Silvia Lopez-Lastra

Les souris humanisées comme modèles d'étude de l'immunité innée humaine et des immunothérapies: Les modèles animaux ont largement contribué à notre compréhension de l’immunologie humaine et des mécanismes pathologiques associés au développement des maladies. Cependant, les modèles murins ne permettent pas de reproduire toute la complexité des pathologies humaines. Les souris à système immunitaire humain (HIS), par leur capacité à récapituler l’hématopoïèse humaine et à être infectées par des pathogènes humains, constituent une solution de choix pour combler ce fossé inter-espèce. Après greffe de cellules souches hématopoïétiques humaines, des souris hôtes sévèrement immunodéprimées permettent un haut niveau de développement du système hémato-lymphoïde humain tout au long de leur vie. Cependant, certains types cellulaires, comme les cellules lymphoïdes innées, ne parviennent pas à se différencier et à fonctionner normalement dans les modèles murins HIS actuels. Ici, nous décrivons le développement d’un modèle souris HIS original, nommé BRGSF, montrant une amélioration de la maturation, de la fonction et de l’homéostasie des cellules natural killer (NK) humaines et des autres ILCs. De plus, en récapitulant les différentes étapes du développement des ILCs humaines, ce modèle souris BRGSF nous a permis d’identifier pour la première fois un précurseur d’ILC (ILCP) présent à la fois dans notre modèle HIS ainsi que dans le sang périphérique et plusieurs organes lymphoïdes et non-lymphoïdes humains. Cette population circulante d’ILCPs pourrait constituer un substrat pour la production d’ILCs matures dans les tissus périphériques en réponse à des stress environnementaux, inflammatoires et/ou infectieux. Dans une seconde partie de ce travail de thèse, nous avons utilisé ces souris BRGS afin de tester l’efficacité de deux immunothérapies reposant sur les lymphocytes innés pour le traitement d’un carcinome colorectal exprimant EGFR et muté pour KRAS. La première approche a consisté en la co-administration des cellules NK dérivées de sang de cordon ombilical et d'anticorps monoclonal cetuximab afin de promouvoir le mécanisme de cytotoxicité cellulaire dépendante des anticorps (ADCC) contre la tumeur. La seconde stratégie a reposé sur l’injection de nanobodies VHH combinant l’inhibition de l’EGFR et l’activation spécifique du récepteur Vγ9Vδ2 des cellules T effectrices. Les résultats de cette étude soulignent l’importance des modèles murins HIS pour la compréhension du développement des lymphocytes innés humains et pour mieux les mettre à profit dans les thérapies anti-tumeurs

Emerging high-throughput sequencing technologies promise lower costs and higher scalability, yet their performance on archival clinical samples remains poorly characterized. Here, we benchmarked Ultima Genomics UG100 against Illumina Novaseq platforms across single-nuclei RNA-seq (snRNA-seq), whole-transcriptome (WTS), whole-exome (WES), and whole-genome sequencing (WGS) using FFPE tissues from oncologic and immune-mediated diseases. Across matched samples, we systematically assessed data quality, coverage profiles, error spectra, variant concordance and transcriptomic reproducibility. UG100 produced highly comparable results to Illumina, capturing key oncogenic and immune-related transcripts, accurately resolving cellular composition in snRNA-seq, and maintaining sensitivity for lowly expressed genes, despite characteristic insertion-biased indels and modest differences in multi-mapping reads. Discrepancies were subtle, largely limited to pseudogene and non-coding transcripts, and did not affect pathway-level conclusions. Ultima UG100 platform prioritized high precision and reduced low-frequency artifacts, offering a cleaner but more conservative variant-calling profile compared to the more sensitive, yet noisier, Illumina/DRAGEN workflow. This multimodal, clinically oriented assessment provides the first comprehensive evaluation of UG100, demonstrating its translational utility in population-scale genomics, and highlighting the potential for emerging sequencing technologies to lower the cost of biomedical research and clinical diagnostics.

