Biology subjects

Benvenuti, F.

Publications and source records attributed to Benvenuti, F..

6 recordsLinked to original sources

bioRxiv2024

DCs targeted therapy expands CD8 T cell responses to bona-fide neoantigens in lung tumors

Cross-presentation by type 1 cDCs (cDC1) is critical to induce and sustain antitumoral CD8 T cell responses to model antigens, in various tumor settings. However, the impact of cross-presenting cDC1 and the potential of DC-based therapies in tumors carrying varied levels of bona-fide neoantigens (neoAgs) remains unclear. Here we generated a non-small cell lung cancer model with distinct ranges of TMB and MHC-I neoepitopes to test immunogenicity and response to Flt3L+CD40 (DC-therapy). We found that cDC1 are required to broaden the pattern of CD8 responses to basal and acquired neoAgs and DC-therapy strongly inhibits the growth of TMBhigh tumors. In contrast, TMBlow tumors induce weaker responses that are not sufficient to block progression. scRNA transcriptional analysis, immune profiling and functional assays show that DC-therapy triggers the accumulation of lung cDC1 with increased immunostimulatory properties and CD8 T cells with enhanced cytotoxic functions and reduced exhaustion, most prominently in neoAgshigh tumors. We conclude that boosting cDC1 activity is critical to broaden the diversity of anti-tumoral CD8 T cell responses and to leverage neoAgs content for therapeutic advantage.

immunology↗

bioRxiv2023

A mitochondrial quality control mechanism reverses the phagosome maturation arrest caused by Mycobacterium tuberculosis

Phagosome maturation arrest (PMA) imposed by Mycobacterium tuberculosis (Mtb) is a classic tool that helps Mtb evade macrophage anti-bacterial responses. The exclusion of RAB7, a small GTPase, from Mtb-phagosomes causes PMA. Here, we report an unexpected mechanism that triggers crosstalk between the mitochondrial quality control (MQC) and the phagosome maturation pathways that reverses the PMA. CRISPR-mediated p62/SQSTM1 depletion (p62KD) does not appear to impact mitochondrial quality. The p62KD cells are restrictive to Mtb growth, triggered by an increasingly oxidative environment and increased lysosomal targeting. The lysosomal targeting of Mtb is facilitated by enhanced TOM20+ mitochondria-derived vesicles (MDVs) biogenesis, a key MQC mechanism. In p62KD cells, TOM20+-MDVs biogenesis is MIRO1/MIRO2-dependent and gets delivered to lysosomes for degradation in a RAB7-dependent manner. Upon infection in p62KD cells, TOM20+-MDVs get extensively targeted to Mtb-phagosomes, inadvertently facilitating RAB7 recruitment, PMA reversal and lysosomal targeting of Mtb; the phenotype also replicated in p62/SQSTM1 knockout cells. Triggering MQC collapse in p62KDcells further diminishes Mtb survival, signifying cooperation between redox- and lysosome-mediated mechanisms. The MQC-anti-bacterial pathway crosstalk could be exploited for host-directed anti-tuberculosis therapies.

microbiology↗

bioRxiv2023

ALCAM-mediated synapses between DC1 and CD8 T cells are inhibited in advanced lung tumors

Conventional type 1 dendritic cells (DC1) control anti-tumoral CD8 T responses, in lymph nodes and tumor tissues. T-cell activation depends on the establishment of a tight physical interaction with antigen-presenting cell, the immunological synapse (IS). The molecular determinants of DC1-CD8 IS in tumor tissues and how they are regulated during cancer progression remain poorly investigated. Using a reporter for DC1 in a genetic model of non-small cell lung cancer (KP-XCR1venus) we show that IS in lung tissues are abundant and productive at early stages of tumor development but progressively diminish in advanced tumors. Transcriptional profiling and flow cytometry of lung resident DC1 identified a module of adhesion molecules downregulated in advanced tumors. We focused on ALCAM and LFA-1, ligands for CD6 and ICAM-1 on T cells, to investigate their role and functional impact. By immobilizing single receptor agonists on artificial cell surfaces, we demonstrate that ALCAM and LFA-1 are sufficient to trigger cytoskeletal remodeling in early tumor DC1, whereas late tumors DC1 are not responsive. Blocking ALCAM-CD6 interactions in functional assays impairs the acquisition of effector functions in CD8 T cells. Together these data highlight that adhesion molecules required to establish IS in early, immune-reactive, tumors are targeted during tumor progression blunting cross-talk within the IS.

immunology↗

bioRxiv2023

Tim4 enables large peritoneal macrophages to cross-present tumor antigens at early stages of tumorigenesis

