Search NASA⌕ Search

SEARCH · Search NASA

Results for “TARGETS”

Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 235 records · Page 13
All sourcesNASA NTRSDOE OSTI
DOE OSTI2024

Development of CD46 targeted alpha theranostics in prostate cancer using 134 Ce/ 225 Ac-Macropa-PEG 4 -YS5

Rationale: 225 Ac, a long-lived α-emitter with a half-life of 9.92 days, has garnered significant attention as a therapeutic radionuclide when coupled with monoclonal antibodies and other targeting vectors. Nevertheless, its clinical utility has been hampered by potential off-target toxicity, a lack of optimized chelators for 225 Ac, and limitations in radiolabeling methods. In a prior study evaluating the effectiveness of CD46-targeted radioimmunotherapy, we found great therapeutic efficacy but also significant toxicity at higher doses. To address these challenges, we have developed a radioimmunoconjugate called 225 Ac-Macropa-PEG 4 -YS5, incorporating a stable PEGylated linker to maximize tumoral uptake and increase tumor-to-background ratios. Our research demonstrates that this conjugate exhibits greater anti-tumor efficacy while minimizing toxicity in prostate cancer 22Rv1 tumors. Methods: We synthesized Macropa.NCS and Macropa-PEG 4/8 -TFP esters and prepared Macropa-PEG 0/4/8 -YS5 (with nearly ~1:1 ratio of macropa chelator to antibody YS5) as well as DOTA-YS5 conjugates. These conjugates were then radiolabeled with 225 Ac in a 2 M NH 4 OAc solution at 30 °C, followed by purification using YM30K centrifugal purification. Subsequently, we conducted biodistribution studies and evaluated antitumor activity in nude mice (nu/nu) bearing prostate 22Rv1 xenografts in both single-dose and fractionated dosing studies. Micro-PET imaging studies were performed with 134 Ce-Macropa-PEG 0/4/8 -YS5 in 22Rv1 xenografts for 7 days. Toxicity studies were also performed in healthy athymic nude mice. Results: As expected, we achieved a >95% radiochemical yield when labeling Macropa-PEG 0/4/8 -YS5 with 225 Ac, regardless of the chelator ratios (ranging from 1 to 7.76 per YS5 antibody). The isolated yield exceeded 60% after purification. Such high conversions were not observed with the DOTA-YS5 conjugate, even at a higher ratio of 8.5 chelators per antibody (RCY of 83%, an isolated yield of 40%). Biodistribution analysis at 7 days post-injection revealed higher tumor uptake for the 225 Ac-Macropa-PEG 4 -YS5 (82.82 ± 38.27 %ID/g) compared to other conjugates, namely 225 Ac-Macropa-PEG 0/8 -YS5 (38.2 ± 14.4/36.39 ± 12.4 %ID/g) and 225 Ac-DOTA-YS5 (29.35 ± 7.76 %ID/g). The PET Imaging of 134 Ce-Macropa-PEG 0/4/8 -YS5 conjugates resulted in a high tumor uptake, and tumor to background ratios. In terms of antitumor activity, 225 Ac-Macropa-PEG 4 -YS5 exhibited a substantial response, leading to prolonged survival compared to 225 Ac-DOTA-YS5, particularly when administered at 4.625 kBq doses, in single or fractionated dose regimens. Chronic toxicity studies observed mild to moderate renal toxicity at 4.625 and 9.25 kBq doses. Conclusions: Our study highlights the promise of 225 Ac-Macropa-PEG 4 -YS5 for targeted alpha particle therapy. The 225 Ac-Macropa-PEG 4 -YS5 conjugate demonstrates improved biodistribution, reduced off-target binding, and enhanced therapeutic efficacy, particularly at lower doses, compared to 225 Ac-DOTA-YS5. Incorporating theranostic 134 Ce PET imaging further enhances the versatility of macropa-PEG conjugates, offering a more effective and safer approach to cancer treatment. Overall, this methodology has a high potential for broader clinical applications.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

Shortwave‐Infrared‐Emitting Nanoprobes for CD8 Targeting and In Vivo Imaging of Cytotoxic T Cells in Breast Cancer

