Subversion of the Complement System by Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an opportunistic pathogen heavily implicated in chronic diseases. Immunocompromised patients that become infected with P. aeruginosa usually are afflicted with a lifelong chronic infection, leading to worsened patient outcomes. The complement system is an integral piece of the first line of defense against invading microorganisms. Gram-negative bacteria are thought to be generally susceptible to attack from complement; however, P. aeruginosa can be an exception, with certain strains being serum resistant. Various molecular mechanisms have been described that confer P. aeruginosa unique resistance to numerous aspects of the complement response. In this review, we summarize the current published literature regarding the interactions of P. aeruginosa and complement, as well as the mechanisms used by P. aeruginosa to exploit various complement deficiencies and the strategies used to disrupt or hijack normal complement activities.

Immune Response Modulation by Pseudomonas aeruginosa Persister Cells.

Bacterial persister cells-a metabolically dormant subpopulation tolerant to antimicrobials-contribute to chronic infections and are thought to evade host immunity. In this work, we studied the ability of Pseudomonas aeruginosa persister cells to withstand host innate immunity. We found that persister cells resist MAC-mediated killing by the complement system despite being bound by complement protein C3b at levels similar to regular vegetative cells, in part due to reduced bound C5b, and are engulfed at a lower rate (10- to 100-fold), even following opsonization. Once engulfed, persister cells resist killing and, contrary to regular vegetative cells which induce a M1 favored (CD80+/CD86+/CD206-, high levels of CXCL-8, IL-6, and TNF-α) macrophage polarization, they initially induce a M2 favored macrophage polarization (CD80+/CD86+/CD206+, high levels of IL-10, and intermediate levels of CXCL-8, IL-6, and TNF-α), which is skewed toward M1 favored polarization (high levels of CXCL-8 and IL-6, lower levels of IL-10) by 24 h of infection, once persister cells awaken. Overall, our findings further establish the ability of persister cells to evade the innate host response and to contribute chronic infections. IMPORTANCE Bacterial cells have a subpopulation-persister cells-that have a low metabolism. Persister cells survive antimicrobial treatment and can regrow to cause chronic and recurrent infections. Currently little is known as to whether the human immune system recognizes and responds to the presence of persister cells. In this work, we studied the ability of persister cells from Pseudomonas aeruginosa to resist the host defense system (innate immunity). We found that this subpopulation is recognized by the defense system, but it is not killed. The lack of killing likely stems from hindering the immune response regulation, resulting in a failure to distinguish whether a pathogen is present. Findings from this work increase the overall knowledge as to how chronic infections are resilient.

Immune response modulation by Pseudomonas aeruginosa persister cells.

Bacterial persister cells - a metabolically dormant subpopulation tolerant to antimicrobials - contribute to chronic infections and are thought to evade host immunity. In this work, we studied the ability of Pseudomonas aeruginosa persister cells to withstand host innate immunity. We found that persister cells resist MAC-mediated killing by the complement system despite being bound by complement protein C3b at levels similar to regular vegetative cells, in part due to reduced bound C5b - and are engulfed at a lower rate (10-100 fold), even following opsonization. Once engulfed, persister cells resist killing and, contrary to regular vegetative cells which induce a M1 favored (CD80+/CD86+/CD206-, high levels of CXCL-8, IL-6, and TNF-α) macrophage polarization, they initially induce a M2 favored macrophage polarization (CD80+/CD86+/CD206+, high levels of IL-10, and intermediate levels of CXCL-8, IL-6, and TNF-α), which is skewed towards M1 favored polarization (high levels of CXCL-8 and IL-6, lower levels of IL-10) by 24 hours of infection, once persister cells awaken. Overall, our findings further establish the ability of persister cells to evade the innate host response and to contribute chronic infections.

A comparative proteomics analysis of the egg secretions of three major schistosome species.

