with scripting support from MATLAB and Python. The Toolkit implements matched, separable footprint CT reconstruction operators for projection and backprojection in two geometries: planar, cone-beam CT (CBCT) and 3rd generation, cylindrical multi-detector row CT (MDCT). Analytical reconstruction is performed using filtered backprojection (FBP) for circular CBCT, weighted FBP (WFBP) for helical CBCT, and cone-parallel projection rebinning followed by WFBP for MDCT. Arbitrary combinations of energy and temporal channels are iteratively reconstructed under a generalized multi-channel signal model for joint reconstruction. We solve this generalized model algebraically using the split Bregman optimization method and the BiCGSTAB(l) linear solver interchangeably for both CBCT and MDCT data. Rank-sparse kernel regression (RSKR) and patch-based singular value thresholding (pSVT) are used to regularize the energy and time dimensions, respectively. Under a Gaussian noise model, regularization parameters are estimated automatically from the input data, dramatically reducing algorithm complexity for end users. Multi-GPU parallelization of the reconstruction operators is supported to manage reconstruction times. Denoising with RSKR and pSVT and post-reconstruction material decomposition are illustrated with preclinical and clinical cardiac photon-counting (PC)CT data. A digital MOBY mouse phantom with cardiac motion is used to illustrate single energy (SE), multi-energy (ME), time resolved (TR), and combined multi-energy and time-resolved (METR) helical, CBCT reconstruction. A fixed set of projection data is used across all reconstruction cases to demonstrate the Toolkit's robustness to increasing data dimensionality. Identical reconstruction code is applied to in vivo cardiac PCCT data acquired in a mouse model of atherosclerosis (METR). Clinical cardiac CT reconstruction is illustrated using the XCAT phantom and the DukeSim CT simulator, while dual-source, dual-energy CT reconstruction is illustrated for data acquired with a Siemens Flash scanner. Benchmarking results with NVIDIA RTX 8000 GPU hardware demonstrate 61%-99% efficiency in scaling computation from one to four GPUs for these reconstruction problems. The MCR Toolkit provides a robust solution for temporal and spectral x-ray CT reconstruction problems and was built from the ground up to facilitate translation of CT research and development between preclinical and clinical applications.

A phase 1/2 open-label, multicenter, dose escalation and expansion study of AVB-001, an intraperitoneally administered, cell-generated, human IL-2 immunotherapy in patients with platinum-resistant, high-grade, serous adenocarcinoma of the ovary, primary peritoneum, or fallopian tube.

TPS5616 Background: The potential of cytokines as cancer therapeutics has been limited by short half-life and severe adverse effects associated with high systemic exposure when delivered intravenously. Many strategies are being explored to overcome these limitations. A locoregional delivery approach to achieve high sustained local concentrations in the tumor microenvironment with minimal systemic exposure could widen the therapeutic window. Early experience with free recombinant human IL2 (rhIL-2) given intraperitoneally (IP) showed meaningful clinical activity in relapsed ovarian cancer. Still, the cumbersome delivery procedure requiring indwelling catheters and need for high volume IP infusions leading to frequent complications and poor patient compliance limited the utility of this approach. To overcome these shortcomings, we developed a clinically translatable localized delivery LOCOcyte platform composed of polymer encapsulated allogeneic epithelial cells engineered to produce immune effector molecules for local delivery with temporal regulation. The first product, AVB-001, produces native hIL-2, for the treatment of ovarian cancer and other peritoneal malignancies. IP AVB-001 inhibited tumor growth and improved survival in an in vivo ID8 ovarian cancer murine model. Sustained IL-2 production with well tolerated high IP concentrations were achieved, with >100x differential concentration vs. systemic blood levels in both mice and non-human primates. Strong local and systemic immune activating effects, optimized T cell profile and immune memory were observed without concomitant increase of T regs. The first in human study of AVB-001 in patients with advanced ovarian cancer (NCT05538624) is described here. Methods: Part 1 dose escalation exploring 4 ascending dose levels (capsules target production of 0.6, 1.2, 2.4, and 3.6 ug hIL2/kg/d) with a Bayesian Optimal Interval 3+3 design, in patients with recurrent high grade serous adenocarcinoma of the ovary, primary peritoneum, or fallopian tube. A minimum of 12 and up to 24 patients will be enrolled to receive one dose of AVB-001 administered IP. The primary objective is to evaluate safety (Incidence and severity of adverse events per NCI CTCAE v5.0), tolerability, and feasibility of delivering AVB-001 IP, and establish the RP2D. Secondary objectives include assessment of antitumor activity (RECIST v1.1, iRECIST), pharmacokinetics, and pharmacodynamic correlates of immune activation. Part 2 will enroll 20 patients at the RP2D with efficacy as the primary objective. This multicenter study will be conducted at 5 sites, currently two sites are open. The first patient was dosed in December 2022. Recruitment in dose level 1 cohort continues. Clinical trial information: NCT05538624 .

