Essential role of pre-existing humoral immunity in TLR9-mediated type I IFN response to recombinant AAV vectors in human whole blood.

Recombinant adeno-associated virus (AAV) vectors have emerged as the preferred platform for gene therapy of rare human diseases. Despite the clinical promise, host immune responses to AAV vectors and transgene remain a major barrier to the development of successful AAV-based human gene therapies. Here, we assessed the human innate immune response to AAV9, the preferred serotype for AAV-mediated gene therapy of the CNS. We showed that AAV9 induced type I interferon (IFN) and IL-6 responses in human blood from healthy donors. This innate response was replicated with AAV6, required full viral particles, but was not observed in every donor. Depleting CpG motifs from the AAV transgene or inhibiting TLR9 signaling reduced type I IFN response to AAV9 in responding donors, highlighting the importance of TLR9-mediated DNA sensing for the innate response to AAV9. Remarkably, we further demonstrated that only seropositive donors with preexisting antibodies to AAV9 capsid mounted an innate immune response to AAV9 in human whole blood and that anti-AAV9 antibodies were necessary and sufficient to promote type I IFN release and plasmacytoid dendritic (pDC) cell activation in response to AAV9. Thus, our study reveals a previously unidentified requirement for AAV preexisting antibodies for TLR9-mediated type I IFN response to AAV9 in human blood.

A lateral approach allows accurate and stable total elbow replacement in dogs.

OBJECTIVE: Evaluate whether total elbow replacement (TER) through a lateral approach is accurate and stable. ANIMALS: 12 skeletally mature large-breed dog cadavers were used. METHODS: Limb alignment, elbow joint motion, and collateral ligament laxity were evaluated preoperatively. The order of surgery (left or right) and the approach (lateral or medial) were randomly selected for TER in each dog. The other approach was used in the contralateral elbow. Intraoperative technical difficulties, duration of surgery, and anatomic complications were recorded. Limb alignment, elbow joint motion, collateral ligament laxity, and prosthetic component alignment were evaluated after surgery. Data were collected from June 11 to 15, 2023. RESULTS: The duration of surgery using a lateral or medial approach did not differ (P = .499). Anatomic complications were not observed. The lateral approach resulted in 8° more elbow extension (P = .003), 1.58° less lateral collateral ligament constraint (P = .033), 2.80° less medial collateral ligament constraint (P = .002), 4.38° less frontal plane constraint (P = .004), 8° greater humeral component inclination (P = .033), and 5.6° greater radioulnar component varus (P = .001) than the medial approach. Varus of the radius, mechanical axis deviation, limb supination, elbow flexion, mediolateral humeral component and craniocaudal radioulnar component orientation did not differ among joints operated using a lateral or medial approach. In normal cadaveric elbows, a lateral approach for TER appears feasible, producing equivalent limb alignment, joint laxity, and joint motion to normal elbows and to TER placed using a medial approach. CLINICAL RELEVANCE: In dogs, TER can be performed using a lateral surgical approach.

[Radiation-induced lymphopenia: Lymphocytes as a new organ at risk]. / Lymphopénie radio-induite : les lymphocytes comme nouvel organe à risque.

Taking the immune system into account in the fight against tumors has upset the cancer treatment paradigm in the 21st century. Combination treatment strategies associating radiotherapy with immunotherapy are being increasingly implemented in clinical practice. In this context, lymphocytes, whether lymphocytes infiltrating the tumour, circulating blood lymphocytes or lymphocytes residing within the lymph nodes, are key players in cellular and humoral anti-tumor immunity. The significant radiosensitivity of lymphocytes was demonstrated in the early 1990s. Along with the cells of the digestive mucosa, lymphocytes are thus among the most radiosensitive cell types in the body. Compared to the old practices of external radiotherapy, current intensity modulated treatments have allowed a considerable improvement in acute and late toxicity, at the cost of a significant increase in the volume irradiated at low doses. This is not without consequence on the incidence of radiation-induced lymphopenia, with prognostic implications for many tumor types. Thus, in order not to hinder the action of antitumor immunity and the efficacy of immunotherapy, it is essential to consider lymphocytes as a new organ at risk in its own right. In this development, based on current data from the literature, we will begin by justifying the necessary prevention of radiation-induced lymphopenia, before providing the tools currently known to apprehend lymphocytes as a new multicompartments. Finally, we will broaden the perspective by outlining ways to develop research in this area.

