Low-dose ionizing γ-radiation elicits the extrusion of neutrophil extracellular traps.

PURPOSE: Cancer patients frequently undergo radiotherapy in their clinical management with unintended irradiation of blood vessels and copiously irrigated organs in which polymorphonuclear leukocytes circulate. Following the observation that such low doses of ionizing radiation are able to induce neutrophils to extrude neutrophil extracellular traps (NETs), we have investigated the mechanisms, consequences and the occurrence of such phenomena in patients undergoing radiotherapy. EXPERIMENTAL DESIGN: NETosis was analyzed in cultures of neutrophils isolated from healthy donors, cancer patients and cancer-bearing mice under confocal microscopy. Cocultures of radiation-induced NETs, immune effector lymphocytes and tumor cells were used to study the effects of irradiation-induced NETs on immune cytotoxicity. Radiation-induced NETs were intravenously injected to mice assessing their effects on metastasis. Circulating NETs in irradiated cancer patients were measured by ELISA methods detecting MPO-DNA complexes and citrullinated H3. RESULTS: Very low γ-radiation doses (0.5-1 Gy) given to neutrophils elicit NET formation in a manner dependent on oxidative stress, NADPH oxidase activity and autocrine interleukin-8. Radiation-induced NETs interfere with NK- and T-cell cytotoxicity. As a consequence, pre-injection of irradiation-induced NETs increases the number of successful metastases in mouse tumor models. Increases in circulating NETs were readily detected in two prospective series of patients following the first fraction of their radiotherapy courses. CONCLUSIONS: NETosis is induced by low-dose ionizing irradiation in a neutrophil-intrinsic fashion and radiation-induced NETs are able to interfere with immune-mediated cytotoxicity. Radiation-induced NETs foster metastasis in mouse models and can be detected in the circulation of patients undergoing conventional radiotherapy treatments.

Safety, Pharmacokinetics, Pharmacodynamics, and Antitumor Activity from a Phase I Study of Simlukafusp Alfa (FAP-IL2v) in Advanced/Metastatic Solid Tumors.

PURPOSE: Simlukafusp alfa (FAP-IL2v), a tumor-targeted immunocytokine, comprising an interleukin-2 variant moiety with abolished CD25 binding fused to human immunoglobulin G1, is directed against fibroblast activation protein-α. This phase I, open-label, multicenter, dose-escalation and extension study (NCT02627274) evaluated the safety, pharmacokinetics, pharmacodynamics, and antitumor activity of FAP-IL2v in patients with advanced/metastatic solid tumors. METHODS: Participants received FAP-IL2v intravenously once weekly. Dose escalation started at 5 mg; flat dosing (≤25 mg) and intra-participant up-titration regimens (15/20 mg, 20/25 mg, 20/20/35 mg, 20/35/35 mg) were evaluated. Primary objectives were dose-limiting toxicities (DLT), maximum tolerated dose (MTD), recommended expansion dose, and pharmacokinetics. RESULTS: Sixty-one participants were enrolled. DLTs included fatigue (flat dose 20 mg: n = 1), asthenia (25 mg: n = 1), drug-induced liver injury (up-titration regimen 20/25 mg: n = 1), transaminase increase (20/25 mg: n = 1), and pneumonia (20/35/35 mg: n = 1). Up-titration regimen 15/20 mg was the MTD and was selected as the recommended expansion dose. Increases in peripheral blood absolute immune cell counts were seen for all tested doses (natural killer cells, 13-fold; CD4+ T cells [including Tregs], 2-fold; CD8+ T cells, 3.5-fold), but without any percentage change in Tregs. Clinical activity was observed from 5 mg (objective response rate, 5.1% [n = 3]; disease control rate, 27.1% [n = 16]). Responses were durable (n = 3; 2.8 [censored], 6.3, and 43.4 months). CONCLUSIONS: FAP-IL2v had a manageable safety profile and showed initial signs of antitumor activity in advanced/metastatic solid tumors.

