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Repeated administration of peroxisome proliferator-activated receptor gamma (PPARγ) agonists reduces neuropathic pain-like behavior and associated changes in glial activation in the spinal cord dorsal horn. As PPARγ is a nuclear receptor, sustained changes in gene expression are widely believed to be the mechanism of pain reduction. However, we recently reported that a single intrathecal (i.t.) injection of pioglitazone, a PPARγ agonist, reduced hyperalgesia within 30 minutes, a time frame that is typically less than that required for genomic mechanisms. To determine the very rapid antihyperalgesic actions of PPARγ activation, we administered pioglitazone to rats with spared nerve injury and evaluated hyperalgesia. Pioglitazone inhibited hyperalgesia within 5 minutes of injection, consistent with a nongenomic mechanism. Systemic or i.t. administration of GW9662, a PPARγ antagonist, inhibited the antihyperalgesic actions of intraperitoneal or i.t. pioglitazone, suggesting a spinal PPARγ-dependent mechanism. To further address the contribution of nongenomic mechanisms, we blocked new protein synthesis in the spinal cord with anisomycin. When coadministered intrathecally, anisomycin did not change pioglitazone antihyperalgesia at an early 7.5-minute time point, further supporting a rapid nongenomic mechanism. At later time points, anisomycin reduced pioglitazone antihyperalgesia, suggesting delayed recruitment of genomic mechanisms. Pioglitazone reduction of spared nerve injury-induced increases in GFAP expression occurred more rapidly than expected, within 60 minutes. We are the first to show that activation of spinal PPARγ rapidly reduces neuropathic pain independent of canonical genomic activity. We conclude that acute pioglitazone inhibits neuropathic pain in part by reducing astrocyte activation and through both genomic and nongenomic PPARγ mechanisms. [ABSTRACT FROM AUTHOR]

Radiation has long been the standard of care for many types of cancer. It is employed to locally eradicate tumor cells as well as alter tumor stroma with either curative or palliative intent. Radiation-induced cell damage is an immunologically active process in which danger signals are released that stimulate immune cells to phagocytose and present locally released tumor-associated antigens (TAAs). Recent studies have indicated that radiotherapy can also alter the phenotype of cancer cells that remain after treatment. These cells upregulate TAAs as well as markers, including major histocompatibility complex and costimulatory molecules, that make them much more immunostimulatory. As our understanding of the immunomodulatory effects of radiation has improved, interest in combining this type of therapy with immune-based therapies for the treatment of cancer has grown. Therapeutic cancer vaccines have been shown to initiate the dynamic process of host immune system activation, culminating in the recognition of host cancer cells as foreign. The environment created after radiotherapy can be exploited by active therapeutic cancer vaccines in order to achieve further, more robust immune system activation. This review highlights preclinical studies that have examined the alteration of the tumor microenvironment with regard to immunostimulatory molecules following different types of radiotherapy, including external beam radiation, radiolabeled monoclonal antibodies, bone-seeking radionuclides, and brachytherapy. We also emphasize how combination therapy with a cancer vaccine can exploit these changes to achieve improved therapeutic benefit. Lastly, we describe how these laboratory findings are translating into clinical benefit for patients undergoing combined radiotherapy and cancer vaccination. [ABSTRACT FROM AUTHOR]

Abstract: Objective: The opioid growth factor (OGF) and its receptor, OGFr, serve as a tonically active inhibitory axis regulating cell proliferation in normal cells and a variety of cancers, including human ovarian cancer. Blockade of OGF and OGFr with the nonselective opioid receptor antagonist naltrexone (NTX) upregulates expression of OGF and OGFr. Administration of a low dosage of NTX (LDN) blocks endogenous opioids from opioid receptors for a short period of time (4–6h) each day, providing a window of 18–20h for the upregulated opioids and receptors to interact. The present study investigated the repercussions of upregulating the OGF–OGFr axis by treatment with OGF or LDN on human ovarian tumorigenesis in vivo. Methods: Female nude mice were transplanted intraperitoneally with SKOV-3 human ovarian cancer cells and treated on a daily basis with OGF (10mg/kg), LDN (0.1mg/kg), or an equivalent volume of vehicle (saline). Tumor burden, as well as DNA synthesis, apoptosis, and angiogenesis was assessed in tumor tissue following 40days of treatment. Results: OGF and LDN markedly reduced ovarian tumor burden (tumor nodule number and weight). The mechanism of action was targeted to an inhibition of tumor cell proliferation and angiogenesis; no changes in cell survival were noted. Conclusions: This study shows that a native opioid pathway can suppress human ovarian cancer in a xenograft model, and provides novel non-toxic therapies for the treatment of this lethal neoplasia. [Copyright &y& Elsevier]

Abstract: Opioid peptides function as immunomodulatory molecules. Reports have linked the opioid growth factor (OGF), [Met5]-enkephalin, and its receptor OGFr to autoimmune diseases. OGF repressed the incidence and magnitude of myelin oligodendrocyte-induced experimental autoimmune encephalomyelitis in mice. Given the extensive connection between the immune system and autoimmune diseases, the present study was conducted to examine the relationship of the OGF–OGFr axis and T lymphocyte proliferation. Splenic-derived mouse lymphocytes were stimulated with phytohemagglutin (PHA). All non-stimulated and PHA-stimulated T lymphocytes had immunoreactivity for OGF-like enkephalin and OGFr. OGF markedly suppressed T lymphocyte number in a dose-dependent manner. However, PHA-stimulated T lymphocytes were not altered in cell number by a variety of natural and synthetic opioid-related compounds, some specific for μ, δ, and κ opioid receptors. Persistent blockade of opioid receptors with the general opioid antagonist naltrexone (NTX), as well as antibody neutralization of OGF-like peptides, had no effect on cell number. Non-stimulated T lymphocytes exhibited no change in cell number when subjected to OGF or NTX. Treatment of T lymphocytes with siRNAs for μ, δ, or κ opioid receptors did not affect cell number, and the addition of OGF to these siRNA-exposed cultures depressed the population of cells. T lymphocytes treated with OGFr siRNA also had a comparable number of cells to control cultures, but the addition of OGF did not alter cell number. DNA synthesis in PHA-stimulated T lymphocytes exposed to OGF was markedly decreased from PHA-stimulated cultures receiving vehicle, but the number of cells undergoing apoptosis or necrosis in these cultures was similar to control levels. T lymphocytes subjected to siRNA for p16 and/or p21 had a comparable number of cells compared to controls, and treatment with OGF did not depress cell number in preparations transfected with both p16 and p21 siRNA. These data reveal that the OGF–OGFr axis is present in T lymphocytes and is capable of suppressing cell proliferation. However, T lymphocytes are not dependent on the regulation of cell proliferation by this system. The results showing that the OGF–OGFr axis is an immunosuppressant, offers explanation for reports that autoimmune diseases can be modulated by this system. [Copyright &y& Elsevier]