A highly selective humanized DDR1 mAb reverses immune exclusion by disrupting collagen fiber alignment in breast cancer.

BACKGROUND: Immune exclusion (IE) where tumors deter the infiltration of immune cells into the tumor microenvironment has emerged as a key mechanism underlying immunotherapy resistance. We recently reported a novel role of discoidin domain-containing receptor 1 (DDR1) in promoting IE in breast cancer and validated its critical role in IE using neutralizing rabbit monoclonal antibodies (mAbs) in multiple mouse tumor models. METHODS: To develop a DDR1-targeting mAb as a potential cancer therapeutic, we humanized mAb9 with a complementarity-determining region grafting strategy. The humanized antibody named PRTH-101 is currently being tested in a Phase 1 clinical trial. We determined the binding epitope of PRTH-101 from the crystal structure of the complex between DDR1 extracellular domain (ECD) and the PRTH-101 Fab fragment with 3.15 Å resolution. We revealed the underlying mechanisms of action of PRTH-101 using both cell culture assays and in vivo study in a mouse tumor model. RESULTS: PRTH-101 has subnanomolar affinity to DDR1 and potent antitumor efficacy similar to the parental rabbit mAb after humanization. Structural information illustrated that PRTH-101 interacts with the discoidin (DS)-like domain, but not the collagen-binding DS domain of DDR1. Mechanistically, we showed that PRTH-101 inhibited DDR1 phosphorylation, decreased collagen-mediated cell attachment, and significantly blocked DDR1 shedding from the cell surface. Treatment of tumor-bearing mice with PRTH-101 in vivo disrupted collagen fiber alignment (a physical barrier) in the tumor extracellular matrix (ECM) and enhanced CD8+ T cell infiltration in tumors. CONCLUSIONS: This study not only paves a pathway for the development of PRTH-101 as a cancer therapeutic, but also sheds light on a new therapeutic strategy to modulate collagen alignment in the tumor ECM for enhancing antitumor immunity.

Closed reduction and fluoroscopic-guided percutaneous pinning of femoral capital physeal or neck fractures: Thirteen fractures in 11 dogs.

OBJECTIVES: To report the short-term clinical outcome and complications in dogs that underwent surgical repair of femoral capital physeal or neck fractures via fluoroscopic-guided percutaneous pinning (FGPP). STUDY DESIGN: Retrospective case series. ANIMALS: Client owned dogs (n = 11) with femoral capital physeal or neck fractures (n = 13). METHODS: Records (July 2018-July 2021) of dogs that underwent surgery for femoral capital physeal or neck fracture repair with FGPP from two hospitals were reviewed. Data collected included signalment, age, weight, preoperative lameness severity, fracture factors (etiology, Salter-Harris classification, time from injury to surgery, radiographic displacement), surgical factors (surgery time, number/size of implants, reduction quality) and outcome (follow-up examination findings, radiographic findings, complications). RESULTS: Most fractures (11/13) occurred secondary to trauma. The median time from injury to surgery was 5.5 days. There was mild radiographic displacement preoperatively in 10/13 fractures. Satisfactory fracture healing and limb function was achieved in 10/13 femurs. Complications occurred in 5/11 cases and included intra-articular implants, implant failure/nonunion, implant migration (2), and malunion. Of these five complications, two resolved with implant removal, and a salvage procedure was recommended in the remaining cases. Of the three cases requiring salvage procedures, two originally presented with radiographic evidence of fracture chronicity. CONCLUSIONS: FGPP can be used to successfully treat femoral head and neck fractures with appropriate case selection and precise surgical technique.

The impact of orthostatic intolerance on early ambulation following abdominal surgery in an enhanced recovery programme.

