Biology subjects

Lange, S.

Publications and source records attributed to Lange, S..

At least 19 recordsLinked to original sources

bioRxiv2026

Cardiomyocyte Perm1 sustains cardiac bioenergetic reserve and exercise capacity

Objective: Mitochondrial oxidative capacity in striated muscle is governed by the PGC-1/ERR transcriptional axis, which controls not only steady-state respiration but the capacity to expand energy production when demand rises. Whether this reserve function requires dedicated downstream effectors, and whether its loss is detectable at rest, remains unclear. We asked whether Perm1, a striated muscle-specific PGC-1/ERR-induced regulator, is required cell-autonomously for cardiac bioenergetic reserve. Methods: We generated cardiomyocyte-specific Perm1 knockout mice using alphaMHC-nCre, complemented by an adult tamoxifen-inducible alphaMHC-MerCreMer model, and assessed cardiac development, structure, and function longitudinally through 12 months of age in both sexes. We evaluated mitochondrial function by high-resolution respirometry, transmission electron microscopy, and molecular profiling. We tested energetic capacity under load by treadmill exercise with tissue ATP quantification and by beta-adrenergic stimulation of cultured neonatal cardiomyocytes. Results: Perm1 deletion did not perturb embryonic cardiac development, and neither constitutive nor adult-onset deletion impaired cardiac structure, systolic function, or diastolic function through 12 months. Despite this preserved resting phenotype, mitochondrial respiration was reduced by 20 - 25% across complex I- and complex I+II-supported conditions, without changes in citrate synthase activity, mtDNA copy number, or OXPHOS subunit abundance. Instead, the MICOS components IMMT (MIC60) and CHCHD3 (MIC19) were selectively reduced, and mitochondrial subpopulations shifted toward fewer subsarcolemmal and more interfibrillar mitochondria, with unchanged total mitochondrial density. Functionally, Perm1-deficient mice of both sexes showed reduced treadmill capacity at 6 and 12 months and failed to maintain cardiac ATP during exercise at a submaximal workload, while skeletal muscle ATP and plasma lactate were unaffected. This deficit was cardiomyocyte-autonomous: beta-adrenergic stimulation increased ATP content in control cardiomyocytes, but Perm1 knockdown reduced this response. Conclusions. Perm1 is a cardiomyocyte-intrinsic determinant of bioenergetic reserve, dispensable for resting cardiac function but required to expand ATP production under increased demand. These findings identify a mode of metabolic regulation that is invisible to resting functional phenotyping.

physiology↗

bioRxiv2026

Dynamic Responses to an Inflammatory Challenge Distinguish Metabolic Health Across Lean and Obese Individuals

Inflammation is a key driver of cardiometabolic disease, yet it remains unclear whether systemic inflammatory markers can distinguish metabolically healthy from unhealthy individuals or capture the temporal dynamics of inflammation. Here, we combined systemic immune profiling with a cantharidin-induced peripheral blister model to investigate dynamic inflammatory regulation across metabolic phenotypes in metabolically healthy lean (MHL), metabolically unhealthy lean (MUL), metabolically healthy obese (MHO), and metabolically unhealthy obese (MUO) individuals. While systemic inflammation was elevated in obesity, differences between metabolically healthy and unhealthy groups were modest, with limited discrimination by plasma proteomics, circulating leukocyte phenotyping, and whole-blood transcriptomics. In contrast, the dynamic response to inflammatory challenge revealed pronounced differences at proteomic, cellular, and transcriptomic levels. Metabolically unhealthy individuals exhibited exaggerated early innate immune responses, impaired inflammatory resolution and tissue repair, reduced recruitment of reparative immune cells, and sustained T cell presence. Transcriptomic analyses further showed blunted dynamic gene regulation and defective epidermal barrier restoration. These findings indicate that metabolic health is better reflected in tissue-level inflammatory dynamics than in systemic measures.

immunology↗

bioRxiv2026

Atomic structure and dynamics of the mechanosensitive channel MscL from E. coli by cryo-EM and solid-state NMR

