Biomarker conversion from primary breast cancer to synchronous axillary lymph node metastasis and neoadjuvant therapy response: a single-center analysis.

PURPOSE: The biomarker characteristics of breast cancer plays an important role in predicting treatment sensitivity. The aim of the present study was to compare immunohistochemical profiles (ER, PR, HER2, and Ki67) between the primary tumor and synchronous axillary lymph node metastasis and investigate the subsequent effects on neoadjuvant therapy response. METHODS: A total of 358 patients with pathologically confirmed synchronous axillary lymph node metastasis at first diagnosis and treated by neoadjuvant therapy at Peking University First Hospital from January 1, 2013 to December 31, 2022 were included in this retrospective study. Clinicopathologic data, especially receptor status in primary and metastatic foci, was collected for each case. RESULTS: Change of ER, PR, HER2, and Ki67 expression was observed in 5.9%, 8.7%, 12.6%, and 17.3% of patients, respectively. HR discordance was observed more frequently when the ER status (p = 0.023) or PR status (p = 0.010) of primary tumor was negative, while HER2 discordance seemed to be more frequent when the HER2 status of primary tumor was HER2-0 or HER2-low (p < 0.001). Patients with loss of HR-positivity (positive to negative) responded to neoadjuvant chemotherapy better compared to those with stable positive HR expression (50% vs. 11.1%, p = 0.0017). A significantly decrease in pCR rate was observed in patients with unstable HER2 status, but not in the HER2-0/HER2-low subgroup. CONCLUSION: Receptor discordance between primary tumor and synchronous axillary LNM appears to already exist before any anti-tumor therapy. This instability has limited clinical impact on the choice of neoadjuvant therapy at current stage, but further investigation is warranted with the incremental application of endocrine drugs and ADCs in neoadjuvant therapy.

Heterogeneity in PD-L1 expression between primary and metastatic lymph nodes: a predictor of EGFR-TKI therapy response in non-small cell lung cancer.

BACKGROUND: There is inconclusive evidence to suggest that the expression of programmed cell death ligand 1 (PD-L1) is a putative predictor of response to EGFR-TKI therapy in advanced EGFR-mutant non-small cell lung cancer (NSCLC). We evaluated the heterogeneity in PD-L1 expression in the primary lung site and metastatic lymph nodes to analyze the association between PD-L1 expression and response for patients treated with EGFR-TKI. METHODS: This study reviewed 184 advanced NSCLC patients with EGFR mutations who received first-generation EGFR-TKI as first-line treatment from 2020 to 2021 at Shanghai Chest Hospital. The patients were divided into the primary lung site group (n = 100) and the metastatic lymph nodes group (n = 84) according to the biopsy site. The patients in each group were divided into TPS < 1%, TPS 1-49%, and TPS ≥ 50% groups according to PD-L1 expression. RESULTS: The median PFS was 7 (95% CI: 5.7-8.3) months, and the median OS was 26 (95% CI: 23.5-28.5) months for all patients. No correlation existed between PFS or OS and PD-L1 expression. The median PFS in the primary lung site group was 11 months (95% CI: 9.6-12.4) in the TPS < 1% group, 8 months (95% CI: 6.6-9.4) in TPS 1-49% group, and 4 months (95% CI: 3.2-4.8) in TPS ≥ 50% group, with statistically significant differences (p = 0.000). The median OS of the TPS < 1% group and TPS ≥ 50% group showed a statistically significant difference (p = 0.008) in the primary lung site group. In contrast, PD-L1 expression in the lymph nodes of EGFR-mutant patients was unrelated to PFS or OS after EGFR-TKI therapy. CONCLUSION: PD-L1 expression from the primary lung site might predict clinical benefit from EGFR-TKI, whereas PD-L1 from metastatic lymph nodes did not. TRIAL REGISTRATION: This retrospective study was approved by the Ethics Committee of Shanghai Chest Hospital (ID: IS23060) and performed following the Helsinki Declaration of 1964 (revised 2008).

[Multi-omics prediction of lymph node metastasis status in breast cancer].