Combination beta-adrenergic blockade and immunotherapy for the treatment of primary and metastatic brain cancer.

e14017 Background: Brain tumors present a unique challenge to immunotherapeutic success, as immune responses in patients with brain tumors are suppressed both within the tumor microenvironment and peripherally. The systemic immunosuppression observed in brain tumor patients limits the capacity of the immune system to respond to immunotherapy. Brain tumor patients, particularly those with primary brain tumors like glioblastoma, represent a population in dire need of more thoughtful, targeted treatment approaches. Through this work we present an immunosuppressive axis specifically activated in the context of intracranial malignancies that inhibits immunotherapeutic success in these patients. We demonstrate that overactive adrenergic signaling is a major barrier to immunotherapeutic success. More specifically, beta-adrenergic signaling is known to suppress immune responses in various contexts. Our data indicate that combining beta-adrenergic blockade with immunotherapy provides a survival benefit in the setting of brain tumors, where immunotherapy alone has proven ineffective. Methods: Using the SEER Medicaid database, we retrospectively examined outcomes in patients with melanoma and lung adenocarcinoma brain metastasis who received checkpoint blockade therapy alone vs those concurrently on beta-blocker therapy. Results: We found that patients receiving combination therapy showed increased overall survival compared to those receiving checkpoint blockade alone. We then validated these findings in preclinical models of both glioma and melanoma brain metastases, demonstrating that combining immunotherapy and propranolol, a highly-prescribed FDA-approved beta-blocker, extended survival. Conclusions: Moving forward, we suggest that combination therapy with propranolol and immunotherapy, particularly checkpoint blockade, represents a promising translational treatment platform for patients with primary or metastatic intracranial malignancies.

Evidence synthesis to policy: development and implementation of an impact-oriented approach from the Eastern Mediterranean Region

BackgroundDespite the importance of evidence syntheses in informing policymaking, their production and use remain limited in the Eastern Mediterranean region (EMR). There is a lack of empirical research on approaches to promote and use policy-relevant evidence syntheses to inform policymaking processes in the EMR.ObjectiveThis study sought to describe the development of an impact-oriented approach to link evidence synthesis to policy, and its implementation through selected case studies in Lebanon, a middle-income country in the EMR.MethodsThis study followed a multifaceted and iterative process that included (i) a review of the literature, (ii) input from international experts in evidence synthesis and evidence-informed health policymaking, and (iii) application in a real-world setting (implementation). We describe four selected case studies of implementation. Surveys were used to assess policy briefs, deliberative dialogues, and post-dialogue activities. Additionally, Kingdon’s stream theory was adopted to further explain how and why the selected policy issues rose to the decision agenda.ResultsThe approach incorporates three interrelated phases: (1) priority setting, (2) evidence synthesis, and (3) uptake. Policy-relevant priorities are generated through formal priority setting exercises, direct requests by policymakers and stakeholders, or a focusing event. Identified priorities are translated into focused questions that can be addressed via evidence synthesis (phase 1). Next, a scoping of the literature is conducted to identify existing evidence syntheses addressing the question of interest. Unless the team identifies relevant, up-to-date and high-quality evidence syntheses, it proceeds to conducting SRs addressing the priority questions of interest (phase 2). Next, the team prepares knowledge translation products (e.g., policy briefs) for undertaking knowledge uptake activities, followed by monitoring and evaluation (phase 3). There are two prerequisites to the application of the approach: enhancing contextual awareness and capacity strengthening. The four case studies illustrate how evidence produced from the suites of activities was used to inform health policies and practices.ConclusionsTo our knowledge, this is the first study to describe both the development and implementation of an approach to link evidence synthesis to policy in the EMR. We believe the approach will be useful for researchers, knowledge translation platforms, governments, and funders seeking to promote evidence-informed policymaking and practice.

Representation and Reception of the Female Images in the English Version of Stories of the Sahara

San Mao (English pen name Echo), a famous wandering female writer from Taiwan, China, has influenced the growth of several generations of Chinese people, especially Chinese women, with her works and life experiences. Taking the English version of San Mao’s most famous work, Stories of the Sahara, as an example, this paper analyzes the English version, especially the translator Mike Fu's representation of San Mao's female image. The article holds that San Mao, as a narrator, bystander and witness of Saharan life, has a rich personal image, which is influenced by both eastern culture and western culture. Translator Mike Fu insists on "faithfulness" and has androgynous translation consciousness, which successfully reproduces the image of San Mao. Through the foreign translation platforms of China literature, such as Paper-Republic, and the promotion of foreign mainstream medias, the English version has gained more readers and comments, and San Mao has formed an irreplaceable female image through the understanding of different readers.