Mechanosensitive channels are central to cellular responses to membrane tension, yet the structural basis of their gating remains incompletely understood. Here, we determine the structures of wild-type and G22S mutants of MscL from E. coli (EcMscL) by cryo-EM in peptide-based lipid nanodiscs and complement them with solid-state NMR measurements in liposomes to capture their dynamics in a native-like membrane environment. The cryo-EM structures reveal a closed conformation, whereas analysis of the low-threshold G22S mutant by NMR uncovers widespread conformational changes in both cytoplasmic and periplasmic regions. These data indicate enhanced dynamics and conformational heterogeneity in the mutant, revealing the early transitions from the closed towards the open state. Together, our results establish a synergistic framework integrating cryo-EM and NMR to resolve both structure and dynamics of mechanosensitive channels, and identify lipid-protein interactions as key determinants of MscL gating and mechanosensitivity. Our study further provides a quantitative benchmark for computational investigations of mechanogating and lays the foundation for the rational design of channels with tunable gating kinetics. TeaserBy integrating cryo-EM and solid-state NMR, we reveal how lipid-coupled dynamics prime MscL for opening, capturing the earliest transitions from closed to active states.

biophysics↗

bioRxiv2026

Blindness reshapes mental time travel: From perceptual scenes to conceptual scaffolds

How can humans remember and imagine without vision? Mental time travel, the ability to re-experience past events and envision future ones, is widely assumed to rely on visual imagery and the construction of mental scenes. Blindness provides a critical test of this assumption. Across behavioral interviews, language analyses, and multimodal neuroimaging in congenitally blind, late-blind, and sighted individuals, we show that blind individuals, even those blind from birth, mentally time travel as vividly as sighted people, but construct their inner worlds differently. Sighted participants relied on perceptual detail and activated classic scene-processing regions, whereas blind participants emphasized thoughts and emotions and recruited reorganized occipital cortex. Connectivity analyses revealed strengthened coupling between occipital and medial temporal regions, indicating adaptive reconfiguration of the episodic system. The brain does not require images to imagine: it flexibly builds internal experiences using the representational resources available. Summary ParagraphHow humans reconstruct events that are no longer available to the senses is a fundamental but unresolved question. Remembering the past and imagining the future, known as mental time travel, is a defining feature of human cognition, shaping identity and guiding decisions{superscript 1},{superscript 2}. Prevailing theories assume that such constructions depend on visual imagery, with the minds eye reconstructing events on a visuospatial stage3,. Blindness provides a critical test of this assumption. Across extensive behavioral interviews and multimodal neuroimaging, we find that mental time travel remains phenomenologically intact in blindness, but its scaffolding changes fundamentally. Sighted individuals rely on perceptual detail and activate regions specialized for visual scenes, whereas blind individuals, whether blind from birth or later in life, emphasize thoughts and emotions and reorganize occipital cortex for conceptual strategies,. These contrasting strategies map onto distinct neural signatures, revealing a dissociation between perceptual and conceptual routes to episodic simulation. Together, these findings reveal that the brains capacity to construct internal experience rests on conceptual scaffolding, not perceptual re-creation.

neuroscience↗

bioRxiv2026

Lipoxins Regulate Intercalated Disk-Associated Signaling and Immune Remodeling in Dilated Cardiomyopathy