Morbidity associated with hepatic and urogenital schistosomiasis stems primarily from the host immune response directed against schistosome eggs. When eggs become entrapped in host tissues, the development of fibrotic plaques drives downstream pathology. These events occur due to the antigenic nature of egg excretory/secretory products (ESPs). Both Schistosoma mansoni and S. japonicum ESPs have been shown to interact with several cell populations in the host liver including hepatocytes, macrophages, and hepatic stellate cells, with both immunomodulatory and pathological consequences. Several protein components of the ESPs of S. mansoni and S. japonicum eggs have been characterised; however, studies into the collective contents of schistosome egg ESPs are lacking. Utilising shotgun mass spectrometry and an array of in silico analyses, we identified 266, 90 and 50 proteins within the S. mansoni, S. japonicum and S. haematobium egg secretomes respectively. We identified numerous proteins with already established immunomodulatory activities, vaccine candidates and vesicle markers. Relatively few common orthologues within the ESPs were identified by BLAST, indicating that the three egg secretomes differ in content significantly. Having a clearer understanding of these components may lead to the identification of new proteins with uncharacterised immunomodulatory potential or pathological relevance. This will enhance our understanding of host-parasite interactions, particularly those occurring during chronic schistosomiasis, and pave the way towards novel therapeutics and vaccines.

Publisher Correction: Programmable multistage drug delivery to lymph nodes.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Programmable multistage drug delivery to lymph nodes.

Therapeutic delivery selectively to lymph nodes has the potential to address a variety of unmet clinical needs. However, owing to the unique structure of the lymphatics and the size-restrictive nature of the lymph node reticular network, delivering cargo to specific cells in the lymph node cortex and paracortex is difficult. Here, we describe a delivery system to overcome lymphatic and intra-lymph node transport barriers by combining nanoparticles that are rapidly conveyed to draining lymph nodes after administration in peripheral tissues with programmable degradable linkers. This platform enables the controlled release of intra-lymph-mobile small-molecular cargo, which can reach vastly more immune cells throughout the lymph node than either the particles or free compounds alone. The release rate can be programmed, allowing access to different lymph node structures and therefore specific lymphocyte subpopulations. We are thus able to alter the subtypes of drugged lymph node cells to improve immunotherapeutic effects.

The effect of isokinetic dynamometer deceleration phase on maximum ankle joint range of motion and plantar flexor mechanical properties tested at different angular velocities.

During range of motion (max-ROM) tests performed on an isokinetic dynamometer, the mechanical delay between the button press (by the participant to signal their max-ROM) and the stopping of joint rotation resulting from system inertia induces errors in both max-ROM and maximum passive joint moment. The present study aimed to quantify these errors by comparing data when max-ROM was obtained from the joint position data, as usual (max-ROMPOS), to data where max-ROM was defined as the first point of dynamometer arm deceleration (max-ROMACC). Fifteen participants performed isokinetic ankle joint max-ROM tests at 5, 30 and 60° s-1. Max-ROM, peak passive joint moment, end-range musculo-articular (MAC) stiffness and area under the joint moment-position curve were calculated. Greater max-ROM was observed in max-ROMPOS than max-ROMACC (P < 0.01) at 5 (0.2 ±â€¯0.15%), 30 (1.8 ±â€¯1.0%) and 60° s-1 (5.9 ±â€¯2.3%), with the greatest error at the fastest velocity. Peak passive moment was greater and end-range MAC stiffness lower in max-ROMPOS than in max-ROMACC only at 60° s-1 (P < 0.01), whilst greater elastic energy storage was found at all velocities. Max-ROM and peak passive moment are affected by the delay between button press and eventual stopping of joint rotation in an angular velocity-dependent manner. This affects other variables calculated from the data. When high data accuracy is required, especially at fast joint rotation velocities (≥30° s-1), max-ROM (and associated measures calculated from joint moment data) should be taken at the point of first change in acceleration rather than at the dynamometer's ultimate joint position.

The NIH-NIAID Schistosomiasis Resource Center at the Biomedical Research Institute: Molecular Redux.

