Targeted chemotherapy via HER2-based chimeric antigen receptor (CAR) engineered T-cell membrane coated polymeric nanoparticles.

Cell membrane-derived nanoparticles (NPs) have recently gained popularity due to their desirable features in drug delivery such as mimicking properties of native cells, impeding systemic clearance, and altering foreign body responses. Besides NP technology, adoptive immunotherapy has emerged due to its promise in cancer specificity and therapeutic efficacy. In this research, we developed a biomimetic drug carrier based on chimeric antigen receptor (CAR) transduced T-cell membranes. For that purpose, anti-HER2 CAR-T cells were engineered via lentiviral transduction of anti-HER2 CAR coding lentiviral plasmids. Anti-HER2 CAR-T cells were characterized by their specific activities against the HER2 antigen and used for cell membrane extraction. Anti-cancer drug Cisplatin-loaded poly (D, l-lactide-co-glycolic acid) (PLGA) NPs were coated with anti-human epidermal growth factor receptor 2 (HER2)-specific CAR engineered T-cell membranes. Anti-HER2 CAR-T-cell membrane-coated PLGA NPs (CAR-T-MNPs) were characterized and confirmed via fluorescent microscopy and flow cytometry. Membrane-coated NPs showed a sustained drug release over the course of 21 days in physiological conditions. Cisplatin-loaded CAR-T-MNPs also inhibited the growth of multiple HER2+ cancer cells in vitro. In addition, in vitro uptake studies revealed that CAR-T-MNPs showed an increased uptake by A549 cells. These results were also confirmed via in vivo biodistribution and therapeutic studies using a subcutaneous lung cancer model in nude mice. CAR-T-MNPs localized preferentially at tumor areas compared to those of other studied groups and consisted of a significant reduction in tumor growth in tumor-bearing mice. In Conclusion, the new CAR modified cell membrane-coated NP drug-delivery platform has demonstrated its efficacy both in vitro and in vivo. Therefore, CAR engineered membrane-coated NP system could be a promising cell-mimicking drug carrier that could improve therapeutic outcomes of lung cancer treatments.

Radiation-Induced Innate Neutrophil Response in Tumor Is Mediated by the CXCLs/CXCR2 Axis.

The early events that lead to the inflammatory and immune-modulatory effects of radiation therapy (RT) in the tumor microenvironment (TME) after its DNA damage response activating the innate DNA-sensing pathways are largely unknown. Neutrophilic infiltration into the TME in response to RT is an early innate inflammatory response that occurs within 24-48 h. Using two different syngeneic murine tumor models (RM-9 and MC-38), we demonstrated that CXCR2 blockade significantly reduced RT-induced neutrophilic infiltration. CXCR2 blockade showed the same effects on RT-induced tumor inhibition and host survival as direct neutrophil depletion. Neutrophils highly and preferentially expressed CXCR2 compared to other immune cells. Importantly, RT induced both gene and protein expression of CXCLs in the TME within 24 h, attracting neutrophils into the tumor. Expectedly, RT also upregulated the gene expression of both cGAS and AIM2 DNA-sensing pathways in cGAS-positive MC-38 tumors but not in cGAS-negative RM-9 tumors. Activation of these pathways resulted in increased IL-1ß, which is known to activate the CXCLs/CXCR2 axis. Gene ontology analysis of mRNA-Seq supported these findings. Taken together, the findings suggest that the CXCLs/CXCR2 axis mediates the RT-induced innate inflammatory response in the TME, likely translating the effects of innate DNA-sensing pathways that are activated in response to RT-induced DNA damage.

Systemic immune parameters after prior radiation therapy in patients receiving immune checkpoint inhibitors.