Dramatically up-regulated in the dorsal horn of the mammalian spinal cord following inflammation or nerve injury, neuropeptide Y (NPY) is poised to regulate the transmission of sensory signals. We found that doxycycline-induced conditional in vivo (Npytet/tet) knockdown of NPY produced rapid, reversible, and repeatable increases in the intensity and duration of tactile and thermal hypersensitivity. Remarkably, when allowed to resolve for several weeks, behavioral hypersensitivity could be dramatically reinstated with NPY knockdown or intrathecal administration of Y1 or Y2 receptor antagonists. In addition, Y2 antagonism increased dorsal horn expression of Fos and phosphorylated form of extracellular signal-related kinase. Taken together, these data establish spinal NPY receptor systems as an endogenous braking mechanism that exerts a tonic, long-lasting, broad-spectrum inhibitory control of spinal nociceptive transmission, thus impeding the transition from acute to chronic pain. NPY and its receptors appear to be part of a mechanism. whereby mammals naturally recover from the hyperalgesia associated with inflammation or nerve injury. [ABSTRACT FROM AUTHOR]

Hepatocellular carcinoma (HCC) is the third leading cause of cancer deaths worldwide, with a mortality rate approximating its incidence. Understanding the biology of these tumors, as well as treatment modalities, has been challenging. The opioid growth factor (OGF; [Met5]-enkephalin) and the OGF receptor (OGFr) form an endogenous growth-regulating pathway in homeostasis and neoplasia. In this investigation, we examined the relationship of the OGF-OGFr axis in HCC and define its presence, function, and mechanism. Using SK-HEP-1, Hep G2, and Hep 3B human HCC cell lines, we found that OGF and OGFr were present and functional. Exogenous OGF was observed to have a dosedependent, reversible, and receptor-mediated inhibitory action on cell proliferation. Endogenous OGF was found to be constitutively produced and tonically active on cell replicative activities, with neutralization of this peptide accelerating cell proliferation. Silencing of OGFr using 5iRNA stimulated cell replication, even when exogenous OGF was added to the cultures, documenting its importance in mediating OGF activity. The mechanism of OGF-OGFr action on cell number was related to inhibition of DNA synthesis and not to apoptotic or necrotic pathways. Both OGF and OGFr were detected in surgical specimens of HCC, and no quantitative differences were recorded in peptide or receptor between pathological and normal specimens. These data are the first to report that the OGF-OGFr system is a native biological regulator of cell proliferation in HCC. The findings may provide important insight in designing treatment strategies for this deadly disease. [ABSTRACT FROM AUTHOR]

The opioid growth factor (OGF) regulates cell proliferation of human cancer cells through the cyclin-dependent kinase inhibitory pathway, with mediation of this action by the OGF receptor (OGFr). The ubiquity of the OGF-OGFr axis in human cancer is unknown. We used 31 human cancer cell lines, representative of more than 90% of neoplasias occurring in humans, and found that OGF and OGFr were detected in the cytoplasm and nucleus by immunohistochemistry. The addition of OGF to cultures depressed cell number up to 41%, whereas naltrexone (NTX) increased cell proliferation by up to 44%, a total of 85% in the modulating capacity for the OGF-OGFr axis. Neutralization of OGF by specific antibodies led to a marked increase in cell number. Knockdown of OGFr by OGFr-siRNA resulted in a significant increase in the number of cells, even in the face of the addition of exogenous OGF. The cultures to which NTX was added and subjected to OGFr-siRNA were similar to those with OGF-siRNA alone. The OGF-OGFr axis, a physiological determinant of cell-proliferative activity, is a ubiquitous feature of human cancer cells. The identification of this native biological system in neoplasia may be important in understanding the pathophysiology of neoplasia, and in designing treatment modalities that utilize the body's own chemistry. [ABSTRACT FROM AUTHOR]

Donahue RN, McLaughlin PJ, Zagon IS. Cell proliferation of human ovarian cancer is regulated by the opioid growth factoropioid growth factor receptor axis. Am J Physiol Regul Integr Comp Physiol 296: R1716-R1725, 2009. First published March 18, 2009; doi:10.1152/ajpregu.00075.2009.-Ovarian cancer is the leading cause of death from gynecological malignancies. Understanding the biology of these tumors, as well as treatment modalities, has been challenging. The opioid growth factor (OGF; [Met[sup5]]-enkephalin) and the OGF receptor (OGFr) form an endogenous growth-regulating pathway in homeostasis and neoplasia. In this investigation, we examined the relationship of the OGF-OGFr axis to ovarian cancer, and defined its presence, function, and mechanisms. Using OVCAR-3 and SKOV-3 ovarian cancer cell lines, we found that OGF and OGFr were present and functional. Exogenous OGF was observed to have a dose-dependent, serum-independent, reversible, and receptor-mediated inhibitory action on cell proliferation that was dependent on RNA and protein synthesis. The repressive effect of OGF on cell proliferation also was observed in SW626, CAOV-3, and HEY ovarian cancer cell lines. Endogenous OGF was found to be constitutively produced and tonically active on cell replicative activities, with neutralization of this peptide accelerating cell proliferation. Silencing of OGFr using siRNA technology stimulated cell replication, documenting its integral role. The mechanism of OGF-OGFr action on DNA synthesis was related to the cyclin-dependent kinase inhibitory pathway because knockdown of p16 or p21 in OVCAR-3 cells, and p21 in SKOV-3 cells, eliminated OGF's inhibitory effect on growth. These data are the first to report that the OGF-OGFr system is a native biological regulator of cell proliferation in human ovarian cancer. This information will be important in designing treatment strategies for this deadly disease. [ABSTRACT FROM AUTHOR]

Background: Ischemic heart disease, often caused by an acute myocardial infarction (AMI) is one of the leading causes of morbidity and mortality worldwide. Despite significant advances in medical and procedural therapies, millions of AMI patients progress to develop heart failure every year. Methods: Here, we examine the combination therapy of human mesenchymal stromal cells (MSCs) and endothelial colony-forming cells (ECFCs) to reduce the early ischemic damage (MSCs) and enhance angiogenesis (ECFCs) in a pre-clinical model of acute myocardial infarction. NOD/SCID mice were subjected to AMI followed by transplantation of MSCs and ECFCs either alone or in combination. Cardiomyocyte apoptosis and cardiac functional recovery were assessed in short- and long-term follow-up studies. Results: At 1 day after AMI, MSC- and ECFC-treated animals demonstrated significantly lower cardiomyocyte apoptosis compared to vehicle-treated animals. This phenomenon was associated with a significant reduction in infarct size, cardiac fibrosis, and improvement in functional cardiac recovery 4 weeks after AMI. Conclusions: The use of ECFCs, MSCs, and the combination of both cell types reduce cardiomyocyte apoptosis, scar size, and adverse cardiac remodeling, compared to vehicle, in a pre-clinical model of AMI. These results support the use of this combined cell therapy approach in future human studies during the acute phase of ischemic cardiac injury. [ABSTRACT FROM AUTHOR]