BACKGROUND: The prevalence of orthostatic intolerance on the day of surgery is more than 50% after abdominal surgery. The impact of orthostatic intolerance on ambulation on the day of surgery has been little studied. We investigated orthostatic intolerance and walking ability after colorectal and bariatric surgery in an enhanced recovery programme. METHODS: Eighty-two patients (colorectal: n = 46, bariatric n = 36) were included and analysed in this prospective study. Walk tests for 2 min (2-MWT) and 6 min (6-MWT) were performed before and 24 h after surgery, and 3 h after surgery for 2-MWT. Orthostatic intolerance characterised by presyncopal symptoms when rising was recorded at the same time points. Multivariate binary logistic regressions modelling the probability of orthostatic intolerance and walking inability were performed taking into account potential risk factors. RESULTS: Prevalence of orthostatic intolerance and walking inability was, respectively, 65% and 18% 3-hour after surgery. The day after surgery, patients' performance had greatly improved: approximately 20% of the patients experienced orthostatic intolerance, whilst only 5% of the patients were unable to walk. Adjusted binary logistic regressions demonstrated that age (p = .37), sex (p = .39), BMI (p = .74), duration of anaesthesia (p = .71) and type of surgery (p = .71) did not significantly influence walking ability. CONCLUSION: Our study confirms that orthostatic intolerance was frequent (~ 60%) 3-hour after abdominal surgery but prevented a 2-MWT only in ~20% of patients. No risk factors for orthostatic intolerance and walking inability were evidenced.

The HLA class-II immunopeptidomes of AAV capsids proteins.

Introduction: Gene therapies are using Adeno-associated viruses (AAVs) as vectors, but immune responses against the capsids pose challenges to their efficiency and safety. Helper T cell recognition of capsid-derived peptides bound to human leukocyte antigen (HLA) class II molecules is an essential step in the AAV-specific adaptive immunity. Methods: Using MHC-associated peptide proteomics, we identified the HLA-DR and HLA-DQ immunopeptidomes of the capsid proteins of three different AAV serotypes (AAV2, AAV6, and AAV9) from a panel of healthy donors selected to represent a majority of allele usage. Results: The identified sequences span the capsids of all serotypes, with AAV2 having the highest peptide count. For all the serotypes, multiple promiscuous peptides were identified and displayed by both HLA-DR and -DQ. However, despite high sequence homology, there were few identical peptides among AAV2, AAV6, and AAV9 immunopeptidomes, and none were promiscuous. Discussion: Results from this work represent a comprehensive immunopeptidomics research of potential CD4+ T cell epitopes and provide the basis for immunosurveillance efforts for safer and more efficient AAV-based gene therapies.

A prevalent and culturable microbiota links ecological balance to clinical stability of the human lung after transplantation.

There is accumulating evidence that the lower airway microbiota impacts lung health. However, the link between microbial community composition and lung homeostasis remains elusive. We combine amplicon sequencing and bacterial culturing to characterize the viable bacterial community in 234 longitudinal bronchoalveolar lavage samples from 64 lung transplant recipients and establish links to viral loads, host gene expression, lung function, and transplant health. We find that the lung microbiota post-transplant can be categorized into four distinct compositional states, 'pneumotypes'. The predominant 'balanced' pneumotype is characterized by a diverse bacterial community with moderate viral loads, and host gene expression profiles suggesting immune tolerance. The other three pneumotypes are characterized by being either microbiota-depleted, or dominated by potential pathogens, and are linked to increased immune activity, lower respiratory function, and increased risks of infection and rejection. Collectively, our findings establish a link between the lung microbial ecosystem, human lung function, and clinical stability post-transplant.

Discovery of Potent and Selective Methylenephosphonic Acid CD73 Inhibitors.

Solid tumors are often associated with high levels of extracellular ATP. Ectonucleotidases catalyze the sequential hydrolysis of ATP to adenosine, which potently suppresses T-cell and NK-cell functions via the adenosine receptors (A2a and A2b). The ectonucleotidase CD73 catalyzes the conversion of AMP to adenosine. Thus, increased CD73 enzymatic activity in the tumor microenvironment is a potential mechanism for tumor immune evasion and has been associated with poor prognosis in the clinic. CD73 inhibition is anticipated to restore immune function by skirting this major mechanism of adenosine generation. We have developed a series of potent and selective methylenephosphonic acid CD73 inhibitors via a structure-based design. Key binding interactions of the known inhibitor adenosine-5'-(α,ß-methylene)diphosphate (AMPCP) with hCD73 provided the foundation for our early designs. The structure-activity relationship study guided by this structure-based design led to the discovery of 4a, which exhibits excellent potency against CD73, exquisite selectivity against related ectonucleotidases, and a favorable pharmacokinetic profile.