INTRODUCTION: Preclinical studies have demonstrated the ability of radiation therapy (RT) to augment immune response and tumor control by immune checkpoint inhibitors (ICI). However, numerous clinical trials combining RT and ICI have yielded relatively disappointing results. To improve understanding of optimal use of these therapies, we assessed systemic immune effects of prior RT in patients receiving ICI. METHODS AND MATERIALS: Pre- and post-ICI blood samples were collected from patients enrolled in a prospective immunotherapy biospecimen protocol. Mutiplex panels of 40 cytokines and 120 autoantibodies (Ab) were analyzed. We identified differences in these parameters according to receipt, timing, and type of prior RT. We calculated P values using the Pearson product-moment correlation coefficient and false discovery rate (FDR) using the Benjamini-Hochberg Procedure. RESULTS: Among 277 total patients, 69 (25%) received RT in the 6 months prior to ICI initiation. Among RT-treated patients, 23 (33%) received stereotactic RT, and 33 (48%) received curative intent RT. There was no significant difference in demographics or type of immunotherapy between patients according to prior RT exposure. Baseline complement C8 Ab and MIP-1d/CCL15 were significantly higher among patients with prior RT. For MIP-1d/CCL15, only prior stereotactic RT was associated with significant differences. CONCLUSIONS: Prior RT is associated with few changes in systemic immune parameters in patients receiving ICI. The underlying mechanisms and optimal approach to harnessing the potential synergy of RT and ICI require further prospective clinical investigation.

Phase 2 Trial of Stereotactic Ablative Radiotherapy for Patients with Primary Renal Cancer.

BACKGROUND: Most renal cell carcinomas (RCCs) are localized and managed by active surveillance, surgery, or minimally invasive techniques. Stereotactic ablative radiation (SAbR) may provide an innovative non-invasive alternative although prospective data are limited. OBJECTIVE: To investigate whether SAbR is effective in the management of primary RCCs. DESIGN, SETTING, AND PARTICIPANTS: Patients with biopsy-confirmed radiographically enlarging primary RCC (≤5 cm) were enrolled. SAbR was delivered in either three (12 Gy) or five (8 Gy) fractions. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: The primary endpoint was local control (LC) defined as a reduction in tumor growth rate (compared with a benchmark of 4 mm/yr on active surveillance) and pathologic evidence of tumor response at 1 yr. Secondary endpoints included LC by the Response Evaluation Criteria in Solid Tumors (RECIST 1.1), safety, and preservation of kidney function. Exploratory tumor cell-enriched spatial protein and gene expression analysis were conducted on pre- and post-treatment biopsy samples. RESULTS AND LIMITATIONS: Target accrual was reached with the enrollment of 16 ethnically diverse patients. Radiographic LC at 1 yr was observed in 94% of patients (15/16; 95% confidence interval: 70, 100), and this was accompanied by pathologic evidence of tumor response (hyalinization, necrosis, and reduced tumor cellularity) in all patients. By RECIST, 100% of the sites remained without progression at 1 yr. The median pretreatment growth rate was 0.8 cm/yr (interquartile range [IQR]: 0.3, 1.4), and the median post-treatment growth rate was 0.0 cm/yr (IQR: -0.4, 0.1, p < 0.002). Tumor cell viability decreased from 4.6% to 0.7% at 1 yr (p = 0.004). With a median follow-up of 36 mo for censored patients, the disease control rate was 94%. SAbR was well tolerated with no grade ≥2 (acute or late) toxicities. The average glomerular filtration rate declined from a baseline of 65.6 to 55.4 ml/min at 1 yr (p = 0.003). Spatial protein and gene expression analyses were consistent with the induction of cellular senescence by radiation. CONCLUSIONS: This clinical trial adds to the growing body of evidence suggesting that SAbR is effective for primary RCC supporting its evaluation in comparative phase 3 clinical trials. PATIENT SUMMARY: In this clinical trial, we investigated a noninvasive treatment option of stereotactic radiation therapy for the treatment of primary kidney cancer and found that it was safe and effective.

Phase II Trial of Sipuleucel-T and Stereotactic Ablative Body Radiation for Patients with Metastatic Castrate-Resistant Prostate Cancer.