Lymph node metastasis status stands as a pivotal prognostic indicator in forecasting the outlook for breast cancer patients. Consequently, precise evaluation of this status holds paramount importance in the staging, treatment, and prognosis of breast cancer. The utilization of radiomics, genomics, proteomics, transcriptomics, and histopathology methodologies has notably enhanced the precision of lymph node metastasis status prediction in breast cancer. This review provides an overview of recent advancements in omics-based lymph node metastasis prediction for breast cancer, elucidating the significance of various omics prediction models and integrated multi-omics models in this predictive endeavor. The overarching goal is to augment the accuracy of preoperative lymph node metastasis status prediction in breast cancer, thereby aiding clinicians in the selection of efficacious personalized treatment strategies, while concurrently averting undertreatment of patients with a heightened risk of metastasis.

Runx3 Regulates CD8+ T Cell Local Expansion and CD43 Glycosylation in Mice by H1N1 Influenza A Virus Infection.

We have recently reported that transcription factor Runx3 is required for pulmonary generation of CD8+ cytotoxic T lymphocytes (CTLs) that play a crucial role in the clearance of influenza A virus (IAV). To understand the underlying mechanisms, we determined the effects of Runx3 knockout (KO) on CD8+ T cell local expansion and phenotypes using an inducible general Runx3 KO mouse model. We found that in contrast to the lungs, Runx3 general KO promoted enlargement of lung-draining mediastinal lymph node (mLN) and enhanced CD8+ and CD4+ T cell expansion during H1N1 IAV infection. We further found that Runx3 deficiency greatly inhibited core 2 O-glycosylation of selectin ligand CD43 on activated CD8+ T cells but minimally affected the cell surface expression of CD43, activation markers (CD44 and CD69) and cell adhesion molecules (CD11a and CD54). Runx3 KO had a minor effect on lung effector CD8+ T cell death by IAV infection. Our findings indicate that Runx3 differently regulates CD8+ T cell expansion in mLNs and lungs by H1N1 IAV infection. Runx3 is required for CD43 core 2 O-glycosylation on activated CD8+ T cells, and the involved Runx3 signal pathway may mediate CD8+ T cell phenotype for pulmonary generation of CTLs.

Interactions between nanoparticles and lymphatic systems: Mechanisms and applications in drug delivery.

The lymphatic system has garnered significant attention in drug delivery research due to the advantages it offers, such as enhancing systemic exposure and enabling lymph node targeting for nanomedicines via the lymphatic delivery route. The journey of drug carriers involves transport from the administration site to the lymphatic vessels, traversing the lymph before entering the bloodstream or targeting specific lymph nodes. However, the anatomical and physiological barriers of the lymphatic system play a pivotal role in influencing the behavior and efficiency of carriers. To expedite research and subsequent clinical translation, this review begins by introducing the composition and classification of the lymphatic system. Subsequently, we explore the routes and mechanisms through which nanoparticles enter lymphatic vessels and lymph nodes. The review further delves into the interactions between nanomedicine and body fluids at the administration site or within lymphatic vessels. Finally, we provide a comprehensive overview of recent advancements in lymphatic delivery systems, addressing the challenges and opportunities inherent in current systems for delivering macromolecules and vaccines.

Quantifying the Lymphatic Transport of Model Therapeutics from the Brain in Rats.

In recent years, the drainage of fluids, immune cells, antigens, fluorescent tracers, and other solutes from the brain has been demonstrated to occur along lymphatic outflow pathways to the deep cervical lymph nodes in the neck. To the best of our knowledge, no studies have evaluated the lymphatic transport of therapeutics from the brain. The objective of this study was to determine the lymphatic transport of model therapeutics of different molecular weights and lipophilicity from the brain using cervical lymph cannulation and ligation models in rats. To do this, anesthetized Sprague-Dawley rats were cannulated at the carotid artery and cannulated, ligated, or left intact at the cervical lymph duct. Rats were administered 14C-ibuprofen (206.29 g/mol, logP 3.84), 3H-halofantrine HCl (536.89 g/mol, logP 8.06), or 3H-albumin (∼65,000 g/mol) via direct injection into the brain striatum at a rate of 0.5 µL/min over 16 min. Plasma or cervical lymph samples were collected for up to 6-8 h following dosing, and brain and lymph nodes were collected at 6 or 8 h. Samples were subsequently analyzed for radioactivity levels via scintillation counting. For 14C-ibuprofen, plasma concentrations over time (plasma AUC0-6h) were >2 fold higher in lymph-ligated rats than in lymph-intact rats, suggesting that ibuprofen is cleared from the brain primarily via nonlymphatic routes (e.g., across the blood-brain barrier) but that this clearance is influenced by changes in lymphatic flow. For 3H-halofantrine, >73% of the dose was retained at the brain dosing site in lymph-intact and lymph-ligated groups, and plasma AUC0-8h values were low in both groups (<0.3% dose.h/mL), consistent with the high retention in the brain. It was therefore not possible to determine whether halofantrine undergoes lymphatic transport from the brain within the duration of the study. For 3H-albumin, plasma AUC0-8h values were not significantly different between lymph-intact, lymph-ligated, and lymph-cannulated rats. However, >4% of the dose was recovered in cervical lymph over 8 h. Lymph/plasma concentration ratios of 3H-albumin were also very high (up to 53:1). Together, these results indicate that 3H-albumin is transported from the brain not only via lymphatic routes but also via the blood. Similar to other tissues, the lymphatics may thus play a significant role in the transport of macromolecules, including therapeutic proteins, from the brain but are unlikely to be a major transport pathway from the brain for small molecule drugs that are not lipophilic. Our rat cervical lymph cannulation model can be used to quantify the lymphatic drainage of different molecules and factors from the brain.