Radiotherapy protocols for mouse cancer model.

Radiotherapy is a crucial treatment modality for cancer patients, with approximately 60% of individuals undergoing ionizing radiation as part of their disease management. In recent years, there has been a growing trend toward minimizing irradiation fields through the use of image-guided dosimetry and innovative technologies. These advancements allow for selective irradiation, delivering higher local doses while reducing the number of treatment sessions. Consequently, computer-assisted methods have significantly enhanced the effectiveness of radiotherapy in the curative and palliative treatment of various cancers. Although radiation therapy alone can effectively achieve local control in some cancer types, it may not be sufficient for others. As a result, further preclinical research is necessary to explore novel approaches including new schedules of radiotherapy treatments. Unfortunately, there is a concerning lack of correlation between clinical outcomes and experiments conducted on mouse models. We hypothesize that this disparity arises from the differences in irradiation strategies employed in preclinical studies compared to those used in clinical practice, which ultimately affects the translatability of findings to patients. In this study, we present two comprehensive radiotherapy protocols for the treatment of orthotopic melanoma and glioblastoma tumors. These protocols utilize a small animal radiation research platform, which is an ideal radiation device for delivering localized and precise X-ray doses to the tumor mass. By employing these platforms, we aim to limit the side effects associated with irradiating healthy surrounding tissues. Our detailed protocols offer a valuable framework for conducting preclinical studies that closely mimic clinical radiotherapy techniques, bridging the gap between experimental results and patient outcomes.

An international phase II trial and immune profiling of SBRT and atezolizumab in advanced pretreated colorectal cancer.

BACKGROUND: Immuno-radiotherapy may improve outcomes for patients with advanced solid tumors, although optimized combination modalities remain unclear. Here, we report the colorectal (CRC) cohort analysis from the SABR-PDL1 trial that evaluated the PD-L1 inhibitor atezolizumab in combination with stereotactic body radiation therapy (SBRT) in advanced cancer patients. METHODS: Eligible patients received atezolizumab 1200 mg every 3 weeks until progression or unmanageable toxicity, together with ablative SBRT delivered concurrently with the 2nd cycle (recommended dose of 45 Gy in 3 fractions, adapted upon normal tissue tolerance constraint). SBRT was delivered to at least one tumor site, with at least one additional measurable lesion being kept from the radiation field. The primary efficacy endpoint was one-year progression-free survival (PFS) rate from the start of atezolizumab. Sequential tumor biopsies were collected for deep multi-feature immune profiling. RESULTS: Sixty pretreated (median of 2 prior lines) advanced CRC patients (38 men [63%]; median age, 59 years [range, 20-81 years]; 77% with liver metastases) were enrolled in five centers (France: n = 4, Spain: n = 1) from 11/2016 to 04/2019. All but one (98%) received atezolizumab and 54/60 (90%) received SBRT. The most frequently irradiated site was lung (n = 30/54; 56.3%). Treatment-related G3 (no G4-5) toxicity was observed in 3 (5%) patients. Median OS and PFS were respectively 8.4 [95%CI:5.9-11.6] and 1.4 months [95%CI:1.2-2.6], including five (9%) patients with PFS > 1 year (median time to progression: 19.2 months, including 2/5 MMR-proficient). Best overall responses consisted of stable disease (n = 38; 64%), partial (n = 3; 5%) and complete response (n = 1; 2%). Immune-centric multiplex IHC and RNAseq showed that SBRT redirected immune cells towards tumor lesions, even in the case of radio-induced lymphopenia. Baseline tumor PD-L1 and IRF1 nuclear expression (both in CD3 + T cells and in CD68 + cells) were higher in responding patients. Upregulation of genes that encode for proteins known to increase T and B cell trafficking to tumors (CCL19, CXCL9), migration (MACF1) and tumor cell killing (GZMB) correlated with responses. CONCLUSIONS: This study provides new data on the feasibility, efficacy, and immune context of tumors that may help identifying advanced CRC patients most likely to respond to immuno-radiotherapy. TRIAL REGISTRATION: EudraCT N°: 2015-005464-42; Clinicaltrial.gov number: NCT02992912.