We investigated whether pro-resolving lipid mediators of the lipoxin family can attenuate fibrosis and inflammation in muscle LIM protein knockout (MLPko) mice, a model of dilated cardiomyopathy (DCM). Male and female MLPko mice received either vehicle or a mix of lipoxin-A4 and lipoxin-B4 three times per week for six weeks. Cardiac function was assessed using echocardiography, and fibrosis and DCM-associated cardiac signaling was evaluated through histology, immunofluorescence and immunoblot analyses. Flow cytometry and RNA sequencing (RNAseq) was performed to identify changes in cardiac gene expression and characterize macrophage subpopulations, respectively. Flow cytometry showed increased inflammatory CD11c+ M1-like macrophages and reduction of CD206+ M2-like macrophages in MLPko hearts compared to wild-type controls. Lipoxin treatment partially reversed the macrophage imbalance and showed mild improvements in cardiac physiology in MLPko males. RNAseq analyses revealed sex-dependent alterations in the expression of pro-fibrotic and inflammation-related genes, suggesting changes in extracellular matrix (ECM) integrity and composition, and to the adaptive immune response. Intriguingly, several ECM proteins showed unexpected localizations at cardiac intercalated disks, which are known to be involved in DCM etiology. Further analysis identified lipoxin-dependent reduction in the DCM-associated expression of intercalated disk components only in lipoxin-treated MLPko males. Lipoxins also modulated key cardiac signaling pathways in a sex-specific manner, including Erk1/2 and PKC-linked Ankrd1/Carp1, which is associated with DCM development in MLPko mice. While lipoxins do not directly reverse cardiac dysfunction or fibrosis in MLPko mice, they may provide sex-specific protective effects by modulating DCM-related cardiac signaling pathways and by influencing immune-cell populations.

cell biology↗

bioRxiv2026

Glycoprotein G enables HSV-2 neuroinvasion and provides protection as a glycosylated vaccine antigen

The function of glycoprotein G (gG-2) of herpes simplex virus 2 (HSV-2) during genital infection is unknown. gG-2 is cleaved into a secreted variant (sgG-2) and a membrane associated variant (mgG-2). This work delineates the glycan profile of mgG-2, and demonstrates that mgG-2 is strongly immunogenic, eliciting glycan dependent humoral and Th1-polarized CD4+ T cell responses in a mouse vaccination model. The N- and O-glycosylation of mgG-2 is important to achieve full protection against HSV-2 infection as immunization with deglycosylated mgG-2 resulted in poor CD4+ T cell activation and viral spread to dorsal root ganglia and spinal cord. Furthermore, an mgG-2 negative HSV-2 mutant virus failed to spread to the dorsal root ganglia and the central nervous system of genitally infected mice, despite viral replication in vaginal cells. Our data demonstrate that mgG-2 is important for HSV-2 propagation and that adaptive immune responses targeting the glycosylated protein can prevent neuronal infection, identifying mgG-2 as a promising vaccine candidate.

microbiology↗

bioRxiv2026

Atomic structure and plasticity of the MthK-CTX complex investigated by cryo-EM, NMR, and MD simulations

Scorpion toxins block potassium channels, disrupting cellular excitability and causing symptoms such as pain, muscle spasms, or paralysis. Here, we use an integrated structural biology approach to uncover the binding mode of the scorpion toxin charybdotoxin (CTX) to the MthK channel, a model system for human large-conductance potassium (BK) channels. Cryo-EM defines the overall architecture of the MthK-CTX complex, while complementary solution- and solid-state NMR experiments identify key binding residues and show that toxin engagement alters the selectivity filter (SF) ion configuration without rearranging the filter itself. NMR and MD simulations further reveal an anchoring lysine residue stably inserted into the SF, while other contacts undergo fast NMR timescale dynamics. Together, these findings explain how CTX-like toxins maintain exceptionally high affinity while tolerating binding across multiple K+ channel subtypes, paving the way for site-specific extracellular modulation.

molecular biology↗

bioRxiv2025

Atomistic Mechanism of Calcium-Mediated Inward Rectification of the MthK Potassium Channel by Solid-State NMR and MD Simulations

Inward rectification is a fundamental but poorly understood phenomenon in potassium channel physiology. Despite its physiological importance, the exact mechanism has remained elusive. In this work, we uncover a previously unrecognized calcium-mediated gating mechanism in the MthK potassium channel that sheds new light on this essential process. By combining state-of-the-art proton-detected solid-state NMR spectroscopy with atomistic molecular dynamics simulations, we reveal that divalent calcium ions bind to a novel site just below the selectivity filter, physically obstructing the outward flow of potassium ions whereas inward flow is still possible - analogous to a molecular ball check valve. Secondly, the binding of Ca2+ to the newly identified site leads to stabilization of the selectivity filter and allows us to directly observe ion-ion interactions in the filter. These results offer direct experimental support for the long-debated "direct knock-on" mechanism, in which potassium ions move through the filter, without water co-transport.