Expression of SOX4 Significantly Predicts the Risk of Lymph Node Metastasis for Patients With Early-Stage Esophageal Squamous Cell Carcinoma.

Esophageal squamous cell carcinoma stands as a notably aggressive malignancy within the digestive system. In cases of early esophageal cancer without lymph node metastasis, endoscopic surgical resection offers a viable alternative, often resulting in improved patient quality of life. However, the paucity of methods to preoperatively ascertain lymph node involvement complicates surgical planning. SOX4 gene was previously found to be highly associated with invasive metastasis in our work through single-cell RNA sequencing on 5 paired tumor/peritumor tissues. This research included the collection of 124 tissue samples from 106 patients (106 tumor and 18 lymph node specimens). Samples were methodically arranged into a tissue microarray and treated with immunohistochemical staining. Statistical analysis was conducted to assess the relationship between them. In the univariate analysis, 3 factors were identified as statistically significant in relation to lymph node metastasis: T category (P = .014), vascular invasion (P < .001), and SOX4 intensity (P = .001). Additionally, when evaluating SOX4 intensity alongside other clinical indicators, SOX4 was shown to independently influence lymph node metastasis. Further, the multivariate analysis revealed that vascular invasion (P < .001) and SOX4 intensity (P = .003) were significantly associated with lymph node metastasis, exhibiting hazard ratios of 10.174 and 7.142, respectively. The results of our study indicate that both SOX4 expression and vascular invasion serve as predictors of lymph node metastasis in patients diagnosed with category T1 esophageal squamous cell carcinoma, underscoring the potential utility of SOX4 in prognostic evaluations.

Impact of conjugation to different lipids on the lymphatic uptake and biodistribution of brush PEG polymers.

Delivery to peripheral lymphatics can be achieved following interstitial administration of nano-sized delivery systems (nanoparticles, liposomes, dendrimers etc) or molecules that hitchhike on endogenous nano-sized carriers (such as albumin). The published work concerning the hitchhiking approach has mostly focussed on the lymphatic uptake of vaccines conjugated directly to albumin binding moieties (ABMs such as lipids, Evans blue dye derivatives or peptides) and their subsequent trafficking into draining lymph nodes. The mechanisms underpinning access and transport of these constructs into lymph fluid, including potential interaction with other endogenous nanocarriers such as lipoproteins, have largely been ignored. Recently, we described a series of brush polyethylene glycol (PEG) polymers containing end terminal short-chain or medium-chain hydrocarbon tails (1C2 or 1C12, respectively), cholesterol moiety (Cho), or medium-chain or long-chain diacylglycerols (2C12 or 2C18, respectively). We evaluated the association of these materials with albumin and lipoprotein in rat plasma, and their intravenous (IV) and subcutaneous (SC) pharmacokinetic profiles. Here we fully detail the association of this suite of polymers with albumin and lipoproteins in rat lymph, which is expected to facilitate lymph transport of the materials from the SC injection site. Additionally, we characterise the thoracic lymph uptake, tissue and lymph node biodistribution of the lipidated brush PEG polymers following SC administration to thoracic lymph cannulated rats. All polymers had moderate lymphatic uptake in rats following SC dosing with the lymph uptake higher for 1C2-PEG, 2C12-PEG and 2C18-PEG (5.8%, 5.9% and 6.7% dose in lymph, respectively) compared with 1C12-PEG and Cho-PEG (both 1.5% dose in lymph). The enhanced lymph uptake of 1C2-PEG, 2C12-PEG and 2C18-PEG appeared related to their association profile with different lipoproteins. The five polymers displayed different biodistribution patterns in major organs and tissues in mice. All polymers reached immune cells deep within the inguinal lymph nodes of mice following SC dosing. The ability to access these immune cells suggests the potential of the polymers as platforms for the delivery of vaccines and immunotherapies. Future studies will focus on evaluating the lymphatic targeting and therapeutic potential of drug or vaccine-loaded polymers in pre-clinical disease models.