From ‘quarantini party’ to academic impact: An early childhood visual arts special interest group becomes a community of practice

ABSTRACT The Community of Practice (CoP) framework has been successfully utilised in a number of sectors to underpin meaningful collaborations. However, to date, the application of CoP in early childhood education (ECE) has been limited, with no studies having explored the unique intersection of early childhood and visual arts CoP. A qualitative case study was conducted with nine geographically dispersed visual arts ECE academics and practitioners who formed an early childhood visual arts special interest group (SIG) to facilitate social and professional connection in response to COVID-19 lockdowns in early 2020. The research collected and thematically analysed, questionnaire and focus group data to investigate how the practice of the virtual, international, ECE visual arts SIG aligned with CoP framework elements. Overall, the practices and experiences of the SIG were well aligned with the CoP framework, particularly in relation to the Domain and Practice elements. In addition, the model of an online, international CoP described in this study proposes that virtual CoP opportunities that foster strong relational connections alongside professional networking sustains the activity of the CoP to offer participants not only rich opportunities for professional learning and collaboration with like-minded colleagues, but also offers a platform for significant research translation and impact.

Translation regulation by a guanidine-II riboswitch is highly tunable in sensitivity, dynamic range, and apparent cooperativity.

Riboswitches function as important translational regulators in bacteria. Comprehensive mutational analysis of transcriptional riboswitches has been used to probe the energetic intricacies of interplay between the aptamer and expression platform, but translational riboswitches have been inaccessible to massively parallel techniques. The guanidine-II (gdm-II) riboswitch is an exclusively translational class. We have integrated RelE cleavage with next-generation sequencing to quantify ligand-dependent changes in translation initiation for all single and double mutations of the Pseudomonas aeruginosa gdm-II riboswitch, a total of more than 23,000 variants. This extensive mutational analysis is consistent with the prominent features of the bioinformatic consensus. These data indicate, unexpectedly, that direct sequestration of the Shine-Dalgarno sequence is dispensable for riboswitch function. Additionally, this comprehensive data set reveals important positions not identified in previous computational and crystallographic studies. Mutations in the variable linker region stabilize alternate conformations. The double mutant data reveal the functional importance of the previously modeled P0b helix formed by the 5' and 3' tails that serves as the basis for translational control. Additional mutations to GU wobble base pairs in both P1 and P2 reveal how the apparent cooperativity of the system involves an intricate network of communication between the two binding sites. This comprehensive examination of a translational riboswitch's expression platform illuminates how the riboswitch is precisely tuned and tunable with regard to ligand sensitivity, the amplitude of expression between ON and OFF states, and the cooperativity of ligand binding.

대학 EFL 학습자들을 위한 인공지능 기반 플랫폼 활용 교양 영어 쓰기 교수⋅학습 방안

Objectives The present study aimed to find out the characteristics of English writing and learners' perception of the use of AI-based platforms after having university EFL learners use AI-based platforms in the English writing teaching and learning process.
 Methods In the 2nd semester of 2022, AI-based English writing scoring/English grammar check platform and machine translation platform were used for the English writing process for 20 students in a general English required course in C university. The English composition drafts in 3rd week (pre-test) and 15th week(post-test) were constructed as a learner corpus and were analyzed by Text Inspector and Jamovi 2.3.21. In addition, a survey and reflection writings were used to investigate learners' perceptions of learning English writing using AI-based platforms.
 Results As a result of analyzing the corpus of pre- and post-test, a significant improvement was shown only in terms of vocabulary diversity in English writing, and no significant difference was shown in quantitative improvement such as the number of sentences and sentence length. In addition, learners were positively aware of the effectiveness of English writing teaching and learning using AI-based platforms, and in particular, reported that it was very helpful in reducing the burden of using English, which has a positive effect on the affective aspect of using AI-based platforms.
 Conclusions Based on these results, it is suggested that guidance for learners, improvement of instructors' awareness, and teaching ability are necessary for the effectiveness of using AI platform and its proper use in English writing teaching and learning.