Lessons Learned: Modified vaccinia Ankara‐Bavarian Nordic (MVA‐BN)‐Brachyury followed by fowlpox virus‐BN‐Brachyury was well tolerated upon administration to patients with advanced cancer.Sixty‐three percent of patients developed CD4+ and/or CD8+ T‐cell responses to brachyury after vaccination.BN‐Brachyury vaccine also induced T‐cell responses against CEA and MUC1, which are cascade antigens, that is, antigens not encoded in the vaccines. Background: Brachyury, a transcription factor, plays an integral role in the epithelial–mesenchymal transition, metastasis, and tumor resistance to chemotherapy. It is expressed in many tumor types, and rarely in normal tissues, making it an ideal immunologic target. Bavarian Nordic (BN)‐Brachyury consists of vaccination with modified vaccinia Ankara (MVA) priming followed by fowlpox virus (FPV) boosting, each encoding transgenes for brachyury and costimulatory molecules. Methods: Patients with metastatic solid tumors were treated with two monthly doses of MVA‐brachyury s.c., 8 × 108 infectious units (IU), followed by FPV‐brachyury s.c., 1 × 109 IU, for six monthly doses and then every 3 months for up to 2 years. The primary objective was to determine safety and tolerability. Results: Eleven patients were enrolled from March 2018 to July 2018 (one patient was nonevaluable). No dose‐limiting toxicities were observed. The most common treatment‐related adverse event was grade 1/2 injection‐site reaction observed in all patients. Best overall response was stable disease in six patients, and the 6‐month progression‐free survival rate was 50%. T cells against brachyury and cascade antigens CEA and MUC1 were detected in the majority of patients. Conclusion: BN‐Brachyury vaccine is well tolerated and induces immune responses to brachyury and cascade antigens and demonstrates some evidence of clinical benefit. [ABSTRACT FROM AUTHOR]

Lessons Learned: Concurrent ETBX‐011, ETBX‐051, and ETBX‐061 can be safely administered to patients with advanced cancer.All patients developed CD4+ and/or CD8+ T‐cell responses after vaccination to at least one tumor‐associated antigen (TAA) encoded by the vaccine; 5/6 patients (83%) developed MUC1‐specific T cells, 4/6 (67%) developed CEA‐specific T cells, and 3/6 (50%) developed brachyury‐specific T cells.The presence of adenovirus 5‐neutralizing antibodies did not prevent the generation of TAA‐specific T cells. Background: A novel adenovirus‐based vaccine targeting three human tumor‐associated antigens—CEA, MUC1, and brachyury—has demonstrated antitumor cytolytic T‐cell responses in preclinical animal models of cancer. Methods: This open‐label, phase I trial evaluated concurrent administration of three therapeutic vaccines (ETBX‐011 = CEA, ETBX‐061 = MUC1 and ETBX‐051 = brachyury). All three vaccines used the same modified adenovirus 5 (Ad5) vector backbone and were administered at a single dose level (DL) of 5 × 1011 viral particles (VP) per vector. The vaccine regimen consisting of all three vaccines was given every 3 weeks for three doses then every 8 weeks for up to 1 year. Clinical and immune responses were evaluated. Results: Ten patients enrolled on trial (DL1 = 6 with 4 in the DL1 expansion cohort). All treatment‐related adverse events were temporary, self‐limiting, grade 1/2 and included injection site reactions and flu‐like symptoms. Antigen‐specific T cells to MUC1, CEA, and/or brachyury were generated in all patients. There was no evidence of antigenic competition. The administration of the vaccine regimen produced stable disease as the best clinical response. Conclusion: Concurrent ETBX‐011, ETBX‐051, and ETBX‐061 can be safely administered to patients with advanced cancer. Further studies of the vaccine regimen in combination with other agents, including immune checkpoint blockade, are planned. [ABSTRACT FROM AUTHOR]

Background: With the rapid adoption of immunotherapy for the treatment of cancer comes the pressing need for readily accessible biomarkers to guide immunotherapeutic strategies and offer insights into outcomes with specific treatments. Regular sampling of solid tumour tissues outside of melanoma for immune monitoring is not often feasible; conversely, routine, frequent interrogation of circulating immune biomarkers is entirely possible. As immunotherapies and immune checkpoint inhibitors, in particular, are more widely used in first‐line, neoadjuvant, and metastatic settings, the discovery and validation of peripheral immune biomarkers are urgently needed across solid tumour types for improved prediction and prognostication of clinical outcomes in response to immunotherapy, as well as elucidation of mechanistic underpinnings of the intervention. Careful experimental design, encompassing both retrospective and prospective studies, is required in such biomarker identification studies, and concerted efforts are essential for their advancement into clinical settings. Conclusion: In this review, we summarize shared immune features between the tumour microenvironment and systemic circulation, evaluate exploratory peripheral immune biomarker studies, and discuss associations between candidate biomarkers with clinical outcomes. We also consider integration of multiple peripheral immune parameters for better prediction and prognostication and discuss considerations in study design to further evaluate the clinical utility of candidate peripheral immune biomarkers for immunotherapy of solid tumours. Highlights: Peripheral immune biomarkers are critical for improved prediction and prognostication of clinical outcomes for patients with solid tumours treated with immune checkpoint inhibition.Candidate peripheral biomarkers, such as cytokines, soluble factors, and immune cells, have potential as biomarkers to guide immunotherapy of solid tumours.Multiple peripheral immune parameters may be integrated to improve prediction and prognostication.The potential of peripheral immune biomarkers to guide immunotherapy of solid tumours requires critical work in biomarker discovery, validation, and standardization. [ABSTRACT FROM AUTHOR]

Background: Thymic epithelial tumors are PD-L1–expressing tumors of thymic epithelial origin characterized by varying degrees of lymphocytic infiltration and a predisposition towards development of paraneoplastic autoimmunity. PD-1–targeting antibodies have been evaluated, largely in patients with thymic carcinoma. We sought to evaluate the efficacy and safety of the anti-PD-L1 antibody, avelumab (MSB0010718C), in patients with relapsed, advanced thymic epithelial tumors and conduct correlative immunological studies. Methods: Seven patients with thymoma and one patient with thymic carcinoma were enrolled in a phase I, dose-escalation trial of avelumab (MSB0010718C), and treated with avelumab at doses of 10 mg/kg to 20 mg/kg every 2 weeks until disease progression or development of intolerable side effects. Tissue and blood immunological analyses were conducted. Results: Two of seven (29%) patients with thymoma had a confirmed Response Evaluation Criteria in Solid Tumors–defined partial response, two (29%) had an unconfirmed partial response and three patients (two thymoma; one thymic carcinoma) had stable disease (43%). Three of four responses were observed after a single dose of avelumab. All responders developed immune-related adverse events that resolved with immunosuppressive therapy. Only one of four patients without a clinical response developed immune-related adverse events. Responders had a higher absolute lymphocyte count, lower frequencies of B cells, regulatory T cells, conventional dendritic cells, and natural killer cells prior to therapy. Conclusion: These results demonstrate anti-tumor activity of PD-L1 inhibition in patients with relapsed thymoma accompanied by a high frequency of immune-related adverse events. Pre-treatment immune cell subset populations differ between responders and non-responders. Trial registration: ClinicalTrials.gov - NCT01772004. Date of registration – January 21, 2013. [ABSTRACT FROM AUTHOR]

