Immune-competent Hosts Research Articles

Genomic and functional analysis of the host response to acute simian varicella infection in the lung.

Varicella Zoster Virus (VZV) is the causative agent of varicella and herpes zoster. Although it is well established that VZV is transmitted via the respiratory route, the host-pathogen interactions during acute VZV infection in the lungs remain poorly understood due to limited access to clinical samples. To address these gaps in our knowledge, we leveraged a nonhuman primate model of VZV infection where rhesus macaques are intrabronchially challenged with the closely related Simian Varicella Virus (SVV). Acute infection is characterized by immune infiltration of the lung airways, a significant up-regulation of genes involved in antiviral-immunity, and a down-regulation of genes involved in lung development. This is followed by a decrease in viral loads and increased expression of genes associated with cell cycle and tissue repair. These data provide the first characterization of the host response required to control varicella virus replication in the lung and provide insight into mechanisms by which VZV infection can cause lung injury in an immune competent host.

Lack of immunoediting in murine pancreatic cancer reversed with neoantigen.

In carcinogen-driven cancers, a high mutational burden results in neoepitopes that can be recognized immunologically. Such carcinogen-induced tumors may evade this immune response through "immunoediting," whereby tumors adapt to immune pressure and escape T cell-mediated killing. Many tumors lack a high neoepitope burden, and it remains unclear whether immunoediting occurs in such cases. Here, we evaluated T cell immunity in an autochthonous mouse model of pancreatic cancer and found a low mutational burden, absence of predicted neoepitopes derived from tumor mutations, and resistance to checkpoint immunotherapy. Spontaneous tumor progression was identical in the presence or absence of T cells. Moreover, tumors arising in T cell-depleted mice grew unchecked in immune-competent hosts. However, introduction of the neoantigen ovalbumin (OVA) led to tumor rejection and T cell memory, but this did not occur in OVA immune-tolerant mice. Thus, immunoediting does not occur in this mouse model - a likely consequence, not a cause, of absent neoepitopes. Because many human tumors also have a low missense mutational load and minimal neoepitope burden, our findings have clinical implications for the design of immunotherapy for patients with such tumors.

Primary intracranial lymphomas.

Background:Primary CNS lymphoma (PCNSL), a rare form of aggressive extranodal non-Hodgkin's lymphoma (NHL), has increased in incidence during the last three decades and occurs in both immune compromised and immune competent hosts. It has an overall poor prognosis.Objective:This study attempts to further delineate the clinico-pathological, immunohistochemical and radiological profile of PCNSL at Jeddah to King Faisal Hospital and Research Center.Methods:Computerized search through the archives of King Faisal Hospital and Research Centre between July 2000- December 2012 identified 15 patients with pathologically confirmed PCNSL. These were analyzed retrospectively. Their clinico-pathological, immunohistochemical and radiological data were analyzed.Results:Of the 15 PCNSL patients, 8 (53.3%) were females and 7 (46.6%) were males. There was female predilection especially in the age group of 40-59 years. Mean age at diagnosis for all patients was 50.4 years. There was no patient in the pediatric age group. The most common location in the brain was the frontal region in 7 patients (46.6%), 7 (46.6%) had multiple intracranial masses; all 15 (100%) were Non Hodgkin B-cell lymphomas, among which 13 (86.6%) were diffuse large B-cell lymphomas. All 15 (100%) cases showed diffuse and strong positivity for CD 45, and CD 20. Fourteen patients were immune competent while one was immune compromised.Conclusions:PCNSL often occurs in middle-aged and aged patients. There is female predilection especially in the middle age. Frontal region is the most common location with diffuse large B-cell lymphoma being the predominant subtype.

Therapeutic Advances in Localized Pancreatic Cancer.

