Intra-arterial Pressure Enabled Drug Delivery Significantly Increases Penetration of Glass Microspheres in a Porcine Liver Tumor Model.

PURPOSE: To test the hypothesis that Pressure Enabled Drug Delivery (PEDD) with a TriNav device (TNV-21120-35, TriSalus Life Sciences, Westminster, CO) would improve the delivery of surrogate therapeutic glass microspheres (GM) via hepatic artery infusion (HAI) to liver tumors when compared to a conventional endhole microcatheter. MATERIALS AND METHODS: The study was conducted in transgenic pigs (Oncopigs) with induced liver tumors. Tumors were infused intra-arterially with fluorescently labeled GM. PEDD with a TriNav device was compared to conventional endhole delivery in both lobar and selective infusions. Near-Infrared (nearIR) imaging was used to detect GM fluorescent signal in tumors. Image analysis with a custom Deep Learning algorithm (Visiopharm A/S) was used to quantitate signal intensity in relation to the tumor border. RESULTS: With lobar infusions, significant increases in GM signal intensity were observed in and around tumors after PEDD (n=10) when compared to conventional delivery (n=7), with PEDD increasing penetration into the tumor by 117% (p = 0.004). In selective infusions, PEDD (n=9) increased penetration into the tumor by 39% relative to conventional delivery (n=8, p =0.032). Lobar PEDD delivery of GM to the tumor was statistically equivalent to conventional selective delivery (p=0.497). CONCLUSIONS: PEDD with a TriNav device significantly improved GM uptake in liver tumors relative to conventional infusion in both lobar and selective procedures. Lobar GM delivery with PEDD was equivalent to conventional selective delivery with an endhole device, suggesting that proximal PEDD infusions may enable effective delivery without selection of distal target vessels.

Blockade of interleukin 10 potentiates antitumour immune function in human colorectal cancer liver metastases.

OBJECTIVE: Programmed cell death protein 1 (PD-1) checkpoint inhibition and adoptive cellular therapy have had limited success in patients with microsatellite stable colorectal cancer liver metastases (CRLM). We sought to evaluate the effect of interleukin 10 (IL-10) blockade on endogenous T cell and chimeric antigen receptor T (CAR-T) cell antitumour function in CRLM slice cultures. DESIGN: We created organotypic slice cultures from human CRLM (n=38 patients' tumours) and tested the antitumour effects of a neutralising antibody against IL-10 (αIL-10) both alone as treatment and in combination with exogenously administered carcinoembryonic antigen (CEA)-specific CAR-T cells. We evaluated slice cultures with single and multiplex immunohistochemistry, in situ hybridisation, single-cell RNA sequencing, reverse-phase protein arrays and time-lapse fluorescent microscopy. RESULTS: αIL-10 generated a 1.8-fold increase in T cell-mediated carcinoma cell death in human CRLM slice cultures. αIL-10 significantly increased proportions of CD8+ T cells without exhaustion transcription changes, and increased human leukocyte antigen - DR isotype (HLA-DR) expression of macrophages. The antitumour effects of αIL-10 were reversed by major histocompatibility complex class I or II (MHC-I or MHC-II) blockade, confirming the essential role of antigen presenting cells. Interrupting IL-10 signalling also rescued murine CAR-T cell proliferation and cytotoxicity from myeloid cell-mediated immunosuppression. In human CRLM slices, αIL-10 increased CEA-specific CAR-T cell activation and CAR-T cell-mediated cytotoxicity, with nearly 70% carcinoma cell apoptosis across multiple human tumours. Pretreatment with an IL-10 receptor blocking antibody also potentiated CAR-T function. CONCLUSION: Neutralising the effects of IL-10 in human CRLM has therapeutic potential as a stand-alone treatment and to augment the function of adoptively transferred CAR-T cells.

Antigen Receptor T Cells (CAR-T) Effectively Control Tumor Growth in a Colorectal Liver Metastasis Model.