Cytotoxicity as a form of immunogenic cell death leading to efficient tumor antigen cross-priming.

Antigen cross-priming of CD8+ T cells is a critical process necessary for the effective expansion and activation of CD8+ T cells endowed with the ability to recognize and destroy tumor cells. The cross-presentation of tumor antigens to cross-prime CD8+ T cells is mainly mediated, if not only, by a subset of professional antigen-presenting cells termed type-1 conventional dendritic cells (cDC1). The demise of malignant cells can be immunogenic if it occurs in the context of premortem stress. These ways of dying are termed immunogenic cell death (ICD) and are associated with biochemical features favoring cDC1 for the efficient cross-priming of tumor antigens. Immunosurveillance and the success of immunotherapies heavily rely on the ability of cytotoxic immune cells, primarily CD8+ T cells and NK cells, to detect and eliminate tumor cells through mechanisms collectively known as cytotoxicity. Recent studies have revealed the significance of NK- and CTL-mediated cytotoxicity as a prominent form of immunogenic cell death, resulting in mechanisms that promote and sustain antigen-specific immune responses. This review focuses on the mechanisms underlying the cross-presentation of antigens released during tumor cell killing by cytotoxic immune cells, with an emphasis on the role of cDC1 cells. Indeed, cDC1s are instrumental in the effectiveness of most immunotherapies, underscoring the significance of tumor antigen cross-priming in contexts of immunogenic cell death. The notion of the potent immunogenicity of cell death resulting from NK or cytotoxic T lymphocyte (CTL)-mediated cytotoxicity has far-reaching implications for cancer immunotherapy.

Parent knowledge regarding food selection for children with PKU: Results of a survey in the United States.

OBJECTIVES: Dietary treatment is the main therapy for most patients with phenylketonuria (PKU). Parental knowledge regarding food selection is crucial to ensure adequate metabolic control and brain development during childhood and to promote lifelong adherence and healthy dietary behavior in the offspring. The aims of this study were to assess whether parental or caregiver knowledge regarding nutritional selection for children with PKU is in accordance with medical recommendations and to evaluate factors that influence their level of knowledge. METHODS: This was a cross-sectional observational study. An online or paper survey (N = 178) was distributed throughout the United States. The survey included a validated food selection questionnaire to assess whether the respondent adequately identified foods that require certain restrictions versus foods that can be consumed freely by an individual with PKU. RESULTS: General knowledge of food selection among the caregivers was very high or high in nearly 60% (60-98th score percentile). Participants with the lowest scores in general knowledge of the PKU diet (quartile 1) were more likely to label allowed foods incorrectly. Respondents with the highest scores (quartile 4) were more likely to label limited foods correctly but incorrectly label allowed items. CONCLUSION: Knowledge of restricted foods is important to avoid poor metabolic control, but knowledge of allowed foods can be just as important to allow for a diet that is diverse, palatable, and nutritionally balanced.

Methods to assess radiation induced abscopal responses in mice.

Radiotherapy (RT) can work together with the immune system to eliminate cancer. It can cause immunogenic cell death and facilitate tumor neoantigen presentation and thereby the cross-priming of tumor-specific T-lymphocytes, turning irradiated tumors into in-situ vaccines. Accumulating preclinical and clinical evidence indicates that RT in conjunction with ICB leads to systemic anti-tumor immune responses, thus stimulating interest in using ICB to overcome primary and acquired cancer resistance to radiotherapy. However, the systemic effects (abscopal effects) obtained to date are far from being acceptable for clinical translation. In this context, multiple preclinical mouse models have demonstrated that a variety of immunotherapy agents can be delivered locally to enhance antitumor immunity both in a local and systemic fashion. Using two slightly asynchronous and anatomically distant subcutaneous B16OVA tumors in syngeneic immunocompetent hosts (C57BL/6), we describe the feasibility of a local immunotherapy treatment given in combination with external beam irradiation, which exerts immune-mediated antitumor effects in mice and humans upon intratumoral delivery. With minor variations, the same technique can be easily applied to a variety of mouse transplantable tumors.