Improved pharmacokinetic and lymphatic uptake of Rose Bengal after transfersome intradermal deposition using hollow microneedles.

The lymphatic system is active in several processes that regulate human diseases, among which cancer progression stands out. Thus, various drug delivery systems have been investigated to promote lymphatic drug targeting for cancer therapy; mainly, nanosized particles in the 10-150 nm range quickly achieve lymphatic vessels after an interstitial administration. Herein, a strategy to boost the lymphotropic delivery of Rose Bengal (RB), a hydrosoluble chemotherapeutic, is proposed, and it is based on the loading into Transfersomes (RBTF) and their intradermal deposition in vivo by microneedles. RBTF of 96.27 ± 13.96 nm (PDI = 0.29 ± 0.02) were prepared by a green reverse-phase evaporation technique, and they showed an RB encapsulation efficiency of 98.54 ± 0.09%. In vitro, RBTF remained physically stable under physiological conditions and avoided the release of RB. In vivo, intravenous injection of RBTF prolonged RB half-life of 50 min in healthy rats compared to RB intravenous injection; the RB half-life in rat body was further increased after intradermal injection reaching 24 h, regardless of the formulation used. Regarding lymphatic targeting, RBTF administered intravenously provided an RB accumulation in the lymph nodes of 12.3 ± 0.14 ng/mL after 2 h, whereas no RB accumulation was observed after RB intravenous injection. Intradermally administered RBTF resulted in the highest RB amount detected in lymph nodes after 2 h from the injection (84.2 ± 25.10 ng/mL), which was even visible to the naked eye based on the pink colouration of the drug. In the case of intradermally administered RB, RB in lymph node was detected only at 24 h (13.3 ± 1.41 ng/mL). In conclusion, RBTF proved an efficient carrier for RB delivery, enhancing its pharmacokinetics and promoting lymph-targeted delivery. Thus, RBTF represents a promising nanomedicine product for potentially facing the medical need for novel strategies for cancer therapy.

Spatially resolved quantification of oxygen consumption rate in ex vivo lymph node slices.

Cellular metabolism has been closely linked to activation state in cells of the immune system, and the oxygen consumption rate (OCR) in particular serves as a valuable metric for assessing metabolic activity. Several oxygen sensing assays have been reported for cells in standard culture conditions. However, none have provided a spatially resolved, optical measurement of local oxygen consumption in intact tissue samples, making it challenging to understand regional dynamics of consumption. Therefore, here we established a system to monitor the rates of oxygen consumption in ex vivo tissue slices, using murine lymphoid tissue as a case study. By integrating an optical oxygen sensor into a sealed perfusion chamber and incorporating appropriate correction for photobleaching of the sensor and of tissue autofluorescence, we were able to visualize and quantify rates of oxygen consumption in tissue. This method revealed for the first time that the rate of oxygen consumption in naïve lymphoid tissue was higher in the T cell region compared to the B cell and cortical regions. To validate the method, we measured OCR in the T cell regions of naïve lymph node slices using the optical assay and estimated the consumption rate per cell. The predictions from the optical assay were similar to reported values and were not significantly different from those of the Seahorse metabolic assay, a gold standard method for measuring OCR in cell suspensions. Finally, we used this method to quantify the rate of onset of tissue hypoxia for lymph node slices cultured in a sealed chamber and showed that continuous perfusion was sufficient to maintain oxygenation. In summary, this work establishes a method to monitor oxygen consumption with regional resolution in intact tissue explants, suitable for future use to compare tissue culture conditions and responses to stimulation.