Schistosomiasis remains a health burden in many parts of the world. The complex life cycle of Schistosoma parasites and the economic and societal conditions present in endemic areas make the prospect of eradication unlikely in the foreseeable future. Continued and vigorous research efforts must therefore be directed at this disease, particularly since only a single World Health Organization (WHO)-approved drug is available for treatment. The National Institutes of Health (NIH)-National Institute of Allergy and Infectious Diseases (NIAID) Schistosomiasis Resource Center (SRC) at the Biomedical Research Institute provides investigators with the critical raw materials needed to carry out this important research. The SRC makes available, free of charge (including international shipping costs), not only infected host organisms but also a wide array of molecular reagents derived from all life stages of each of the three main human schistosome parasites. As the field of schistosomiasis research rapidly advances, it is likely to become increasingly reliant on omics, transgenics, epigenetics, and microbiome-related research approaches. The SRC has and will continue to monitor and contribute to advances in the field in order to support these research efforts with an expanding array of molecular reagents. In addition to providing investigators with source materials, the SRC has expanded its educational mission by offering a molecular techniques training course and has recently organized an international schistosomiasis-focused meeting. This review provides an overview of the materials and services that are available at the SRC for schistosomiasis researchers, with a focus on updates that have occurred since the original overview in 2008.

Promising oncolytic agents for metastatic breast cancer treatment.

New therapies for metastatic breast cancer patients are urgently needed. The long-term survival rates remain unacceptably low for patients with recurrent disease or disseminated metastases. In addition, existing therapies often cause a variety of debilitating side effects that severely impact quality of life. Oncolytic viruses constitute a developing therapeutic modality in which interest continues to build due to their ability to spare normal tissue while selectively destroying tumor cells. A number of different viruses have been used to develop oncolytic agents for breast cancer, including herpes simplex virus, adenovirus, vaccinia virus, measles virus, reovirus, and others. In general, clinical trials for several cancers have demonstrated excellent safety records and evidence of efficacy. However, the impressive tumor responses often observed in preclinical studies have yet to be realized in the clinic. In order for the promise of oncolytic virotherapy to be fully realized for breast cancer patients, effectiveness must be demonstrated in metastatic disease. This review provides a summary of oncolytic virotherapy strategies being developed to target metastatic breast cancer.

Histone deacetylase inhibitors improve the replication of oncolytic herpes simplex virus in breast cancer cells.

New therapies are needed for metastatic breast cancer patients. Oncolytic herpes simplex virus (oHSV) is an exciting therapy being developed for use against aggressive tumors and established metastases. Although oHSV have been demonstrated safe in clinical trials, a lack of sufficient potency has slowed the clinical application of this approach. We utilized histone deacetylase (HDAC) inhibitors, which have been noted to impair the innate antiviral response and improve gene transcription from viral vectors, to enhance the replication of oHSV in breast cancer cells. A panel of chemically diverse HDAC inhibitors were tested at three different doses (<,  = , and >LD50) for their ability to modulate the replication of oHSV in breast cancer cells. Several of the tested HDAC inhibitors enhanced oHSV replication at low multiplicity of infection (MOI) following pre-treatment of the metastatic breast cancer cell line MDA-MB-231 and the oHSV-resistant cell line 4T1, but not in the normal breast epithelial cell line MCF10A. Inhibitors of class I HDACs, including pan-selective compounds, were more effective for increasing oHSV replication compared to inhibitors that selectively target class II HDACs. These studies demonstrate that select HDAC inhibitors increase oHSV replication in breast cancer cells and provides support for pre-clinical evaluation of this combination strategy.

Evaluation of the safety and biodistribution of M032, an attenuated herpes simplex virus type 1 expressing hIL-12, after intracerebral administration to aotus nonhuman primates.