Intratumoral neoadjuvant immunotherapy based on the BO-112 viral RNA mimetic.

BO-112 is a poly I:C-based viral mimetic that exerts anti-tumor efficacy when intratumorally delivered in mouse models. Intratumoral BO-112 synergizes in mice with systemic anti-PD-1 mAbs and this combination has attained efficacy in PD1-refractory melanoma patients. We sought to evaluate the anti-tumor efficacy of BO-112 pre-surgically applied in neoadjuvant settings to mouse models. We have observed that repeated intratumoral injections of BO-112 prior to surgical excision of the primary tumor significantly reduced tumor metastasis from orthotopically implanted 4T1-derived tumors and subcutaneous MC38-derived tumors in mice. Such effects were enhanced when combined with systemic anti-PD-1 mAb. The anti-tumor efficacy of this neoadjuvant immunotherapy approach depended on the presence of antigen-specific effector CD8 T cells and cDC1 antigen-presenting cells. Since BO-112 has been successful in phase-two clinical trials for metastatic melanoma, these results provide a strong rationale for translating this pre-surgical strategy into clinical settings, especially in combination with standard-of-care checkpoint inhibitors.

Intratumoral BO-112 in combination with radiotherapy synergizes to achieve CD8 T-cell-mediated local tumor control.

BACKGROUND: Radioimmunotherapy combines irradiation of tumor lesions with immunotherapy to achieve local and abscopal control of cancer. Most immunotherapy agents are given systemically, but strategies for delivering immunotherapy locally are under clinical scrutiny to maximize efficacy and avoid toxicity. Local immunotherapy, by injecting various pathogen-associated molecular patterns, has shown efficacy both preclinically and clinically. BO-112 is a viral mimetic based on nanoplexed double-stranded RNA (poly I:C) which exerts immune-mediated antitumor effects in mice and humans on intratumoral delivery. BO-112 and focal irradiation were used to make the proof-of-concept for local immunotherapy plus radiation therapy combinations. METHODS: Murine transplantable tumor cell lines (TS/A, MC38 and B16-OVA) were used to show increased immunogenic features under irradiation, as well as in bilateral tumor models in which only one of the lesions was irradiated or/and injected with BO-112. Flow cytometry and multiplex tissue immunofluorescence were used to determine the effects on antitumor immunity. Depletions of immune cell populations and knockout mice for the IFNAR and BATF-3 genes were used to delineate the immune system requirements for efficacy. RESULTS: In cultures of TS/A breast cancer cells, the combination of irradiation and BO-112 showed more prominent features of immunogenic tumor cell death in terms of calreticulin exposure. Injection of BO-112 into the tumor lesion receiving radiation achieved excellent control of the treated tumor and modest delays in contralateral tumor progression. Local effects were associated with more prominent infiltrates of antitumor cytotoxic tumor lymphocytes (CTLs). Importantly, local irradiation plus BO-112 in one of the tumor lesions that enhanced the therapeutic effects of radiotherapy on distant irradiated lesions that were not injected with BO-112. Hence, this beneficial effect of local irradiation plus BO-112 on a tumor lesion enhanced the therapeutic response to radiotherapy on distant non-injected lesions. CONCLUSION: This study demonstrates that local BO-112 immunotherapy and focal irradiation may act in synergy to achieve local tumor control. Irradiation plus BO-112 in one of the tumor lesions enhanced the therapeutic effects on distant irradiated lesions that were not injected with BO-112, suggesting strategies to treat oligometastatic patients with lesions susceptible to radiotherapy and with at least one tumor accessible for repeated BO-112 intratumoral injections.

Flow cytometry-assisted quantification of immune cells infiltrating irradiated tumors in mice.

