Reconstruction of Orbital Exenteration Defects with Cross-median Forehead Flaps.

The aim of this study was to examine the role of forehead flaps in the reconstruction of orbital defects after exenteration. Patients undergoing orbital exenteration and reconstruction using median forehead flaps from January 2002 to August 2019 were enrolled in this retrospective study. All the patients were evaluated for wound complications, functional results, and tumor relapse. In total, 105 patients completed the study. Twenty-nine complications in 24 of these patients were assessed. The most common complications were wound dehiscence (11%), partial necrosis of the flap (6%), and bleeding (4%). A major complication occurred in two patients (2%), necessitating surgical correction. Only one patient had a local relapse. Sixty patients received osseointegrated implants, and 58 of them were treated with facial prostheses. The follow-up periods ranged from 6 to 189 months. Thus, forehead flap reconstruction after exenteration is a reliable method with a low complication rate. This technique can be used for primary or secondary reconstruction, and excellent long-term functional results can be achieved. A relapse-oriented follow-up is certainly possible.

nf-core/nanostring: a pipeline for reproducible NanoString nCounter analysis.

MOTIVATION: The NanoString™ nCounter® technology platform is a widely used targeted quantification platform for the analysis of gene expression of up to ∼800 genes. Whereas the software tools by the manufacturer can perform the analysis in an interactive and GUI driven approach, there is no portable and user-friendly workflow available that can be used to perform reproducible analysis of multiple samples simultaneously in a scalable fashion on different computing infrastructures. RESULTS: Here, we present the nf-core/nanostring open-source pipeline to perform a comprehensive analysis including quality control and additional features such as expression visualization, annotation with additional metadata and input creation for differential gene expression analysis. The workflow features an easy installation, comprehensive documentation, open-source code with the possibility for further extensions, a strong portability across multiple computing environments and detailed quality metrics reporting covering all parts of the pipeline. nf-core/nanostring has been implemented in the Nextflow workflow language and supports Docker, Singularity, Podman container technologies as well as Conda environments, enabling easy deployment on any Nextflow supported compatible system, including most widely used cloud computing environments such as Google GCP or Amazon AWS. AVAILABILITY AND IMPLEMENTATION: The source code, documentation and installation instructions as well as results for continuous tests are freely available at https://github.com/nf-core/nanostring and https://nf-co.re/nanostring.

Solitary Fibrous Tumours of the Orbit - Clinical Pathology, Therapy and Prognosis. / Solitär fibröse Tumoren der Orbita ­ klinisch-pathologische Charakteristik, Therapie und Prognose.

BACKGROUND: Solitary fibrous tumours are rare. The aim of this study is to describe the clinical features, therapy and outcome of affected patients and to identify factors associated with recurrence. METHODS: Retrospective study of a cohort of 20 patients who underwent surgery for orbital solitary fibrous tumour at the University Department of Oral and Maxillofacial Surgery between 2002 and 2023. Demographic, clinical, and therapeutic data as well as tumour follow-up results were collected. Tumour volume and molecular genetic mutations were retrospectively determined. RESULTS: The median patient age was 49.5 years at initial surgery. The left orbit was affected in 65% of cases. The most common clinical symptom was proptosis (80%). This was reported with a mean lateral difference of 3.9 mm (range: 1 - 10 mm). The tumours were localised predominantly in the intra- and extraconal space, craniolateral quadrant and middle third. The median tumour volume was 7.66 cm³ (range 2.15 - 12.57 cm³). In all patients, the diagnosis was made by pathological examination. All tumours investigated showed a NAB2-STAT6 mutation. The most frequently detected mutation was the fusion NAB2 exon 4 - STAT6 exon 2. All patients were initially managed with frontolateral orbitotomy. Incomplete resection (R1-status) occurred in 35% (n = 7). The recurrence rate was 25% (n = 5), with a median disease-free interval of 45.5 months (range 23 - 130). 80% (n = 4) of recurrences were initially R1-resected. CONCLUSION: Orbital solitary fibrous tumours are rare tumours and are clinically manifested by signs of displacement of orbital structures. Diagnosis is made by histology and immunohistochemistry and can be proven with the molecular genetic detection of the NAB2-STAT6 mutation. The therapy of choice is complete surgical resection. R1-resection is more likely in the intraconal location as well as in location in the posterior third of the orbit - due to difficult surgical accessibility. The greatest risk factor for the development of recurrence is incomplete surgical excision. Late recurrences are possible, which is why a long-term connection to a specialised clinic is necessary.