(1) We hypothesized that adding concurrent stereotactic ablative radiotherapy (SAbR) would increase the time to progression in patients with metastatic castrate-resistant prostate cancer (mCRPCA) treated with sipuleucel-T. (2) Patients with a history of prostate cancer (PC), radiographic evidence of metastatic disease, and rising prostate-specific antigen (PSA) > 0.2 ng/dL on castrate testosterone levels were enrolled in this single-arm phase II clinical trial and treated with sipuleucel-T and SAbR. The primary endpoint was time to progression (TTP). Cellular and humoral responses were measured using ELISpot and Luminex multiplex assays, respectively. (3) Twenty patients with mCRPC were enrolled and treated with SAbR to 1−3 sites. Treatment was well tolerated with 51, 8, and 4 treatment-related grade 1, 2, and 3 toxicities, respectively, and no grade 4 or 5 adverse events. At a median follow-up of 15.5 months, the median TTP was 11.2 weeks (95% CI; 6.8−14.0 weeks). Median OS was 76.8 weeks (95% CI; 41.6−130.8 weeks). This regimen induced both humoral and cellular immune responses. Baseline M-MDSC levels were elevated in mCRPC patients compared to healthy donors (p = 0.004) and a decline in M-MDSC was associated with biochemical response (p = 0.044). Responders had lower baseline uric acid levels (p = 0.05). No clear correlation with radiographic response was observed. (4) While the regimen was safe, the PC-antigen-specific immune response induced by SAbR did not yield a synergistic clinical benefit for patients treated with sipuleucel-T compared to the historically reported outcomes.

Outcome and Immune Correlates of a Phase II Trial of High-Dose Interleukin-2 and Stereotactic Ablative Radiotherapy for Metastatic Renal Cell Carcinoma.

PURPOSE: This phase II clinical trial evaluated whether the addition of stereotactic ablative radiotherapy (SAbR), which may promote tumor antigen presentation, improves the overall response rate (ORR) to high-dose IL2 (HD IL2) in metastatic renal cell carcinoma (mRCC). PATIENTS AND METHODS: Patients with pathologic evidence of clear cell renal cell carcinoma (RCC) and radiographic evidence of metastasis were enrolled in this single-arm trial and were treated with SAbR, followed by HD IL2. ORR was assessed based on nonirradiated metastases. Secondary endpoints included overall survival (OS), progression-free survival (PFS), toxicity, and treatment-related tumor-specific immune response. Correlative studies involved whole-exome and transcriptome sequencing, T-cell receptor sequencing, cytokine analysis, and mass cytometry on patient samples. RESULTS: Thirty ethnically diverse mRCC patients were enrolled. A median of two metastases were treated with SAbR. Among 25 patients evaluable by RECIST v1.1, ORR was 16% with 8% complete responses. Median OS was 37 months. Treatment-related adverse events (AE) included 22 grade ≥3 events that were not dissimilar from HD IL2 alone. There were no grade 5 AEs. A correlation was observed between SAbR to lung metastases and improved PFS (P = 0.0165). Clinical benefit correlated with frameshift mutational load, mast cell tumor infiltration, decreased circulating tumor-associated T-cell clones, and T-cell clonal expansion. Higher regulatory/CD8+ T-cell ratios at baseline in the tumor and periphery correlated with no clinical benefit. CONCLUSIONS: Adding SAbR did not improve the response rate to HD IL2 in patients with mRCC in this study. Tissue analyses suggest a possible correlation between frameshift mutation load as well as tumor immune infiltrates and clinical outcomes.

Renal Lipid Metabolism Abnormalities in Obesity and Clear Cell Renal Cell Carcinoma.