BACKGROUND: Effective treatment of solid tumors requires multi-modality approaches. In many patients with stage IV liver disease, current treatments are not curative. Chimeric antigen receptor T cells (CAR-T) are an intriguing option following success in hematological malignancies, but this has not been translated to solid tumors. Limitations include sub-optimal delivery and elevated interstitial fluid pressures. We developed a murine model to test the impact of high-pressure regional delivery (HPRD) on trafficking to liver metastases (LM) and tumor response. MATERIALS AND METHODS: CAR-T were generated from CD45.1 mice and adoptively transferred into LM-bearing CD45.2 mice via regional or systemic delivery (RD, SD). Trafficking, tumor growth, and toxicity were evaluated with flow cytometry, tumor bioluminescence (TB, photons/sec log2-foldover baseline), and liver function tests (LFTs). RESULTS: RD of CAR-T was more effective at controlling tumor growth versus SD from post-treatment days (PTD) 2-7 (P = 0.002). HPRD resulted in increased CAR-T penetration versus low-pressure RD (LPRD, P = 0.004), suppression of tumor proliferation (P = 0.03), and trended toward improved long-term control at PTD17 (TB=3.7 versus 6.1, P = 0.47). No LFT increase was noted utilizing HPRD versus LPRD (AST/ALT P = 0.65/0.84) while improved LFTs in RD versus SD groups suggested better tumor control (HPRD AST/ALT P = 0.04/0.04, LPRD AST/ALT P = 0.02/0.02). CONCLUSIONS: Cellular immunotherapy is an emerging option for solid tumors. Our model suggests RD and HPRD improved CAR-T penetration into solid tumors with improved short-term tumor control. Barriers associated with SD can be overcome using RD techniques to maximize therapeutic delivery and HPRD may further augment efficacy without increased toxicity.

Multidisciplinary Management of Angiosarcoma - A Review.

Angiosarcomas (AS) are a diverse group of soft tissue sarcomas, arising from blood and lymphatic vessels. They frequently present in the elderly, and in patients with previous radiation or lymphedema. A wide range of genetic derangements contribute to their development, and AS histology is often high-grade in keeping with aggressive disease biology. The clinical presentation, while often innocuous, is marked by its infiltrative and aggressive nature, with a proclivity for metastatic spread, and outcomes are often poor. Surgery is performed for localized, resectable cases. A multidisciplinary approach, appropriately employing surgery, radiation, chemotherapy, or potentially recently approved immune-oncology agents, can result in positive outcomes.

Trends in Incidence and Factors Affecting Survival of Patients With Cholangiocarcinoma in the United States.

Background: Cholangiocarcinoma (CCA) includes cancers arising from the intrahepatic and extrahepatic bile ducts. The etiology and pathogenesis of CCA remain poorly understood. This is the first study investigating both incidence patterns of CCA from 1973 through 2012 and demographic, clinical, and treatment variables affecting survival of patients with CCA. Patients and Methods: Using the SEER database, age-adjusted incidence rates were evaluated from 1973-2012 using SEER*Stat software. A retrospective cohort of 26,994 patients diagnosed with CCA from 1973-2008 was identified for survival analysis. Cox proportional hazards models were used to perform multivariate survival analysis. Results: Overall incidence of CCA increased by 65% from 1973-2012. Extrahepatic CCA (ECC) remained more common than intrahepatic CCA (ICC), whereas the incidence rates for ICC increased by 350% compared with a 20% increase seen with ECC. Men belonging to non-African American and non-Caucasian ethnicities had the highest incidence rates of CCA. This trend persisted throughout the study period, although African Americans and Caucasians saw 50% and 59% increases in incidence rates, respectively, compared with a 9% increase among other races. Median overall survival (OS) was 8 months in patients with ECC compared with 4 months in those with ICC. Our survival analysis found Hispanic women to have the best 5-year survival outcome (P<.0001). OS diminished with age (P<.0001), and ECC had better survival outcomes compared with ICC (P<.0001). Patients who were married, were nonsmokers, belonged to a higher income class, and underwent surgery had better survival outcomes compared with others (P<.0001). Conclusions: This is the most up-to-date study of CCA from the SEER registry that shows temporal patterns of increasing incidence of CCA across different races, sexes, and ethnicities. We identified age, sex, race, marital status, income, smoking status, anatomic location of CCA, tumor grade, tumor stage, radiation, and surgery as independent prognostic factors for OS in patients with CCA.

Mincle Signaling Promotes Con A Hepatitis.