biophysics↗

bioRxiv2025

The titin N2A-MARP signalosome constrains muscle longitudinal hypertrophy in response to stretch

Titin-based mechanosensing is a key driver of trophic signaling in muscle, yet the downstream pathways linking titin sensing to muscle remodeling remain poorly understood. To investigate these signaling mechanisms, we utilized unilateral diaphragm denervation (UDD), an in vivo model that induces titin-stiffness-dependent hypertrophy via mechanical stretch. Using UDD in rats and mice, we characterized the longitudinal hypertrophic response and distinguished stretch-induced signaling from denervation effects by performing global transcriptomic and proteomic analyses following UDD and bilateral diaphragm denervation (BDD) in rats. Our findings identified upregulation of titin-associated muscle ankyrin repeat proteins (MARPs). Subsequent phosphorylation enrichment mass spectrometry in mouse diaphragm highlighted the involvement of the N2A-element. UDD in MARP knockout (KO) mice resulted in enhanced longitudinal hypertrophy, with Western blot analysis revealing activation of the mTOR pathway. Furthermore, pharmacological inhibition of mTORC1 with rapamycin suppressed longitudinal hypertrophy, demonstrating that mTOR signaling regulates titin-mediated hypertrophic growth in a MARP-dependent manner. These findings establish MARPs as key modulators of titin-based mechanotransduction and highlight mTORC1 as a central regulator of longitudinal muscle hypertrophy.

physiology↗

bioRxiv2024

KRas, in addition to Tp53 is a driver for early carcinogenesis and a molecular target in a mouse model of invasive gastro-esophageal adenocarcinoma

ObjectiveThe incidence of gastro-esophageal adenocarcinoma (GEAC) has increased dramatically and is associated with Barretts Esophagus (BE). Gastric cardia progenitors are the likely origin for BE and GEAC. Here we analyze p53, Rb1 and Kras alterations in Lgr5 progenitor cells during carcinogenesis. DesignWe introduced single and combined genetic alterations (p53, Rb1 and Kras) in Lgr5-expressing progenitor cells at the inflamed gastroesophageal junction in the L2-IL1b (L2) mouse model crossed to Lgr5-CreERTmice. For in-vitro treatment we utilized mouse and human 3D organoids. ResultsInactivation of Tp53 or Rb1 alone (L2-LP and L2-LR mice) resulted in metaplasia, and mild dysplasia, while expression of KrasG12D (L2-LK) accelerated dysplasia in L2-IL1b mice. Dual induction of genetic alteration in L2-LPR, L2-LKP and L2-LKR mice confirmed the accelerating role of mutant Kras, with the development of invasive cancer in mice with combined Tp53 and Kras alteration. All three genetic events in cardia progenitor cells generated invasive cancer at 6 months of age, with chromosomal instability (CNV). The dominant role of Kras prompted us to treat with a SHP2 inhibitor in combination with an ERK or MEK inhibitor, leading to reduced growth in Kras mutant organoids. SHP2 and MEK inhibition in-vivo reduced Kras dependent tumor formation. ConclusionIn the first invasive GEAC mouse model, Kras mutation in combination with loss of tumor suppressor genes Tp53 or Rb1 emerges as a key player in GEAC and with importance of p53 and Rb1 in promoting metaplasia. Targeting this SHP2/MEK/KRAS pathway represents a promising therapeutic option for Kras altered GEAC. What is already known on this topicThe increased incidence of GEAC is challenging current screening and surveillance strategies. Therapeutic and preventive options are limited due to a lack of knowledge on the role of genetic alterations commonly associated with GEAC and their function during progression to dysplasia. What this study addsWe generate the first invasive GEAC model and show that KRAS at least in combination with a second genetic alterations (Tp53 and/or Rb1) may be a driver of tumorigenesis, and targeting KRAS alterations could be a promising now treatment substitution. How this study might affect research, practice or policyTargeting KRAS alterations will be important for GEAC, especially as specific KRAS inhibitor are on the horizon. In addition, a concept of single genetic alteration inducing metaplasia as an adaptation to chronic inflammation might emerge as an important factor for surveillance.