Background: HuMax-IL8 (now known as BMS-986253) is a novel, fully human monoclonal antibody that inhibits interleukin-8 (IL-8), a chemokine that promotes tumor progression, immune escape, epithelial-mesenchymal transition, and recruitment of myeloid-derived suppressor cells. Studies have demonstrated that high serum IL-8 levels correlate with poor prognosis in many malignant tumors. Preclinical studies have shown that IL-8 blockade may reduce mesenchymal features in tumor cells, making them less resistant to treatment. Methods: Fifteen patients with metastatic or unresectable locally advanced solid tumors were enrolled in this 3 + 3 dose-escalation trial at four dose levels (4, 8, 16, or 32 mg/kg). HuMax-IL8 was given IV every 2 weeks, and patients were followed for safety and immune monitoring at defined intervals up to 52 weeks. Results: All enrolled patients (five chordoma, four colorectal, two prostate, and one each of ovarian, papillary thyroid, chondrosarcoma, and esophageal) received at least one dose of HuMax-IL8. Eight patients had received three or more prior lines of therapy and five patients had received prior immunotherapy. Treatment-related adverse events occurred in five patients (33%), mostly grade 1. Two patients receiving the 32 mg/kg dose had grade 2 fatigue, hypophosphatemia, and hypersomnia. No dose-limiting toxicities were observed, and maximum tolerated dose was not reached. Although no objective tumor responses were observed, 11 patients (73%) had stable disease with median treatment duration of 24 weeks (range, 4–54 weeks). Serum IL-8 was significantly reduced on day 3 of HuMax-IL8 treatment compared to baseline (p = 0.0004), with reductions in IL-8 seen at all dose levels. Conclusions: HuMax-IL8 is safe and well-tolerated. Ongoing studies are evaluating the combination of IL-8 blockade and other immunotherapies. Trial registration: NCTN, NCT02536469. Registered 23 August 2015, https://clinicaltrials.gov/ct2/show/NCT02536469?term=NCT02536469&rank=1. [ABSTRACT FROM AUTHOR]

Background: Checkpoint inhibitors have not been effective for prostate cancer as single agents. Durvalumab is a human IgG1-K monoclonal antibody that targets programmed death ligand 1 and is approved by the U.S. Food and Drug Administration for locally advanced or metastatic urothelial cancer and locally advanced, unresectable stage 3 non-small cell lung cancer. Olaparib, a poly (ADP-ribose) polymerase inhibitor, has demonstrated an improvement in median progression-free survival (PFS) in select patients with metastatic castration-resistant prostate cancer (mCRPC). Data from other trials suggest there may be improved activity in men with DNA damage repair (DDR) mutations treated with checkpoint inhibitors. This trial evaluated durvalumab and olaparib in patients with mCRPC with and without somatic or germline DDR mutations. Methods: Eligible patients had received prior enzalutamide and/or abiraterone. Patients received durvalumab 1500 mg i.v. every 28 days and olaparib 300 mg tablets p.o. every 12 h until disease progression or unacceptable toxicity. All patients had biopsies of metastatic lesions with an evaluation for both germline and somatic mutations. Results: Seventeen patients received durvalumab and olaparib. Nausea was the only nonhematologic grade 3 or 4 toxicity occurring in > 1 patient (2/17). No patients were taken off trial for toxicity. Median radiographic progression-free survival (rPFS) for all patients is 16.1 months (95% CI: 4.5–16.1 months) with a 12-month rPFS of 51.5% (95% CI: 25.7–72.3%). Activity is seen in patients with alterations in DDR genes, with a median rPFS of 16.1 months (95% CI: 7.8–18.1 months). Nine of 17 (53%) patients had a radiographic and/or PSA response. Patients with fewer peripheral myeloid-derived suppressor cells and with alterations in DDR genes were more likely to respond. Early changes in circulating tumor cell counts and in both innate and adaptive immune characteristics were associated with response. Conclusions: Durvalumab plus olaparib has acceptable toxicity, and the combination demonstrates efficacy, particularly in men with DDR abnormalities. Trial registration: ClinicalTrials.gov identifier: NCT02484404. [ABSTRACT FROM AUTHOR]

Introduction: Acute myocardial infarction (AMI) and resulting cardiac damage and heart failure are leading causes of morbidity and mortality worldwide. Multiple studies have examined the utility of CD34+ cells for the treatment of acute and ischemic heart disease. However, the optimal strategy to enrich CD34 cells from clinical sources is not known. We examined the efficacy of fluorescence activated cell sorting (FACS) and magnetic beads cell sorting (MACS) methods for CD34 cell isolation from mobilized human mononuclear peripheral blood cells (mhPBMNCs). Methods: mhPBCs were processed following acquisition using FACS or MACS according to clinically established protocols. Cell viability, CD34 cell purity and characterization of surface marker expression were assessed using a flow cytometer. For in vivo characterization of cardiac repair, we conducted LAD ligation surgery on 8–10 weeks female NOD/SCID mice followed by intramyocardial transplantation of unselected mhPBMNCs, FACS or MACS enriched CD34+ cells. Results: Both MACS and FACS isolation methods achieved high purity rates, viability, and enrichment of CD34+ cells. In vivo studies following myocardial infarction demonstrated retention of CD34+ in the peri-infarct region for up to 30 days after transplantation. Retained CD34+ cells were associated with enhanced angiogenesis and reduced inflammation compared to unselected mhPBMNCs or PBS treatment arms. Cardiac scar and fibrosis as assessed by immunohistochemistry were reduced in FACS and MACS CD34+ treatment groups. Finally, reduced scar and augmented angiogenesis resulted in improved cardiac functional recovery, both on the global and regional function and remodeling assessments by echocardiography. Conclusion: Cell based therapy using enriched CD34+ cells sorted by FACS or MACS result in better cardiac recovery after ischemic injury compared to unselected mhPBMNCs. Both enrichment techniques offer excellent recovery and purity and can be equally used for clinical applications. [ABSTRACT FROM AUTHOR]

Multiple sclerosis (MS) is an autoimmune-inflammatory neurodegenerative disease that is often accompanied by a debilitating neuropathic pain. Disease-modifying agents slow down the progression of multiple sclerosis and prevent relapses, yet it remains unclear if they yield analgesia. We explored the analgesic potential of fingolimod (FTY720), an agonist and/or functional antagonist at the sphingosine-1-phosphate receptor 1 (S1PR1), because it reduces hyperalgesia in models of peripheral inflammatory and neuropathic pain. We used a myelin oligodendrocyte glycoprotein 35 to 55 (MOG35-55) mouse model of experimental autoimmune encephalomyelitis, modified to avoid frank paralysis, and thus, allow for assessment of withdrawal behaviors to somatosensory stimuli. Daily intraperitoneal fingolimod reduced behavioral signs of central neuropathic pain (mechanical and cold hypersensitivity) in a dose-dependent and reversible manner. Both autoimmune encephalomyelitis and fingolimod changed hyperalgesia before modifying motor function, suggesting that pain-related effects and clinical neurological deficits were modulated independently. Fingolimod also reduced cellular markers of central sensitization of neurons in the dorsal horn of the spinal cord: glutamate-evoked Ca signaling and stimulus-evoked phospho-extracellular signal-related kinase ERK (pERK) expression, as well as upregulation of astrocytes (GFAP) and macrophage/microglia (Iba1) immunoreactivity. The antihyperalgesic effects of fingolimod were prevented or reversed by the S1PR1 antagonist W146 (1 mg/kg daily, i.p.) and could be mimicked by either repeated or single injection of the S1PR1-selective agonist SEW2871. Fingolimod did not change spinal membrane S1PR1 content, arguing against a functional antagonist mechanism. We conclude that fingolimod behaves as an S1PR1 agonist to reduce pain in multiple sclerosis by reversing central sensitization of spinal nociceptive neurons. [ABSTRACT FROM AUTHOR]