MRI-based long-term follow-up of indolent orbital lymphomas after curative radiotherapy: imaging remission criteria and volumetric regression kinetics.

We systematically analyzed the kinetics of tumor regression, the impact of residual lesions on disease control and the applicability of the Lugano classification in follow-up MRI of orbital non-Hodgkin lymphomas that were irradiated with photons. We retrospectively analyzed a total of 154 pre- and post-irradiation MRI datasets of 36 patients with low-grade, Ann-Arbor stage I, orbital non-Hodgkin lymphomas. Patients with restricted conjunctival involvement were excluded. Lymphoma lesions were delineated and volumetrically analyzed on T1-weighted sequences. Tumor residues were present in 91.2% of all cases during the first six months after treatment. Volumetric partial response rates (> 50% volume reduction) were 75%, 69.2%, and 50% at 12-24 months, 36-48 months and > 48 months after the end of treatment. The corresponding complete response (CR) rates according to the Lugano classification were 20%, 23.1% and 50%. During a median clinical follow-up of 37 months no significant differences in progression free survival (PFS) rates were observed between the CR and non-CR group (p = 0.915). A residual tumor volume below 20% of the pretreatment volume should be expected at long-term follow-up beyond one year after radiotherapy.

Novel Insights into Pathophysiology of Orbital Inflammatory Diseases and Progression to Orbital Lymphoma by Pathway Enrichment Analysis.

Non-specific orbital inflammation (NSOI) and IgG4-related orbital disease (IgG4-ROD) are currently treated with non-specific immunosuppressive agents based on non-randomized, uncontrolled studies. Therefore, relapses and prolongated courses are common and remain challenging. For a more specific therapy, a better understanding of the underlying pathophysiology is crucial. Therefore, we aimed to analyze signaling pathways to expand the knowledge on the pathophysiology and possibly identify specific targets in the future, as occurred recently in Graves' orbitopathy with the IGF-1 receptor. Furthermore, we analyzed potential mechanisms for the described potential progression to orbital MALT (mucosa-associated lymphoid tissue) lymphoma. The investigation cohort for this screening study comprised of 12 patients with either typical NSOI (n = 6), IgG4-ROD or MALT lymphoma (n = 3 each). Mean age was 56.4 ± 17 years. MALT samples, in contrast with IgG4-ROD and NSOI, showed overall upregulation for extracellular matrix receptor interaction (ECM) and adipocytokine signaling. Investigating signaling compounds for MALT samples, differentially expressed genes were re-identified as targets with relevant expression. Even though pathway analysis showed differentially altered products when comparing IgG4-ROD with MALT, main conductors of differentiation in B- and T-cell signaling were commonly altered when observing the microenvironment of examined tissues. Our data reveal the characteristic differences and similarities in genetic-expression-based pathway profiles between MALT lymphoma, IgG4-ROD and NSOI, which may be useful for elucidating the associated pathogenic mechanisms and developing specific treatments for these orbital diseases.

Modeling receptor flexibility in the structure-based design of KRASG12C inhibitors.