Dynamic Proteomic Changes in Tumor and Immune Organs Reveal Systemic Immune Response to Tumor Development.

In orthotopic mouse tumor models, tumor progression is a complex process, involving interactions among tumor cells, host cell-derived stromal cells, and immune cells. Much attention has been focused on the tumor and its tumor microenvironment, while the host's macroenvironment including immune organs in response to tumorigenesis is poorly understood. Here, we report a temporal proteomic analysis on a subcutaneous tumor and three immune organs (LN, MLN, and spleen) collected on Days 0, 3, 7, 10, 14, and 21 after inoculation of mouse forestomach cancer cells in a syngeneic mouse model. Bioinformatics analysis identified key biological processes during distinct tumor development phases, including an initial acute immune response, the attack by the host immune system, followed by the adaptive immune activation, and the build-up of extracellular matrix. Proteomic changes in LN and spleen largely recapitulated the dynamics of the immune response in the tumor, consistent with an acute defense response on D3, adaptive immune response on D10, and immune evasion by D21. In contrast, the immune response in MLN showed a gradual and sustained activation, suggesting a delayed response from a distal immune organ. Combined analyses of tumors and host immune organs allowed the identification of potential therapeutic targets. A proof-of-concept experiment demonstrated that significant growth reduction can be achieved by dual inhibition of MEK and DDR2. Together, our temporal proteomic dataset of tumors and immune organs provides a useful resource for understanding the interaction between tumors and the immune system and has the potential for identifying new therapeutic targets for cancer treatment.

Endoplasmic reticulum stress response in immune cells contributes to experimental autoimmune encephalomyelitis pathogenesis in rats.

We examined the role of endoplasmic reticulum (ER) stress and the ensuing unfolded protein response (UPR) in the development of the central nervous system (CNS)-directed immune response in the rat model of experimental autoimmune encephalomyelitis (EAE). The induction of EAE with syngeneic spinal cord homogenate in complete Freund's adjuvant (CFA) caused a time-dependent increase in the expression of ER stress/UPR markers glucose-regulated protein 78 (GRP78), X-box-binding protein 1 (XBP1), C/EBP homologous protein (CHOP), and phosphorylated eukaryotic initiation factor 2α (eIF2α) in the draining lymph nodes of both EAE-susceptible Dark Agouti (DA) and EAE-resistant Albino Oxford (AO) rats. However, the increase in ER stress markers was more pronounced in AO rats. CFA alone also induced ER stress, but the effect was weaker and less sustained compared to full immunization. The ultrastructural analysis of DA lymph node tissue by electron microscopy revealed ER dilatation in lymphocytes, macrophages, and plasma cells, while immunoblot analysis of CD3-sorted lymph node cells demonstrated the increase in ER stress/UPR markers in both CD3+ (T cell) and CD3- (non-T) cell compartments. A positive correlation was observed between the levels of ER stress/UPR markers in the CNS-infiltrated mononuclear cells and the clinical activity of the disease. Finally, the reduction of EAE clinical signs by ER stress inhibitor ursodeoxycholic acid was associated with the decrease in the expression of mRNA encoding pro-inflammatory cytokines TNF and IL-1ß, and encephalitogenic T cell cytokines IFN-γ and IL-17. Collectively, our data indicate that ER stress response in immune cells might be an important pathogenetic factor and a valid therapeutic target in the inflammatory damage of the CNS.

Fatty Acid Oxidation Supports Lymph Node Metastasis of Cervical Cancer via Acetyl-CoA-Mediated Stemness.

Accumulating evidence indicates that metabolic reprogramming of cancer cells supports the energy and metabolic demands during tumor metastasis. However, the metabolic alterations underlying lymph node metastasis (LNM) of cervical cancer (CCa) have not been well recognized. In the present study, it is found that lymphatic metastatic CCa cells have reduced dependency on glucose and glycolysis but increased fatty acid oxidation (FAO). Inhibition of carnitine palmitoyl transferase 1A (CPT1A) significantly compromises palmitate-induced cell stemness. Mechanistically, FAO-derived acetyl-CoA enhances H3K27 acetylation (H3K27Ac) modification level in the promoter of stemness genes, increasing stemness and nodal metastasis in the lipid-rich nodal environment. Genetic and pharmacological loss of CPT1A function markedly suppresses the metastatic colonization of CCa cells in tumor-draining lymph nodes. Together, these findings propose an effective method of cancer therapy by targeting FAO in patients with CCa and lymph node metastasis.