Clear cell renal cell carcinoma is the most common and deadly type of cancer affecting the kidney, and is characterized histologically by large intracellular lipid deposits. These deposits are thought to result from lipid metabolic reprogramming occurring in tumor cells, but the exact mechanisms and implications of these metabolic alterations are incompletely understood. Obesity is an independent risk factor for clear cell renal cell carcinoma, and is also associated with lipid accumulation in noncancerous epithelial cells of the proximal tubule, where clear cell renal cell carcinoma originates. This article explores the potential link between obesity-associated renal lipid metabolic disturbances and lipid metabolic reprogramming in clear cell renal cell carcinoma, and discusses potential implications for future research.

Transcriptomic analysis links hepatocellular carcinoma (HCC) in HZE ion irradiated mice to a human HCC subtype with favorable outcomes.

High-charge, high-energy ion particle (HZE) radiations are extraterrestrial in origin and characterized by high linear energy transfer (high-LET), which causes more severe cell damage than low-LET radiations like γ-rays or photons. High-LET radiation poses potential cancer risks for astronauts on deep space missions, but the studies of its carcinogenic effects have relied heavily on animal models. It remains uncertain whether such data are applicable to human disease. Here, we used genomics approaches to directly compare high-LET radiation-induced, low-LET radiation-induced and spontaneous hepatocellular carcinoma (HCC) in mice with a human HCC cohort from The Cancer Genome Atlas (TCGA). We identified common molecular pathways between mouse and human HCC and discovered a subset of orthologous genes (mR-HCC) that associated high-LET radiation-induced mouse HCC with a subgroup (mrHCC2) of the TCGA cohort. The mrHCC2 TCGA cohort was more enriched with tumor-suppressing immune cells and showed a better prognostic outcome than other patient subgroups.

Personalized Ultrafractionated Stereotactic Adaptive Radiotherapy (PULSAR) in Preclinical Models Enhances Single-Agent Immune Checkpoint Blockade.

PURPOSE: Harnessing the immune-stimulatory effects of radiation by combining it with immunotherapy is a promising new treatment strategy. However, more studies characterizing immunotherapy and radiation dose scheduling for the optimal therapeutic effect is essential for designing clinical trials. METHODS AND MATERIALS: A new ablative radiation dosing scheme, personalized ultrafractionated stereotactic adaptive radiation therapy (PULSAR), was tested in combination with α-PD-L1 therapy in immune-activated and resistant syngeneic immunocompetent mouse models of cancer. Specifically, tumor growth curves comparing immunotherapy and radiation therapy dose sequencing were evaluated in immunologically cold and hot tumor models. The response relative to cytotoxic killer T cells was evaluated using an α-CD8 depleting antibody, and immunologic memory was tested by tumor rechallenge of cured mice. RESULTS: We report that both radiation and immunotherapy sequencing, as well as radiation therapy fraction spacing, affect the combination treatment response. Better tumor control was achieved by giving α-PD-L1 therapy during or after radiation, and spacing fractions 10 days apart (PULSAR) achieved better tumor control than traditional daily fractions. We showed that CD8+ depleting antibody abrogated tumor control in the PULSAR combination treatment, and certain treatment schedules induced immunologic memory. CONCLUSIONS: These results illustrate that radiation therapy dosing and scheduling affect tumor control, in combination with checkpoint blockade therapies. PULSAR-style radiation dosing is more complementary in combination with single-agent immunotherapy than traditional daily fractions in this preclinical model. Preclinical investigation could prove helpful in designing clinical trials investigating combination therapy.

Neoadjuvant SABR for Renal Cell Carcinoma Inferior Vena Cava Tumor Thrombus-Safety Lead-in Results of a Phase 2 Trial.