cancer biology↗

bioRxiv2023

Mechanism of human PINK1 activation at the TOM complex in a reconstituted system

Loss of function mutations in PTEN-induced kinase 1 (PINK1) are a frequent cause of early-onset Parkinsons disease (PD). Stabilisation of PINK1 at the Translocase of Outer Membrane (TOM) complex of damaged mitochondria is a critical step for its activation. To date the mechanism of how PINK1 is activated in the TOM complex is unclear. Herein we report co-expression of human PINK1 and all seven TOM subunits in Saccharomyces cerevisiae is sufficient for PINK1 activation. We use this reconstitution system to systematically assess the role of each TOM subunit towards PINK1 activation. We unambiguously demonstrate that the TOM20 and TOM70 receptor subunits are required for optimal PINK1 activation and map their sites of interaction with PINK1 using AlphaFold structural modelling and mutagenesis. We also demonstrate an essential role of the pore-containing subunit TOM40 and its structurally associated subunits TOM7 and TOM22 for PINK1 activation. These molecular findings will aid in the development of small molecule activators of PINK1 as a therapeutic strategy for PD.

biochemistry↗

bioRxiv2023

In-situ spectroscopic detection of large-scale reorientations of transmembrane α-helices during viroporin channel opening

Viroporins are small ion channels in membranes of enveloped viruses that play key roles during viral life cycles. To use viroporins as drug targets against viral infection requires in-depth mechanistic understanding and, with that, methods that enable investigations under in-situ conditions. Here, we apply surface-enhanced infrared absorption (SEIRA) spectroscopy to Influenza A M2 reconstituted within a solid-supported membrane, to shed light on the mechanics of its viroporin function. M2 is a paradigm of pH-activated proton channels and controls the proton flux into the viral interior during viral infection. We use SEIRA to track the large-scale reorientation of M2s transmembrane -helices in-situ during pH-activated channel opening. We quantify this event as a helical tilt from 26{degrees} to 40{degrees} by correlating the experimental results with solid-state NMR-informed computational spectroscopy. This mechanical motion is impeded upon addition of the inhibitor rimantadine, giving a direct spectroscopic marker to test antiviral activity. The presented approach provides a spectroscopic tool to quantify large-scale structural changes and to track the function and inhibition of the growing number of viroporins from pathogenic viruses in future studies.

biophysics↗

bioRxiv2023

Community-based Reconstruction and Simulation of a Full-scale Model of Region CA1 of Rat Hippocampus

The CA1 region of the hippocampus is one of the most studied regions of the rodent brain, thought to play an important role in cognitive functions such as memory and spatial navigation. Despite a wealth of experimental data on its structure and function, it has been challenging to reconcile information obtained from diverse experimental approaches. To address this challenge, we present a community-driven, full-scale in silico model of the rat CA1 that integrates a broad range of experimental data, from synapse to network, including the reconstruction of its principal afferents, the Schaffer collaterals, and a model of the effects that acetylcholine has on the system. We tested and validated each model component and the final network model, and made input data, assumptions, and strategies explicit and transparent. The unique flexibility of the model allows scientists to address a range of scientific questions. In this article, we describe the methods used to set up simulations that reproduce and extend in vitro and in vivo experiments. Among several applications in the article, we focus on theta rhythm, a prominent hippocampal oscillation associated with various behavioral correlates and use our computer model to reproduce and reconcile experimental findings. Finally, we make data, code and model available through the hippocampushub.eu portal, which also provides an extensive set of analyses of the model and a user-friendly interface to facilitate adoption and usage. This neuroscience community-driven model represents a valuable tool for integrating diverse experimental data and provides a foundation for further research into the complex workings of the hippocampal CA1 region.

neuroscience↗

bioRxiv2023

An Atypical E3 Ligase Module in UBR4 Mediates Destabilization of N-degron Substrates