Checkpoint immunotherapy has made great strides in the treatment of solid tumors, but many patients do not respond to immune checkpoint inhibitors. Identification of tumor‐infiltrating cytotoxic T cells (CTLs) has the potential to stratify patients and monitor immunotherapy responses. In this study, the design of cluster of differentiation (CD8 + ) T cell‐targeted nanoprobes that emit shortwave infrared (SWIR) light in the second tissue‐transparent window for noninvasive, real‐time imaging of CTLs in murine models of breast cancer is presented. SWIR‐emitting rare‐earth nanoparticles encapsulated in human serum albumin are conjugated with anti‐CD8α to target CTLs with high specificity. CTL targeting is validated in vitro through binding of nanoprobes to primary mouse CTLs. The potential for the use of SWIR fluorescence intensity to determine CTL presence is validated in two syngeneic mammary fat pad tumor models, EMT6 and 4T1, which differ in immune infiltration. SWIR imaging using CD8‐targeted nanoprobes successfully identifies the presence of CTLs in the more immunogenic EMT6 model, while imaging confirms the lack of substantial immune infiltration in the nonimmunogenic 4T1 model. In this work, the opportunity for SWIR imaging using CD8‐targeted nanoprobes to assess CTL infiltration in tumors for the stratification and monitoring of responders to checkpoint immunotherapy is highlighted.

Shah, Jay V.↗
DOE OSTI2024

“Off‐Label Use” of the Siderophore Enterobactin Enables Targeted Imaging of Cancer with Radioactive Ti (IV)

Abstract The development of inert, biocompatible chelation methods is required to harness the emerging positron emitting radionuclide 45 Ti for radiopharmaceutical applications. Herein, we evaluate the Ti (IV) ‐coordination chemistry of four catechol‐based, hexacoordinate chelators using synthetic, structural, computational, and radiochemical approaches. The siderophore enterobactin (Ent) and its synthetic mimic TREN‐CAM readily form mononuclear Ti (IV) species in aqueous solution at neutral pH. Radiolabeling studies reveal that Ent and TREN‐CAM form mononuclear complexes with the short‐lived, positron‐emitting radionuclide 45 Ti (IV) , and do not transchelate to plasma proteins in vitro and exhibit rapid renal clearance in naïve mice. These features guide efforts to target the 45 Ti isotope to prostate cancer tissue through the design, synthesis, and evaluation of Ent‐DUPA, a small molecule conjugate composed of a prostate specific membrane antigen (PSMA) targeting peptide and a monofunctionalized Ent scaffold. The [ 45 Ti][Ti(Ent‐DUPA)] 2− complex forms readily at room temperature. In a tumor xenograft model in mice, selective tumor tissue accumulation (8±5 %, n =5), and low off‐target uptake in other organs is observed. Overall, this work demonstrates targeted imaging with 45 Ti (IV) , provides a foundation for advancing the application of 45 Ti in nuclear medicine, and reveals that Ent can be repurposed as a 45 Ti‐complexing cargo for targeted nuclear imaging applications.

Koller, Angus J.↗
DOE OSTI2024

“Off‐Label Use” of the Siderophore Enterobactin Enables Targeted Imaging of Cancer with Radioactive Ti (IV)

The development of inert, biocompatible chelation methods is required to harness the emerging positron emitting radionuclide 45 Ti for radiopharmaceutical applications. Herein, we evaluate the Ti (IV) -coordination chemistry of four catechol-based, hexacoordinate chelators using synthetic, structural, computational, and radiochemical approaches. The siderophore enterobactin (Ent) and its synthetic mimic TREN-CAM readily form mononuclear Ti (IV) species in aqueous solution at neutral pH. Radiolabeling studies reveal that Ent and TREN-CAM form mononuclear complexes with the short-lived, positron-emitting radionuclide 45 Ti (IV) , and do not transchelate to plasma proteins in vitro and exhibit rapid renal clearance in naïve mice. These features guide efforts to target the 45 Ti isotope to prostate cancer tissue through the design, synthesis, and evaluation of Ent-DUPA, a small molecule conjugate composed of a prostate specific membrane antigen (PSMA) targeting peptide and a monofunctionalized Ent scaffold. Additionally, the [ 45 Ti][Ti(Ent-DUPA)] 2− complex forms readily at room temperature. In a tumor xenograft model in mice, selective tumor tissue accumulation (8±5 %, n=5), and low off-target uptake in other organs is observed. Overall, this work demonstrates targeted imaging with 45 Ti (IV) , provides a foundation for advancing the application of 45 Ti in nuclear medicine, and reveals that Ent can be repurposed as a 45 Ti-complexing cargo for targeted nuclear imaging applications.