The immunomodulatory properties of local hypofractionated radiotherapy are known to promote the generation of anti-tumor immune responses. Such responses are largely due to the infiltration of cytotoxic lymphocytes (TILs) into the tumors that are able to destroy malignant lesions. In this context, characterizing the tumor immune microenvironment following radiotherapy is crucial for the study of its mechanism of action. Flow cytometry-based analyses are frequently used to elucidate changes in the tumor immune microenvironment. The use of a fluorochrome-conjugated antibody panel is currently a standard technique to assess the number and phenotype of immune cell populations infiltrating the tumors. Here, we describe a method to isolate and quantify TILs based on flow-cytometry in mammary carcinoma-bearing mice that undergo a local hypofractionated radiotherapy regimen consisting of 3 consecutive doses of 8 Gy. With some adaptations, this protocol can be successfully applied to a diverse range of transplantable and inducible solid mouse tumors of different origins.

A simple method to assess clonogenic survival of irradiated cancer cells.

The clonogenic assay is an in vitro method based on the ability of a single cell to proliferate indefinitely into a colony. This assay is the gold standard method to analyze cell viability and quantify reproductive cell survival fraction after treatment with ionizing radiation and other cytotoxic agents in vitro. After the cytotoxic effect, only some cells retain their ability to grow from one cell and form colonies. The colony is defined to consist of at least 50 cells. The radiosensitivity of each cell line may vary. Thus, characterizing cell sensitivity following radiation is crucial to choose the optimum radiotherapy dose. Here, we describe a method to test the in vitro capability of cell lines to form colonies following radiation treatment. This assay allows to analyze the efficacy of specific treatments on the cell reproductivity of cell lines With some adaptations, this protocol can be essentially applied to analyze the cell proliferation rate after different doses of irradiation on many different cell lines.

Assessment of hypoxia by pimonidazole staining following radiotherapy.

The rapid proliferation of cancer cells and the aberrant vasculature present in most solid tumors frequently result in the lack of oxygen generating a hypoxic tumor microenvironment. Low levels of oxygen not only affect the tumor cell biology and tumorigenesis, but also the other components of the tumor microenvironment such as the tumor stroma and the immune infiltrate, promoting a more suppressive environment. In addition, tumor hypoxia has been associated with reduced sensitivity to chemotherapy (CH) and radiotherapy (RT), leading to poor outcomes in cancer patients. Therefore, the evaluation of tumor oxygen status has become clinically relevant. Tumor hypoxia can be assessed by different methods that include the analysis of the oxygen concentration or the expression of endogenous markers directly related to hypoxia. In this paper, we focus on the use of the hypoxia-specific marker pimonidazole as a straightforward way to measure tumor hypoxia following radiotherapy in a preclinical melanoma model.

Charting roadmaps towards novel and safe synergistic immunotherapy combinations.

Checkpoint inhibitor-based cancer immunotherapy is often combined in the clinic with other immunotherapy strategies, targeted therapies, chemotherapy or standard-of-care treatments to achieve superior therapeutic efficacy. The large number of immunotherapy combinations that are currently undergoing clinical testing necessitate the establishment of faithful criteria to prioritize optimal combinations with evidence of synergy, to determine their safety and optimal sequence of administration and to identify biomarkers of therapy resistance and response. In this review, we focus on recent developments in immunotherapy combinations and reflect on how combinations should be optimized to maximize the impact of immunotherapy in clinical oncology.

Soluble CD137 as a dynamic biomarker to monitor agonist CD137 immunotherapies.