Con A hepatitis is regarded as a T cell-mediated model of acute liver injury. Mincle is a C-type lectin receptor that is critical in the immune response to mycobacteria and fungi but does not have a well-defined role in preclinical models of non-pathogen-mediated inflammation. Because Mincle can ligate the cell death ligand SAP130, we postulated that Mincle signaling drives intrahepatic inflammation and liver injury in Con A hepatitis. Acute liver injury was assessed in the murine Con A hepatitis model using C57BL/6, Mincle(-/-), and Dectin-1(-/-) mice. The role of C/EBPß and hypoxia-inducible factor-1α (HIF-1α) signaling was assessed using selective inhibitors. We found that Mincle was highly expressed in hepatic innate inflammatory cells and endothelial cells in both mice and humans. Furthermore, sterile Mincle ligands and Mincle signaling intermediates were increased in the murine liver in Con A hepatitis. Most significantly, Mincle deletion or blockade protected against Con A hepatitis, whereas Mincle ligation exacerbated disease. Bone marrow chimeric and adoptive transfer experiments suggested that Mincle signaling in infiltrating myeloid cells dictates disease phenotype. Conversely, signaling via other C-type lectin receptors did not alter disease course. Mechanistically, we found that Mincle blockade decreased the NF-κß-related signaling intermediates C/EBPß and HIF-1α, both of which are necessary in macrophage-mediated inflammatory responses. Accordingly, Mincle deletion lowered production of nitrites in Con A hepatitis and inhibition of both C/EBPß and HIF-1α reduced the severity of liver disease. Our work implicates a novel innate immune driver of Con A hepatitis and, more broadly, suggests a potential role for Mincle in diseases governed by sterile inflammation.

Tumor-Associated Macrophage Infiltration in Colorectal Cancer Liver Metastases is Associated With Better Outcome.

BACKGROUND: Tumor-associated macrophages (TAMs) support growth in most human cancers, with the notable exception of colorectal adenocarcinoma, in which TAM infiltration of primary tumors is correlated with a better outcome. The importance of TAMs in colorectal liver metastases (CLM) is unknown. METHODS: Using a tissue microarray of CLM resected at their institution from 1998 to 2000, the authors quantified immune marker expression by immunohistochemistry (IHC) using Metamorph Image Analysis software. Findings showed that CD68, CD3, CD4, CD8, FoxP3, and MHC-I were correlated with overall survival (OS) and disease-free survival (DFS). RESULTS: Tumor cores from 158 patients were analyzed. The median follow-up period was 117 months for survivors (n = 39). The univariate analysis showed a significant positive association between DFS and CD4+ (p = 0.025) and CD68+ (p = 0.007). The findings showed a significant positive correlation of OS with CD4+ (p = 0.042), whereas the correlation with CD68+ was not significant (p = 0.17). Cutoffs were determined to dichotimize each marker for the highest log-rank statistic. Patients with CD4high had a median OS of 115 months and DFS of 41 months (p = 0.007 compared with 40 and 16 months, respectively, for patients with CD4low (p = 0.022). Patients with CD68high had a median OS of 50 months and a median DFS of 25 months (p = 0.67) compared with 43 and 15 months (p = 0.028). In the multivariate analysis of factors affecting DFS, high CD68 was associated with longer DFS (hazard ratio [HR], 0.63, 95% confidence interval [CI], 0.43-0.94; p = 0.02), independently of clinicopathologic variables and CD4. CONCLUSIONS: High TAM infiltration in resected CLM is associated with better outcome, independently of known clinicopathologic and immune predictors. This suggests that TAM depletion, which is being tested clinically in other cancers, may be detrimental in CLM.

Regional immunotherapy for liver and peritoneal metastases.

Pancreatic adenocarcinoma is a biologically aggressive disease, with liver and peritoneal metastases being a frequent cause of death. We examine how the pancreatic carcinoma microenvironment and immunosuppressive landscape favor tumor progression. Immunotherapy has shown promise in select solid tumors, yet challenges remain in applying these gains to stage IV pancreatic adenocarcinoma. We discuss how regional therapy strategies may be leveraged to open new avenues for treating pancreatic carcinoma metastases with immunotherapy.