Rumen function in goats, an example of adaptive capacity.

The aim of this Research Reflection is to describe the basic rumen function of goats and its modification in response to environmental factors, as well as to discuss similarities and differences when compared to other ruminants. In so doing we shall reveal the adaptive capacity of goats to harsh environments. The basic rumen function in goats is similar to other species of ruminants, as stressed by the opportunity to apply the updates of feeding systems for ruminants to goats. The rumen epithelium acts as a protective barrier between the rumen and the host, but it can be damaged by toxic compounds or acidosis. The rumen also plays an important role in water balance, both for dehydration and rehydration. Recent studies show that the microbiota exhibits a high fractional stability due to functional redundancy and resilience, but this needs more investigation. The microbial community structure differs between goats and cows, which explains the difference in sensitivity to milk fat depression following intake of high lipid diets. Goats also differ from other ruminants by their enhanced ability to feed-sort, but as with cows they can suffer from acidosis. Nevertheless, goats can be considered to be very resistant to environmental factors such as water stress, salt stress or heat stress, and this is especially so in some endogenous breeds. They also are able to detoxify tannins, polyphenols and other secondary metabolites. Some new trials involving feeding behaviour, microbiota and omics or approaches by meta-analyses or modelling will improve our knowledge of rumen function in goats.

Retinal relaxation following membrane peeling: Effect on vision, central macular thickness, and vector analysis of motion.

BACKGROUND: Epiretinal membranes (ERM) form as a result of an inward displacement of retinal structures. Removal of an ERM leads to the outward displacement of retinal vessels and visual improvement. PURPOSE: The aim of the study was to evaluate the direction and extent of displacement of retinal/superficial vascular structures after a membrane peeling procedure by means of image comparison and in selected cases, a vector analysis of displacement. METHODOLOGY: Scanning laser ophthalmoscope images of the retina of eyes undergoing ERM peeling were compared before and 6 months after surgery. Stratification was made between prominent and limited displacement, with assessment of visual acuity (VA), and central macular thickness (CMT). In three cases, using the optic nerve as reference, 50 landmarks were chosen within the posterior pole along large and small vascular structures allowing the construction of a vector map of displacement over 1 year. RESULTS: Nine eyes with prominent and six with limited displacement were assessed. Improvement in VA was similar for both groups, while CMT drop was greatest for the worst group. Vector analysis showed that most vascular movement occurs over the first 6 months, covers most of the posterior retina, is centered around distinct nodes, and may lead to several hundred micrometers of displacement. CONCLUSIONS: Superficial retinal relaxation has no direct implication on visual recovery. It originates in nodes of retinal contraction. Its extent can be significant, covering most of the posterior pole. RELEVANCE FOR PATIENTS: A better understanding of retinal relaxation following the peeling of ERM may help better understand when intervention is required and which part of the membrane is critical to surgical success.

Total hip replacement in dogs with contralateral pelvic limb amputation: A retrospective evaluation of 13 cases.

OBJECTIVE: To report the clinical characteristics, surgical management, and medium-term outcomes of total hip replacement (THR) performed in dogs with previous contralateral pelvic limb amputation. ANIMALS: Thirteen client-owned dogs. STUDY DESIGN: Multi-institutional retrospective clinical study. METHODS: Data recorded from medical records included signalment, indication for amputation and THR, and surgical complications. Implant positioning and complications were assessed on radiographs. Clinical outcomes were evaluated during follow-up examinations by one of the authors and through a mobility- and lifestyle-based questionnaire completed by owners. RESULTS: All 13 dogs had satisfactory clinical results at follow-up a median of 3 months (range, 2-36) after THR. No postoperative luxation was recorded. Four dogs had minor complications that did not require additional treatment. The only major complication was one failure of osseointegration of a cementless acetabular cup, and it was successfully revised. CONCLUSION: Total hip replacement resulted in satisfactory clinical results and acceptable morbidity in this population. CLINICAL SIGNIFICANCE: Total hip replacement should be considered in dogs with severe coxofemoral joint disease and contralateral pelvic limb amputation.

Radiotherapy for primary tumor in lung cancer with synchronous metastases: Overview from the past and proposal for the future.