It is estimated that pancreatic cancer (PC) will become the second leading cause of cancer-related death in the United States by 2030. Clinical and preclinical data support the understanding that PC metastases occur early in the pathogenesis of this disease, even before the primary tumor can be detected. This has important implications for the clinical management of patients with localized PC, as surgery alone is unlikely to be curative for most patients. The delivery of postoperative adjuvant therapy is problematic in this disease because of the magnitude of the operation needed to remove the primary tumor, which can affect patient recovery and delay (sometimes indefinitely) the delivery of systemic therapy. For these reasons, the use of chemotherapy and/or chemoradiation prior to surgery (neoadjuvant therapy) is increasingly recognized as the preferred strategy for treatment sequencing. Neoadjuvant therapy is recommended for patients with borderline resectable PC and, at some centers, neoadjuvant therapy has been extended to patients with resectable PC as well. Importantly, therapeutic advances in multidrug systemic therapy and radiation therapy have already been adopted in the neoadjuvant setting where treatment toxicity will not be compounded by surgical recovery. In addition, the use of local-regional therapies in highly selected patients with locally advanced PC, following a prolonged period of induction systemic therapy, will be an area of intense scrutiny. Future improvements in diagnostic biomarkers may allow for real-time sequencing of multimodality therapy for individual patients based on a more accurate and timely assessment of treatment response. Neoadjuvant treatment sequencing allows patients to receive multimodality therapy in a manner that prioritizes early exposure to systemic therapy to maximize the treatment of micrometastatic disease in an immune-competent host prior to surgical intervention. Patients who complete all intended neoadjuvant therapy, including surgery, experience an overall survival benefit that is unmatched by a surgery-first approach.

Assessment of Immune Isolation of Allogeneic Mouse Pancreatic Progenitor Cells by a Macroencapsulation Device.

Embryonic stem cell (ESC)-derived β cells hold the promise of providing a renewable source of tissue for the treatment of insulin-dependent diabetes. Encapsulation may allow ESC-derived β cells to be transplanted without immunosuppression, thus enabling wider application of this therapy. In this study, we investigated the immunogenicity of mouse pancreatic progenitor cells and efficacy of a new macroencapsulation device in protecting these cells against alloimmune and autoimmune responses in mouse models. Mouse pancreatic progenitor cells activated the indirect but not the direct pathway of alloimmune response and were promptly rejected in immune competent hosts. The new macroencapsulation device abolished T cell activation induced by allogeneic splenocytes and protected allogeneic MIN6 β cells and pancreatic progenitors from rejection even in presensitized recipients. In addition, the device was effective in protecting MIN6 cells in spontaneously diabetic nonobese diabetic recipients against both alloimmune and recurring autoimmune responses. Our results demonstrate that macroencapsulation can effectively prevent immune sensing and rejection of allogeneic pancreatic progenitor cells in fully sensitized and autoimmune hosts.

T cell responses to cytomegalovirus.

Human cytomegalovirus (HCMV) establishes a latent infection that generally remains asymptomatic in immune-competent hosts for decades but can cause serious illness in immune-compromised individuals. The long-term control of CMV requires considerable effort from the host immune system and has a lasting impact on the profile of the immune system. One hallmark of CMV infection is the maintenance of large populations of CMV-specific memory CD8(+) T cells - a phenomenon termed memory inflation - and emerging data suggest that memory inflation is associated with impaired immunity in the elderly. In this Review, we discuss the molecular triggers that promote memory inflation, the idea that memory inflation could be considered a natural pathway of T cell maturation that could be harnessed in vaccination, and the broader implications of CMV infection and the T cell responses it elicits.

Broncholaveolar lavage to detect cytomegalovirus infection, latency, and reactivation in immune competent hosts.

Roughly 1/3rd of immune competent patients will reactivate latent cytomegalovirus (CMV) during critical illness. There are no standard methods to detect reactivation, and some investigators have postulated that presence of DNA in BAL fluid is indicative of viral replication. To test this hypothesis, we used a murine model that allows inclusion of matched healthy controls which is not possible in human studies. BALB/c mice infected with Smith-murine CMV or PBS (mock) had BAL evaluated 7, 14, or 21 days after acute infections, during latency, or during bacterial sepsis. Plaque assay, PCR, and rtPCR were performed on BALs and concomitantly obtained lung tissue. BAL cellular compositions, including tetramer evaluation of CMV-specific T cells were evaluated by flow cytometry. CMV DNA were detected in BAL at all time-points during acute infection, becoming undetectable in all mice during latency, then were detected again during bacterial sepsis, peaking 3 weeks after onset. mCMV specific T-cells were most numerous in BAL after acute viral infections, decreasing to low levels during latency, then fluctuating during bacterial sepsis. Specifically, mCMV-specific T-cells contracted at sepsis onset, expanding 2-4 weeks post-sepsis, presumably in response to increased viral loads at that time point. Altogether, our results support the use of BAL PCR for the diagnosis of CMV replication in immune competent hosts. Additionally, we demonstrate dynamic changes in CMV-specific T cells that occur in BAL during CMV infection and during sepsis induced viral reactivation. J. Med. Virol. 88:1408-1416, 2016. © 2016 Wiley Periodicals, Inc.