62 RADIOLOGY AND NUCLEAR MEDICINE↗
DOE OSTI2022

Applications of targeted proteomics in metabolic engineering: advances and opportunities

Optimization of metabolically engineered organisms requires good understanding of producing balanced level of pathway proteins. Targeted proteomics via selected-reaction monitoring (SRM) has been increasingly used in metabolic engineering studies to detect and quantify sets of proteins with high selectivity, multiplexity, and reproducibility. In combination with metabolomics and other omics tools, targeted proteomics has helped optimize the production of many bio-based chemicals in various metabolic engineering cell factories. In this review, we present recent applications of targeted proteomics in metabolic engineering studies and highlight several successful cases of targeted proteomics in boosting production of commodity and high value chemicals. Additionally, we also discuss challenges and limitations of current targeted proteomics and map opportunities for future research.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Elevated-Mn ChemCam targets illuminating Mn redox cycling and diagenesis in the Bradbury Rise, Gale Crater, Mars

Manganese plays a crucial role as a paleo-environmental and geological indicator due to its sensitivity to redox potential and pH variations in the environment. On Earth, the association between the rise of atmospheric oxygen during the Great Oxidation Event and the presence of Mn in the sedimentary rock record underscores its significance. Here, in this study, we reexamined ChemCam targets from the first 600 sols of the Mars Science Laboratory mission, focusing on identifying instances of above-average Mn within these targets. These elevated-Mn targets were categorized into distinct geologic classes, revealing a pattern linking heightened Mn levels with diagenetically altered materials, such as calcium-sulfate veins and concretions, as well as clay minerals within the same targets, indicating a compelling relationship between Mn enrichment and diagenetic processes. High concentrations of Mn were observed in chemically altered targets, suggesting the occurrence of multiple fluid events: the first to alter the material and the second to deposit Mn. The observed patterns suggest multiple diagenetic events and redox cycling that facilitated the deposition and transport of Mn subsequent to the initial dissolution of basaltic materials. This research sheds light on the complexity of martian diagenetic processes and their implications for the planet’s environmental evolution.

58 GEOSCIENCES↗
DOE OSTI2024

Comparison of machine learning and electrical resistivity arrays to inverse modeling for locating and characterizing subsurface targets

Here, this study evaluates the performance of multiple machine learning (ML) algorithms and electrical resistivity (ER) arrays for inversion with comparison to a conventional Gauss-Newton numerical inversion method. Four different ML models and four arrays were used for the estimation of only six variables for locating and characterizing hypothetical subsurface targets. The combination of dipole-dipole with Multilayer Perceptron Neural Network (MLP-NN) had the highest accuracy. Evaluation showed that both MLP-NN and Gauss-Newton methods performed well for estimating the matrix resistivity while target resistivity accuracy was lower, and MLP-NN produced sharper contrast at target boundaries for the field and hypothetical data. Both methods exhibited comparable target characterization performance, whereas MLP-NN had increased accuracy compared to Gauss-Newton in prediction of target width and height, which was attributed to numerical smoothing present in the Gauss-Newton approach. MLP-NN was also applied to a field dataset acquired at U.S. DOE Hanford site.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2023

To what extent can decommissioning options for marine artificial structures move us toward environmental targets?

Switching from fossil fuels to renewable energy is key to international energy transition efforts and the move toward net zero. For many nations, this requires decommissioning of hundreds of oil and gas infrastructure in the marine environment. Current international, regional and national legislation largely dictates that structures must be completely removed at end-of-life although, increasingly, alternative decommissioning options are being promoted and implemented. Yet, a paucity of real-world case studies describing the impacts of decommissioning on the environment make decision-making with respect to which option(s) might be optimal for meeting international and regional strategic environmental targets challenging. To address this gap, we draw together international expertise and judgment from marine environmental scientists on marine artificial structures as an alternative source of evidence that explores how different decommissioning options might ameliorate pressures that drive environmental status toward (or away) from environmental objectives. Synthesis reveals that for 37 United Nations and Oslo-Paris Commissions (OSPAR) global and regional environmental targets, experts consider repurposing or abandoning individual structures, or abandoning multiple structures across a region, as the options that would most strongly contribute toward targets. This collective view suggests complete removal may not be best for the environment or society. However, different decommissioning options act in different ways and make variable contributions toward environmental targets, such that policy makers and managers would likely need to prioritise some targets over others considering political, social, economic, and ecological contexts. Current policy may not result in optimal outcomes for the environment or society.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2024