BACKGROUND: On the basis of efficacy in mouse tumor models, multiple CD137 (4-1BB) agonist agents are being preclinically and clinically developed. The costimulatory molecule CD137 is inducibly expressed as a transmembrane or as a soluble protein (sCD137). Moreover, the CD137 cytoplasmic signaling domain is a key part in approved chimeric antigen receptors (CARs). Reliable pharmacodynamic biomarkers for CD137 ligation and costimulation of T cells will facilitate clinical development of CD137 agonists in the clinic. METHODS: We used human and mouse CD8 T cells undergoing activation to measure CD137 transcription and protein expression levels determining both the membrane-bound and soluble forms. In tumor-bearing mice plasma sCD137 concentrations were monitored on treatment with agonist anti-CD137 monoclonal antibodies (mAbs). Human CD137 knock-in mice were treated with clinical-grade agonist anti-human CD137 mAb (Urelumab). Sequential plasma samples were collected from the first patients intratumorally treated with Urelumab in the INTRUST clinical trial. Anti-mesothelin CD137-encompassing CAR-transduced T cells were stimulated with mesothelin coated microbeads. sCD137 was measured by sandwich ELISA and Luminex. Flow cytometry was used to monitor CD137 surface expression. RESULTS: CD137 costimulation upregulates transcription and protein expression of CD137 itself including sCD137 in human and mouse CD8 T cells. Immunotherapy with anti-CD137 agonist mAb resulted in increased plasma sCD137 in mice bearing syngeneic tumors. sCD137 induction is also observed in human CD137 knock-in mice treated with Urelumab and in mice transiently humanized with T cells undergoing CD137 costimulation inside subcutaneously implanted Matrigel plugs. The CD137 signaling domain-containing CAR T cells readily released sCD137 and acquired CD137 surface expression on antigen recognition. Patients treated intratumorally with low dose Urelumab showed increased plasma concentrations of sCD137. CONCLUSION: sCD137 in plasma and CD137 surface expression can be used as quantitative parameters dynamically reflecting therapeutic costimulatory activity elicited by agonist CD137-targeted agents.

Tumor ENPP1 (CD203a)/Haptoglobin Axis Exploits Myeloid-Derived Suppressor Cells to Promote Post-Radiotherapy Local Recurrence in Breast Cancer.

ABSTRACT: Locoregional failure (LRF) in patients with breast cancer post-surgery and post-irradiation is linked to a dismal prognosis. In a refined new model, we identified ectonucleotide pyrophosphatase/phosphodiesterase 1/CD203a (ENPP1) to be closely associated with LRF. ENPP1hi circulating tumor cells (CTC) contribute to relapse by a self-seeding mechanism. This process requires the infiltration of polymorphonuclear myeloid-derived suppressor cells and neutrophil extracellular trap (NET) formation. Genetic and pharmacologic ENPP1 inhibition or NET blockade extends relapse-free survival. Furthermore, in combination with fractionated irradiation, ENPP1 abrogation obliterates LRF. Mechanistically, ENPP1-generated adenosinergic metabolites enhance haptoglobin (HP) expression. This inflammatory mediator elicits myeloid invasiveness and promotes NET formation. Accordingly, a significant increase in ENPP1 and NET formation is detected in relapsed human breast cancer tumors. Moreover, high ENPP1 or HP levels are associated with poor prognosis. These findings unveil the ENPP1/HP axis as an unanticipated mechanism exploited by tumor cells linking inflammation to immune remodeling favoring local relapse. SIGNIFICANCE: CTC exploit the ENPP1/HP axis to promote local recurrence post-surgery and post-irradiation by subduing myeloid suppressor cells in breast tumors. Blocking this axis impairs tumor engraftment, impedes immunosuppression, and obliterates NET formation, unveiling new opportunities for therapeutic intervention to eradicate local relapse and ameliorate patient survival. This article is highlighted in the In This Issue feature, p. 1171.

Three-dimensional colon cancer organoids model the response to CEA-CD3 T-cell engagers.