Bacteremia with an Unusual Pathogen: Mycobacterium neoaurum.

Mycobacterium neoaurum (M. neoaurum) is an infrequently encountered cause of infection in humans. It is a member of the rapidly growing mycobacteria family. It predominately afflicts those with a compromised immune status and a chronically indwelling vascular access. Isolation of this organism is challenging yet the advent of 16s ribosomal sequencing paved the way for more sensitive detection. No treatment guidelines are available and treatment largely depends on the experience of the treating physician and nature of the isolate. We report a case of M. neoaurum bacteremia in an immune competent host, with a chronically placed peripherally inserted central catheter (PICC line).

Disseminated varicella zoster in an immunocompromised patient: A case report

Introduction: Human Herpes Virus 3 (HHV- 3) is a human-specific virus known to be the cause behind chicken pox and varicella zoster. Presentation of this virus can vary depending on the immune status of the host. Typically, a uniform rash in a single dermatomal distribution is all that manifests. Alternatively, as in our case, a widespread and painful vesicular rash can erupt causing significant impairment requiring hospitalization. Case Report: An HIV-positive immunocompromised male presented with disseminated varicella zoster (VZV) despite adequate compliance with HAART therapy. The painful vesicles sporadically covered all areas of the body (feet, abdomen, back, face, etc.) and did not present in the classic dermatomal distribution found in immune competent hosts. Despite adequate therapy with a new combination HIV medication, the patient's CD4 count had diminished and he was admitted for suspected disseminated varicella zoster. PCR was later able to confirm the diagnosis. Conclusion: Varicella Zoster can have varying presentations depending on the immune status of the host. In an immunocompromised state, such as in our patient, the presentation can be widespread and more severe. Early diagnosis with polymerase chain reaction (PCR) and treatment with anti- virals and analgesics is a top priority for the patient's well-being and clinical outcome. In our case, early detection and treatment allowed only for aminimal duration of the disseminated virus with prompt recovery and discharge from the hospital. Further investigation of the virus' interaction with medications, such as HAART therapy, that potentially alters the expression of the virus despite vaccination, as some studies suggest, could be beneficial.

Aspergillus Lymphadenitis Mimicking Cervical Lymph Node Recurrence in a Breast Cancer Patient

Invasive aspergillosis mostly occurs in severely immune compromised patient and infrequently in an immunecompetent host. The usual site of infection is the pulmonary system. Extra pulmonary sites such as isolated lymphadenopathy without systemic symptoms are rare. We present a case of aspergillosis in the cervical lymph node of breast cancer patient six years after completing chemotherapy. Diagnosis was excision biopsy of an enlarged cervical lymph node and the patient was successfully treated with oral Itraconazole for three months without disease recurrence 3 years following completion of medication.

Improved Treatment Efficacy of Antiangiogenic Therapy when Combined with Picornavirus Vaccination in the GL261 Glioma Model.

The addition of antiangiogenic therapy to the standard-of-care treatment regimen for recurring glioblastoma has provided some clinical benefits while also delineating numerous caveats, prompting evaluation of the elicited alterations to the tumor microenvironment. Of critical importance, given the steadily increasing incorporation of immunotherapeutic approaches clinically, is an enhanced understanding of the interplay between angiogenic and immune response pathways within tumors. In the present study, the GL261 glioma mouse model was used to determine the effects of antiangiogenic treatment in an immune-competent host. Following weekly systemic administration of aflibercept, an inhibitor of vascular endothelial growth factor, tumor volume was assessed by magnetic resonance imaging and changes to the tumor microenvironment were determined. Treatment with aflibercept resulted in reduced tumor burden and increased survival compared with controls. Additionally, decreased vascular permeability and preservation of the integrity of tight junction proteins were observed. Treated tumors also displayed hallmarks of anti-angiogenic evasion, including marked upregulation of vascular endothelial growth factor expression and increased tumor invasiveness. Aflibercept was then administered in combination with a picornavirus-based antitumor vaccine and tumor progression was evaluated. This combination therapy significantly delayed tumor progression and extended survival beyond that observed for either therapy alone. As such, this work demonstrates the efficacy of combined antiangiogenic and immunotherapy approaches for treating established gliomas and provides a foundation for further evaluation of the effects of antiangiogenic therapy in the context of endogenous or vaccine-induced inflammatory responses.