UBR4 is an E3 ligase (E3) of the N-degron pathway and is involved in neurodevelopment, age-associated muscular atrophy and cancer progression. The location and mechanistic classification of the E3 module within the 600 kDa protein UBR4 remains unknown. Herein, we identify and characterize, at a biochemical and structural level, a distinct E3 module within human UBR4 consisting of a novel "hemiRING" zinc finger, a helical-rich UBR Zinc-finger Interacting (UZI) subdomain, and a predicted backside interacting N-terminal helix. A structure of an E2 conjugating enzyme (E2)-E3 complex provides atomic level insight into the exquisite specificity of the hemiRING towards the E2s UBE2A/B. The UZI subdomain can be considered a component of the E3 module as it has a modest activating effect on the ubiquitin loaded E2 (E2[~]Ub), which is complemented by the intrinsically high lysine reactivity of UBE2A. These findings reveal the mechanistic underpinnings of a neuronal N-degron E3 ligase, its specific recruitment of UBE2A, and highlight the underappreciated architectural diversity of cross-brace domains associated with ubiquitin E3 activity.

biochemistry↗

bioRxiv2022

Proteogenomic analysis reveals RNA as an important source for tumor-agnostic neoantigen identification correlating with T-cell infiltration

Systemic pan-tumor analyses may reveal the significance of common features implicated in cancer immunogenicity and patient survival. Here, we provide a comprehensive multi-omics data set for 32 patients across 25 tumor types by combining proteogenomics with phenotypic and functional analyses. By using an optimized computational approach, we discovered a large number of novel tumor-specific and tumor-associated antigens including shared common target candidates. To create a pipeline for the identification of neoantigens in our cohort, we combined deep DNA and RNA sequencing with MS- based immunopeptidomics of tumor specimens, followed by the assessment of their immunogenicity. In fact, we could detect a broad variety of non-wild type HLA-binding peptides in the majority of patients and confirmed the immunogenicity of 24 neoantigens. Most interestingly, the majority of total and immunogenic neoantigens originated from variants identified in the RNA dataset, illustrating the importance of RNA as a still understudied source of cancer antigens. Moreover, the amount of these mainly RNA-based immunogenic neoantigens correlated positively with overall CD8+ tumor-infiltrating T cells. This study therefore underlines the importance of RNA-centered variant detection for the identification of shared biomarkers and potentially relevant neoantigen candidates. Statement of significanceThe significance of this study lies not only in the potential of our optimized proteogenomic workflow for the discovery of neoantigens (in particular RNA-derived neoantigens) for clinical application, but sheds light on the entity-agnostic prevalence of HLA class I peptide presentation of RNA processing events to be used for tumor targeting.

cancer biology↗

bioRxiv2022

Interaction with C21ORF2 controls the cellular functions of the NEK1 kinase

NEK1 is a pleiotropic protein kinase implicated in mitosis, ciliogenesis and DNA repair but little is known about its regulation or targets. Its relevance for human health is underscored by the association of NEK1 mutations with human diseases including axial spondylometaphyseal dyplasia (SMD) and amyotrophic lateral sclerosis (ALS). Mutations in the C21ORF2 gene are associated with a similar pattern of human diseases, suggesting close functional links with NEK1. Here we report that in unperturbed, untransformed cells, endogenous NEK1 and C21ORF2 form a tight complex that does not appear to contain other proteins. A small acidic domain "CID: C21ORF2 interaction domain" at the C-terminus of NEK1 is necessary and sufficient to interact with C21ORF2, and pathogenic mutations in this region disrupt the complex. AlphaFold modelling predicts with high confidence an extended binding interface between a leucine-rich repeat (LRR) domain in the N-terminal half of C21ORF2 and a stretch of the NEK1-CID; mutating residues mediating electrostatic interactions within this interface disrupts the NEK1-C21ORF2 interaction. This model also explains why pathogenic mutations disrupt the complex. We go on to show that the kinase activity of NEK1 and its interaction with C21ORF2 is critical for NEK1 function in cells. These data reveal C21ORF2 as a regulatory subunit of NEK1, illuminating our understanding of how this kinase is regulated and NEK1-C21ORF2-associated diseases.

biochemistry↗

bioRxiv2022

Combined loss of obscurin and obscurin-like 1 in murine hearts results in impaired diastolic dysfunction, altered metabolism and deregulated mitophagy.