Rationale: The CEA-CD3 T cell bispecific antibody cibisatamab (CEA-TCB) is currently undergoing clinical trials. Here we study its performance against three-dimensional tumor organoids in cocultures with T cells as compared to a higher affinity CEACAM5-CD3 (CEACAM5-TCB) bispecific antibody using time-lapse confocal microscopy. Methods: Pre-labelled spheroids derived from colon cancer cell lines and primary organoids derived from four colorectal cancer surgical specimens, which expressed different graded levels of CEA, were exposed in cocultures to T lymphocytes. Cocultures were treated with CEA-CD3 T-cell engagers and were followed by live confocal microscopy. Caspase 3 activation detected in real-time was used as an indicator of tumor cell death. Co-cultures were also set up with autologous tumor-associated fibroblasts to test the co-stimulatory effect of a fibroblast activated protein (FAP)- targeted 4-1BBL bispecific antibody fusion protein currently undergoing clinical trials. Results: Tumor-cell killing of 3D colon carcinoma cultures was dependent on the levels of surface CEA expression, in such a way that the lower affinity agent (CEA-TCB) did not mediate killing by human preactivated T cells below a certain CEA expression threshold, while the high affinity construct (CEACAM5-TCB) remained active on the low CEA expressing organoids. Modelling heterogeneity in the levels of CEA expression by coculturing CEA high and low organoids showed measurable but weak bystander killing. Cocultures of tumor organoids, autologous fibroblasts and T cells allowed to observe a costimulatory effect of anti-FAP-4-1BBL both to release IFNγ and to attain more efficacious tumor cell killing. Conclusion: Three-dimensional tumor cocultures with T cells using live confocal microscopy provide suitable models to test the requirements for colon-cancer redirected killing as elicited by CEA-targeted T-cell engagers undergoing clinical trials and treatment allow combinations to be tested in a relevant preclinical system.

Preclinical Characterization and Phase I Trial Results of a Bispecific Antibody Targeting PD-L1 and 4-1BB (GEN1046) in Patients with Advanced Refractory Solid Tumors.

Checkpoint inhibitors (CPI) have revolutionized the treatment paradigm for advanced solid tumors; however, there remains an opportunity to improve response rates and outcomes. In preclinical models, 4-1BB costimulation synergizes with CPIs targeting the programmed cell death protein 1 (PD-1)/programmed cell death ligand 1 (PD-L1) axis by activating cytotoxic T-cell-mediated antitumor immunity. DuoBody-PD-L1×4-1BB (GEN1046) is an investigational, first-in-class bispecific immunotherapy agent designed to act on both pathways by combining simultaneous and complementary PD-L1 blockade and conditional 4-1BB stimulation in one molecule. GEN1046 induced T-cell proliferation, cytokine production, and antigen-specific T-cell-mediated cytotoxicity superior to clinically approved PD-(L)1 antibodies in human T-cell cultures and exerted potent antitumor activity in transplantable mouse tumor models. In dose escalation of the ongoing first-in-human study in heavily pretreated patients with advanced refractory solid tumors (NCT03917381), GEN1046 demonstrated pharmacodynamic immune effects in peripheral blood consistent with its mechanism of action, manageable safety, and early clinical activity [disease control rate: 65.6% (40/61)], including patients resistant to prior PD-(L)1 immunotherapy. SIGNIFICANCE: DuoBody-PD-L1×4-1BB (GEN1046) is a first-in-class bispecific immunotherapy with a manageable safety profile and encouraging preclinical and early clinical activity. With its ability to confer clinical benefit in tumors typically less sensitive to CPIs, GEN1046 may fill a clinical gap in CPI-relapsed or refractory disease or as a combination therapy with CPIs. See related commentary by Li et al., p. 1184. This article is highlighted in the In This Issue feature, p. 1171.

The Lymphatic Endothelium in the Context of Radioimmuno-Oncology.