KRAS has long been referred to as an 'undruggable' target due to its high affinity for its cognate ligands (GDP and GTP) and its lack of readily exploited allosteric binding pockets. Recent progress in the development of covalent inhibitors of KRASG12C has revealed that occupancy of an allosteric binding site located between the α3-helix and switch-II loop of KRASG12C-sometimes referred to as the 'switch-II pocket'-holds great potential in the design of direct inhibitors of KRASG12C. In studying diverse switch-II pocket binders during the development of sotorasib (AMG 510), the first FDA-approved inhibitor of KRASG12C, we found the dramatic conformational flexibility of the switch-II pocket posing significant challenges toward the structure-based design of inhibitors. Here, we present our computational approaches for dealing with receptor flexibility in the prediction of ligand binding pose and binding affinity. For binding pose prediction, we modified the covalent docking program CovDock to allow for protein conformational mobility. This new docking approach, termed as FlexCovDock, improves success rates from 55 to 89% for binding pose prediction on a dataset of 10 cross-docking cases and has been prospectively validated across diverse ligand chemotypes. For binding affinity prediction, we found standard free energy perturbation (FEP) methods could not adequately handle the significant conformational change of the switch-II loop. We developed a new computational strategy to accelerate conformational transitions through the use of targeted protein mutations. Using this methodology, the mean unsigned error (MUE) of binding affinity prediction were reduced from 1.44 to 0.89 kcal/mol on a set of 14 compounds. These approaches were of significant use in facilitating the structure-based design of KRASG12C inhibitors and are anticipated to be of further use in the design of covalent (and noncovalent) inhibitors of other conformationally labile protein targets.

Long-term follow-up and health-related quality of life among cancer survivors with stage IEA orbital-type lymphoma after external photon-beam radiotherapy: Results from a longitudinal study.

We assessed the long-term outcomes and treatment-related adverse effects of patients with Stage I, "orbital-type" lymphomas that were uniformly treated with photons. All consecutive patients diagnosed with low-grade, Ann Arbor Stage IEA orbital lymphoma treated between 1999 and 2020 at our department were retrospectively reviewed. We excluded patients with exclusive conjunctival involvement, typically treated with en face electrons. In order to quantify radiotherapy related side effects we applied the CTCAE criteria, analyzed changes in visual acuity, quantified dry eye symptoms by use of the Ocular Surface Disease Index (OSDI) score and applied the EORTC QLQ-C30 questionnaire for quality of life (QoL) assessment. In total 66 eyes of 62 patients were irradiated with a median dose of 30.6 Gy. The median follow-up was 43.5 months. The predominant histological subtype were MALT lymphomas. No local failure occurred in this cohort. Of nine outfield relapses, six solely occurred in the contralateral eye. The 5- and 10- years distant progression free survival rates (PFS) were 81.4% and 63.5%. The 5- and 10-years overall survival rates were 85.1% and 71.9% without any tumor related death. Of the acute toxicities none was higher than CTCAE grade 1. The predominant late toxicities were dry eyes (21.2%) of CTCAE Grade <2 and radiation induced cataracts (19.7%). During long-term follow up the average visual acuity did not deteriorate. The global QoL was worst before treatment and improved significantly after 24 months (p = 0.007). External beam radiotherapy of "orbital-type" lymphomas with photons is an effective and gentle treatment option with excellent local control rates. From the high control rates the trend to use slightly lower total doses of 24-27 Gy with conventional fractionation is supported. As non-coplanar radiotherapy techniques improved and total doses can slightly be reduced, the current status of radiotherapy as first line therapy is provided.

Conjunctival and uveal melanoma: Survival and risk factors following orbital exenteration.

PURPOSE: This study aims to analyse disease-free survival, overall survival and risk factors after orbital exenteration in patients with conjunctival and uveal melanoma. METHODS: Patients who underwent orbital exenteration due to conjunctival and uveal melanoma were included in this retrospective study (March 2000 to March 2018). RESULTS: A total of 76 patients were enrolled in this study: 60 patients had a conjunctival melanoma and 16 had a uveal melanoma. In conjunctival melanoma, the mean age was 68.4 years. The overall survival rate was 82% after 1 year and 52% after 5 years. Univariate analysis of overall survival found that the following parameters were predictive of a worse prognosis: gender, extent of the primary tumour, lymph node metastases, distant metastases, adjuvant chemotherapy or radiotherapy and relapse. In multivariate analysis, relapse and adjuvant radiotherapy appeared to contribute to a significantly worse prognosis. In uveal melanoma, the mean age was 63.6 years. Eleven patients died during follow-up (mean follow up 30.7 months). The overall survival and disease-free survival rates after 1 year were 62% and 57%, respectively. An analysis of risk factors was not possible due to the small number of cases. CONCLUSION: Orbital exenterations in conjunctival and uveal melanoma are rarely necessary, but can be performed as an ultima ratio treatment with curative intent. Disease-free survival and overall survival are significantly lower for both groups due to the advanced stage of the disease compared to patients treated without exenteration in the literature. If a recurrence occurs after exenteration, the prognosis is poor in both groups.