PURPOSE: To evaluate the feasibility, safety, oncologic outcomes, and immune effect of neoadjuvant stereotactic radiation (Neo-SAbR) followed by radical nephrectomy and thrombectomy (RN-IVCT). METHODS AND MATERIALS: These are results from the safety lead-in portion of a single-arm phase 1 and 2 trial. Patients with kidney cancer (renal cell carcinoma [RCC]) and inferior vena cava (IVC) tumor thrombus (TT) underwent Neo-SAbR (40 Gy in 5 fractions) to the IVC-TT followed by open RN-IVCT. Absence of grade 4 to 5 adverse events (AEs) within 90 days of RN-IVCT was the primary endpoint. Exploratory studies included pathologic and immunologic alterations attributable to SAbR. RESULTS: Six patients were included in the final analysis. No grade 4 to 5 AEs were observed. A total of 81 AEs were reported within 90 days of surgery: 73% (59/81) were grade 1, 23% (19/81) were grade 2, and 4% (3/81) were grade 3. After a median follow-up of 24 months, all patients are alive. One patient developed de novo metastatic disease. Of 3 patients with metastasis at diagnosis, 1 had a complete and another had a partial abscopal response without the concurrent use of systemic therapy. Neo-SABR led to decreased Ki-67 and increased PD-L1 expression in the IVC-TT. Inflammatory cytokines and autoantibody titers reflecting better host immune status were observed in patients with nonprogressive disease. CONCLUSIONS: Neo-SAbR followed by RN-IVCT for RCC IVC-TT is feasible and safe. Favorable host immune environment correlated with abscopal response to SABR and RCC relapse-free survival, though direct causal relation to SABR has yet to be established.

Renal lipid accumulation, oxidative stress and uric acid handling in a rodent model of obesity and metabolic syndrome.

Hyperuricemia is more prevalent among people with obesity and metabolic syndrome, and is associated with adverse clinical outcomes. We hypothesized that increased renal reabsorption of uric acid (UA) in obesity and metabolic syndrome may be an adaptive response of the kidney when faced with fatty acid-induced oxidative stress. To test this hypothesis, we examined lipid accumulation, markers of oxidative stress, and renal UA handling in Zucker diabetic fatty (ZDF) rats, and in matched lean control animals. Rats were randomized to either normal rodent chow or a diet supplemented with antioxidants (α-tocopheryl acetate, sodium selenite, zinc sulfate, and ascorbic acid), and were followed up for either 4 or 20 weeks after randomization. Dietary antioxidant supplementation had no significant effects in lean control rats but led to partial improvement in markers of elevated oxidative stress in the kidney of ZDF rats. Renal UA handling was not affected by antioxidant supplementation. We observed robust correlations between renal lipid content and oxidative stress markers in the pooled experimental groups, particularly in older animals after 20 weeks on the study diets. Dietary antioxidant supplementation did not prevent the gradual decline in renal function observed in older ZDF rats. These findings suggest that hyperuricemia in the ZDF rat model of obesity and the metabolic syndrome is not caused by renal oxidative stress, that there may be a pathophysiological link between lipid accumulation and oxidative stress in the kidney, and that antioxidant supplementation does not prevent age-related decline in renal function in ZDF rats.

Type I Interferon Response in Radiation-Induced Anti-Tumor Immunity.

The anti-tumor activity of interferons (IFNs) was first appreciated about half a century ago, and IFN-α2 was the first cancer immunotherapy approved by the US Food and Drug Administration. Radiation therapy (RT), one of the pillars of cancer treatment, directly causes DNA damage, which can lead to senescence and cell death in tumor cells. In recent years, however, RT-induced immunomodulatory effects have been recognized to play an indispensable role in achieving the optimum therapeutic effect of RT. Increasing evidence indicates that RT enhances adaptive anti-tumor immunity by augmenting the innate immune sensing of tumors in a type I IFN-dependent matter. This review briefly introduces the role of type I interferon in cancer and the available evidence on the overall effects of RT on tumor immunity mediated via type I IFN. Recent advances in deciphering the molecular mechanisms underlying the induction of type I IFNs triggered by RT, their clinical implications, and therapeutic opportunities will be highlighted.

A randomized trial comparing the efficacy and safety of treating patients with type 2 diabetes and highly elevated HbA1c levels with basal-bolus insulin or a glucagon-like peptide-1 receptor agonist plus basal insulin: The SIMPLE study.