Predictors of surgical non-referral for colorectal liver metastases.

BACKGROUND: Surgical resection is the only curative option for patients with colorectal liver metastases (CRLM). The objective of our study was to identify factors associated with failure to refer patients with CRLM to a surgeon with oncologic and hepatobiliary expertise. MATERIALS AND METHODS: Data were retrospectively reviewed on 75 patients with CRLM treated at our institution. Patients were divided into referred and nonreferred groups for comparison. Quantitative assessment of association was tabulated using the odds ratio (OR). Statistical comparison was performed using the chi-square test and multiple regression models. Overall survival (OS) was calculated using the Kaplan-Meier method. Multivariate analysis was done using Cox regression. RESULTS: Factors independently associated with lower surgical referral rates included age ≥ 65 y (OR 0.29, 95% confidence interval [CI] 0.09-0.89, P = 0.032), bilobar CRLM (OR 0.35, 95% CI 0.09-0.97, P = 0.048), and presence of >3 CRLM (OR 0.33, 95% CI 0.11-0.94, P = 0.044). The 5-y OS for referred patients was 33% compared with only 8% in patients who were not referred (P < 0.001). Factors independently associated with worse OS included age ≥ 65 y (hazard ratio [HR] 2.01, 95% CI 1.12-3.59, P = 0.019), bilobar hepatic metastases (HR 3.04, 95% CI 1.62-5.70, P < 0.001), and the presence of extrahepatic metastases (HR 2.11, 95% CI 1.02-4.16, P = 0.011). Referral to a surgeon was associated with improved OS (HR 0.42, 95% CI 0.24-0.74, P = 0.003). CONCLUSIONS: Failure to refer CRLM patients for surgical evaluation is associated with aggressive biologic features that do not necessarily preclude resection. Determination of resectability should be made with input from appropriately trained surgical experts.

Liver myeloid-derived suppressor cells expand in response to liver metastases in mice and inhibit the anti-tumor efficacy of anti-CEA CAR-T.

Chimeric antigen receptor-modified T cell (CAR-T) technology, a promising immunotherapeutic tool, has not been applied specifically to treat liver metastases (LM). While CAR-T delivery to LM can be optimized by regional intrahepatic infusion, we propose that liver CD11b+Gr-1+ myeloid-derived suppressor cells (L-MDSC) will inhibit the efficacy of CAR-T in the intrahepatic space. We studied anti-CEA CAR-T in a murine model of CEA+ LM and identified mechanisms through which L-MDSC expand and inhibit CAR-T function. We established CEA+ LM in mice and studied purified L-MDSC and responses to treatment with intrahepatic anti-CEA CAR-T infusions. L-MDSC expanded threefold in response to LM, and their expansion was dependent on GM-CSF, which was produced by tumor cells. L-MDSC utilized PD-L1 to suppress anti-tumor responses through engagement of PD-1 on CAR-T. GM-CSF, in cooperation with STAT3, promoted L-MDSC PD-L1 expression. CAR-T efficacy was rescued when mice received CAR-T in combination with MDSC depletion, GM-CSF neutralization to prevent MDSC expansion, or PD-L1 blockade. As L-MDSC suppressed anti-CEA CAR-T, infusion of anti-CEA CAR-T in tandem with agents targeting L-MDSC is a rational strategy for future clinical trials.

Chemotherapy does not adversely impact outcome following post-incisional hernia repair with biomaterial mesh.

BACKGROUND: Patients receiving chemotherapy are at increased risk for developing recurrent or post-incisional hernias (PIH). Biological materials are an alternative to synthetic mesh in contaminated fields. The impact of chemotherapy on biomaterial tissue ingrowth and integration has not been well studied. METHODS: From 2008 to 2011 patients who underwent PIH repair with biomaterial mesh (Biodesign®) were selected. Patients were divided into two groups: those receiving chemotherapy (CT) and those not receiving chemotherapy (NCT). RESULTS: Forty-five patients were identified, 28 (62%) in the NCT group and 17 (38%) in the CT group. Median follow up for NCT and CT groups were 27 and 17 months, respectively. A total of 9/45 (20%) surgical site infections (SSI) were diagnosed, with 6/28 (21%) in the NCT and 3/17 (18%) in the CT group (P = 0.53). Seroma formation was seen in 5/28 (18%) of NCT patients and 4/17 (23%) in CT group (P = 0.46). Overall hernia recurrence rate was 22%, and the rates of recurrence were similar among the CT 3/17 (18%) and NCT 7/28 (25%) groups (P = 0.42). CONCLUSION: The use of perioperative chemotherapy did not increase the rate of wound complications following PIH repair with biologic mesh in this group of patients.