The study of lymphatic tumor vasculature has been gaining interest in the context of cancer immunotherapy. These vessels constitute conduits for immune cells' transit toward the lymph nodes, and they endow tumors with routes to metastasize to the lymph nodes and, from them, toward distant sites. In addition, this vasculature participates in the modulation of the immune response directly through the interaction with tumor-infiltrating leukocytes and indirectly through the secretion of cytokines and chemokines that attract leukocytes and tumor cells. Radiotherapy constitutes the therapeutic option for more than 50% of solid tumors. Besides impacting transformed cells, RT affects stromal cells such as endothelial and immune cells. Mature lymphatic endothelial cells are resistant to RT, but we do not know to what extent RT may affect tumor-aberrant lymphatics. RT compromises lymphatic integrity and functionality, and it is a risk factor to the onset of lymphedema, a condition characterized by deficient lymphatic drainage and compromised tissue homeostasis. This review aims to provide evidence of RT's effects on tumor vessels, particularly on lymphatic endothelial cell physiology and immune properties. We will also explore the therapeutic options available so far to modulate signaling through lymphatic endothelial cell receptors and their repercussions on tumor immune cells in the context of cancer. There is a need for careful consideration of the RT dosage to come to terms with the participation of the lymphatic vasculature in anti-tumor response. Here, we provide new approaches to enhance the contribution of the lymphatic endothelium to radioimmuno-oncology.

Evaluación de adherencia a la guía de práctica clínica para el manejo de faringoamigdalitis aguda en la atención de urgencias del Hospital Universitario San Ignacio, Bogotá, Colombia / Evaluation of adherence to the clinical practice guideline for the management of acute pharyngotonsillitis in the emergency care of the University Hospital San Ignacio, Bogota, Colombia

Introducción: la faringoamigdalitis aguda, independientemente de su origen, es de los diagnósticos más frecuentes en los servicios de urgencias. Aunque se ha establecido que en la mayoría de casos su origen es viral, ante la dificultad de identificar su agente etiológico con hallazgos clínicos se ha aumentado la formulación indiscriminada de antibióticos, principalmente en los servicios de urgencias, lo que contribuye con la emergencia de resistencias bacterianas y la aparición de efectos secundarios. Se propone evaluar la adherencia a la guía de práctica clínica para el manejo de faringoamigdalitis aguda en urgencias en el Hospital Universitario San Ignacio en Bogotá, Colombia. Materiales y métodos: se realizó un estudio de tipo descriptivo retrospectivo. Se revisaron 7762 historias clínicas de pacientes que consultaron por urgencias entre 2016 y 2019 por dolor de garganta. Se analizaron los datos de formulación de analgésicos, antibióticos, solicitud de la prueba rápida de detección de Estreptococo betahemolítico del grupo A (SBHGA) y el registro de la presencia de exudados al examen físico. Resultados: se incluyeron 7762 pacientes. Del total, 74,2 % recibieron antibiótico y 98 % analgesia. Se solicitó la prueba rápida de detección de SBHGA al 11,53 % de los pacientes. La presencia de exudados es el principal factor asociado a la formulación de antibióticos, y la solicitud de una prueba rápida de detección disminuye significativamente su formulación, dado que 21 % de dichos estudios fueron positivos

Objective: Sore throat is one of the most frequent complaints in the ER, both in children and adults. Although it has been established that most cases of acute tonsillitis are caused by viruses, given the difficulty in identifying its etiology based exclusively upon clinical signs, the indiscriminate prescription of antibiotics in the emergency setting has become very frequent. This practice may lead to the emergence of antibiotic resistance and secondary effects. We evaluated the adherence of ER physicians to clinical practice guidelines for the management of acute tonsillitis at Hospital Universitario San Ignacio in Bogotá, Colombia. Methods: A retrospective descriptive study was carried out. Clinical records for emergency visits between the years of 2016 and 2019 were reviewed. Data regarding rapid antigen detection test for GABHS (RAD), antibiotic and analgesic prescription and tonsillar exudates on physical exam, were recorded. Results: 7.762 patients with acute tonsillitis were included in the study. 74,2% were prescribed antibiotics and 98% received pain medication. For 11,53% of cases the rapid antigen test for GABHS was requested. Although tonsillar exudates are the main factor associated with the formulation of antibiotics, only 21% of rapid antigen detection tests were positive. As such, the systemic use of rapid diagnostic tests may help reduce unnecessary antibiotic prescription, bacterial resistance and drug side effects