AIM: To compare the efficacy and safety of a glucagon-like peptide-1 receptor agonist (GLP1RA) plus basal insulin versus basal-bolus insulin treatment in patients with very uncontrolled type 2 diabetes. MATERIALS AND METHODS: The SIMPLE study was a 6-month pragmatic, randomized, open-label trial testing the effectiveness of two approaches to treat patients with type 2 diabetes and HbA1c ≥10%. We randomized patients to detemir plus liraglutide or detemir plus aspart (before each meal). The primary endpoint was change in HbA1c; changes in body weight, insulin dose, hypoglycaemia and diabetes-related quality-of-life were secondary outcomes. RESULTS: We randomized 120 participants aged 47.4 ± 9.5 years, Hispanic 40%, African American 42%, diabetes duration 10 [25th-75th percentile (6 to 15)] years, body mass index 37.2 ± 10.3 kg/m2 . HbA1c decreased more with GLP1RA plus basal insulin [12.2% (95% CI 11.8% to 12.6%) to 8.1% (95% CI 7.4% to 8.7%)] compared with basal-bolus insulin [11.8% (95% CI 11.5% to 12.2%) to 8.8% (95% CI 88.1% to 9.55%)]; estimated treatment difference (ETD) of -1.1% (95% CI -2.0% to -0.1%) (non-inferiority margin 0.4% and P = .0001, superiority P = .026). Compared with basal-bolus insulin, treatment with GLP1RA plus basal insulin led to a body weight ETD of -3.7 kg (95% CI -5.8 to -1.5; P = .001), fewer patients experiencing hypoglycaemia [66.1% vs 35.2% (P = .002)], and greater improvements in general/current health perception, treatment satisfaction, and fear of hypoglycaemia, while taking a lower total daily dose of insulin [estimated treatment ratio 0.68 (95% CI 0.55 to 0.84)]. CONCLUSIONS: In patients with HbA1c ≥10% treatment with GLP1RA plus basal insulin, compared with basal-bolus insulin, resulted in better glycaemic control and body weight, lower insulin dosage and hypoglycaemia, and improved quality of life. This treatment strategy is an effective and safe alternative to a basal-bolus insulin regimen.

Roux-en-Y gastric bypass compared with equivalent diet restriction: Mechanistic insights into diabetes remission.

AIMS: To investigate the physiological mechanisms leading to rapid improvement in diabetes after Roux-en-Y gastric bypass (RYGB) and specifically the contribution of the concurrent peri-operative dietary restrictions, which may also alter glucose metabolism. MATERIALS AND METHODS: In order to assess the differential contributions of diet and surgery to the mechanisms leading to the rapid improvement in diabetes after RYGB we enrolled 10 patients with type 2 diabetes scheduled to undergo RYGB. All patients underwent a 10-day inpatient supervised dietary intervention equivalent to the peri-operative diet (diet-only period), followed by, after a re-equilibration (washout) period, an identical period of pair-matched diet in conjunction with RYGB (diet and RYGB period). We conducted extensive metabolic assessments during a 6-hour mixed-meal challenge test, with stable isotope glucose tracer infusion performed before and after each intervention. RESULTS: Similar improvements in glucose levels, ß-cell function, insulin sensitivity and post-meal hepatic insulin resistance were observed with both interventions. Both interventions led to significant reductions in fasting and postprandial acyl ghrelin. The diet-only intervention induced greater improvements in basal hepatic glucose output and post-meal gastric inhibitory polypeptide (GIP) secretion. The diet and RYGB intervention induced significantly greater increases in post-meal glucagon-like peptide-1 (GLP-1), peptide YY (PYY) and glucagon levels. CONCLUSIONS: Strict peri-operative dietary restriction is a main contributor to the rapid improvement in glucose metabolism after RYGB. The RYGB-induced changes in the incretin hormones GLP-1 and PYY probably play a major role in long-term compliance with such major dietary restrictions through central and peripheral mechanisms.