LMB‐100 is a novel immune‐conjugate (immunotoxin) that targets mesothelin. A phase 1/2 clinical trial was conducted (NCT02810418) with primary objectives assessing the safety and efficacy of LMB‐100 ± nab‐paclitaxel. Participant blood samples were analyzed for changes in serum cytokines and circulating immune cell subsets associated with response or toxicity. On Arm A, participants (n = 20) received standard 30‐minute LMB‐100 infusion with nab‐paclitaxel. Although clinical efficacy was observed, the combination caused intolerable capillary leak syndrome (CLS), a major toxicity of unclear etiology that affects many immunotoxin drugs. Participants developing CLS experienced rapid elevations in IFNγ and IL‐8 compared to those without significant CLS, along with midcycle increases in Ki‐67‐ CD4 T cells that were CD38, HLA‐DR, or TIM3 positive. Additionally, a strong increase in activated CD4 and CD8 T cells and a concurrent decrease in Tregs were seen in the single Arm A patient achieving a partial response. In Arm B, administration of single agent LMB‐100 to participants (n = 20) as a long infusion given over 24–48 h was investigated based on pre‐clinical data that this format could reduce CLS. An optimal dose and schedule of long infusion LMB‐100 were identified, but no clinical efficacy was observed even in patients receiving LMB‐100 in combination with nab‐paclitaxel. Despite this, both Arm A and B participants experienced increases in specific subsets of proliferating CD4 and CD8 T cells following Cycle 1 treatment. In summary, LMB‐100 treatment causes systemic immune activation. Inflammatory and immune changes that accompany drug associated CLS were characterized for the first time. [ABSTRACT FROM AUTHOR]

Prime-boost vaccination with recombinant (r) vaccinia(V)-CEA(6D)-TRICOM (triad of co-stimulatory molecules B7.1, ICAM-1 and LFA-3) and rFowlpox(F)-CEA(6D)-TRICOM infect antigen-presenting cells and direct expression of co-stimulatory molecules. We hypothesized that co-administration of vaccine with GM-CSF and interferon alpha (IFN-α) would have efficacy in CEA-expressing cancers. Patients with CEA-expressing cancers received the rV-CEA(6D)-TRICOM vaccine subcutaneously (s.c.) on day 1 followed by GM-CSF s.c. to the injection site on days 1-4. In Cycle 1, patients received thrice weekly s.c. injections of IFN-α-2b the week after rV-CEA(6D)-TRICOM. In Cycles 2-4, patients received thrice weekly s.c. injections of IFN-α-2b the same week that rF-CEA(6D)-TRICOM was given. The first cohort received no IFN followed by dose escalation of IFN-α in subsequent cohorts. Thirty-three patients were accrued (mean 59.8 years). Grade 3 toxicities included fatigue and hyperglycemia. Grade 4-5 adverse events (unrelated to treatment) were confusion (1), elevated aspartate transaminase (AST)/alanine transaminase (ALT) (1), and sudden death (1). No patients had a partial response, and eight patients exhibited stable disease of ≥3 months. Median progression-free survival and overall survival (OS) were 1.8 and 6.3 months, respectively. Significantly higher serum CD27 levels were observed after vaccine therapy ( p = 0.006 post 1-2 cycles, p = 0.003 post 3 cycles, p = 0.03 post 4-7 cycles) and 42 % of patients assayed developed CEA-specific T cell responses. Pre-treatment levels of myeloid-derived suppressor cells correlated with overall survival ( p = 0.04). Administration of IFN-α led to significantly increased OS ( p = 0.02) compared to vaccine alone. While the vaccine regimen produced no clinical responses, IFN-α administration was associated with improved survival. [ABSTRACT FROM AUTHOR]

Acute myocardial infarction (AMI) results in weakening of the heart muscle and an increased risk for chronic heart failure. Therapeutic stem cells have been shown to reduce inflammatory signaling and scar tissue expansion, despite most of these studies being limited by poor retention of cells. Gelatin methacrylate (GelMA) coatings have been shown to increase the retention of these therapeutic cells near the infarct. In this work, we evaluate two different potential binding partners for GelMA-coated bone marrow cells (BMCs) and myocardial tissue: the extracellular matrix (ECM) and interstitial non-cardiomyocytes. While cells containing β1 integrins mediate cell-ECM adhesion in vivo, these cells do not promote binding to our collagen-degraded, GelMA coating. Specifically, microscopic imagining shows that even with high integrin expression, GelMA-coated BMCs do not bind to cells within the myocardium. Alternatively, BMC incubation with decellularized heart tissue results in higher adhesion of coated cells versus uncoated cells supporting our GelMA-ECM binding mode. To further evaluate the ECM binding mode, cells were incubated on slides modified with one of three different major heart ECM components: collagen, laminin, or fibronectin. While all three components promoted higher adhesion than unmodified glass, collagen-coated slides resulted in a significantly higher adhesion of GelMA-coated BMCs over laminin and fibronectin. Incubation with unmodified BMCs confirmed that without a GelMA coating minimal adhesion of BMCs occurred. We conclude that GelMA cellular coatings significantly increase the binding of cells to collagen within the ECM. Our results provide progress towards a biocompatible and easily translatable method to enhance the retention of transplanted cells in human studies. [ABSTRACT FROM AUTHOR]

Acute myocardial infarction (AMI) initiates pathological inflammation which aggravates tissue damage and causes heart failure. Lysophosphatidic acid (LPA), produced by autotaxin (ATX), promotes inflammation and the development of atherosclerosis. The role of ATX/LPA signaling nexus in cardiac inflammation and resulting adverse cardiac remodeling is poorly understood. We assessed autotaxin activity and LPA levels in relation to cardiac and systemic inflammation in AMI patients and C57BL/6 (WT) mice. Human and murine peripheral blood and cardiac tissue samples showed elevated levels of ATX activity, LPA, and inflammatory cells following AMI and there was strong correlation between LPA levels and circulating inflammatory cells. In a gain of function model, lipid phosphate phosphatase-3 (LPP3) specific inducible knock out (Mx1-Plpp 3 Δ) showed higher systemic and cardiac inflammation after AMI compared to littermate controls (Mx1-Plpp3fl/fl); and a corresponding increase in bone marrow progenitor cell count and proliferation. Moreover, in Mx1- Plpp3Δ mice, cardiac functional recovery was reduced with corresponding increases in adverse cardiac remodeling and scar size (as assessed by echocardiography and Masson's Trichrome staining). To examine the effect of ATX/LPA nexus inhibition, we treated WT mice with the specific pharmacological inhibitor, PF8380, twice a day for 7 days post AMI. Inhibition of the ATX/LPA signaling nexus resulted in significant reduction in post-AMI inflammatory response, leading to favorable cardiac functional recovery, reduced scar size and enhanced angiogenesis. ATX/LPA signaling nexus plays an important role in modulating inflammation after AMI and targeting this mechanism represents a novel therapeutic target for patients presenting with acute myocardial injury. Schematic summarizing the role of autotaxin/lysophosphatidic acid signaling in post-AMI inflammation and cardiac dysfunction. Damage associated molecular pattern molecules released from dying cardiomyocytes and cardiac fibroblasts initiate an inflammatory cascade resulting in the activation of Autotaxin/lysophosphatidic acid signaling; which in turn augments local and systemic inflammation through the release of inflammatory cytokines and chemokines. Systemically, activation of bone marrow progenitors results in increased production of inflammatory cells. Locally, increased chemoattractants such as MCP-1 results in increased infiltration of inflammatory cells to the damaged myocardium. Collectively, these events result in prolonged and exacerbated inflammatory response and impaired cardiac functional recovery Unlabelled Image • LPA-ATX signaling nexus modulated post-myocardial infarction inflammation. • Pharmacological inhibition of ATX/LPA signaling attenuates cardiac and systemic inflammation post-AMI. • ATX inhibition post-AMI enhances cardiac recovery, reduces adverse remodeling and scar size. [ABSTRACT FROM AUTHOR]