Radiation damage study of POCO ZXF-5Q graphite for neutrino production targets using 4.5 MeV helium ions

To address the challenges of increased beam power and target survivability associated with next-generation particle production beam lines, high dose, high-energy proton beam conditions are simulated using irradiation from low-energy ion beams. A low-energy ion irradiation study of POCO ZXF-5Q graphite under conditions similar to those of the NuMI NT-02 neutrino production target at the Fermi National Accelerator Laboratory is reported. Helium ion irradiation was performed at 100 ∘ C to a maximum damage level of 0.9 displacements per atom (DPA). Irradiation induced hardening, swelling of the irradiated region, inter-plane lattice expansion, and intraplane lattice contraction with increasing ion fluence was observed using micromechanical (nanoindentation, atomic force microscopy) and electron microscopy (high-resolution imaging, selected area diffraction) characterization. Similar changes were also observed in post irradiation examination of the NT-02 target indicating that ion irradiation can be a valuable tool for estimating radiation damage in proton beam targets. Caution must be exercised though, because the hardening, lattice alteration, and swelling occur to different magnitudes for a given damage level. The observed hardening and embrittlement were greater for ion irradiated graphite. For He ion irradiated samples the lattice spacing changes were smaller at low damage levels (78% less expansion and 71% less contraction at 0.1 DPA) and larger at high damage levels (38% more expansion and 5% more contraction at 0.9 DPA) relative to that observed in the NT-02 target. The magnitude of swelling was 8.5× greater under ion irradiation which is influenced by the differing damage gradients and inclusion of implanted He ions in the region of interest.

43 PARTICLE ACCELERATORS↗
DOE OSTI2023

Correlated characterization of 20 Ne-implanted targets using nuclear reaction analysis, Rutherford backscattering spectrometry, and ion transport modeling

In this article, we present the preparation and characterization of a large sample of implanted noble gas targets for use in precision nuclear astrophysics measurements with intense proton beams. Tantalum and titanium backings were prepared using wet-acid etching, outgassed via resistive heating, and implanted with 20 Ne + beams from differing ion sources. These experimental targets were investigated using both nuclear reaction analysis techniques on the 1169-keV resonance in 20 Ne(p, $\gamma$) 21 Na and Rutherford backscattering spectrometry analysis with 2-MeV α-particle beams. Results from these analyses reveal small target-to-target variations in stoichiometry, while exhibiting excellent agreement independent of ion-beam analysis method. We also present a self-consistent validation of the nuclear reaction analysis results using ion transport simulations in TRIM-2013 that rely on input parameters from SIMNRA scattering-yield fits to Rutherford backscattering spectra. In addition to a complete description of the implantation profile, this analysis method provides an alternate solution for characterizing a large sample of implanted targets when no suitable resonances are available for nuclear reaction analysis.

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND ↗
DOE OSTI2022

Model calibration of the liquid mercury spallation target using evolutionary neural networks and sparse polynomial expansions

The mercury constitutive model predicting the strain and stress in the target vessel plays a central role in improving the lifetime prediction and future target designs of the mercury targets at the Spallation Neutron Source. We leverage the experiment strain data collected over multiple years to improve the mercury constitutive model through a combination of large scale simulations of the target behavior and the use of machine learning tools for parameter estimation. We present two interdisciplinary approaches for surrogate-based model calibration of expensive simulations using evolutionary neural networks and sparse polynomial expansions. The newly calibrated simulations achieve 7% average improvement on the prediction accuracy and 8% reduction in mean absolute error compared to previously reported reference parameters, with some individual sensors experiencing up to 30% improvement. The calibrated simulations can aid in fatigue analysis to estimate the mercury target lifetime, which reduces abrupt failure and saves tremendous amount of costs.