Next generation Fc scaffold for multispecific antibodies.

Bispecific antibodies (Bispecifics) demonstrate exceptional clinical potential to address some of the most complex diseases. However, Bispecific production in a single cell often requires the correct pairing of multiple polypeptide chains for desired assembly. This is a considerable hurdle that hinders the development of many immunoglobulin G (IgG)-like bispecific formats. Our approach focuses on the rational engineering of charged residues to facilitate the chain pairing of distinct heavy chains (HC). Here, we deploy structure-guided protein design to engineer charge pair mutations (CPMs) placed in the CH3-CH3' interface of the fragment crystallizable (Fc) region of an antibody (Ab) to correctly steer heavy chain pairing. When used in combination with our stable effector functionless 2 (SEFL2.2) technology, we observed high pairing efficiency without significant losses in expression yields. Furthermore, we investigate the relationship between CPMs and the sequence diversity in the parental antibodies, proposing a rational strategy to deploy these engineering technologies.

Orbital exenteration: Symptoms, indications, tumour localizations, pathologies, reconstruction, complications and survival.

OBJECTIVE: This study aims to evaluate malignant and benign diseases that lead to orbital exenteration. PATIENTS: From December 1999 to September 2017, patients undergoing orbital exenteration were included in this retrospective study. All of them were evaluated on clinical symptoms, indications, tumour localizations, pathologies, reconstruction techniques, complications, recurrences, and survival. RESULTS: Of the 205 patients enrolled in this study, 94 had a carcinoma, 73 melanoma, 9 a sarcoma, 14 some other malignant disease, and 15 a benign medical condition. Sixteen patients underwent reconstruction using a local eyelid skin flap (7.8%), 6 with a split-thickness graft (2.9%), 144 with a local flap (70.2%), and 25 with a microvascular graft (12.2%), whereas 14 patients did not undergo reconstruction (6.8%). The most common complications were wound dehiscences (25 cases), pain (17 cases), and partial flap necroses (13 cases). Moreover, 62% of the patients were treated with different facial prostheses or artificial eyes. Given these results, it appears that lymph nodes and distant metastases, as well as lymphatic invasion into vessels, perineural invasion, and non-cleared resection margins, seem to affect overall survival after orbital exenteration. CONCLUSION: Different reconstruction techniques can be used to provide the patient with maximum functionality and aesthetics after orbital exenteration. Individual concepts should be discussed at the beginning of the treatment. Using primary reconstruction and providing osseointegrated implant-retained prostheses remain the gold standard.

Reconstruction of Orbital Exenteration Defects with Cheek Rotation Flaps: Indications, Technique, Complications, Rehabilitation, and Survival.

The objective of this study was to examine the role of cheek rotation flaps in the reconstruction of orbital defects after exenteration. From January 2000 to August 2018, patients undergoing orbital exenteration and reconstruction with cheek rotation flaps were enrolled in this retrospective study. All patients were evaluated for wound complications, orbital rehabilitation, tumor relapse, and survival. Thirty patients completed the study. Fourteen complications allocated to 11 patients were assessed. The most common complications were seroma (13%), temporary facial nerve weakness (13%), and partial necrosis of the flap (10%). A major complication occurred in a total of two patients (7%), so that surgical correction was necessary. Eleven patients had a relapse; 15 patients died as part of the follow-up. Fifteen patients were treated with facial prostheses. The overall survival rate was 61% after 1 year and 42% after 5 years. Follow-up periods ranged from 6 to 95 months. Cheek rotation flap reconstruction after exenteration is a reliable method with a low rate of major complications. It is indicated when an approach to the parotid gland or the neck region is necessary because of suspected lymph node metastasis and in elderly patients because of their skin's laxity. It can be performed as primary or secondary reconstruction. Good esthetic results can be achieved, especially after endosseous implantation.