Immune checkpoint inhibitors, particularly T cell targeting anti-PD(L)1 therapies, have revolutionised the treatment landscape for solid malignancies, but challenges related to non-responsiveness and the development of treatment resistance continue to be observed. An additional immunosuppressive axis relates to prostaglandin signaling downstream of cyclooxygenase-2 (COX2), where COX2 inhibitors have shown clinical promise in re-engaging both T and non-T cell immune compartments, yet have suffered from toxicity concerns. We report here the preclinical characterization of OKN4395, a highly potent and specific first-in-class triple antagonist of EP2, EP4, and DP1, major tumor immunosuppressive receptors downstream of COX2. OKN4395 restores immune function on both T cells and NK cells in vitro. Additionally, OKN4395 acts synergistically with anti-PD1 to increase speed and depth of antitumor activity. Overall, these findings robustly support the clinical investigation of OKN4395 in an ongoing Phase 1 trial ( NCT06789172 ) as an innovative cancer immunotherapy for solid tumors, as a single agent and in combination with anti-PD1 therapy.

Recent advancements in probe-based, full-transcriptome, high-resolution technologies for Formalin-Fixed Paraffin-Embedded (FFPE) tissues, such as Visium CytAssist, Chromium Flex (10X Genomics), and GeoMx DSP (Nanostring), have opened new opportunities for studying decades-old archival samples in biobanks, facilitating the generation of data from extensive cohorts. However, the experimental protocols can be labor-intensive and costly; therefore, it is thus essential for researchers to carefully evaluate the strengths and limitations of each technology in relation to their specific research objectives. Here, we report the results of a comparative analysis of the three methods mentioned above on FFPE archival tumor samples from four non-small cell lung cancer, four breast cancer and six diffuse large B-cell lymphoma. We highlight some relative advantages and disadvantages of each method in the context of operational challenges, bioinformatic analysis and biological discovery. Our results show that: 1) all three methods yielded good-quality, highly reproducible transcriptomic data from serial sections of the same FFPE block; 2) GeoMx data contained mixtures of cell types, even when pre-selecting areas with cell type-specific markers; 3) high-throughput spot-level (Visium) or cell-level (Chromium) data enabled the identification of tumor heterogeneity within and between patients, which could be used to identify targeted therapies. Our data support the use of Visium and Chromium for high-throughput and discovery-driven projects, while the GeoMx platform could be suited for addressing specialized questions on targeted regions. All data generated from this study, including GeoMx, Visium, Chromium, H&E, and expert annotations are publicly available.

The thymus is the primary lymphoid organ where naïve T cells are generated; however, with the exception of age, the parameters that govern its function in healthy humans remain unknown. We characterized the variability of thymic function among 1000 age- and sex-stratified healthy adults of the Milieu Intérieur cohort, using quantification of T cell receptor excision circles (TRECs) in peripheral blood T cells as a surrogate marker of thymopoiesis. Age and sex were the only nonheritable factors identified that affect thymic function. TREC amounts decreased with age and were higher in women compared to men. In addition, a genome-wide association study revealed a common variant (rs2204985) within the T cell receptor TCRA-TCRD locus, between the DD2 and DD3 gene segments, which associated with TREC amounts. Strikingly, transplantation of human hematopoietic stem cells with the rs2204985 GG genotype into immunodeficient mice led to thymopoiesis with higher TRECs, increased thymocyte counts, and a higher TCR repertoire diversity. Our population immunology approach revealed a genetic locus that influences thymopoiesis in healthy adults, with potentially broad implications in precision medicine.