Abstract CT122: A phase 1/2 open-label, multicenter, dose escalation and expansion study of AVB-001, an intraperitoneally administered, cell-generated, human IL-2 immunotherapy in patients with platinum-resistant, high-grade, serous adenocarcinoma of the ovary, primary peritoneum, or fallopian tube

Abstract Background: The potential of cytokines as cancer therapeutics has been limited by short half-life and severe adverse effects associated with high systemic exposure when delivered intravenously. Many strategies are being explored to overcome these limitations. A locoregional delivery approach to achieve high sustained local concentrations in the tumor microenvironment with minimal systemic exposure could widen the therapeutic window. Early experience with free recombinant human IL2 (rhIL-2) given intraperitoneally (IP) showed meaningful clinical activity in relapsed ovarian cancer. Still, the cumbersome delivery procedure requiring indwelling catheters and need for high volume IP infusions leading to frequent complications and poor patient compliance limited the utility of this approach. To overcome these shortcomings, we developed a clinically translatable localized delivery LOCOcyteTM platform composed of polymer encapsulated allogeneic epithelial cells engineered to produce immune effector molecules for local delivery with temporal regulation. The first product, AVB-001, produces native hIL-2, for the treatment of ovarian cancer and other peritoneal malignancies. IP AVB-001 inhibited tumor growth and improved survival in an in vivo ID8 ovarian cancer murine model. Sustained IL-2 production with well tolerated high IP concentrations were achieved, with >100x differential concentration vs. systemic blood levels in both mice and non-human primates. Strong local and systemic immune activating effects, optimized T cell profile and immune memory were observed without concomitant increase of T regs. The first in human study of AVB-001 in patients with advanced ovarian cancer (NCT05538624) is described here. Methods: Part 1 dose escalation exploring 4 ascending dose levels (capsules target production of 0.6, 1.2, 2.4, and 3.6 ug hIL2/kg/d) with a Bayesian Optimal Interval 3+3 design, in patients with recurrent high grade serous adenocarcinoma of the ovary, primary peritoneum, or fallopian tube. A minimum of 12 and up to 24 patients will be enrolled to receive one dose of AVB-001 administered IP. The primary objective is to evaluate safety (Incidence and severity of adverse events per NCI CTCAE v5.0), tolerability, and feasibility of delivering AVB-001 IP, and establish the RP2D. Secondary objectives include assessment of antitumor activity (RECIST v1.1, iRECIST), pharmacokinetics, and pharmacodynamic correlates of immune activation. Part 2 will enroll 20 patients at the RP2D with efficacy as the primary objective. This multicenter study will be conducted at 5 sites, currently one site is open. The first patient was dosed in December 2022. Recruitment in dose level 1 cohort continues. Citation Format: Shannon N. Westin, Travis Sims, Karen Andreas, Eric Soliman, Manish Jain, Claudio A. Dansky Ullmann, Amir A. Jazaeri. A phase 1/2 open-label, multicenter, dose escalation and expansion study of AVB-001, an intraperitoneally administered, cell-generated, human IL-2 immunotherapy in patients with platinum-resistant, high-grade, serous adenocarcinoma of the ovary, primary peritoneum, or fallopian tube [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT122.

QuNex-An integrative platform for reproducible neuroimaging analytics.

Neuroimaging technology has experienced explosive growth and transformed the study of neural mechanisms across health and disease. However, given the diversity of sophisticated tools for handling neuroimaging data, the field faces challenges in method integration, particularly across multiple modalities and species. Specifically, researchers often have to rely on siloed approaches which limit reproducibility, with idiosyncratic data organization and limited software interoperability. To address these challenges, we have developed Quantitative Neuroimaging Environment & Toolbox (QuNex), a platform for consistent end-to-end processing and analytics. QuNex provides several novel functionalities for neuroimaging analyses, including a "turnkey" command for the reproducible deployment of custom workflows, from onboarding raw data to generating analytic features. The platform enables interoperable integration of multi-modal, community-developed neuroimaging software through an extension framework with a software development kit (SDK) for seamless integration of community tools. Critically, it supports high-throughput, parallel processing in high-performance compute environments, either locally or in the cloud. Notably, QuNex has successfully processed over 10,000 scans across neuroimaging consortia, including multiple clinical datasets. Moreover, QuNex enables integration of human and non-human workflows via a cohesive translational platform. Collectively, this effort stands to significantly impact neuroimaging method integration across acquisition approaches, pipelines, datasets, computational environments, and species. Building on this platform will enable more rapid, scalable, and reproducible impact of neuroimaging technology across health and disease.