Periorbital, conjunctival and primary intraorbital carcinomas: Survival and risk factors after orbital exenteration.

OBJECTIVES: This study aimed to analyse the disease-free survival (DFS), overall survival (OS) and risk factors after orbital exenteration in patients with periorbital, conjunctival and primary intraorbital carcinomas. METHODS: Patients undergoing orbital exenteration due to a primary carcinoma between March 2000 and March 2018 were included in this retrospective study. Risk factors in all the patients were evaluated using univariate and multivariate analyses. RESULTS: In total, 97 patients were enroled in this study. The most common tumours were conjunctival carcinoma (35 cases), squamous cell carcinoma of the skin (27 cases) and basal cell carcinoma (20 cases). The median follow-up period was 36 months. The average age of the patients was 67.3 years (range, 29-93 years). In all the patients, OS was 85% after 1 year and 69% after 5 years, while DFS was 71% after 1 year and 55% after 5 years. Univariate analysis of OS revealed that the following parameters were predictive of a poor prognosis: localisation, neck dissection, lymph node metastases, lymphatic invasion, perineural invasion, resection margins and immunosuppression. Multivariate analysis revealed resection margins as the only independent risk factor. CONCLUSION: Orbital exenteration is rarely necessary in patients with periorbital, conjunctival and primary intraorbital carcinomas; however, it can be performed as an ultima ratio treatment with a curative intent. Clear margins can be achieved in most cases. OS and DFS are not significantly different in the subgroups. In most cases, recurrence occurs within the first 2 years.

Modified types of orbital exenteration, survival, and reconstruction.

PURPOSE: This study aims to analyse the various modifications of orbital exenteration. METHODS: Patients undergoing orbital exenteration from March 1978 to October 2019 were included in this retrospective study. The patients were evaluated on the basis of the indication, type of exenteration, reconstruction technique, overall survival (OS), and disease-free survival (DFS). RESULTS: In total, 300 patients were enrolled in this study. As many as 24 patients had lid and conjunctiva sparing anterior exenteration, 16 had lid sparing anterior exenteration, 83 had anterior exenteration, 14 had lid and conjunctiva sparing total exenteration, seven had lid sparing total exenteration, 44 had total exenteration, one had lid and conjunctiva sparing extended exenteration, 23 had lid sparing extended exenteration, and 88 had extended exenteration. As many as 39 patients had a primary wound closure. Six patients underwent reconstruction with a split-thickness skin graft, 177 patients with a local or regional flap, and 40 patients with a microvascular flap. A total of 38 patients did not undergo reconstruction. The mean follow-up was 40 months (range 6-216 months). The OS rate was 82.2% after 1 year, 58.5% after 5 years, and 49% after 10 years for all patients with malignant tumours. The DFS rate was 67.7% after 1 year, 45.6% after 5 years, and 31.7% after 10 years. CONCLUSION: Individual types of orbital exenteration allow patient-adapted therapies. The preservation of uninvolved orbital tissue facilitates orbital reconstruction. The type of exenteration did not have any influence on overall survival.

Treatment of odontogenic abscesses: comparison of primary and secondary removal of the odontogenic focus and antibiotic therapy.

PURPOSE: To examine the time of removal of the odontogenic focus, antibiotic therapy and risk factors in odontogenic abscesses. PATIENTS: From January 2012 to December 2015, inpatients undergoing incision due to odontogenic abscesses were identified in a retrospective study. All the patients were evaluated for time of removal of the odontogenic focus, antibiotic therapy, germ spectrum, complications and risk factors. RESULTS: Two hundred ten patients completed the study. In 89 cases (42.4%), the odontogenic focus was removed as part of the abscess treatment (group A). In 121 cases (57.6%), the focus was secondarily removed (group B). On average, 2 ± 4 teeth were removed in group A, and 6 ± 5 teeth in group B (p < 0.0001). An average of 1.2 ± 0.4 surgical interventions were performed in group A, and 2 ± 0.2 operations in group B (p < 0.0001). Microbiological examination was positive in one-third of the cases (70 cases). Most commonly, streptococci (27%) were isolated. A resistance screening was possible in 57 of the detected germs (68.7%). In 89% of these patients, the combination of ampicillin-sulbactam was effective. The hospital stay was 4.8 ± 2 days for group A and 7.6 ± 3 days for group B (p < 0.0001). The clinical evaluation revealed 12 intermediate (5.7%) and three long-term (1.4%) complications. The long-term complications included a recurrence in two cases (1%) and an osteomyelitis in one case (0.5%). A logistic regression analysis identified no significant risk factor in relation to these complications. CONCLUSION: The study shows that a primary removal of the odontogenic focus may have advantages over a secondary removal: (1) fewer operations, (2) shorter hospital stay and (3) shorter antibiotic therapy. Broad-spectrum penicillins in combination with beta-lactamase inhibitors are a possible, sufficient antibiotic regimen. Long-term complications are rare. No risk factors are identified in relation to these complications.