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND ↗
DOE OSTI2021

The NRG3/ERBB4 signaling cascade as a novel therapeutic target for canine glioma

Highlights: • miR-190a was downregulated in canine brain tumors. • miR-190a functioned as a tumor-suppressor through targeting NRG3. • A pan-ERBB inhibitor exhibited a tumor suppressive effect in canine glioma cells. Canine glioma is a common brain tumor with poor prognosis despite surgery and/or radiation therapy. Therefore, newer and more effective treatment modalities are needed. Neuregulin 3 (NRG3) has known to be a ligand of ERBB4. This study aimed to investigate the usefulness of the NRG3/ERBB4 signaling cascade as a novel therapeutic target in canine glioma. We found out that microRNA (miR)-190a was downregulated in canine brain tumor tissues, including glioma and meningioma. miR-190a directly targeted NRG3 and inhibited the growth of canine glioma cells. The level of p-Akt, which is a downstream target of ERBB4 signaling, was decreased by transfection with miR-190a. NRG3 silencing also suppressed cell growth and decreased the levels of p-Akt and p-ERK1/2, and NRG3 overexpression exhibited opposed effects in canine glioma J3T-1 cells. The mRNA level of erbb4 was significantly upregulated in glioma tissues compared with that in normal brain tissues and meningioma tissues. Furthermore, compared with gefitinib and lapatinib, afatinib exerted a greater inhibitory effect on the growth of canine glioma cells. In conclusion, NRG3/ERBB4 signaling is negatively regulated by miR-190a and contributes to the growth of canine glioma cells, indicating that it may be a promising therapeutic target in canine glioma.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

Bifunctional Janus Particles as Multivalent Synthetic Nanoparticle Antibodies (SNAbs) for Selective Depletion of Target Cells

Monoclonal antibodies (mAb) have had a transformative impact on treating cancers and immune disorders. SNAbs However, their use is limited by high development time and monetary cost, manufacturing complexities, suboptimal pharmacokinetics, and availability of disease-specific targets. To address some of these challenges, we developed an entirely synthetic, multivalent, Janus nanotherapeutic platform, called Synthetic Nanoparticle Antibodies (SNAbs). SNAbs, with phage-display-identified cell-targeting ligands on one "face" and Fc-mimicking ligands on the opposite "face", were synthesized using a custom, multistep, solid-phase chemistry method. SNAbs efficiently targeted and depleted myeloid-derived immune-suppressor cells (MDSCs) from mouse-tumor and rat-trauma models, ex vivo. Systemic injection of MDSC-targeting SNAbs efficiently depleted circulating MDSCs in a mouse triple-negative breast cancer model, enabling enhanced T cell and Natural Killer cell infiltration into tumors. Finally, our results demonstrate that SNAbs are a versatile and effective functional alternative to mAbs, with advantages of a plug- and-play, cell-free manufacturing process, and high-throughput screening (HTS)-enabled library of potential targeting ligands.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2025

Utilizing Machine Learning to Improve Neutralization Potency of an HIV-1 Antibody Targeting the gp41 N-Heptad Repeat

The N-heptad repeat (NHR) of the HIV-1 gp41 prehairpin intermediate (PHI) is an attractive potential vaccine target with high sequence conservation across diverse strains. However, despite the potency of NHR-targeting peptides and clinical efficacy of the NHR-targeting entry inhibitor enfuvirtide, no potently neutralizing NHR-directed monoclonal antibodies (mAbs) nor antisera have been identified or elicited to date. The lack of potent NHR-binding mAbs both dampens enthusiasm for vaccine development efforts at this target and presents a barrier to performing passive immunization experiments with NHR-targeting antibodies. To address this challenge, we previously developed an improved variant of the NHR-directed mAb D5, called D5_AR, which is capable of neutralizing diverse tier-2 viruses. Building on that work, here we present the 2.7Å-crystal structure of D5_AR bound to NHR mimetic peptide IQN17. We then utilize protein language models and supervised machine learning to generate small (n < 100) libraries of D5_AR variants that are subsequently screened for improved neutralization potency. We identify a variant with 5-fold improved neutralization potency, D5_FI, which is the most potent NHR-directed monoclonal antibody characterized to date and exhibits broad neutralization of tier-2 and −3 pseudoviruses as well as replicating R5 and X4 challenge strains. Additionally, our work highlights the ability of protein language models to efficiently identify improved mAb variants from relatively small libraries.