Purpose: Despite the strong interest in combining stereotactic ablative radiation therapy (SAR) with immunotherapy, limited data characterizing the systemic immune response after SAR are available. We hypothesized that the systemic immune response to SAR would differ by irradiated site owing to inherent differences in the microenvironment of various organs.Methods and Materials: Patients receiving SAR to any organ underwent prospective blood banking before and 1 to 2 weeks after SAR. Peripheral blood mononuclear cells (PBMCs) and serum were isolated. PBMCs were stained with fluorophore-conjugated antibodies against T and natural killer (NK) cell markers. Cells were interrogated by flow cytometry, and the results were analyzed using FlowJo software. Serum cytokine and chemokine levels were measured using Luminex. We analyzed the changes from before to after therapy using paired t tests or 1-way analysis of variance (ANOVA) with Bonferroni's post-test.Results: A total of 31 patients had evaluable PBMCs for flow cytometry and 37 had evaluable serum samples for Luminex analysis. The total number of NK cells and cytotoxic (CD56dimCD16+) NK cells decreased (P = .02) and T-cell immunoglobulin- and mucin domain-containing molecule-3-positive (TIM3+) NK cells increased (P = .04) after SAR to parenchymal sites (lung and liver) but not to bone or brain. The total memory CD4+ T cells, activated inducible co-stimulator-positive and CD25+CD4+ memory T cells, and activated CD25+CD8+ memory T cells increased after SAR to parenchymal sites but not bone or brain. The circulating levels of tumor necrosis factor-α (P = .04) and multiple chemokines, including RANTES (P = .04), decreased after SAR to parenchymal sites but not bone or brain.Conclusions: Our data suggest SAR to parenchymal sites induces systemic immune changes, including a decrease in total and cytotoxic NK cells, an increase in TIM3+ NK cells, and an increase in activated memory CD4+ and CD8+ T cells. SAR to nonparenchymal sites did not induce these changes. By comparing the immune response after radiation to different organs, our data suggest SAR induces systemic immunologic changes that are dependent on the irradiated site. [ABSTRACT FROM AUTHOR]

Background: Avelumab (MSB0010718C) is a human IgG1 monoclonal antibody that binds to PD-L1, inhibiting its binding to PD-1, which inactivates T cells. We aimed to establish the safety and pharmacokinetics of avelumab in patients with solid tumours while assessing biological correlatives for future development.Methods: This open-label, single-centre, phase 1a, dose-escalation trial (part of the JAVELIN Solid Tumor trial) assessed four doses of avelumab (1 mg/kg, 3 mg/kg, 10 mg/kg, and 20 mg/kg), with dose-level cohort expansions to provide additional safety, pharmacokinetics, and target occupancy data. This study used a standard 3 + 3 cohort design and assigned patients sequentially at trial entry according to the 3 + 3 dose-escalation algorithm and depending on the number of dose-limiting toxicities during the first 3-week assessment period (the primary endpoint). Patient eligibility criteria included age 18 years or older, Eastern Cooperative Oncology Group performance status 0-1, metastatic or locally advanced previously treated solid tumours, and adequate end-organ function. Avelumab was given as a 1-h intravenous infusion every 2 weeks. Patients in the dose-limiting toxicity analysis set were assessed for the primary endpoint of dose-limiting toxicity, and all patients enrolled in the dose-escalation part were assessed for the secondary endpoints of safety (treatment-emergent and treatment-related adverse events according to National Cancer Institute Common Terminology Criteria for Adverse Events version 4.0), pharmacokinetic and pharmacodynamic profiles (immunological effects), best overall response by Response Evaluation Criteria, and antidrug antibody formation. The population for the pharmacokinetic analysis included a subset of patients with rich pharmacokinetic samples from two selected disease-specific expansion cohorts at the same study site who had serum samples obtained at multiple early timepoints. This trial is registered with ClinicalTrials.gov, number NCT01772004. Patient recruitment to the dose-escalation part reported here is closed.Findings: Between Jan 31, 2013, and Oct 8, 2014, 53 patients were enrolled (four patients at 1 mg/kg, 13 at 3 mg/kg, 15 at 10 mg/kg, and 21 at 20 mg/kg). 18 patients were analysed in the dose-limiting toxicity analysis set: three at dose level 1 (1 mg/kg), three at dose level 2 (3 mg/kg), six at dose level 3 (10 mg/kg), and six at dose level 4 (20 mg/kg). Only one dose-limiting toxicity occurred, at the 20 mg/kg dose, and thus the maximum tolerated dose was not reached. In all 53 enrolled patients (the safety analysis set), common treatment-related adverse events (occurring in >10% of patients) included fatigue (21 patients [40%]), influenza-like symptoms (11 [21%]), fever (8 [15%]), and chills (6 [11%]). Grade 3-4 treatment-related adverse events occurred in nine (17%) of 53 patients, with autoimmune disorder (n=3), increased blood creatine phosphokinase (n=2), and increased aspartate aminotransferase (n=2) each occurring in more than one patient (autoimmune disorder in two patients at 10 mg/kg and one patient at 20 mg/kg, increased blood creatine phosphokinase in two patients at 20 mg/kg, and increased aspartate aminotransferase in one patient at 1 mg/kg, and one patient at 10 mg/kg). Six (11%) of 53 patients had a serious treatment-related adverse event: autoimmune disorder (two [13%]), lower abdominal pain (one [7%]), fatigue (one [7%]), and influenza-like illness (one [7%]) in three patients treated at 10 mg/kg dose level, and autoimmune disorder (one [5%]), increased amylase (one [5%]), myositis (one [5%]), and dysphonia (one [5%]) in three patients who received the 20 mg/kg dose. We recorded some evidence of clinical activity in various solid tumours, with partial confirmed or unconfirmed responses in four (8%) of 53 patients; 30 (57%) additional patients had stable disease. Pharmacokinetic analysis (n=86) showed a dose-proportional exposure between doses of 3 mg/kg and 20 mg/kg and a half-life of 95-99 h (3·9-4·1 days) at the 10 mg/kg and 20 mg/kg doses. Target occupancy was greater than 90% at doses of 3 mg/kg and 10 mg/kg. Antidrug antibodies were detected in two (4%) of 53 patients. No substantial differences were found in absolute lymphocyte count or multiple immune cell subsets, including those expressing PD-L1, after treatment with avelumab. 31 (58%) of 53 patients in the overall safety population died; no deaths were related to treatment on study.Interpretation: Avelumab has an acceptable toxicity profile up to 20 mg/kg and the maximum tolerated dose was not reached. Based on pharmacokinetics, target occupancy, and immunological analysis, we chose 10 mg/kg every 2 weeks as the dose for further development and phase 3 trials are ongoing.Funding: National Cancer Institute and Merck KGaA. [ABSTRACT FROM AUTHOR]