Herpes simplex virus type 1 (HSV-1) mutants lacking the γ(1)34.5 neurovirulence loci are promising agents for treating malignant glioma. Arming oncolytic HSV-1 to express immunostimulatory genes may potentiate therapeutic efficacy. We have previously demonstrated improved preclinical efficacy, biodistribution, and safety of M002, a γ(1)34.5-deleted HSV-1 engineered to express murine IL-12. Herein, we describe the safety and biodistribution of M032, a γ(1)34.5-deleted HSV-1 virus that expresses human IL-12 after intracerebral administration to nonhuman primates, Aotus nancymae. Cohorts were administered vehicle, 10(6), or 10(8) pfu of M032 on day 1 and subjected to detailed clinical observations performed serially over a 92-day trial. Animals were sacrificed on days 3, 31, and 91 for detailed histopathologic assessments of all organs and to isolate and quantify virus in all organs. With the possible exception of one animal euthanized on day 16, neither adverse clinical signs nor sex- or dose-related differences were attributed to M032. Elevated white blood cell and neutrophil counts were observed in virus-injected groups on day 3, but no other significant changes were noted in clinical chemistry or coagulation parameters. Minimal to mild inflammation and fibrosis detected, primarily in meningeal tissues, in M032-injected animals on days 3 and 31 had mostly resolved by day 91. The highest viral DNA levels were detected at the injection site and motor cortex on day 3 but decreased in central nervous system tissues over time. These data demonstrate the requisite safety of intracerebral M032 administration for consideration as a therapeutic for treating malignant brain tumors.

Popliteus strain with concurrent deltoid ligament sprain in an elite soccer athlete: a case report.

STUDY DESIGN: Case Report (Differential diagnosis). BACKGROUND AND PURPOSE: Differential diagnosis of knee pathology after trauma may be difficult when diagnosing an isolated popliteus strain and concurrent medial deltoid ligament sprain. Upon a thorough search of the published literature, the authors found no reports delineating a popliteus strain in professional soccer in the United States. The joints most affected by injury in soccer players are the knee and ankle joints. The purpose of this case report is to describe the presentation of and difficulties encountered in diagnosing a popliteus strain in a Major League Soccer athlete. CASE DESCRIPTION: During an in-season away game, an outside defender was slide-tackled from behind when his right shank was caught in an externally rotated position underneath himself and the opposing player. The initial point of contact was made to the proximal third of the posterior right shank with an anteromedially directed force. The medial longitudinal arch of the foot was forced into a more midfoot pronated position and the subtalar joint was forced into eversion. DIAGNOSIS: The athlete was diagnosed with a moderate strain of the right popliteus muscle with a concurrent medial deltoid ligament sprain of the right ankle. This mechanism of injury, pain with passive knee flexion and internal rotation during McMurray's test, pain with Garrick's Test and magnetic resonance imaging (MRI) study confirmed the diagnosis. The athlete returned to full ninety-minute game participation after an intensive 15-day rehabilitation program. DISCUSSION: This case is unique because the injury manifested itself at multiple joints and specifically involved the popliteus muscle. The mechanism of injury can be associated with many other soft tissue injuries to the knee, and thus, may not lead the clinician initially to consider the diagnosis of a popliteus strain. Diagnosis of this entity may be difficult due to the possible shared attachment of the popliteus muscle to the lateral meniscus, and the lack of available testing methods to assess damage to the popliteus muscle. LEVEL OF EVIDENCE: Level 5.

Expression of osteoprotegerin from a replicating adenovirus inhibits the progression of prostate cancer bone metastases in a murine model.

Metastatic involvement of the skeleton is a frequent consequence of advanced prostate cancer. These skeletal metastases cause a number of debilitating complications and are refractory to current treatments. New therapeutic options are being explored, including conditionally replicating adenoviruses (CRAds). CRAds are engineered to selectively replicate in and destroy tumor cells and can be 'armed' with exogenous transgenes for enhanced potency. We hypothesized that a CRAd armed with osteoprotegerin (OPG), an inhibitor of osteoclastogenesis, would inhibit the progression of prostate cancer bone metastases by directly lysing tumor cells and by reducing osteoclast activity. Although prostate cancer bone metastases are predominantly osteoblastic in nature, increased osteoclast activity is critical for the growth of these lesions. Ad5-Δ24-sOPG-Fc-RGD is a CRAd that carries a fusion of the ligand-binding domains of OPG and the Fc region of human IgG1 in place of the viral E3B genes. To circumvent low tumor cell expression of the native adenoviral receptor, an arginine-glycine-aspartic acid (RGD) peptide insertion within the viral fiber knob allows infection of cells expressing α(v) integrins. A 24-base pair deletion (Δ24) within viral E1A limits replication to cells with aberrant retinoblastoma cell cycle regulator/tumor suppressor expression. We have confirmed that Ad5-Δ24-sOPG-Fc-RGD replicates within and destroys prostate cancer cells and, in both murine and human coculture models, that infection of prostate cancer cells inhibits osteoclastogenesis in vitro. In a murine model, progression of advanced prostate cancer bone metastases was inhibited by treatment with Ad5-Δ24-sOPG-Fc-RGD but not by an unarmed control CRAd.