Cross-presentation of tumor antigens by subsets of macrophages at different stages along tumor progression may have divergent impacts on anti-tumoral immune responses. Here we show that TIM4+ large peritoneal macrophages (LPM) avidly capture tumor cells and cross-present tumor-associated antigens at early stages of peritoneal infiltration by ovarian cancer cells. The phosphatidylserine (PS) receptor TIM4 promotes maximal uptake and triggers inflammatory and metabolic gene programs in combination with cytoskeletal remodeling and upregulation of transcriptional signatures related to antigen processing. At the cellular levels, TIM4 is recruited with F-actin at the phagocytic cup and translocates in nascent phagosomes, controlling the kinetic of phagosomal acidification and cargo degradation. TIM4 deletion abrogates cross-presentation of tumor-associated antigens and blunts expansion of effector CD8 T cells at tumor inception. In addition, targeting tumor antigens to LPM by PS liposomes can trigger CD8 T cell activation. Together these results suggest that TIM4 enables LPMs to scan the antigenic content of incoming tumor cells to promote immune surveillance by CD8 T cells and, at defined temporal windows, may be exploited for therapeutic purposes.

immunology↗

bioRxiv2023

Cebranopadol, a novel long-acting opioid agonist with low abuse liability, to treat opioid use disorder: Preclinical evidence of efficacy

The gold standard pharmacological treatment for opioid use disorder (OUD) consists of maintenance therapy with long-acting opioid agonists such as buprenorphine and methadone. Despite these compounds having demonstrated substantial efficacy, a significant number of patients do not show optimal therapeutic responses. Moreover, the abuse liability of these medications remains a major concern. Cebranopadol, is a new, long-acting pan-opioid agonist that also activates the nociception/orphanin FQ NOP receptor. Here we used rats to explore the therapeutic potential of this agent in OUD. First, in operant intravenous self-administration experiments we compared the potential abuse liability of cebranopadol with the prototypical opioid heroin. Under a fixed ratio 1 (FR1) contingency, rats maintained responding for heroin (1, 7, 20, 60 g/inf) to a larger extent than cebranopadol (0.03, 0.1, 0.3, 1.0, 6.0 g/inf). When the contingency was switched to a progressive ratio (PR) reinforcement schedule, heroin maintained responding at high levels at all except the lowest dose. Conversely, in the cebranopadol groups responding decreased drastically and the break point (BP) did not differ from saline controls. Next, we demonstrated that oral administration of cebranopadol (0, 25, 50 g/kg) significantly attenuated drug self-administration independent of heroin dose (1, 7, 20, 60 g/inf). Cebranopadol also reduced the break point for heroin (20 g/inf). Furthermore, in a heroin self-administration training extinction/reinstatement paradigm, pretreatment with cebranopadol significantly attenuated yohimbine stress-induced reinstatement of drug seeking. Together, these data indicate that cebranopadol has limited abuse liability compared to heroin and is highly efficacious in attenuating opioid self-administration and stress-induced reinstatement, suggesting clinical potential of this compound for OUD treatment.

pharmacology and toxicology↗

bioRxiv2021

SiglecFhigh neutrophils in lung tumor tissues suppress local CD8 T cell responses and limit the efficacy of anti PD-L1 antibodies

BackgroundTumor infiltrating neutrophils generally correlates to worst prognosis and refractoriness to immunotherapy yet the complexity and significance of diverse subsets resident in tumor tissues has just begun to emerge. In lung tumors, a network of neutrophils states with graded protumorigenic properties is conserved between mouse and humans and include a subset of mature, long lived cells expressing the sialic-acid-binding protein SiglecF (SiglecFhigh neu). The mechanism of recruitment of SiglecFhigh neu into tumor tissues and the impact on local anti-tumor T cell responses and interference with immunotherapy is still elusive. MethodsWe used an immunogenic model of KrasG12D Tp53 null adenocarcinoma of the lung to screen for factors inducing the recruitment of SiglecFhigh neu, followed by gene editing to delete selected candidates. We analyzed frequencies and effector functions of endogenous CD8 T cell responses in controls and SiglecFhigh neu depleted tumors by flow cytometry and functional assays. Tissues fluorescence and confocal imaging of lung sections was used to explore the relative distribution of neu and CD8 T cells. To establish the impact of SiglecFhigh neu on anti-tumoral immune responses we treated cohort of animals with anti-PD-L1 antibodies to evaluate tumor growth in control conditions and under therapy. ResultsWe found that tumor tissues express high levels of CXCL5, mapping to cancer cells. Upon deletion of chemokine expression by gene editing, the recruitment of SiglecFhigh neu was almost entirely abrogated. In tumors depleted of SiglecFhigh neu, the density of tumor specific endogenous CD8 T cells was 3-fold higher than in controls and showed significantly enhanced activation and effector functions. Importantly, checkpoint blockade with anti PD-L1 antibodies was ineffective in control tumors but showed a significant benefit in SiglecFhigh neu depleted tumors. ConclusionThis study demonstrates that SiglecFhigh neu differentiating in lung tumor tissues inhibit local CD8 T cell responses and interfere with the success of checkpoint blockade. These data suggest that blocking selectively tissue resident neu may promote better responses to immunotherapy.

immunology↗