• The effect on multiple components of the peripheral immunome from the first-in-human trial of the tumor targeting immune-cytokine NHS-IL12. • Greater immune activation is seen with both a higher dose and a more frequent dosing schedule of NHS-IL12. • Immune analytes of patients at both baseline and early after treatment with NHS-IL12 associate with clinical response. • These findings will help guide future schedule and dosing regimens of NHS-IL12. The immunocytokine NHS-IL12 delivers IL-12 to the tumor microenvironment by targeting DNA/histones in necrotic areas. The first-in-human clinical trial administered NHS-IL12 subcutaneously in 59 patients treated every-four weeks (Q4W), with a maximum tolerated dose of 16.8 mcg/kg. The phase I study was expanded to include a high-exposure cohort that received bi-weekly treatment (Q2W) with two dose levels of NHS-IL12: 12.0 mcg/kg and 16.8 mcg/kg. Here, patients given NHS-IL12 were analyzed both prior to and early after treatment for effects on 10 serum soluble analytes, complete blood counts, and 158 peripheral immune subsets. Higher levels of immune activation were seen with a dose of 16.8 mcg/kg versus 12.0 mcg/kg in patients in the high-exposure cohort, as evidenced by greater increases in serum IFNγ, TNFα, and soluble PD-1, and greater increases in frequencies of peripheral ki67+ mature natural killer (NK), CD8+ T, and NKT cells. Greater immune activation was also seen in the Q2W versus Q4W cohort, as demonstrated by greater increases in pro-inflammatory serum analytes, ki67+ CD8+ T, NK, and NKT cells, intermediate monocytes, and a greater decrease in CD73+ T cells. Specific immune analytes at baseline including lower levels of monocytes and plasmacytoid dendritic cells, and early changes after treatment such as an increase in refined NK cell subsets and total CD8+ T cells, associated with better clinical response. These findings may help to guide future schedule and dosing regimens of clinical studies of NHS-IL12 as monotherapy and in combination therapies. [ABSTRACT FROM AUTHOR]

Abstract: The persistence of pain after surgery increases the recovery interval from surgery to a normal quality of life. AYX1 is a DNA-decoy drug candidate designed to prevent post-surgical pain following a single intrathecal injection. Tissue injury causes a transient activation of the transcription factor EGR1 in the dorsal root ganglia–dorsal horn network, which then triggers changes in gene expression that induce neuronal hypersensitivity. AYX1 is a potent, specific inhibitor of EGR1 activity that mimics the genomic EGR1-binding sequence. Administered in the peri-operative period, AYX1 dose dependently prevents mechanical hypersensitivity in models of acute incisional (plantar), inflammatory (CFA), and chronic neuropathic pain (SNI) in rats. Furthermore, in a knee surgery model evaluating functional measures of postoperative pain, AYX1 improved weight-bearing incapacitance and spontaneous rearing compared to control. These data illustrate the potential clinical therapeutic benefits of AYX1 for preventing the transition of acute to chronic post-surgical pain. [Copyright &y& Elsevier]

Abstract: Objective: The opioid growth factor (OGF) and its receptor (OGFr), serve as inhibitory axis regulating cell proliferation in normal cells and cancer. We investigated the presence and relative expression of OGF and OGFr in normal human ovarian surface epithelial (HOSE) cells, benign ovarian cysts, and ovarian cancers. Methods: Surgical samples of 16 patients with ovarian cancer and 27 patients with ovarian benign cysts were obtained intraoperatively. HOSE were collected by scraping the surface of normal ovaries of 10 post menopausal women undergoing hysterectomy and oophorectomy. Semiquantitative immunohistochemistry was used to assess the presence, distribution, and levels of OGF and OGFr. Receptor binding assays measured binding capacity and affinity of OGFr for radiolabeled OGF. Results: OGF and OGFr were present in HOSE cells, ovarian cysts, and ovarian cancers. Compared to HOSE cells, OGF and OGFr protein levels were reduced 29% and 34% (p<0.001), respectively, in ovarian cysts, and decreased 58% and 48% (p<0.001), respectively, in ovarian cancers. Binding assays revealed 5.4 fold fewer OGFr binding sites in cancers than cysts (p<0.05). Levels of OGF and OGFr were comparable in primary, metastatic, or recurrent ovarian cancers. Conclusion: We have shown that a native opioid pathway, the OGF–OGFr axis, is present in human ovarian cancer. Importantly, the expression of OGF and OGFr is diminished in human ovarian cancer. As OGF and OGFr normally function in maintaining cell proliferation, therapy to harness OGF/OGFr function could provide a useful biologic-based treatment for human ovarian cancer. [Copyright &y& Elsevier]

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a potential cancer therapeutic that induces apoptosis in cancer cells while sparing the non-malignant cells in preclinical models. However, its efficacy in clinical trials has been limited, suggesting unknown modulatory mechanisms responsible for the lack of TRAIL activity in patients. Here, we hypothesized that TRAIL treatment elicits transcriptional changes in triple negative breast cancer (TNBC) cells that alter the immune milieu. To test this, we performed an RNAseq analysis of MDA-MB-231 cells treated with TRAIL, followed by validation in additional TNBC cell lines. TRAIL significantly induces expression of multiple cytokines such as CXCLs 1, 2, 3, 8,11 and IL-6, which are known to modify neutrophil function. Mechanistically, the induction of these cytokines was predominantly mediated by death receptor 5, caspase 8 (but not caspase 8 enzymatic activity), and the non-canonical NFKB2 pathway. The cytokines produced by the TRAIL-treated TNBC cells enhanced chemotaxis of healthy human donor isolated neutrophils. In vivo , TRAIL treated TNBC murine xenograft tumors showed activation of the NFKB2 pathway, elevated production of CXCLs and IL-6, and increased neutrophil recruitment into the tumors. Moreover, donor isolated neutrophils preincubated in supernatants from TRAIL-treated TNBC cells exhibited impaired cytotoxic effect against TNBC cells. Transcriptomic analysis of neutrophils incubated with either TRAIL alone or supernatant of TRAIL-treated TNBC cells revealed increased expression of inflammatory cytokines, immune modulatory genes, immune checkpoint genes, and genes implicated in delayed neutrophil apoptosis. Functional studies with these neutrophils confirmed their suppressive effect on T cell proliferation and an increase in Treg suppressive phenotype. Collectively, our study demonstrates a novel role of TRAIL-induced NFKB2-dependent cytokine production that promotes neutrophil chemotaxis and immune suppression.