Preclinical evaluation of a genetically engineered herpes simplex virus expressing interleukin-12.

Herpes simplex virus 1 (HSV-1) mutants that lack the γ(1)34.5 gene are unable to replicate in the central nervous system but maintain replication competence in dividing cell populations, such as those found in brain tumors. We have previously demonstrated that a γ(1)34.5-deleted HSV-1 expressing murine interleukin-12 (IL-12; M002) prolonged survival of immunocompetent mice in intracranial models of brain tumors. We hypothesized that M002 would be suitable for use in clinical trials for patients with malignant glioma. To test this hypothesis, we (i) compared the efficacy of M002 to three other HSV-1 mutants, R3659, R8306, and G207, in murine models of brain tumors, (ii) examined the safety and biodistribution of M002 in the HSV-1-sensitive primate Aotus nancymae following intracerebral inoculation, and (iii) determined whether murine IL-12 produced by M002 was capable of activating primate lymphocytes. Results are summarized as follows: (i) M002 demonstrated superior antitumor activity in two different murine brain tumor models compared to three other genetically engineered HSV-1 mutants; (ii) no significant clinical or magnetic resonance imaging evidence of toxicity was observed following direct inoculation of M002 into the right frontal lobes of A. nancymae; (iii) there was no histopathologic evidence of disease in A. nancymae 1 month or 5.5 years following direct inoculation; and (iv) murine IL-12 produced by M002 activates A. nancymae lymphocytes in vitro. We conclude that the safety and preclinical efficacy of M002 warrants the advancement of a Δγ(1)34.5 virus expressing IL-12 to phase I clinical trials for patients with recurrent malignant glioma.

Preclinical evaluation of oncolytic δγ(1)34.5 herpes simplex virus expressing interleukin-12 for therapy of breast cancer brain metastases.

The metastasis of breast cancer to the brain and central nervous system (CNS) is a problem of increasing importance. As improving treatments continue to extend patient survival, the incidence of CNS metastases from breast cancer is on the rise. New treatments are needed, as current treatments are limited by deleterious side effects and are generally palliative. We have previously described an oncolytic herpes simplex virus (HSV), designated M002, which lacks both copies of the γ(1)34.5 neurovirulence gene and carries a murine interleukin 12 (IL-12) expression cassette, and have validated its antitumor efficacy in a variety of preclinical models of primary brain tumors. However, M002 has not been yet evaluated for use against metastatic brain tumors. Here, we demonstrate the following: both human breast cancer and murine mammary carcinoma cells support viral replication and IL-12 expression from M002; M002 replicates in and destroys breast cancer cells from a variety of histological subtypes, including "triple-negative" and HER2 overexpressing; M002 improves survival in an immunocompetent model more effectively than does a non-cytokine control virus. Thus, we conclude from this proof-of-principle study that a γ(1)34.5-deleted IL-12 expressing oncolytic HSV may be a potential new therapy for breast cancer brain metastases.

A simplified method for the generation of human osteoclasts in vitro.