Abstract 6377: A translatable tumor-killing platform with robust sensitivity towards immune checkpoint inhibitor targeted therapeutics

Abstract The cancer treatment landscape has transformed following the clinical success of early immune checkpoint inhibitors (ICI) which demonstrate the remarkable power of harnessing the immune system to treat a variety of metastatic cancers. Despite the success of Pembrolizumab, Nivolumab and Atezolizumab, the programmed cell death protein-1 (PD1)-programmed death ligand-1/2 (PD-L1/L2) pathway remains a tractable target for many drug discovery programs, particularly for bi-specific or combinational therapies. The PD1 - PD-L1/L2 pathway is important in protecting the host from uncontrolled immune responses, but can be hijacked by tumors, rendering them resistant to immune attack. Antibodies which antagonize this pathway potentiate T-cell mediated tumor destruction yet do not show efficacy in all patients or guarantee long-term protection, in part due to tumor-associated ICI-resistance mechanisms. There is a distinct need to develop novel ICI therapeutics and robust, complex co-culture platforms to better predict clinical efficacy. Here we outline a series of translationally relevant co-culture assays using Pembrolizumab as a benchmark therapeutic alongside other ICI. PD-1 expression levels are known to be low on resting T cells, as indeed are many checkpoint receptors, and so to enhance the therapeutic window, CD3+T cells from healthy human donors were polyclonally stimulated before addition to SK-OV-3 target cell cultures. The SK-OV-3 tumor line was stably transduced with NucLight Red (NLR) to allow co-localization of the caspase 3/7 signal to tumor cells. Autologous, mature monocyte-derived dendritic cells (moDC), expressing high levels of PD-L1/PD-L2 were added into tri-cultures in a 2D tumor killing assay. Tumor killing kinetics were quantified via live cell imaging using an Incucyte S3. Although Pembrolizumab enhanced immune-mediated direct (pre-stimulated) T cell killing, the therapeutic window increased in the presence of autologous moDC. Using a modified 3D (tumor spheroid) format over a longer time course, Pembrolizumab was also shown to enhance PBMC-mediated killing and the ability of moDC to enhance this effect further was determined. In recent years, the development and implementation of combinatorial approaches has resulted in significant improvements in patient responsiveness and disease outcomes for various cancers. Here, we describe a suite of tumor killing assays which can be adapted to multiple tumor or therapeutic targets, with a focus on immune involvement. ICI are shown to work robustly and efficacy tracks expression of the relevant checkpoint receptors, making this a powerful format in which to test combination therapies or novel ICI. Citation Format: Marie Carkill, Kaitlin Mitchell, Lauren Kelsey, Jezrom Self-Fordham, Nunan Robert, Louise Brackenbury. A translatable tumor-killing platform with robust sensitivity towards immune checkpoint inhibitor targeted therapeutics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6377.

Abstract 5084: Targeted delivery of low-dose radiation alleviates tumor resistance to CAR-T cell therapy

Abstract Purpose: Current understanding of resistance to CAR-T cell therapy in solid tumors implicates inadequate CAR-T cell potency in the immunosuppressive tumor microenvironment (TME). We have previously developed a platform using somatostatin receptor 2 (SSTR2) as a positron emission tomography (PET) reporter to detect CAR-T cell expansion and trafficking. The current study aimed to leverage SSTR2 for low-dose targeted radionuclide therapy (177Lu-DOTATATE, Lutathera), which under dosimetry guidance might enhance antitumor immunity by reprograming the TME and promoting T-cell reinvigoration. Methods: Using intercellular adhesion molecule 1 (ICAM-1) as a model antigen, we evaluated the immunomodulatory effects of low-dose radiation in a gastric cancer animal model. NSG mice were inoculated subcutaneously with firefly luciferase-expressing Hs 746T cells (0.1 × 106 per mouse) and treated 5 days later with 10 × 106 SSTR2-expressing ICAM-1 CAR-T cells. CAR-T cell expansion was monitored weekly by PET scan using 18F-NOTA-Octreotide, a radiotracer targeting SSTR2. Three weeks post-T cell infusion, a cohort of mice were injected with 7.4 MBq of 177Lu-DOTATATE via the tail vein. SPECT imaging was performed for dosimetry analysis. Tumor growth was monitored by bioluminescence imaging and tumor size measurement. Serum cytokines were analyzed. Results: Single dose 177Lu-DOTATATE treatment (delivering 1-6 Gy to tumor) improved response of established gastric cancer tumor (>1,000 mm3) that was not responsive to CAR-T cell treatment alone and significantly prolonged survival. All mice receiving CAR-T cells plus 177Lu-DOTATATE displayed rapid tumor shrinkage, with 83% of mice achieving complete remission within 3 weeks of 177Lu-DOTATATE treatment. SPECT imaging confirmed specific delivery of 177Lu-DOTATATE by tumor-infiltrating CAR-T cells, with tumor uptake of 0.43 ± 0.24 and 0.19 ± 0.08 MBq/g at 24 and 144 hours post 177Lu-DOTATATE injection, respectively. Most radioactivity reduction over time (73%) is explained by physical decay of 177Lu, indicating persistent tumor retention of 177Lu-DOTATATE. In contrast, rapid clearance of 177Lu-DOTATATE was observed in liver and kidneys. Importantly, longitudinal CAR-T cell imaging using 18F-NOTA-Octreotide revealed increased CAR-T cell expansion induced by low-dose radiation. Furthermore, we detected high levels of IFN-γ and perforin in serum after 177Lu-DOTATATE treatment, which were >10 folds higher than that in control mice receiving CAR-T cell only, indicating activation of T cells by low-dose radiation. Conclusions: We developed a translatable radioimmunotherapy platform that incorporates a FDA-approved theranostic endoradiotherapy (Lutathera) to improve tumor response to CAR-T cell therapy. The next steps would be to explore the immunomodulatory effects of low-dose radiation systematically and elucidate the mechanism of action. Citation Format: Yanping Yang, Yago Alcaina, Yogindra Vedvyas, Maria Cristina Riascos, Edward K. Fung, Brett Vaughn, Sarah M. Cheal, Irene M. Min, Claire Vanpouille-Box, Moonsoo M. Jin. Targeted delivery of low-dose radiation alleviates tumor resistance to CAR-T cell therapy. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5084.