The development of vaccines, especially RNA-based, directed against patient-specific tumor neoepitopes is an active and productive area of cancer immunotherapy. Promising clinical results in melanoma and other solid tumor types are emerging. As with all cancer therapy modalities, neoepitope vaccine development and delivery also has some drawbacks, including the level of effort to develop a patient-specific product, accuracy of algorithms to predict neoepitopes, and with the exception of melanoma and some other tumor types, biopsies of metastatic lesions of solid tumors are often not available. We hypothesize that in some circumstances the use of rationally designed combinations of "off-the-shelf" agents may prove an additional path to enable the patient to produce his/her own "neoepitope vaccine" in situ. These combination therapies may consist of agents to activate a tumor-associated T-cell response, potentiate that response, reduce or eliminate immunosuppressive entities in the tumor microenvironment, and/or alter the phenotype of tumor cells to render them more susceptible to immune-mediated lysis. Examples are provided in both preclinical and clinical studies in which combinations of "off-the-shelf" agents lead to the generation of T cells directed against tumor-derived neoepitopes with consequent antitumor activity., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Longitudinal sampling of tumor tissue from patients with solid cancers, aside from melanoma and a few other cases, is often unfeasible, and thus may not capture the plasticity of interactions between the tumor and immune system under selective pressure of a given therapy. Peripheral blood analyses provide salient information about the human peripheral immunome while offering technical and practical advantages over traditional tumor biopsies, and should be utilized where possible alongside interrogation of the tumor. Some common blood-based biomarkers used to study the immune response include immune cell subsets, circulating tumor DNA, and protein analytes such as cytokines. With the recent explosion of immune checkpoint inhibitors (ICI) as a modality of treatment in multiple cancer types, soluble immune checkpoints have become a relevant area of investigation for peripheral immune-based biomarkers. However, the exact functions of soluble immune checkpoints and their roles in cancer for the most part remain unclear. This review discusses current literature on the production, function, and expression of nine soluble immune checkpoints - sPD-L1, sPD-1, sCTLA4, sCD80, sTIM3, sLAG3, sB7-H3, sBTLA, and sHVEM - in patients with solid tumors, and explores their role as biomarkers of response to ICI as well as to conventional therapies (chemotherapy, radiotherapy, targeted therapy, and surgery) in cancer patients., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Chemotherapy and immune checkpoint inhibitors have a role in the post-neoadjuvant setting in patients with triple-negative breast cancer (TNBC). However, the effects of nivolumab, a checkpoint inhibitor, capecitabine, or the combination in changing peripheral immunoscore (PIS) remains unclear. This open-label randomized phase II OXEL study (NCT03487666) aimed to assess the immunologic effects of nivolumab, capecitabine, or the combination in terms of the change in PIS (primary endpoint). Secondary endpoints included the presence of ctDNA, toxicity, clinical outcomes at 2-years and association of ctDNA and PIS with clinical outcomes. Forty-five women with TNBC and residual invasive disease after standard neoadjuvant chemotherapy were randomized to nivolumab, capecitabine, or the combination. Here we show that a combination of nivolumab plus capecitabine leads to a greater increase in PIS from baseline to week 6 (91%) compared with nivolumab (47%) or capecitabine (53%) alone (log-rank p = 0.08), meeting the pre-specified primary endpoint. In addition, the presence of circulating tumor DNA (ctDNA) is associated with disease recurrence, with no new safety signals in the combination arm. Our results provide efficacy and safety data on this combination in TNBC and support further development of PIS and ctDNA analyses to identify patients at high risk of recurrence., (© 2024. The Author(s).)

The paradigm of non-small cell lung cancer (NSCLC) treatment has been profoundly influenced by the development of immune checkpoint inhibitors (ICI), but the range of clinical responses observed among patients poses significant challenges. To date, analyses of tumor biopsies are the only parameter used to guide prognosis to ICI therapy. Tumor biopsies, however, are often difficult to obtain and tissue-based biomarkers are limited by intratumoral heterogeneity and temporal variability. In response, there has been a growing emphasis on the development of "liquid biopsy"‒ derived biomarkers, which offer a minimally invasive means to dynamically monitor the immune status of NSCLC patients either before and/or during the course of treatment. Here we review studies in which multiple blood-based biomarkers encompassing circulating soluble analytes, immune cell subsets, circulating tumor DNA, blood-based tumor mutational burden, and circulating tumor cells have shown promising associations with the clinical response of NSCLC patients to ICI therapy. These investigations have unveiled compelling correlations between the peripheral immune status of patients both before and during ICI therapy and patient outcomes, which include response rates, progression-free survival, and overall survival. There is need for rigorous validation and standardization of these blood-based assays for broader clinical application. Integration of multiple blood-based biomarkers into comprehensive panels or algorithms also has the potential to enhance predictive accuracy. Further research aimed at longitudinal monitoring of circulating biomarkers is also crucial to comprehend immune dynamics and resistance mechanisms and should be used alongside tissue-based methods that interrogate the tumor microenvironment to guide treatment decisions and may inform on the development of novel therapeutic strategies. The data reviewed here reinforce the opportunity to refine patient stratification, optimize treatments, and improve outcomes not only in NSCLC but also in the wider spectrum of solid tumors undergoing immunotherapy., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Background: Bintrafusp alfa, a first-in-class bifunctional fusion protein targeting transforming growth factor-β (TGF-β) and programmed cell death ligand 1, has demonstrated encouraging efficacy as second-line treatment in patients with non-small cell lung cancer (NSCLC) in a dose expansion cohort of the phase 1, open-label clinical trial (NCT02517398). Here, we report the safety, efficacy, and biomarker analysis of bintrafusp alfa in a second expansion cohort of the same trial (biomarker cohort)., Methods: Patients with stage IIIb/IV NSCLC who were either immune checkpoint inhibitor (ICI)-naïve (n=18) or ICI-experienced (n=23) were enrolled. The primary endpoint was the best overall response. Paired biopsies (n=9/41) and peripheral blood (n=14/41) pretreatment and on-treatment were studied to determine the immunological effects of treatment and for associations with clinical activity., Results: Per independent review committee assessment, objective responses were observed in the ICI-naïve group (overall response rate, 27.8%). No new or unexpected safety signals were identified. Circulating TGF-β levels were reduced (>97%; p<0.001) 2 weeks after initiation of treatment with bintrafusp alfa and remained reduced up to 12 weeks. Increases in lymphocytes and tumor-associated macrophages (TAMs) were observed in on-treatment biospies, with an increase in the M2 (tumor trophic TAMs)/M1 (inflammatory TAMs) ratio associated with poor outcomes. Specific peripheral immune analytes at baseline and early changes after treatment were associated with clinical response., Conclusions: Bintrafusp alfa was observed to have modest clinical activity and manageable safety, and was associated with notable immunologic changes involving modulation of the tumor immune microenvironment in patients with advanced NSCLC., Competing Interests: Competing interests: AR reports research funding to his institution from Promontory Therapeutics Inc. OR reports leadership roles as vice president and global clinical strategy head of gastrointestinal oncology at AstraZeneca and reports having stocks at AstraZeneca. RI reports employment with Merck Biopharma Co., Ltd., Tokyo, Japan, an affiliate of Merck KGaA and has attended the independent data monitoring committee/steering committee as part of employment responsibilities. YV reports employment with EMD Serono Research & Development Institute, Inc., Billerica, Massachusetts, USA, an affiliate of Merck KGaA and patent applications. MS reports employment with Merck Biopharma Co., Ltd., Tokyo, Japan, an affiliate of Merck KGaA. AM reports employment with EMD Serono Research & Development Institute, Inc., Billerica, Massachusetts, USA, an affiliate of Merck KGaA. JS reports patents. JLG reports patents and is vice president of the Society for Immunotherapy of Cancer. The remaining authors declare no conflicts of interests., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)