Acute stress transiently activates macrophages and chemokines in cervical lymph nodes.

Acute restraint stress (RS) is routinely used to study the effects of psychological and/or physiological stress. We evaluated the impact of RS on cervical lymph nodes in rats at molecular and cellular levels. Male Sprague-Dawley rats were subjected to stress by immobilization for 30, 60, and 120 min (RS30, RS60, and RS120, respectively) and compared with rats of a no-stress control (C) group. The expression of genes encoding chemokines CXCL1/CXCL2 (Cxcl1 and Cxcl2) and their receptor CXCR2 (Cxcr2) was analyzed using reverse transcription-quantitative PCR (RT-qPCR) and microarray analyses. Immunohistochemistry and in situ hybridization were performed to determine the expression of these proteins and the macrophage biomarker CD68. Microarray analysis revealed that the expression of 514 and 496 genes was upregulated and downregulated, respectively, in the RS30 group. Compared with the C group, the RS30 group exhibited a 23.0-, 13.0-, and 1.6-fold increase in Cxcl1, Cxcl2, and Cxcr2 expression. Gene Ontology analysis revealed the involvement of these three upregulated genes in the cytokine network, inflammation, and leukocyte chemotaxis and migration. RT-qPCR analysis indicated that the mRNA levels of Cxcl1 and Cxcl2 were significantly increased in the RS30 group but were reverted to normal levels in the RS60 and RS120 groups. Cxcr2 mRNA level was significantly increased in the RS30 and RS120 groups compared with that in the C group. RS-induced CXCL1-immunopositive cells corresponded to B/plasma cells, whereas CXCL2-immunopositive cells corresponded to endothelial cells of the high endothelial venules. Stress-induced CXCR2-immunopositive cells corresponded to macrophages. Psychological and/or physiological stress induces an acute stress response and formation of an immunoreactive microenvironment in cervical lymph nodes, with the CXCL1/CXCL2-CXCR2 axis being pivotal in the acute stress response.

Case report: A case of rare metastasis of gastric cancer to the axillary lymph node metastasis treated with combination immunotherapy.

Lymph node (LN) metastasis is a common mode of metastasis in advanced gastric cancer (GC), while axillary LN metastasis infrequently occurs in GC. There are few reports on this rare type of metastasis - especially its clinicopathological features - and systemic treatment are unclear. We describe a case of GC with extensive metastasis, including the rare axillary LN metastasis. The patient achieved partial response of optimal efficacy, who was treated with combination immunotherapy as second-line treatment for nearly two years. The potential mechanisms were revealed by clinical and immune characteristics, such as high expression of PD-L1, high tumor mutational burden (TMB-H), Epstein-Barr virus (EBV) positive and CD8+ tumor-infiltrating lymphocyte positive.

Prevention of experimental autoimmune encephalomyelitis by targeting 6-sulfo sialyl Lewis X glycans involved in lymphocyte homing.

Lymphocyte homing to peripheral lymph nodes (PLN) is critical for immune surveillance. However, autoimmune diseases such as multiple sclerosis (MS) can occur due to excessive immune responses in the PLN. Here we show that 6-sulfo sialyl Lewis X (6-sulfo sLex) glycans on high endothelial venules that function as ligands for l-selectin on lymphocytes play a critical role in the pathogenesis of experimental autoimmune encephalomyelitis (EAE), an animal model of MS. In N-acetylglucosamine-6-O-sulfotransferase (GlcNAc6ST)-1 and GlcNAc6ST-2 double-knockout mice lacking the expression of 6-sulfo sLeX glycans, the EAE symptoms and the numbers of effector Th1 and Th17 cells in the draining lymph nodes (dLN) and spinal cords (SC) were significantly reduced. To determine whether 6-sulfo sLeX could serve as a target for MS, we also examined the effects of anti-glycan monoclonal antibody (mAb) SF1 against 6-sulfo sLeX in EAE. Administration of mAb SF1 significantly reduced EAE symptoms and the numbers of antigen-specific effector T cells in the dLN and SC in association with suppression of critical genes including Il17a and Il17f that are involved in the pathogenesis of EAE. Taken together, these results suggest that 6-sulfo sLeX glycan would serve as a novel target for MS.