Abstract 4956: Discovering novel targetable pathways by combining functional and multi-omic data from primary ovarian cancer samples

Abstract Background: There is a critical necessity to reveal novel and tractable targets for anti-cancer treatments in indications with high unmet medical need, such as high grade serous ovarian cancer (HGSOC). However, standard process for target discovery using models such as outgrown cell lines and well-averaged readouts has yielded a less than 5% approval rate for drugs entering trials (Thomas et al. 2016 Bio.org). Here, we describe patient-centric target discovery through the use of disease relevant primary OC samples and single cell functional characterization using a platform with proven hemonc translatability (Kornauth et al. 2021, Snijder et al. 2017). We integrate data from our functional drug testing platform under multiple drug perturbations with matching genomic and transcriptomic data to reveal associations with novel downstream regulators of sensitivity. Methods: Sensitivity of the cancer cell compartment in primary malignant ascites samples (n = 20; 75% HGSOC) to 85 small molecule drugs, was evaluated using a proprietary and translatable deep learning-driven single cell imaging platform (Vladimer et al. 2017). Cancer cell sensitivity from the drugs was combined with WES, bulk-RNAseq and drug induced changes in phosphoproteome, and single cell RNAseq transcriptome to identify perturbed targets and pathways. Results: Here we describe a family of TKIs including ALKi that induce cytotoxicity of cancer cells in primary samples, not previously captured in publicly available cell line drug sensitivity screening data (Iorio et al. 2016). We report novel sensitivity of OC driven by non-canonical targets of ceritinib such as FAK1 or IGF1R, mediated by the downstream signaling hub YBX1 (Kuenzi et al. 2017), involved in NFB pathway regulation (Motolani et al. 2021). Indeed, transcriptomic scRNA analysis upon ceritinib treatment of primary OC cells revealed rapid perturbation of numerous NFB pathway members, alongside YBX1 inactivation. Conclusions: Combining functional endpoints and single cell-based differential expression analysis of primary OC samples, we have identified the NFB pathway and the regulator YBX1 as a promising novel sensitivity for HGSOC treatment development. These and several other important targetable nodes identified, sit outside the recently suggested JAK/STAT pathway (Izar et al. 2020), thereby demonstrating a pipeline towards novel drug target and pathway discovery driven by patient-centric, disease relevant models of high-need indications. Citation Format: Irene Gutierrez-Perez, Bekir Ergüner, Pisanu Buphamalai, Joost Van Ham, Paul Heinz, Valentin Aranha, Rin Okumura, Elisabeth Waltenberger, Isabella Alt, Claudia Baumgaertler, Maja Stulic, Edgar Petru, Christoph Minichsdorfer, Judith Lafleur, Lukas Hefler, Laudia Hadjari, Lucia Dzurillova, Jozef Sufliarsky, Nikolaus Krall, Thorsten Füreder, Gregory Ian Vladimer, Bojan Vilagos, Robert Sehlke. Discovering novel targetable pathways by combining functional and multi-omic data from primary ovarian cancer samples. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4956.