The role of ICAM-2 in neuroblastoma

The role of immunoglobulin superfamily cell adhesion molecules (CAMs) in facilitating immune responses in normal and tumor cells is well established. Cell adhesion molecule-1 (CADM1), for example, suppresses development of mouse mammary tumor cell metastasis by interacting with CD8+ T cells in immune-competent hosts [1]. Similarly, co-expression of Intercellular Adhesion Molecule-2 (ICAM-2) and chemokine C-X-C motif ligand 17 (CXCL17) elicits anti-tumor immune responses and suppresses tumor growth [2]. Although controversial, current literature suggests that proteins of the immunoglobulin superfamily CAMs also have functions that may well be distinct from their roles in mediating immune responses. Junctional adhesion molecule-A (JAM-A), for example, negatively regulates breast cancer cell invasiveness by disrupting tight junctions [3]; and a member of the B7 family of the immunoglobulin superfamily proteins, B7-H3, impairs osteogenic differentiation in vitro and in vivo [4]. Our lab demonstrated that ICAM-2 inhibited the development of disseminated neuroblastoma tumors in a preclinical model of metastatic neuroblastoma [5-7]. This inhibition depended on the interaction of ICAM-2 with the actin cytoskeletal linker protein α-actinin, an interaction that inhibited cell motility [7]. Ectopic expression of ICAM-2 did not affect the tumorigenic potential of neuroblastoma cells [7]. Importantly, immunohistochemical analyses of primary neuroblastoma tumor specimens demonstrated that neuroblastoma cells expressing ICAM-2 are phenotypically and histologically those recognized clinically to have limited metastatic potential [5]. Since metastatic disease is responsible for >90% of cancer-related deaths for multiple types of solid tumors, we suggest that elucidation of the molecular mechanism by which ICAM-2 suppresses the metastatic potential of neuroblastoma cells would identify proteins or pathways that might be exploited therapeutically to prevent metastatic disease progression. In normal tissues ICAM-2 is expressed predominantly by neovascular endothelial cells, and at lower levels by established vasculature and some leukocytes. The 202 amino acids comprising its extracellular domain mediate binding of ICAM-2 on endothelial cells to β2-integrins on the surface of leukocytes, to facilitate migration of neutrophils through the vascular endothelium as a component of immune reactions [8]. In neuroblastoma cells ICAM-2 inhibits cell motility independent of immune response, as we observed this inhibition in wound healing and modified Boyden chamber assays in vitro, assays that clearly lack an immune component [5,7]. In silico modeling indicated that ICAM-2 with mutations in the proposed α-actinin binding domain had a more ‘closed’ configuration than the wild type protein, and predicted that these ICAM-2 mutants would not interact with α-actinin [7]. Co-immunoprecipitation experiments confirmed in silico predictions [7]. In support of these findings, the interaction of ICAM-2 with α-actinin was essential to inhibit development of disseminated tumors in vivo [7]. As mentioned, ICAM-2 did not affect tumorigenic potential, and unpublished data showed no effect of ICAM-2 on expression of epithelial-mesenchymal transition (EMT) or stemness markers. Although we regard the inhibition of tumor cell motility by ICAM-2 in vitro to be compelling evidence that ICAM-2 participates in cell functions distinct from immune responses, we acknowledge the complex nature of metastatic tumor progression and propose that our data suggest multiple hypotheses with respect to the mechanism by which ICAM-2-inhibits neuroblastoma cell motility. The first hypothesis is that the intracellular interaction of ICAM-2 with α-actinin initiates ‘inside out’ signaling, and causes conformational changes in the extracellular domain of ICAM-2 that facilitate (or inhibit) interactions with specific extracellular matrix proteins that play an integral role in cell motility. The second is that the interaction of ICAM-2 with α-actinin alters the conformation of α-actinin to facilitate (or inhibit) the association of this actin cytoskeletal protein with alternative binding partners, with ICAM-2 acting as an ‘activator’ (or inhibitor) of α-actinin rather than as a membrane anchor protein. The third hypothesis, based on unpublished microarray data, is that ICAM-2 expression indirectly up-regulates protein tyrosine phosphatases (PTPs) involved in the formation and maintenance of focal adhesions such as focal adhesion kinase (FAK), Src, or Rac and Rho GTPases, each of which has crucial role in tumor cell motility. Our data show that ICAM-2 inhibits tumor cell motility and suppresses the metastatic potential of neuroblastoma cells. We propose that elucidating molecular events associated with ICAM-2 expression will identify key protein interactions that regulate the metastatic process in this cell type.