Animal models have extensively contributed to our understanding of human immunobiology and to uncover the underlying pathological mechanisms occurring in the development of diseases. However, mouse models do not reproduce the genetic and molecular complexity inherent in human disease conditions. Human immune system (HIS) mouse models that are susceptible to human pathogens and can recapitulate human hematopoiesis and tumor immunobiology provide one means to bridge the interspecies gap. Natural killer cells are the founding member of the innate lymphoid cell (ILC) family. They exert a rapid and strong immune response against tumor and pathogen-infected cells. Their anti-tumor features have long been exploited for therapeutic purposes in the context of cancer. In this review we detail the development of highly immunodeficient mouse strains and the models currently used in cancer research. We summarize the latest improvements in adoptive NK cell therapies and the development of novel NK cell sources. Finally we discuss the advantages of HIS mice to study the interactions between human NK cells and human cancers and to develop new therapeutic strategies.

Therapeutic monoclonal antibodies (mAbs) against the epidermal growth factor receptor (EGFR) act by inhibiting EGFR downstream signaling and by eliciting an NK cell-mediated anti-tumor response. The IgG1 mAb cetuximab has been used for treatment of RASwt metastatic colorectal cancer (mCRC) patients, showing limited efficacy. In the present study we address the potential of adoptive NK cell therapy to overcome these limitations investigating two allogeneic NK cell products, i.e. activated peripheral blood NK cells (A-PBNK) and umbilical cord blood stem cell derived NK cells (UCB-NK). While cetuximab monotherapy was not effective against EGFR- RASwt, EGFR+ RASmut and EGFR+ BRAFmut cells, A-PBNK were able to initiate lysis of EGFR+ colon cancer cells irrespective of RAS or BRAF status. Cytotoxic effects of A-PBNK (but not UCB-NK) were further potentiated significantly by coating EGFR+ colon cancer cells with cetuximab. Of note, a significantly higher cytotoxicity was induced by UCB-NK in EGFR-RASwt (42 ± 8% versus 67 ± 7%), EGFR+ RASmut (20 ± 2% versus 37 ± 6%) and EGFR+ BRAFmut (23 ± 3% versus 43 ± 7%) colon cancer cells compared to A-PBNK and equaled the cytotoxic efficacy of the combination of A-PBNK and cetuximab. The anti-tumor efficacy of UCB-NK cells against cetuximab resistant human EGFR+ RASmut colon cancer cells was further confirmed in an in vivo preclinical mouse model where UCB-NK showed enhanced anti-tumor cytotoxicity against colon cancer independent of EGFR and RAS status. As UCB-NK have been proven safe in a recently conducted phase I clinical trial in acute myeloid leukemia, a fast translation into clinical proof of concept for mCRC could be considered.

The objectives of the study were to determine the validity and reliability of peak velocity (PV), average velocity (AV), peak power (PP) and average power (AP) measurements were made using a linear position transducer. Validity was assessed by comparing measurements simultaneously obtained using the Tendo Weightlifting Analyzer Systemi and T-Force Dynamic Measurement Systemr (Ergotech, Murcia, Spain) during two resistance exercises, bench press (BP) and full back squat (BS), performed by 71 trained male subjects. For the reliability study, a further 32 men completed both lifts using the Tendo Weightlifting Analyzer Systemz in two identical testing sessions one week apart (session 1 vs. session 2). Intraclass correlation coefficients (ICCs) indicating the validity of the Tendo Weightlifting Analyzer Systemi were high, with values ranging from 0.853 to 0.989. Systematic biases and random errors were low to moderate for almost all variables, being higher in the case of PP (bias ±157.56 W; error ±131.84 W). Proportional biases were identified for almost all variables. Test-retest reliability was strong with ICCs ranging from 0.922 to 0.988. Reliability results also showed minimal systematic biases and random errors, which were only significant for PP (bias -19.19 W; error ±67.57 W). Only PV recorded in the BS showed no significant proportional bias. The Tendo Weightlifting Analyzer Systemi emerged as a reliable system for measuring movement velocity and estimating power in resistance exercises. The low biases and random errors observed here (mainly AV, AP) make this device a useful tool for monitoring resistance training. Key pointsThis study determined the validity and reliability of peak velocity, average velocity, peak power and average power measurements made using a linear position transducerThe Tendo Weight-lifting Analyzer Systemi emerged as a reliable system for measuring movement velocity and power.