Abstract 2151: Identification of transcript adenosine fingerprint to enrich for A2AR and PD-1 inhibition responders

Abstract Background: Precision cancer medicine aims to identify the right drug for the right patient, enriching for patients more likely to respond to a particular treatment. This paradigm is gaining importance during the early clinical lifecycle of a new potential drug to improve patient-centric trial designs, drive clinical success and eventually increase approval rates - enlarging the therapeutic arsenal available for oncology patients. To optimize the chance of success with our A2AR-selective antagonist, EXS-21546 (546; NCT04727138, discovered in collaboration with Evotec), we have identified an adenosine-induced immunosuppression biomarker signature (adenosine burden score or ABS) for clinical trial patient selection that also correlates with checkpoint inhibitor (CI) response prediction in ex vivo primary models. Here we present transcriptional and functional data mapping adenosine burden at the single cell level, and investigate subsequent modulation through antagonism of A2AR with 546, combination effects with CI, to prioritize patients for 546+CI therapy. Methods: By leveraging disease-relevant primary human tissues together with matched single cell and bulk transcriptomics, we assess adenosine-induced anticancer immune suppression and show initial biological confirmation of patient selection methodology and combination therapy effects with a translatable high content imaging platform (Kornauth et al 2021 & Snidjer et al 2017). Results: The ABS detects adenosine rich microenvironments with greater specificity and sensitivity than other published signatures. Validating the ABS in TGCA, we found the ABS anti-correlates with a validated predictor of anti-PD-1 therapy success (TIS, Damotte et al 2019), unraveling that high-adenosine/ABS cases are among patients least likely to respond to immunotherapy (low TIS). A2AR antagonism with 546 demonstrated a reduction of the adenosine burden, and restored the CI response potential as addressed by the ABS and TIS, respectively. Further immune reactivation was seen with antagonism of adenosine signaling by ‘546/CI combination ex vivo in primary tissues pre-selected with our ABS signature. Conclusions: Combining deep learning of single cell functional and multi-omics profiling data of disease relevant primary model systems, we model the association of the immune response potential to A2AR antagonism in cancer to define a biomarker signature to predict patients likely to benefit from A2AR antagonism and CI. This will be confirmed and validated retrospectively in an ongoing clinical study of 546 in two cancer indications. Citation Format: Isabella Alt, Robert Sehlke, Anna Lobley, Claudia Baumgaertler, Maja Stulic, Klaus Hackner, Lucia Dzurillova, Edgar Petru, Laudia Hadjari, Judith Lafleur, Josef Singer, Nikolaus Krall, Jozef Šufliarsky, Lukas Hefler, Thorsten Füreder, Christina Taubert, Andrew Payne, Christophe Boudesco, Gregory Ian Vladimer. Identification of transcript adenosine fingerprint to enrich for A2AR and PD-1 inhibition responders [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2151.

Further Investigations of Nitroheterocyclic Compounds as Potential Antikinetoplastid Drug Candidates.

Due to the lack of specific vaccines, management of the trypanosomatid-caused neglected tropical diseases (sleeping sickness, Chagas disease and leishmaniasis) relies exclusively on pharmacological treatments. Current drugs against them are scarce, old and exhibit disadvantages, such as adverse effects, parenteral administration, chemical instability and high costs which are often unaffordable for endemic low-income countries. Discoveries of new pharmacological entities for the treatment of these diseases are scarce, since most of the big pharmaceutical companies find this market unattractive. In order to fill the pipeline of compounds and replace existing ones, highly translatable drug screening platforms have been developed in the last two decades. Thousands of molecules have been tested, including nitroheterocyclic compounds, such as benznidazole and nifurtimox, which had already provided potent and effective effects against Chagas disease. More recently, fexinidazole has been added as a new drug against African trypanosomiasis. Despite the success of nitroheterocycles, they had been discarded from drug discovery campaigns due to their mutagenic potential, but now they represent a promising source of inspiration for oral drugs that can replace those currently on the market. The examples provided by the trypanocidal activity of fexinidazole and the promising efficacy of the derivative DNDi-0690 against leishmaniasis seem to open a new window of opportunity for these compounds that were discovered in the 1960s. In this review, we show the current uses of nitroheterocycles and the novel derived molecules that are being synthesized against these neglected diseases.

Shape matters: Biodegradable anisotropic nanoparticle artificial antigen presenting cells for cancer immunotherapy.