P81: Recovery of distinct T cell subsets under severe lymphopenic conditions in hemoblastosis patients

Numerous studies have shown that high-dose chemotherapy and autologous hematopoietic stem cell transplantation (AHSCT) led to a profound and long-lasting state of immunodeficiency characterized by persisting low levels of T cells in hemoblastosis patients. Well-timed T-cell reconstitution is crucial for early restoration of anti-infectious and anti- tumor immune response. Lymphocyte recovery is mediated through the two main mechanisms – a homeostatic proliferation of T cells and generation of new naive T cells via thymopoiesis. It is known, that homeostatic proliferation is important for the restoration of T cell count in immune competent host during the 1st year following AHSCT. Thymus begins to fill up T cell repertoire approximately from the 6th month following AHSCT. We have investigated dynamics of CD4+FOXP3+ Treg recovery following AHSCT and possible relationship between Tregs and clinical outcomes since the suppressive activity of Tregs under lymphopenic conditions may influence on peripheral expansion of T cells. Thymic activity following AHSCT has been evaluated by measuring amounts of CD4+ CD45RA+CD31+ naive T cells, i.e. “recent thymic emigrants” (RTEs).109 patients with non-Hodgkin’s lymphomas, Hodgkin’s lymphoma and multiple myeloma underwent AHSCT in 2009–2014. The content of circulating CD4+FOXP3+ Tregs and CD4+CD45RA+CD31+ T cells was evaluated using flow cytometry before AHSCT, at the day of engraftment, and following 6 and 12 months. Pre-transplant count of CD4+FOXP3+ Tregs was significantly higher compared to healthy controls (5.4 ± 2.9 vs 3.8 ± 1.9%; pU = 0.011; here and below data presented as Mean ± SD). Percentage of Tregs restored rapidly and reached initially high level at the time of engraftment, and then subsequently decreased within a year until it lowered to healthy donors‘ values. CD4+FOXP3+ Tregs at the time of engraftment were increased in patients with relapse or progression of disease within 6 and 12 months following AHSCT compared to non-relapsed patients (11.0 ± 6.1 vs 6.2 ± 3.0%; pU = 0.016, and 10.1 ± 5.2 vs 6.1 ± 3.8%; pU = 0.008). Pre-transplant count of CD4+CD45RA+CD31+ T cells was significantly lower compared to healthy controls (17.1 ± 11.4 vs 30.3 ± 11.2%, pU = 0.0005) and did not reach donors‘ values following 12 month (23.1 ± 13.5%, pU = 0.032). Relapsed patients had the same quantity of RTEs as the patients with remission within the 1st year following AHSCT. There was no any significant association between RTEs and Tregs counts. Surprisingly, we have found high levels of circulating CD4+CD45RA- T cells co-expressing CD31 molecule in patients before AHSCT, since this molecule is infrequent on memory subsets in healthy controls (20.7 ± 12.0 vs 8.2 ± 2.1%,pU Our data of Tregs reconstitution may confirm the earlier assumption that the presence of Tregs during the period of immune recovery preserves optimal T cell receptors diversity. However, the excess of these cells leads to the inhibition of proliferative activity and immune response and is associated with early relapse. Conversely, relatively slow recovery of RTEs determines theirlack of influence on survival within the 1st post-transplant year. The biological role and the way of appearance of CD31 molecule on T cell memory subset (CD4+CD45RA- and/or CD4+CD45RO+) still remain unclear. Further studies are required to enlighten the role of CD31+ memory T cells on lymphoproliferative disorders pathogenesis.