The Infamous, Famous Sulfonylureas and Cardiovascular Safety: Much Ado About Nothing?

PURPOSE OF REVIEW: Sulfonylureas (SUs) are one of the most commonly used glucose-lowering agents worldwide. While their efficacy is undisputed, their cardiovascular safety has been debated since the 1970's. RECENT FINDINGS: With no dedicated cardiovascular studies to definitively answer this question, observational studies and meta-analyses abound and have reported divergent results, fueling the controversy. Studies that compared SUs to metformin or newer agents, like GLP-1 agonists and SGLT2 inhibitors, suggest a difference in cardiovascular events, yet this is likely the result of beneficial effects of the latter. Studies comparing SUs to other agents have been reassuring. SUs remain a common choice of treatment for patients with type 2 diabetes due to their exceptional value. They are effective at lowering glucose and thus contributing to the prevention of microvascular complications. Weight gain and hypoglycemia are their main side effects, although less severe when compared to insulin treatment. Their cardiovascular safety will remain a controversial topic due to lack of conclusive data, but there is no definitive evidence of harm with the second-generation agents.

The Role of Regulatory B Cell-Like Malignant Cells and Treg Cells in the Mouse Model of BCL1 Tumor Dormancy.

Cancer dormancy is a clinical state in which residual tumor cells persist for long periods of time but do not cause detectable disease. In the mouse B cell lymphoma model (BCL1), dormancy can be induced and maintained by immunizing mice with a soluble form of the IgM expressed on the surface of the tumor cells. Immunization induces an anti-idiotype antibody response that maintains dormancy. Mice with dormant tumor have low numbers of BCL1 cells in their spleens that divide and are killed at the same rate. When the anti-Id antibodies wane, the tumor cells grow rapidly and kill the host. Spleens from tumor-bearing mice contain both effector (CD4+ and CD8+) and regulatory T cells (Tregs). In other tumor models, it has been reported that Tregs promote tumor progression by preventing effector cells from killing the tumor. In this report, we demonstrate that the tumor site with rapidly dividing BCL1 cells has fewer Tregs than the tumor site harboring dormant BCL1 cells. In both cases, the Tregs were equally suppressive in vitro. In spleens from mice with actively growing tumor, CD8+ but not CD4+ T cells were virtually absent. In vitro analysis demonstrated a tumor-mediated elimination of CD8+ T cells that was contact dependent and involved the caspase-3 pathway. Most importantly, we found that the BCL1 cells expressed characteristics of B10 regulatory B cells, i.e., they were CD1dhiCD5+ and secreted high levels of IL-10. These BCL1 tumor cells can inhibit anti-tumor immune responses by depleting CD8+ effector T cells.

Key role for neutrophils in radiation-induced antitumor immune responses: Potentiation with G-CSF.

Radiation therapy (RT), a major modality for treating localized tumors, can induce tumor regression outside the radiation field through an abscopal effect that is thought to involve the immune system. Our studies were designed to understand the early immunological effects of RT in the tumor microenvironment using several syngeneic mouse tumor models. We observed that RT induced sterile inflammation with a rapid and transient infiltration of CD11b+Gr-1high+ neutrophils into the tumors. RT-recruited tumor-associated neutrophils (RT-Ns) exhibited an increased production of reactive oxygen species and induced apoptosis of tumor cells. Tumor infiltration of RT-Ns resulted in sterile inflammation and, eventually, the activation of tumor-specific cytotoxic T cells, their recruitment into the tumor site, and tumor regression. Finally, the concurrent administration of granulocyte colony-stimulating factor (G-CSF) enhanced RT-mediated antitumor activity by activating RT-Ns. Our results suggest that the combination of RT and G-CSF should be further evaluated in preclinical and clinical settings.

The Immunogenicity of Peptoid-Protein Conjugates.