Molecular Insight into Recognition of the CGRPR Complex by Migraine Prevention Therapy Aimovig (Erenumab).

Calcitonin-gene-related peptide (CGRP) plays a key role in migraine pathophysiology. Aimovig (erenumab; erenumab-aooe in the United States) is the only US Food and Drug Administration (FDA)-approved monoclonal antibody (mAb) therapy against the CGRP receptor (CGRPR) for the prevention of migraine. Aimovig is also the first FDA-approved mAb against a G-protein-coupled receptor (GPCR). Here, we report the architecture and functional attributes of erenumab critical for its potent antagonism against CGRPR. The crystal structure of erenumab in complex with CGRPR reveals a direct ligand-blocking mechanism, enabled by a remarkable 21-residue-long complementary determining region (CDR)-H3 loop, which adopts a tyrosine-rich helix-turn tip and projects into the deep interface of the calcitonin receptor-like receptor (CLR) and RAMP1 subunits of CGRPR. Furthermore, erenumab engages with residues specific to CLR and RAMP1, providing the molecular basis for its exquisite selectivity. Such structural insights reveal the drug action mechanism of erenumab and shed light on developing antibody therapeutics targeting GPCRs.

Discovery of a Covalent Inhibitor of KRASG12C (AMG 510) for the Treatment of Solid Tumors.

KRASG12C has emerged as a promising target in the treatment of solid tumors. Covalent inhibitors targeting the mutant cysteine-12 residue have been shown to disrupt signaling by this long-"undruggable" target; however clinically viable inhibitors have yet to be identified. Here, we report efforts to exploit a cryptic pocket (H95/Y96/Q99) we identified in KRASG12C to identify inhibitors suitable for clinical development. Structure-based design efforts leading to the identification of a novel quinazolinone scaffold are described, along with optimization efforts that overcame a configurational stability issue arising from restricted rotation about an axially chiral biaryl bond. Biopharmaceutical optimization of the resulting leads culminated in the identification of AMG 510, a highly potent, selective, and well-tolerated KRASG12C inhibitor currently in phase I clinical trials (NCT03600883).

The clinical KRAS(G12C) inhibitor AMG 510 drives anti-tumour immunity.

KRAS is the most frequently mutated oncogene in cancer and encodes a key signalling protein in tumours1,2. The KRAS(G12C) mutant has a cysteine residue that has been exploited to design covalent inhibitors that have promising preclinical activity3-5. Here we optimized a series of inhibitors, using novel binding interactions to markedly enhance their potency and selectivity. Our efforts have led to the discovery of AMG 510, which is, to our knowledge, the first KRAS(G12C) inhibitor in clinical development. In preclinical analyses, treatment with AMG 510 led to the regression of KRASG12C tumours and improved the anti-tumour efficacy of chemotherapy and targeted agents. In immune-competent mice, treatment with AMG 510 resulted in a pro-inflammatory tumour microenvironment and produced durable cures alone as well as in combination with immune-checkpoint inhibitors. Cured mice rejected the growth of isogenic KRASG12D tumours, which suggests adaptive immunity against shared antigens. Furthermore, in clinical trials, AMG 510 demonstrated anti-tumour activity in the first dosing cohorts and represents a potentially transformative therapy for patients for whom effective treatments are lacking.