Abstract 469: mPGES1 deletion and the mechanisms of tumor growth suppression

Abstract The pro-tumor contributions of prostaglandin (PG) E2 are established, as is the clinical efficacy of pharmacological inhibition of COX-2, the rate-limiting enzyme in PGE2 synthesis. In addition to the desired suppression of tumor PGE2, collateral loss of endothelial COX-2-derived PGI2 imposes a cardiovascular hazard that limits clinical use of COX-2 inhibitors. mPGES1, the terminal enzyme in the PGE2 synthesis pathway, is an alternative target to interrupt COX-2-driven events in tumors without elevating cardiovascular risk. We engineered mice transgenic for an activated HER2/neu oncogene to lack mPGES1 globally (mPGES1 KOgl) or only in mammary epithelial cells (mPGES1 KOepi). Abdominal mammary glands from wild type (WT), heterozygous (het), mPGES1 KOgl, and mPGES1 KOepi mice were harvested at 22 weeks of age, paraffin embedded, H&E stained and scanned for whole slide imaging (WSI). Tumor multiplicity was calculated based on the number of lesions on WSI. Adjacent sections were immunostained for Ki67, caspase 3, Factor VIII, CD3, and F4/80, to assess proliferation, apoptosis, angiogenesis, and tumor lymphocyte and macrophage infiltration. Tumor multiplicity was significantly higher in WT compared to either mPGES1 KOgl or mPGES1 KOepi mice. Concordantly, shRNA knock down (KD) of mPGES1 in mammary tumor cells dramatically suppressed their growth as orthotopic tumors in syngeneic immune competent WT hosts: 4 weeks after the injections, 6 out of 6 non-target shRNA control tumors grew successfully while 6 out of 10 mPGES1 KD cells failed to grow. In separate experiments, 14 of 14 non-target shRNA cells formed tumors after 2 weeks while 12 out of 18 mPGES1 KD tumors failed. By immunohistochemistry, no difference was observed in proliferation and apoptosis markers, as well as in the number of tumor infiltrating macrophages and lymphocytes, between WT, mPGES1 KOgl, and mPGES1 KOepi tumors. In contrast to our previous study in mice lacking epithelial COX-2 (COX-2 KOepi), vascularization was not different between mPEGS-1 KOepi and WT tumors suggesting divergence in the anti-tumorigenic mechanisms in mPEGS-1 KOepi and COX-2 KOepi tumors. By flow cytometry there was no difference in infiltrating macrophage and lymphocyte density or functional phenotypes, between WT, mPGES1 KOepi, and mPGES1 KD orthotopic tumors. Similarly, no difference was observed in the number of macrophages and myeloid derived suppressor cells in spleens from mPGES1 sufficient and mPGES1 KD orthotopic tumor bearing mice. In contrast, decreased co-inhibitory molecule PD-1 expressing CD4+ and CD8+ lymphocyte numbers were evident in draining lymph nodes in mice injected with mPGES1 KD cells compared to WT. This finding indicates that the absence of mPGES1-derived PGE2 renders tumor cells less efficient in creating and maintaining an immunosuppressive tumor microenvironment and that targeting mPGES1 may enhance anti-tumor immune mechanisms. Citation Format: Nune Markosyan, Andrew Rech, Robert H. Vonderheide, Garret A. FitzGerald, Emer M. Smyth. mPGES1 deletion and the mechanisms of tumor growth suppression. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 469. doi:10.1158/1538-7445.AM2015-469

The 13th International Workshops on Opportunistic Protists (IWOP13).