The management of metastatic lung cancers, either of the small-cell (SCLC) or the non-small cell (NSCLC) subtype, largely based on systemic treatments so far, has been the subject of breakthrough advances over the past few years, with notably the wide use of immunotherapy changing the landscape of these harmful prognosis diseases. In parallel with this major progress, the increasing use of radiotherapy (RT) for the treatment of the primary thoracic lesion±the distant lesions, may contribute to improving the condition of these metastatic patients, both in terms of progression-free survival (PFS) and overall survival (OS). This review proposes to summarize and explain the findings provided by the different studies published in the last years experiencing RT of the primary tumor in metastatic lung cancers, either associated or not with the local ablative treatment of a low number of distant lesions. It will also expose the respective limits encountered in these studies and, in the light of all these elements, suggests various promising issues and fields of research for the future.

In-vivo and in-vitro impact of high-dose rate radiotherapy using flattening-filter-free beams on the anti-tumor immune response.

INTRODUCTION: Modern accelerators have the "flattening filter-free" (FFF) technique to deliver RT with a moderate high-dose rate, currently used in limited clinical indications. No scientifically established data are currently available on the possible effects of this high dose rate on the anti-tumor immune response. We therefore propose here to study these effects in a preclinical CT26 murine colorectal tumor model. MATERIAL AND METHODS: In-vitro, CT26 cells were irradiated on a Varian TrueBeam® linac at 3 different dose rates (4; 12 or 24 Gy/min) using the FFF mode. Activation of the anti-tumor immune response was evaluated by the analysis of induction of genes of the type I interferon pathway by RT-qPCR, and by the study of the induction of immunogenic death biomarkers. In-vivo, an efficacy study of RT delivering 16.5 Gy at 2 different dose rates was performed in immunocompetent Balb/c mice carrying CT26 syngeneic tumors, as well as an immunomonitoring analysed by flow cytometry and a transcriptomic analysis using RNA sequencing. Statistical analyzes were performed using non-parametric tests. RESULTS: In-vitro, no significant influence of an increase in FFF dose rate was shown for the induction of genes of the type I interferon pathway as well as for the studied immunogenic death markers (HMGB1 secretion). In-vivo, no difference in terms of tumor growth retardation between the 2 dose rates used was demonstrated, as well as for the composition of immune cell infiltrates within tumor microenvironment and the expression of immune checkpoints in immunomonitoring and RNAseq. CONCLUSION: In this study involving the CT26 model, no influence of a moderate high dose rate in FFF technique on the anti-tumor immune response was demonstrated, which would make studies of associations between RT and checkpoint inhibitors fit with this technique of RT. However, further explorations using other cellular models seem to be of interest.

PneumoLaus: Prévalence de troubles respiratoires fonctionnels dans un échantillon de la population lausannoise.

PneumoLaus: Prevalence of Lung Function Abnormalities in a Sample of the General Population of Lausanne Abstract. Reduced lung function predicts increased mortality. The prevalence of spirometric abnormalities depends on their definition, the references values used and the use or not of bronchodilation. In the PneumoLaus study, conducted between 2014 and 2017 in a sample of the general population of Lausanne, prevalence of chronic obstruction was 3,8 %, of reversible obstruction 2,5 % and of possible restriction 2,2 %. These numbers are lower than in other population studies. Men had more abnormal spirometry results than women, and ever-smokers more than never-smokers. Two thirds of participants with chronic obstruction, most of which without respiratory symptoms, were not aware of any lung disease.

Résumé. Une fonction pulmonaire réduite s'associe à une mortalité accrue. La prévalence de troubles respiratoires fonctionnels dépend de sa définition, des références employées et de l'utilisation ou non de bronchodilatateur. Dans l'étude PneumoLaus, conduite entre 2014 et 2017 dans un échantillon de la population de Lausanne, la prévalence d'obstruction chronique était de 3,8 %, d'obstruction réversible de 2,5 % et de possible restriction de 2,2 %, proportions plus basses que dans d'autres études populationnelles. Les hommes présentaient plus souvent des anomalies spirométriques comparé aux femmes et les fumeurs plus souvent que les non-fumeurs. Deux tiers des sujets avec obstruction chronique, pour la plupart sans symptômes respiratoires, ne se connaissaient pas de maladie pulmonaire.