We demonstrate that a peptoid composed of five monomers and attached via a maleimide linker to a carrier protein elicits anti-peptoid, anti-linker and anti-carrier antibodies in rabbits. Specific anti-peptoid antibodies were affinity purified and used to reproducibly retrieve three specific peptoid-coupled beads from 20,000 irrelevant peptoid-beads using magnetic screening.

Mechanisms of Action of Liraglutide in Patients With Type 2 Diabetes Treated With High-Dose Insulin.

CONTEXT: The mechanisms of action of incretin mimetics in patients with long-standing type 2 diabetes (T2D) and high insulin requirements have not been studied. OBJECTIVE: To evaluate changes in ß-cell function, glucagon secretion, and fat distribution after addition of liraglutide to high-dose insulin. DESIGN: A single-center, randomized, double-blind, placebo-controlled trial. SETTING: University of Texas Southwestern and Parkland Memorial Hospital clinics. PATIENTS: Seventy-one patients with long-standing (median, 17 years) T2D requiring high-dose insulin treatment (>1.5 U/kg/d; average, 2.2 ± 0.9 U/kg/d). INTERVENTION: Patients were randomized to liraglutide 1.8 mg/d or matching placebo for 6 months. MAIN OUTCOME MEASURES: We measured changes in insulin and glucagon secretion using a 4-hour mixed-meal challenge test. Magnetic resonance-based techniques were used to estimate sc and visceral fat in the abdomen and ectopic fat in the liver and pancreas. RESULTS: Glycosylated hemoglobin improved significantly with liraglutide treatment, with an end-of-trial estimated treatment difference between groups of −0.9% (95% confidence interval, −1.5, −0.4%) (P = .002). Insulin secretion improved in the liraglutide group vs placebo, as measured by the area under the curve of C-peptide (P = .002) and the area under the curves ratio of C-peptide to glucose (P = .003). Insulin sensitivity (Matsuda index) and glucagon secretion did not change significantly between groups. Liver fat and sc fat decreased in the liraglutide group vs placebo (P = .0006 and P = .01, respectively), whereas neither visceral nor pancreatic fat changed significantly. CONCLUSIONS: Treatment with liraglutide significantly improved insulin secretion, even in patients with long-standing T2D requiring high-dose insulin treatment. Liraglutide also decreased liver and sc fat, but it did not alter glucagon secretion.

Thermostable ricin vaccine protects rhesus macaques against aerosolized ricin: Epitope-specific neutralizing antibodies correlate with protection.

Ricin toxin (RT) is the second most lethal toxin known; it has been designated by the CDC as a select agent. RT is made by the castor bean plant; an estimated 50,000 tons of RT are produced annually as a by-product of castor oil. RT has two subunits, a ribotoxic A chain (RTA) and galactose-binding B chain (RTB). RT binds to all mammalian cells and once internalized, a single RTA catalytically inactivates all of the ribosomes in a cell. Administered as an aerosol, RT causes rapid lung damage and fibrosis followed by death. There are no Food and Drug Administration-approved vaccines and treatments are only effective in the first few hours after exposure. We have developed a recombinant RTA vaccine that has two mutations V76M/Y80A (RiVax). The protein is expressed in Escherichia coli and is nontoxic and immunogenic in mice, rabbits, and humans. When vaccinated mice are challenged with injected, aerosolized, or orally administered (gavaged) RT, they are completely protected. We have now developed a thermostable, aluminum-adjuvant-containing formulation of RiVax and tested it in rhesus macaques. After three injections, the animals developed antibodies that completely protected them from a lethal dose of aerosolized RT. These antibodies neutralized RT and competed to varying degrees with a panel of neutralizing and nonneutralizing mouse monoclonal antibodies known to recognize specific epitopes on native RTA. The resulting antibody competition profile could represent an immunologic signature of protection. Importantly, the same signature was observed using sera from RiVax-immunized humans.