The 13th International Workshops on Opportunistic Protists (IWOP-13) was held November 13-15, 2014 in Seville, Spain. The objectives of the IWOP meetings are to: (1) serve as a forum for exchange of new information among active researchers concerning the basic biology, molecular genetics, immunology, biochemistry, pathogenesis, drug development, therapy, and epidemiology of these immunodeficiency-associated pathogenic eukaryotic microorganisms that are seen in patients with AIDS and; (2) to foster the entry of new and young investigators into these underserved research areas. The IWOP meeting focuses on opportunistic protists; e.g. the free-living amoebae, Pneumocystis, Cryptosporidium, Toxoplasma, the Microsporidia, and kinetoplastid flagellates. This conference represents the major conference which brings together research groups working on these opportunistic pathogens. Progress has been achieved on understanding the biology of these pathogenic organisms, their involvement in disease causation in both immune deficient and immune competent hosts and is providing important insights into these emerging and reemerging pathogens. A continuing concern of the participants is the ongoing loss of scientific expertise and diversity in this research community. This decline is due to the small size of these research communities and an ongoing lack of understanding by the broader scientific community of the challenges and limitations faced by researchers working on these organisms, which makes these research communities very sensitive to declines in research funding.

Lister strain vaccinia virus with thymidine kinase gene deletion is a tractable platform for development of a new generation of oncolytic virus.

Vaccinia virus (VV) has many attractive characteristics as a potential cancer therapeutic. There are several strains of VV. The nonvaccine strain Western Reserve VV with deletion of both the thymidine kinase and the viral growth factor genes (known as WRDD) has been reported as the most potent tumor-targeted oncolytic VV. Other strains, such as the European vaccine Lister strain, are largely untested. This study evaluated the antitumor potency and biodistribution of different VV strains using in vitro and in vivo models of cancer. Lister strain virus with thymidine kinase gene deletion (VVΔTK) demonstrated superior antitumor potency and cancer-selective replication in vitro and in vivo, compared with WRDD, especially in human cancer cell lines and immune-competent hosts. Further investigation of functional mechanisms revealed that Lister VVΔTK presented favorable viral biodistribution within the tumors, with lower levels of proinflammatory cytokines compared with WRDD, suggesting that Lister strain may induce a diminished host inflammatory response. This study indicates that the Lister strain VVΔTK may be a particularly promising VV strain for the development of the next generation of tumor-targeted oncolytic therapeutics.

Interleukin-17D Mediates Tumor Rejection through Recruitment of Natural Killer Cells

The process of cancer immunoediting generates a repertoire of cancer cells that can persist in immune-competent hosts. In its most complex form, this process begins with the elimination of highly immunogenic unedited tumor cells followed by the escape of less immunogenic edited cells. Although edited tumors can release immunosuppressive factors, it is unknown whether unedited tumors produce cytokines that enhance antitumor function. Utilizing gene microarray analysis, we found the cytokine interleukin 17D (IL-17D) was highly expressed in certain unedited tumors but not in edited mouse tumor cell lines. Moreover, forced expression of IL-17D in edited tumor cells induced rejection by stimulating MCP-1 production from tumor endothelial cells, leading to the recruitment of natural killer (NK) cells. NK cells promoted M1 macrophage development and adaptive immune responses. IL-17D expression was also decreased in certain high-grade and metastatic human tumors, suggesting that it can be targeted for tumor immune therapy.

Where are we with Wolbachia and doxycycline: An in-depth review of the current state of our knowledge

Dirofilaria immitis, the cause of canine and feline heartworm disease, was the first filarial nematode described to harbour the bacterial endosymbiont Wolbachia. This ground-breaking discovery has led to intense research aimed at unravelling the nature of the endosymbiotic relationship; genomic studies have revealed how the bacteria may interact with the parasite and help explain why each is so dependent on the other. Analysis of the immune response to these bacteria may elucidate the mechanisms through which filarial parasites are able to survive for long periods of time in otherwise immune-competent hosts. Finally, studies aimed at the removal of the bacteria using specific antibiotic treatment in infected hosts is leading to the development of novel approaches for interrupting the transmission cycle and for the treatment and control of heartworm disease.