Immunotherapy-Induced Airway Disease: A New Pattern of Lung Toxicity of Immune Checkpoint Inhibitors.

Immune checkpoint inhibitors (ICIs) have been shown to improve overall and progression-free survival in various cancers but have been associated with various immune-related adverse events (IRAEs), including interstitial lung disease, especially organizing pneumonia. We report 2 cases of isolated severe airway disease attributable to ICIs, a rarely reported pattern of lung toxicity. The first patient received nivolumab with or without ipilimumab in a randomized double-blind trial for locoregional metastatic melanoma. The second patient was treated with nivolumab for lung adenocarcinoma. An IRAE was suspected in both cases due to a temporal relationship between ICI initiation and symptom onset. ICIs were stopped, and high-dose prednisone, inhaled corticosteroids, and bronchodilators were administered, allowing a rapid clinical and functional improvement in Patient 1. In Patient 2, despite prolonged high-dose prednisone, only a stabilization of forced expiratory volume in 1 s could be achieved, and the disease course was complicated by respiratory infections resulting in further loss of lung function. The patient died 1 year later due to progression of metastatic disease. These 2 cases suggest that pulmonary IRAEs secondary to ICIs may present as isolated bronchitis or bronchiolitis, with variable outcomes following ICI withdrawal and systemic corticosteroids.

Interlocking Nails and Minimally Invasive Osteosynthesis.

Reviews of clinical outcomes led to the foundation of a new approach in fracture management known as biological osteosynthesis. As intramedullary rods featuring cannulations and locking devices at both extremities, interlocking nails are well suited for bridging osteosynthesis. Unique biological and mechanical benefits make them ideal for minimally invasive nail osteosynthesis and an attractive, effective alternative to plating, particularly in revisions of failed plate osteosynthesis. Thanks to a new angle-stable locking design, interlocking nailing indications have been expanded to osteosynthesis of epi-metaphyseal fractures, including those with articular involvement and angular deformities such as distal femoral varus and associated patellar luxations.

[Lower limb postoperative analgesia evolution : Time for sensitive blocks]. / Evolution de l'analgésie après chirurgie du membre inférieur. La place des blocs sensitifs.

For about twenty years, peripheral nervous blocks have become the gold standard in postoperative analgesia for knee and foot surgery. The development of echography adapted to anesthesia gave anesthesiologists the opportunity to perform distal, mostly sensitive nerve blocks. Proximal nervous blocks associated analgesia used to be better than general (intravenous or oral) analgesia. Most of all it was associated with fewer side effects (nauseas, vomiting, drowsiness…). They reached an analgesia level comparable to central nerve blocks (epidural) with also fewer side effects (hypotension, bradycardia, lateralization...). A main problem with those blocks was that they involved motor blockade, impeding fast revalidation and walking restart. Distal nerve blocks, without motor blockade, offer as efficient analgesia as produced by proximal nerve blocks, without impeding motricity. Using those new techniques, it is now possible, with optimal analgesia, to allow almost immediate walking after a total knee replacement, and to perform most surgical foot operations in a one-day procedure.

Depuis une vingtaine d'années, les blocs nerveux périphériques sont devenus les techniques de choix pour l'analgésie de la chirurgie osseuse du genou et du pied. L'avènement de l'échographie adaptée à l'anesthésie permet désormais la réalisation de blocs nerveux distaux essentiellement sensitifs. Les blocs périphériques proximaux offraient une antalgie supérieure par rapport à l'antalgie intraveineuse ou per os traditionnelle, avec moins d'effets secondaires généraux (nausées, vomissements, somnolence…). Ils procuraient une antalgie semblable aux blocs centraux (épidurale en particulier), avec beaucoup moins d'effets secondaires (hypotension, bradycardie, latéralisation). Le défaut principal de ces blocs est qu'ils s'accompagnaient de blocs moteurs, constituant un obstacle à la rééducation rapide et à la reprise de la marche. La réalisation de blocs distaux, dépourvus d'effet moteur permet d'obtenir une antalgie aussi efficace que les celle des blocs proximaux, mais sans entraver la motricité. Grâce au développement de ces techniques, il est désormais possible, avec une antalgie optimale, de permettre la reprise quasi immédiate de la marche après mise en place d'une prothèse de genou, et de pratiquer la majorité de la chirurgie du pied en hôpital de jour.