The prognostic value of liver tumor T cell infiltrates.

Tumor infiltrating lymphocytes (TIL) have been demonstrated to predict oncologic outcomes following resection of primary intrahepatic neoplasms and metastatic liver tumors. Despite strong immunosuppressive factors within the intrahepatic space, TIL are frequently demonstrated in liver tumors. The presence of TIL within liver tumors provides evidence of a host immune response that may be protective, but often is rendered ineffective by tumor induced immune dysfunction. In this review, we discuss techniques involved in studying TIL and subsets of TIL commonly identified. We emphasize the unique nature of the intrahepatic milieu that promotes immunosuppression, and how liver TIL and TIL ratios can be used as indicators of prognosis. Several types of primary and metastatic liver tumors are considered to highlight the similarities and important differences in TIL responses, which likely reflect how intrahepatic immunity is influenced by tumor biology. The studies we discuss indicate that tumor infiltration by suppressor cells and expression of immunoinhibitory molecules by TIL limits the anti-tumor immune function of effector T cells. Most patients fail to mount an adequate immune response to liver tumors, which provides compelling rationale for clinical study of immunotherapy for intrahepatic neoplasms.

Immunotherapy for solid tumors--a review for surgeons.

Immunotherapy has evolved considerably in the last decade and is becoming an integral component of the armamentarium for the treatment of patients with advanced solid tumors. It is important for clinicians, especially surgeons, to understand the basic principles of novel immunotherapies and the immune system. This review summarizes the evolution of the most relevant immunotherapies, their mechanisms of action, the data supporting their clinical use, and integration of immunotherapy into multidisciplinary management of solid tumors. This review should serve as a primer for clinicians and surgeons to understand the rapidly evolving field of immunotherapy.

Real-world evidence of Pressure-Enabled Drug Delivery for trans-arterial chemoembolization and radioembolization among patients with hepatocellular carcinoma and liver metastases.

OBJECTIVES: Pressure-Enabled Drug Delivery (PEDD), a method using pressure to advance catheter-delivered drug distribution, can improve treatment for hepatocellular carcinoma (HCC) and liver metastases, but real-world evidence is limited. We compared baseline patient characteristics, clinical complexity, and post-procedure healthcare resource utilization (HRUs) and clinical complications for PEDD and non-PEDD procedures. METHODS: This study used a retrospective, longitudinal, cohort design of claims data from Clarivate's Real World Data Repository, which includes 98% of US payers with over 300 million unique patients from all US states. We identified patients with a trans-arterial chemoembolization (TACE) or trans-arterial radioembolization (TARE) from 1 January 2019 to 31 December 2022. Subsamples grouped patients with HCC receiving a TARE procedure at their first embolization and patients with metastatic colorectal cancer (CRC) that received a TARE procedure. We reported descriptive comparisons of our full sample of patients with HCC and liver metastases receiving PEDD versus non-PEDD procedures. We then conducted a matching-adjusted comparison of HRUs and clinical complications for PEDD and non-PEDD patients among our subsamples (HCC receiving a TARE procedure at their first embolization and patients with metastatic CRC that received a TARE procedure). Matching was based on baseline demographic and clinical characteristics using coarsened exact matching and propensity-score matching. HRUs included inpatient, outpatient, and emergency department visits. Clinical complications included ascites, cholecystitis, fatigue, gastric ulcer, gastritis, jaundice, LFT increase, lymphopenia, portal hypertension, and post-embolization syndrome. RESULTS: PEDD procedures were used on patients with worse baseline disease burdens: baseline Charlson comorbidity index (mean of 6.5 vs. 5.8), any prior clinical complication related to underlying disease (33.7 vs. 31.0%), and prior systemic therapy (22.1% vs. 16.2%). PEDD patients had a greater number of procedural codes indicative of technical complexity for TACE (PEDD mean = 226.3; non-PEDD mean = 134.5; p value <.01) and TARE (PEDD mean = 205.56; non-PEDD mean = 94.8; p value <0.01). Matching-adjusted analyses of patients with HCC and CRC demonstrated comparable HRU and clinical complications for PEDD and non-PEDD procedures post-index. CONCLUSION: Despite higher baseline disease burden and complexity, post-procedure HRU and clinical complications for PEDD patients were similar to non-PEDD patients. The complex baseline clinical profile may reflect selection of challenging cases for PEDD use. Future studies should validate the benefits observed with PEDD embolization in larger samples with greater statistical power.

