Dual TLR9 and PD-L1 targeting unleashes dendritic cells to induce durable antitumor immunity.

BACKGROUND: Although immune checkpoint inhibitors have been a breakthrough in clinical oncology, these therapies fail to produce durable responses in a significant fraction of patients. This lack of long-term efficacy may be due to a poor pre-existing network linking innate and adaptive immunity. Here, we present an antisense oligonucleotide (ASO)-based strategy that dually targets toll-like receptor 9 (TLR9) and programmed cell death ligand 1 (PD-L1), aiming to overcome resistance to anti-PD-L1 monoclonal therapy. METHODS: We designed a high-affinity immunomodulatory IM-TLR9:PD-L1-ASO antisense oligonucleotide (hereafter, IM-T9P1-ASO) targeting mouse PD-L1 messenger RNA and activating TLR9. Then, we performed in vitro and in vivo studies to validate the IM-T9P1-ASO activity, efficacy, and biological effects in tumors and draining lymph nodes. We also performed intravital imaging to study IM-T9P1-ASO pharmacokinetics in the tumor. RESULTS: IM-T9P1-ASO therapy, unlike PD-L1 antibody therapy, results in durable antitumor responses in multiple mouse cancer models. Mechanistically, IM-T9P1-ASO activates a state of tumor-associated dendritic cells (DCs), referred to here as DC3s, which have potent antitumor potential but express the PD-L1 checkpoint. IM-T9P1-ASO has two roles: it triggers the expansion of DC3s by engaging with TLR9 and downregulates PD-L1, thereby unleashing the antitumor functions of DC3s. This dual action leads to tumor rejection by T cells. The antitumor efficacy of IM-T9P1-ASO depends on the antitumor cytokine interleukin-12 (IL-12), produced by DC3s, and Batf3, a transcription factor required for DC development. CONCLUSIONS: By simultaneously targeting TLR9 and PD-L1, IM-T9P1-ASO amplifies antitumor responses via DC activation, leading to sustained therapeutic efficacy in mice. By highlighting differences and similarities between mouse and human DCs, this study could serve to develop similar therapeutic strategies for patients with cancer.

Deletion of SNX9 alleviates CD8 T cell exhaustion for effective cellular cancer immunotherapy.

Tumor-specific T cells are frequently exhausted by chronic antigenic stimulation. We here report on a human antigen-specific ex vivo model to explore new therapeutic options for T cell immunotherapies. T cells generated with this model resemble tumor-infiltrating exhausted T cells on a phenotypic and transcriptional level. Using a targeted pooled CRISPR-Cas9 screen and individual gene knockout validation experiments, we uncover sorting nexin-9 (SNX9) as a mediator of T cell exhaustion. Upon TCR/CD28 stimulation, deletion of SNX9 in CD8 T cells decreases PLCγ1, Ca2+, and NFATc2-mediated T cell signaling and reduces expression of NR4A1/3 and TOX. SNX9 knockout enhances memory differentiation and IFNγ secretion of adoptively transferred T cells and results in improved anti-tumor efficacy of human chimeric antigen receptor T cells in vivo. Our findings highlight that targeting SNX9 is a strategy to prevent T cell exhaustion and enhance anti-tumor immunity.

Admixture and population structure in Mexican-Mestizos based on paternal lineages.

In the nonrecombining region of the Y-chromosome, there are single-nucleotide polymorphisms (Y-SNPs) that establish haplogroups with particular geographical origins (European, African, Native American, etc.). The complex process of admixture that gave rise to the majority of the current Mexican population (~93%), known as Mestizos, can be examined with Y-SNPs to establish their paternal ancestry and population structure. We analyzed 18 Y-SNPs in 659 individuals from 10 Mexican-Mestizo populations from different regions of the country. In the total population sample, paternal ancestry was predominately European (64.9%), followed by Native American (30.8%) and African (4.2%). However, the European ancestry was prevalent in the north and west (66.7-95%) and, conversely, Native American ancestry increased in the center and southeast (37-50%), whereas the African ancestry was low and relatively homogeneous (0-8.8%). Although this paternal landscape concurs with previous studies based on genome-wide SNPs and autosomal short tandem repeats (STRs), this pattern contrasts with the maternal ancestry, mainly of Native American origin, based on maternal lineages haplogroups. In agreement with historical records, these results confirm a strong gender-biased admixture history between European males and Native American females that gave rise to Mexican-Mestizos. Finally, pairwise comparisons and analysis of molecular variance tests demonstrated significant population structure (F(ST)=4.68%; P<0.00005), delimiting clusters that were geographically defined as the following: north-west, center-south and southeast.

Role of Eu2+ and Dy3+ Concentration in the Persistent Luminescence of Sr2MgSi2O7 Glass-Ceramics.

In this study, glass-ceramics based on Sr2MgSi2O7 phosphor co-doped with Eu/Dy were obtained from the sintering and crystallisation of glass powders. The glasses were melted in a gas furnace to simulate an industrial process, and the dopant concentration was varied to optimise the luminescence persistence times. The doped parent glasses showed red emission under UV light excitation due to the doping of Eu3+ ions, while the corresponding glass-ceramics showed persistent blue emission corresponding to the presence of Eu2+ in the crystalline environment. The dopant concentration had a strong impact on the sintering/crystallisation kinetics affecting the final glass-ceramic microstructure. The microstructures and morphology of the crystals responsible for the blue emission were observed by scanning electron microscopy-cathodoluminescence. The composition of the crystallised phases and the distribution of rare-earth (RE) ions in the crystals and in the residual glassy phase were determined by X-ray diffraction and energy dispersive X-ray analysis. The emission and persistence of phosphorescence were studied by photoluminescence.

GEF-H1 Signaling upon Microtubule Destabilization Is Required for Dendritic Cell Activation and Specific Anti-tumor Responses.

Dendritic cell (DC) activation is a critical step for anti-tumor T cell responses. Certain chemotherapeutics can influence DC function. Here we demonstrate that chemotherapy capable of microtubule destabilization has direct effects on DC function; namely, it induces potent DC maturation and elicits anti-tumor immunity. Guanine nucleotide exchange factor-H1 (GEF-H1) is specifically released upon microtubule destabilization and is required for DC activation. In response to chemotherapy, GEF-H1 drives a distinct cell signaling program in DCs dominated by the c-Jun N-terminal kinase (JNK) pathway and AP-1/ATF transcriptional response for control of innate and adaptive immune responses. Microtubule destabilization, and subsequent GEF-H1 signaling, enhances cross-presentation of tumor antigens to CD8 T cells. In absence of GEF-H1, anti-tumor immunity is hampered. In cancer patients, high expression of the GEF-H1 immune gene signature is associated with prolonged survival. Our study identifies an alternate intracellular axis in DCs induced upon microtubule destabilization in which GEF-H1 promotes protective anti-tumor immunity.