MHCquant: Automated and Reproducible Data Analysis for Immunopeptidomics.

Personalized multipeptide vaccines are currently being discussed intensively for tumor immunotherapy. In order to identify epitopes-short, immunogenic peptides-suitable for eliciting a tumor-specific immune response, human leukocyte antigen-presented peptides are isolated by immunoaffinity purification from cancer tissue samples and analyzed by liquid chromatography-coupled tandem mass spectrometry (LC-MS/MS). Here, we present MHCquant, a fully automated, portable computational pipeline able to process LC-MS/MS data automatically and generate annotated, false discovery rate-controlled lists of (neo-)epitopes with associated relative quantification information. We could show that MHCquant achieves higher sensitivity than established methods. While obtaining the highest number of unique peptides, the rate of predicted MHC binders remains still comparable to other tools. Reprocessing of the data from a previously published study resulted in the identification of several neoepitopes not detected by previously applied methods. MHCquant integrates tailor-made pipeline components with existing open-source software into a coherent processing workflow. Container-based virtualization permits execution of this workflow without complex software installation, execution on cluster/cloud infrastructures, and full reproducibility of the results. Integration with the data analysis workbench KNIME enables easy mining of large-scale immunopeptidomics data sets. MHCquant is available as open-source software along with accompanying documentation on our website at https://www.openms.de/mhcquant/ .

Discovery of N-(1-Acryloylazetidin-3-yl)-2-(1H-indol-1-yl)acetamides as Covalent Inhibitors of KRASG12C.

KRAS regulates many cellular processes including proliferation, survival, and differentiation. Point mutants of KRAS have long been known to be molecular drivers of cancer. KRAS p.G12C, which occurs in approximately 14% of lung adenocarcinomas, 3-5% of colorectal cancers, and low levels in other solid tumors, represents an attractive therapeutic target for covalent inhibitors. Herein, we disclose the discovery of a class of novel, potent, and selective covalent inhibitors of KRASG12C identified through a custom library synthesis and screening platform called Chemotype Evolution and structure-based design. Identification of a hidden surface groove bordered by H95/Y96/Q99 side chains was key to the optimization of this class of molecules. Best-in-series exemplars exhibit a rapid covalent reaction with cysteine 12 of GDP-KRASG12C with submicromolar inhibition of downstream signaling in a KRASG12C-specific manner.

Low mutational load in pediatric medulloblastoma still translates into neoantigens as targets for specific T-cell immunotherapy.

BACKGROUND: Medulloblastoma is the most common malignant brain tumor in childhood and adolescence. Although some patients present with distinct genetic alterations, such as mutated TP53 or MYC amplification, pediatric medulloblastoma is a tumor entity with minimal mutational load and low immunogenicity. METHODS: We identified tumor-specific mutations using next-generation sequencing of medulloblastoma DNA and RNA derived from primary tumor samples from pediatric patients. Tumor-specific mutations were confirmed using deep sequencing and in silico analyses predicted high binding affinity of the neoantigen-derived peptides to the patients' human leukocyte antigen molecules. Tumor-specific peptides were synthesized and used to induce a de novo T-cell response characterized by interferon gamma and tumor necrosis factor alpha release of CD8+ cytotoxic T cells in vitro. RESULTS: Despite low mutational tumor burden, at least two immunogenic tumor-specific peptides were identified in each patient. T cells showed a balanced CD4/CD8 ratio and mostly effector memory phenotype. Induction of a CD8-specific T-cell response was achieved for the neoepitopes derived from Histidine Ammonia-Lyase (HAL), Neuraminidase 2 (NEU2), Proprotein Convertase Subtilisin (PCSK9), Programmed Cell Death 10 (PDCD10), Supervillin (SVIL) and tRNA Splicing Endonuclease Subunit 54 (TSEN54) variants. CONCLUSION: Detection of patient-specific, tumor-derived neoantigens confirms that even in tumors with low mutational load a molecular design of targets for specific T-cell immunotherapy is possible. The identified neoantigens may guide future approaches of adoptive T-cell transfer, transgenic T-cell receptor transfer or tumor vaccination.