Anti-KIT designer T cells for the treatment of gastrointestinal stromal tumor.

BACKGROUND: Imatinib mesylate is an effective treatment for metastatic gastrointestinal stromal tumor (GIST). However, most patients eventually develop resistance and there are few other treatment options. Immunotherapy using genetically modified or designer T cells (dTc) has gained increased attention for several malignancies in recent years. The aims of this study were to develop and test novel anti-KIT dTc engineered to target GIST cells. METHODS: Human anti-KIT dTc were created by retroviral transduction with novel chimeric immune receptors (CIR). The gene for stem cell factor (SCF), the natural ligand for KIT, was cloned into 1st generation (SCF-CD3ζ, 1st gen) and 2nd generation (SCF-CD28-CD3ζ, 2nd gen) CIR constructs. In vitro dTc proliferation and tumoricidal capacity in the presence of KIT+ tumor cells were measured. In vivo assessment of dTc anti-tumor efficacy was performed by treating immunodeficient mice harboring subcutaneous GIST xenografts with dTc tail vein infusions. RESULTS: We successfully produced the 1st and 2nd gen anti-KIT CIR and transduced murine and human T cells. Average transduction efficiencies for human 1st and 2nd gen dTc were 50% and 42%. When co-cultured with KIT+ tumor cells, both 1st and 2nd gen dTc proliferated and produced IFNγ. Human anti-KIT dTc were efficient at lysing GIST in vitro compared to untransduced T cells. In mice with established GIST xenografts, treatment with either 1st or 2nd gen human anti-KIT dTc led to significant reductions in tumor growth rates. CONCLUSIONS: We have constructed a novel anti-KIT CIR for production of dTc that possess specific activity against KIT+ GIST in vitro and in vivo. Further studies are warranted to evaluate the therapeutic potential and safety of anti-KIT dTc.

Regulatory T cell infiltration predicts outcome following resection of colorectal cancer liver metastases.

BACKGROUND: Tumor-infiltrating lymphocyte (TIL) counts in colorectal cancer liver metastases (CRCLM) predict survival following resection. While CD4 and CD8 T cells have been correlated with outcome following CRCLM resection, the role of regulatory T cells (Treg) is not well defined. METHODS: TIL in 188 patients who underwent CRCLM resection between 1998 and 2000 were analyzed by immunohistochemistry using tissue microarrays. Correlation between TIL composition and outcome was determined while controlling for established prognostic factors. Total T cells (CD3), helper T cells (CD4), cytotoxic T cells (CD8), and Treg (FoxP3) were analyzed. RESULTS: Median follow-up time was 40 months for all patients and 95 months for survivors. Overall survival (OS) at 5 and 10 years was 40 and 25%, respectively. The CD4 T cell count correlated with OS (p = .02) and recurrence-free survival (p = .04). A high number of CD8 T cells relative to total T cells (CD8:CD3 ratio) predicted longer OS times (p = .05). Analysis of Treg revealed that high FoxP3:CD4 (p = .03) and FoxP3:CD8 (p = .05) ratios were independent predictors of shorter OS. Patients with a high clinical risk score (CRS) were more likely to have a high number of intratumoral Treg, and patients ≥65 years old had a less robust CRCLM T cell infiltration. CONCLUSIONS: A high number of Treg relative to CD4 or CD8 T cells predicted poor outcome, suggesting an immunosuppressive role for FoxP3 + TIL. The intratumoral immune response was an independent predictor of outcome in patients with colorectal liver metastases.