Artificial antigen presenting cells are biomimetic particles that recapitulate the signals presented by natural antigen presenting cells in order to stimulate T cells in an antigen-specific manner using an acellular platform. We have engineered an enhanced nanoscale biodegradable artificial antigen presenting cell by modulating particle shape to achieve a nanoparticle geometry that allows for increased radius of curvature and surface area for T cell contact. The non-spherical nanoparticle artificial antigen presenting cells developed here have reduced nonspecific uptake and improved circulation time compared both to spherical nanoparticles and to traditional microparticle technologies. Additionally, the anisotropic nanoparticle artificial antigen presenting cells efficiently engage with and activate T cells, ultimately leading to a marked anti-tumor effect in a mouse melanoma model that their spherical counterparts were unable to achieve. STATEMENT OF SIGNIFICANCE: Artificial antigen presenting cells (aAPC) can activate antigen-specific CD8+ T cells but have largely been limited to microparticle-based platforms and ex vivo T cell expansion. Although more amenable to in vivo use, nanoscale aAPC have traditionally been ineffective due to limited surface area available for T cell interaction. In this work, we engineered non-spherical biodegradable nanoscale aAPC to investigate the role of particle geometry and develop a translatable platform for T cell activation. The non-spherical aAPC developed here have increased surface area and a flatter surface for T cell engagement and, therefore, can more effectively stimulate antigen-specific T cells, resulting in anti-tumor efficacy in a mouse melanoma model.

Zebrafish Cancer Avatars: A Translational Platform for Analyzing Tumor Heterogeneity and Predicting Patient Outcomes.

The increasing number of available anti-cancer drugs presents a challenge for oncologists, who must choose the most effective treatment for the patient. Precision cancer medicine relies on matching a drug with a tumor's molecular profile to optimize the therapeutic benefit. However, current precision medicine approaches do not fully account for intra-tumoral heterogeneity. Different mutation profiles and cell behaviors within a single heterogeneous tumor can significantly impact therapy response and patient outcomes. Patient-derived avatar models recapitulate a patient's tumor in an animal or dish and provide the means to functionally assess heterogeneity's impact on drug response. Mouse xenograft and organoid avatars are well-established, but the time required to generate these models is not practical for clinical decision-making. Zebrafish are emerging as a time-efficient and cost-effective cancer avatar model. In this review, we highlight recent developments in zebrafish cancer avatar models and discuss the unique features of zebrafish that make them ideal for the interrogation of cancer heterogeneity and as part of precision cancer medicine pipelines.

Electrophysiological signatures of reward learning in the rodent touchscreen-based Probabilistic Reward Task.

Blunted reward learning and reward-related activation within the corticostriatal-midbrain circuitry have been implicated in the pathophysiology of anhedonia and depression. Unfortunately, the search for more efficacious interventions for anhedonic behaviors has been hampered by the use of vastly different preclinical and clinical assays. In a first step in addressing this gap, in the current study, we used event-related potentials and spectral analyses in conjunction with a touchscreen version of the rodent Probabilistic Reward Task (PRT) to identify the electrophysiological signatures of reward learning in rats. We trained 11 rats (5 females and 6 males) on the rodent touchscreen-based PRT and subsequently implanted them with deep electrodes in the anterior cingulate cortex (ACC) and nucleus accumbens (NAc) for local field potentials recordings during the PRT. Behaviorally, the expected responsivity-to-reward profile was observed. At the electrophysiological level, we identified a negative amplitude deflection 250-500 ms after feedback in the ACC and NAc electrodes, as well as power increase in feedback-locked delta (1-5 Hz) and alpha/beta (9-17 Hz) bands in both electrodes for rewarded trials. Using a reverse-translational approach, we identified electrophysiological signatures of reward learning in rats similar to those described in humans. These findings and approaches might provide a useful translational platform to efficiently evaluate novel therapeutics targeting anhedonia.

Defined, Simplified, Scalable, and Clinically Compatible Hydrogel-Based Production of Human Brain Organoids

Human brain organoids present a new paradigm for modeling human brain organogenesis, providing unprecedented insight to the molecular and cellular processes of brain development and maturation. Other potential applications include in vitro models of disease and tissue trauma, as well as three-dimensional (3D) clinically relevant tissues for pharmaceuticals development and cell or tissue replacement. A key requirement for this emerging technology in both research and medicine is the simple, scalable, and reproducible generation of organoids using reliable, economical, and high-throughput culture platforms. Here we describe such a platform using a defined, clinically compliant, and readily available hydrogel generated from gelatin methacrylate (GelMA). We demonstrate the efficient production of organoids on GelMA from human induced pluripotent stem cells (iPSCs), with scalable production attained using 3D printed GelMA-based multiwell arrays. The differentiation of iPSCs was systematic, rapid, and direct to enable iPSCs to form organoids in their original position following seeding on GelMA, thereby avoiding further cell and organoid disruption. Early neural precursors formed by day 5, neural rosettes and early-stage neurons by day 14, and organoids with cellular and regional heterogeneity, including mature and electrophysiologically active neurons, by day 28. The optimised method provides a simplified and well-defined platform for both research and translation of iPSCs and derivative brain organoids, enabling reliable 3D in vitro modelling and experimentation, as well as the provision of clinically relevant cells and tissues for future therapeutics.