Muscle proteins of the obscurin protein family play important roles in sarcomere organization, sarcoplasmic reticulum (SR) and T-tubule architecture and function. However, their precise molecular functions and redundancies between protein family members as well as their involvement in cardiac diseases remain to be fully understood. To investigate the functional roles of obscurin and its close homologue obscurin-like 1 (Obsl1) in the heart, we generated and analyzed knockout mice for obscurin, Obsl1, as well as obscurin/Obsl1 double-knockouts (dKO). We show that dKO mice are viable but show postnatal deficits in cardiac muscle SR and mitochondrial architecture and function at the microscopic, biochemical and cellular level. Altered SR structure resulted in perturbed calcium cycling, while mitochondrial ultrastructure deficits were linked to decreased levels of Chchd3, a Micos complex protein. Hearts of dKO mice also show increased expression of Atg4d, a novel Obsl1 interacting protein, resulting in abnormal mitophagy and increased unfolded protein response. At the physiological level, loss of obscurin and Obsl1 resulted in a profound delay of cardiac relaxation, associated with metabolic signs of heart failure. Taken together, our data suggest that obscurin and Obsl1 play crucial roles in cardiac SR structure, calcium cycling, mitochondrial function, turnover and metabolism.

cell biology↗

bioRxiv2021

Spatial and temporal plasticity of neoantigen-specific T-cell responses bases on characteristics associated to antigen and TCR

Neoantigens derived from somatic mutations have been demonstrated to correlate with therapeutic responses mediated by treatment with immune checkpoint inhibitors. Neoantigens are therefore highly attractive targets for the development of personalized medicine approaches although their quality and associated immune responses is not yet well understood. In a case study of metastatic malignant melanoma, we performed an in-depth characterization of neoantigens and respective T-cell responses in the context of immunotherapy with Ipilimumab. Three neoantigens identified either by immunopeptidomics or in silico prediction were investigated using binding affinity analyses and structural simulations. We isolated seven T-cell receptors (TCRs) from the patient immune repertoire recognizing these antigens. TCRs were compared in-vitro and in-vivo with multi-parametric analyses. Identified immunogenic peptides showed similar binding affinities to the human leukocyte antigen (HLA) complex and comparable differences to their wildtype counterparts in molecular dynamic simulations. Nevertheless, isolated TCRs differed substantially in functionality and frequency. In fact, TCRs with comparably lower functional avidity and higher potential for cross-reactivity provided at least equal anti-tumor immune responses in vivo. Of note, these TCRs showed a reduced activation pattern upon primary in vitro stimulation. Exploration of the TCR-{beta} repertoire in blood and in different tumor-related tissues over three years, offered insights on the high frequency and particular long-term persistence of low-avidity TCRs. These data indicate that qualitative differences of neoantigen-specific TCRs and their impact on function and longevity need to be considered for neoantigen targeting by adoptive T-cell therapy using TCR-transgenic T cells. Statement of translational relevanceImmunotherapy has demonstrated high efficacy in diverse malignancies. Neoantigens derived from mutations provide promising targets for safe and highly tumor-specific therapeutic approaches. Yet, single determinants of an effective and enduring T-cell mediated tumor rejection are still not well understood. We analyzed in detail seven neoantigen-specific T-cell receptors (TCRs) derived from a melanoma patient targeting three different altered peptide ligands identified by mass spectrometry and prediction analyses. Functional characterization of these TCRs revealed potent anti-tumor reactivity of all TCRs. Of special interest, TCRs with comparably lower affinity demonstrated effective in vivo activity as well as dominant spatial and temporal distribution in blood and tissue. Functional differences of TCR may require further T-cell and/or TCR engineering and should be considered for future clinical trial designs.

immunology↗