Obstructive jaundice expands intrahepatic regulatory T cells, which impair liver T lymphocyte function but modulate liver cholestasis and fibrosis.

Although obstructive jaundice has been associated with a predisposition toward infections, the effects of bile duct ligation (BDL) on bulk intrahepatic T cells have not been clearly defined. The aim of this study was to determine the consequences of BDL on liver T cell phenotype and function. After BDL in mice, we found that bulk liver T cells were less responsive to allogeneic or syngeneic Ag-loaded dendritic cells. Spleen T cell function was not affected, and the viability of liver T cells was preserved. BDL expanded the number of CD4(+)CD25(+)Foxp3(+) regulatory T cells (Treg), which were anergic to direct CD3 stimulation and mediated T cell suppression in vitro. Adoptively transferred CD4(+)CD25(-) T cells were converted into Treg within the liver after BDL. In vivo depletion of Treg after BDL restored bulk liver T cell function but exacerbated the degrees of inflammatory cytokine production, cholestasis, and hepatic fibrosis. Thus, BDL expands liver Treg, which reduce the function of bulk intrahepatic T cells yet limit liver injury.

Pressure-Enabled Drug Delivery (PEDD) of a class C TLR9 agonist in combination with checkpoint inhibitor therapy in a murine pancreatic cancer model.

BACKGROUND: Systemic immunotherapy has had limited clinical benefit in pancreatic ductal adenocarcinoma. This is thought to be due to its desmoplastic immunosuppressive tumor microenvironment in addition to high intratumoral pressures that limit drug delivery. Recent preclinical cancer models and early-phase clinical trials have demonstrated the potential of toll-like receptor 9 agonists, including the synthetic CpG oligonucleotide SD-101, to stimulate a wide range of immune cells and eliminate suppressive myeloid cells. We hypothesized that Pressure-Enabled Drug Delivery via Pancreatic Retrograde Venous Infusion of toll-like receptor 9 agonist would improve responsiveness to systemic anti-programmed death receptor-1 checkpoint inhibitor therapy in a murine orthotopic pancreatic ductal adenocarcinoma model. METHODS: Murine pancreatic ductal adenocarcinoma (KPC4580P) tumors were implanted into the pancreatic tails of C57BL/6J mice and treated 8 days after implantation. Mice were assigned to one of the following treatment groups: Pancreatic Retrograde Venous Infusion delivery of saline, Pancreatic Retrograde Venous Infusion delivery of toll-like receptor 9 agonist, systemic anti-programmed death receptor-1, systemic toll-like receptor 9 agonist, or the combination of Pancreatic Retrograde Venous Infusion delivery of toll-like receptor 9 agonist and systemic anti-programmed death receptor-1 (Combo). Fluorescently labeled toll-like receptor 9 agonist (radiant efficiency) was used to measure uptake of the drug on day 1. Changes in tumor burden were evaluated by necropsy at 2 different time points, 7 and 10 days after toll-like receptor 9 agonist treatment. Blood and tumors were collected at necropsy 10 days after toll-like receptor 9 agonist treatment for flow cytometric analysis of tumor-infiltrating leukocytes and plasma cytokines. RESULTS: All mice analyzed survived to necropsy. Site of tumor fluorescence measurements revealed 3-fold higher intensity fluorescence in Pancreatic Retrograde Venous Infusion delivery of toll-like receptor 9 agonist compared to systemic toll-like receptor 9 agonist mice. Tumor weights were significantly lower in the Combo group compared to Pancreatic Retrograde Venous Infusion delivery of saline. Flow cytometry of the Combo group demonstrated significantly increased overall T-cell number, specifically CD4+ T-cells, and a trend toward increased CD8+ T-cells. Cytokine analysis showed significantly decreased IL-6 and CXCL1. CONCLUSION: Pressure-Enabled Drug Delivery of toll-like receptor 9 agonist by Pancreatic Retrograde Venous Infusion with systemic anti-programmed death receptor-1 demonstrated improved pancreatic ductal adenocarcinoma tumor control in a murine pancreatic ductal adenocarcinoma model. These results support study of this combination therapy in pancreatic ductal adenocarcinoma patients and expansion of ongoing Pressure-Enabled Drug Delivery clinical trials.