Biopolymers↗
DOE OSTI2020

Marker-free carotenoid-enriched rice generated through targeted gene insertion using CRISPR-Cas9

Targeted insertion of transgenes at pre-determined plant genomic safe harbors provides a desirable alternative to insertions at random sites achieved through conventional methods. Most existing cases of targeted gene insertion in plants have either relied on the presence of a selectable marker gene in the insertion cassette or occurred at low frequency with relatively small DNA fragments (<1.8 kb). Here, we report the use of an optimized CRISPR-Cas9-based method to achieve the targeted insertion of a 5.2 kb carotenoid biosynthesis cassette at two genomic safe harbors in rice. We obtain marker-free rice plants with high carotenoid content in the seeds and no detectable penalty in morphology or yield. Whole-genome sequencing reveals the absence of off-target mutations by Cas9 in the engineered plants. These results demonstrate targeted gene insertion of marker-free DNA in rice using CRISPR-Cas9 genome editing, and offer a promising strategy for genetic improvement of rice and other crops.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Design of diverse, functional mitochondrial targeting sequences across eukaryotic organisms using variational autoencoder

Mitochondria play a key role in energy production and metabolism, making them a promising target for metabolic engineering and disease treatment. However, despite the known influence of passenger proteins on localization efficiency, only a few protein-localization tags have been characterized for mitochondrial targeting. To address this limitation, we leverage a Variational Autoencoder to design novel mitochondrial targeting sequences. In silico analysis reveals that a high fraction of the generated peptides (90.14%) are functional and possess features important for mitochondrial targeting. We characterize artificial peptides in four eukaryotic organisms and, as a proof-of-concept, demonstrate their utility in increasing 3-hydroxypropionic acid titers through pathway compartmentalization and improving 5-aminolevulinate synthase delivery by 1.62-fold and 4.76-fold, respectively. Moreover, we employ latent space interpolation to shed light on the evolutionary origins of dual-targeting sequences. Overall, our work demonstrates the potential of generative artificial intelligence for both fundamental research and practical applications in mitochondrial biology.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Targeting of intracellular oncoproteins with peptide-centric CARs

The majority of oncogenic drivers are intracellular proteins, constraining their immunotherapeutic targeting to mutated peptides (neoantigens) presented by individual human leukocyte antigen (HLA) allotypes. However, most cancers have a modest mutational burden that is insufficient for generating responses using neoantigen-based therapies. Neuroblastoma is a paediatric cancer that harbours few mutations and is instead driven by epigenetically deregulated transcriptional networks. Here we show that the neuroblastoma immunopeptidome is enriched with peptides derived from proteins essential for tumorigenesis. We focused on targeting the unmutated peptide QYNPIRTTF discovered on HLA-A*24:02, which is derived from the neuroblastoma-dependency gene and master transcriptional regulator PHOX2B. To target QYNPIRTTF, we developed peptide-centric chimeric antigen receptors (PC-CARs) through a counter panning strategy using predicted potentially cross-reactive peptides. We further proposed that PC-CARs can recognize peptides on additional HLA allotypes when presenting a similar overall molecular surface. Informed by our computational modelling results, we show that PHOX2B PC-CARs also recognize QYNPIRTTF presented by HLA-A*23:01, the most common non-A2 allele in people with African ancestry. Finally, we demonstrate potent and specific killing of neuroblastoma cells expressing these HLAs in vitro and complete tumour regression in mice. These data suggest that PC-CARs have the potential to expand the pool of immunotherapeutic targets to include non-immunogenic intracellular oncoproteins and allow targeting through additional HLA allotypes in a clinical setting.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

Template and target-site recognition by human LINE-1 in retrotransposition

The long interspersed element-1 (LINE-1, hereafter L1) retrotransposon has generated nearly one-third of the human genome and serves as an active source of genetic diversity and human disease. L1 spreads through a mechanism termed target-primed reverse transcription, in which the encoded enzyme (ORF2p) nicks the target DNA to prime reverse transcription of its own or non-self RNAs. Here we purified full-length L1 ORF2p and biochemically reconstituted robust target-primed reverse transcription with template RNA and target-site DNA. We report cryo-electron microscopy structures of the complete human L1 ORF2p bound to structured template RNAs and initiating cDNA synthesis. The template polyadenosine tract is recognized in a sequence-specific manner by five distinct domains. Among them, an RNA-binding domain bends the template backbone to allow engagement of an RNA hairpin stem with the L1 ORF2p C-terminal segment. Moreover, structure and biochemical reconstitutions demonstrate an unexpected target-site requirement: L1 ORF2p relies on upstream single-stranded DNA to position the adjacent duplex in the endonuclease active site for nicking of the longer DNA strand, with a single nick generating a staggered DNA break. Our research provides insights into the mechanism of ongoing transposition in the human genome and informs the engineering of retrotransposon proteins for gene therapy.

59 BASIC BIOLOGICAL SCIENCES↗