Lymph node and tumor-associated PD-L1+ macrophages antagonize dendritic cell vaccines by suppressing CD8+ T cells.

Current immunotherapies provide limited benefits against T cell-depleted tumors, calling for therapeutic innovation. Using multi-omics integration of cancer patient data, we predict a type I interferon (IFN) responseHIGH state of dendritic cell (DC) vaccines, with efficacious clinical impact. However, preclinical DC vaccines recapitulating this state by combining immunogenic cancer cell death with induction of type I IFN responses fail to regress mouse tumors lacking T cell infiltrates. Here, in lymph nodes (LNs), instead of activating CD4+/CD8+ T cells, DCs stimulate immunosuppressive programmed death-ligand 1-positive (PD-L1+) LN-associated macrophages (LAMs). Moreover, DC vaccines also stimulate PD-L1+ tumor-associated macrophages (TAMs). This creates two anatomically distinct niches of PD-L1+ macrophages that suppress CD8+ T cells. Accordingly, a combination of PD-L1 blockade with DC vaccines achieves significant tumor regression by depleting PD-L1+ macrophages, suppressing myeloid inflammation, and de-inhibiting effector/stem-like memory T cells. Importantly, clinical DC vaccines also potentiate T cell-suppressive PD-L1+ TAMs in glioblastoma patients. We propose that a multimodal immunotherapy and vaccination regimen is mandatory to overcome T cell-depleted tumors.

IGH repertoire analysis at scale: deciphering the complexity of B cell infiltration and migration in esophageal squamous cell carcinoma.

Tumor-infiltrating B-lineage cells have become predictors of prognosis and immunotherapy responses in various cancers. However, limited knowledge about their infiltration and migration patterns has hindered the understanding of their anti-tumor functions. Here, we examined the immunoglobulin heavy chain (IGH) repertoires in 496 multi-regional tumor, 107 normal tissue, and 48 metastatic lymph node samples obtained from 107 patients with esophageal squamous cell carcinoma (ESCC). Our study revealed higher IgG-type B-lineage cells infiltration in tumors than in healthy tissue, which was associated with improved patient outcomes. Genes such as ACTN1, COL6A5, and pathways like focal adhesion, which shapes the physical structure of tumors, could affect B-lineage cell infiltration. Notably, the IGH sequence was used as an identity-tag to monitor B cell migration, and their infiltration schema within the tumor were depicted based on our multi-regional tumor specimens. This analysis revealed an escalation in B cell clones overlapped between metastatic lymph nodes and tumors. Therefore, the Lymph Node Activation Index was defined, which could predict the outcomes of patients with lymph node metastasis. This research introduces a novel framework for probing B cell infiltration and migration within the tumor microenvironment using large-scale transcriptome data, while simultaneously providing fresh perspectives on B cell immunology within ESCC.

Para- and Transcellular Transport Kinetics of Nanoparticles across Lymphatic Endothelial Cells.

Lymphatic vessels have received significant attention as drug delivery targets, as they shuttle materials from peripheral tissues to the lymph nodes, where adaptive immunity is formed. Delivery of immune modulatory materials to the lymph nodes via lymphatic vessels has been shown to enhance their efficacy and also improve the bioavailability of drugs when delivered to intestinal lymphatic vessels. In this study, we generated a three-compartment model of a lymphatic vessel with a set of kinematic differential equations to describe the transport of nanoparticles from the surrounding tissues into lymphatic vessels. We used previously published data and collected additional experimental parameters, including the transport efficiency of nanoparticles over time, and also examined how nanoparticle formulation affected the cellular transport mechanisms using small molecule inhibitors. These experimental data were incorporated into a system of kinematic differential equations, and nonlinear, least-squares curve fitting algorithms were employed to extrapolate transport coefficients within our model. The subsequent computational framework produced some of the first parameters to describe transport kinetics across lymphatic endothelial cells and allowed for the quantitative analysis of the driving mechanisms of transport into lymphatic vessels. Our model indicates that transcellular mechanisms, such as micro- and macropinocytosis, drive transport into lymphatics. This information is crucial to further design strategies that will modulate lymphatic transport for drug delivery, particularly in diseases like lymphedema, where normal lymphatic functions are impaired.