Tumor-derived inducible heat-shock protein 70 (HSP70) is an essential component of anti-tumor immunity

The anti-apoptotic function and tumor-associated expression of heat-shock protein 70 (HSP70) is consistent with HSP70 functioning as a survival factor to promote tumorigenesis. However, its immunomodulatory activities to induce anti-tumor immunity predict the suppression of tumor growth. Using the Hsp70.1/3(-/-)(Hsp70(-/-)) mouse model, we observed that tumor-derived HSP70 was neither required for cellular transformation nor for in vivo tumor growth. Hsp70(-/-)murine embryonic fibroblasts (MEFs) were transformed by E1A/Ras and generated tumors in immunodeficient hosts as efficiently as wild-type (WT) transformants. Comparison of Bcr-Abl-mediated transformation of WT and Hsp70(-/-)bone marrow and progression of B-cell leukemogenesis in vivo revealed no differences in disease onset or survival rates, and Eμ-Myc-driven lymphoma in Hsp70(-/-)mice was phenotypically indistinguishable from that in WT Eμ-Myc mice. However, Hsp70(-/-)E1A/Ras MEFs generated significantly larger tumors than their WT counterparts in C57BL/6 J immune-competent hosts. Concurrent with this was a reduction in intra-tumoral infiltration of innate and adaptive immune cells, including macrophages and CD8(+) T cells. Evaluation of several potential mechanisms revealed an HSP70-chemokine-like activity to promote cellular migration. These observations support a role for tumor-derived HSP70 in facilitating anti-tumor immunity to limit tumor growth and highlight the potential consequences of anti-HSP70 therapy as an efficacious anti-cancer strategy.

Neoadjuvant Chemotherapy for Localized Pancreatic Cancer: Too Little or Too Long?

For a ‘‘systemic’’ disease such as pancreatic cancer, neoadjuvant chemotherapy is a logical approach that thus far has been limited largely by a lack of effective systemic agents. For example, when considering treatment sequencing for patients with resectable disease, the concept of offering a treatment with an objective response rate of less than 10 % (as for gemcitabine alone) has been unacceptable to most clinicians. (Chemo)radiation has therefore been the backbone of most neoadjuvant approaches, with the goal of improving local/regional disease control (preventing local recurrence) and avoiding operation in those with progressive disease found on post-treatment, preoperative restaging. A few neoadjuvant studies to date have included relatively short courses of chemotherapy—without any obvious improvement in outcomes compared with chemoradiation alone. To further complicate treatment sequencing controversy, surgeons have cautioned that many patients (and referring physicians) want their tumors resected tomorrow and can find a surgeon who will accommodate them. Concerns about increased perioperative morbidity (following neoadjuvant therapy) still exist but have been allayed by several studies, including a recent NSQIP analysis. Importantly, an ‘‘effective’’ neoadjuvant chemotherapy regimen could theoretically improve longterm outcomes by successfully treating (possibly even eradicating) micrometastatic disease in the setting of an immune-competent host who is not attempting to recover from a large operation. The duration of neoadjuvant chemotherapy is a critical variable. If the right drug is selected for the right patient, treatment will need to be long enough to effectively treat micrometastatic disease. If the wrong drug(s) is selected (we only learn this in retrospect when restaging is performed), treatment should be short enough to avoid losing a window of resectability due to local disease progression. This is an important consideration in patients with resectable and borderline resectable disease (in contrast to those patients with locally advanced pancreatic cancer—thus the importance of accurate pretreatment staging). At present, there is little evidence to support any specific neoadjuvant treatment duration, although opinions exist that are based largely on personal experience, extrapolation from other solid tumor sites, and translational laboratory science. Rose and coauthors from Virginia Mason Medical Center are to be commended for demonstrating that patients are willing and able to undergo almost 6 months (on average) of neoadjuvant chemotherapy with good results, both shortterm and long-term . To what degree this was the result of the well-known experience and talent of this multidisciplinary group will be reflected in the ability of others to duplicate these results. Although patients were not treated on a clinical trial, they were maintained in a prospective registry, which is a practice that should be encouraged. The major strength of the study was the relatively large (n = 64) and homogeneously staged patient population, all radiographically ‘‘borderline resectable’’ by the SSO/AHPBA consensus criteria and with no evidence of metastatic disease by pretreatment staging that included laparoscopy with peritoneal washings—which did introduce an added selection bias as most such reports have not included laparoscopic staging. The outcomes in this study are not entirely due to increased ‘‘selection’’ provided by the extended duration of therapy. There clearly was a treatment effect demonstrated at the site of the primary tumor. Almost half of patients underwent resection of the primary, with R0 Society of Surgical Oncology 2014