Osteoclasts are large, multinucleated cells responsible for the resorption of mineralized bone matrix. These cells are critical players in the bone turnover involved in bone homeostasis. Osteoclast activity is connected to the establishment and expansion of skeletal metastases from a number of primary neoplasms. Thus, the formation and activation of osteoclasts is an area of research with many potential avenues for clinical translation. Past studies of osteoclast biology have utilized primary murine cells cultured in vitro. Recently, techniques have been described that involve the generation of osteoclasts from human precursor cells. However, these protocols are often time-consuming and insufficient for generating large numbers of osteoclasts. We therefore developed a simplified protocol by which human osteoclasts may be easily and reliably generated in large numbers in vitro. In this study, osteoclasts were differentiated from bone marrow cells that had been aliquotted and frozen. Cells were generated by culture with recombinant macrophage colony-stimulating factor (M-CSF) and receptor activator of NF-κB ligand (RANKL). Both human and murine RANKL were shown to efficiently generate osteoclasts, although higher concentrations of murine RANKL were required. Formation of osteoclasts was demonstrated qualitatively by tartrate-resistant acid phosphatase (TRAP) staining. These cells were fully functional, as confirmed by their ability to form resorption pits on cortical bone slices. Functional human osteoclasts can be difficult to generate in vitro by current protocols. We have demonstrated a simplified system for the generation of human osteoclasts in vitro that allows for large numbers of osteoclasts to be obtained from a single donor.

Conditionally replicating adenovirus expressing TIMP2 increases survival in a mouse model of disseminated ovarian cancer.

Ovarian cancer remains difficult to treat mainly due to presentation of the disease at an advanced stage. Conditionally-replicating adenoviruses (CRAds) are promising anti-cancer agents that selectively kill the tumor cells. The present study evaluated the efficacy of a novel CRAd (Ad5/3-CXCR4-TIMP2) containing the CXCR4 promoter for selective viral replication in cancer cells together with TIMP2 as a therapeutic transgene, targeting the matrix metalloproteases (MMPs) in a murine orthotopic model of disseminated ovarian cancer. An orthotopic model of ovarian cancer was established in athymic nude mice by intraperitonal injection of the human ovarian cancer cell line, SKOV3-Luc, expressing luciferase. Upon confirmation of peritoneal dissemination of the cells by non-invasive imaging, mice were randomly divided into four treatment groups: PBS, Ad-ΔE1-TIMP2, Ad5/3-CXCR4, and Ad5/3-CXCR4-TIMP2. All mice were imaged weekly to monitor tumor growth and were sacrificed upon reaching any of the predefined endpoints, including high tumor burden and significant weight loss along with clinical evidence of pain and distress. Survival analysis was performed using the Log-rank test. The median survival for the PBS cohort was 33 days; for Ad-ΔE1-TIMP2, 39 days; for Ad5/3-CXCR4, 52.5 days; and for Ad5/3-CXCR4-TIMP2, 63 days. The TIMP2-armed CRAd delayed tumor growth and significantly increased survival when compared to the unarmed CRAd. This therapeutic effect was confirmed to be mediated through inhibition of MMP9. Results of the in vivo study support the translational potential of Ad5/3-CXCR4-TIMP2 for treatment of human patients with advanced ovarian cancer.

Posttranscriptional control of type I interferon genes by KSRP in the innate immune response against viral infection.

Inherently unstable mRNAs contain AU-rich elements (AREs) in the 3' untranslated regions. Expression of ARE-containing type I interferon transcripts is robustly induced upon viral infection and rapidly shut off thereafter. Their transient accumulation is partly mediated through posttranscriptional regulation. Here we show that mouse embryonic fibroblasts derived from knockout mice deficient in KH-type splicing regulatory protein (KSRP), an RNA-binding protein required for ARE-mediated mRNA decay, produce higher levels of Ifna and Ifnb mRNAs in response to viral infection as a result of decreased mRNA decay. Functional analysis showed that KSRP is required for the decay of Ifna4 and Ifnb mRNAs by interaction with AREs. The increased IFN expression renders Ksrp(-)(/)(-) cells refractory to herpes simplex virus type 1 and vesicular stomatitis virus infection. These findings support a role of a posttranscriptional mechanism in the control of type I IFN expression and highlight the function of KSRP in innate immunity by negatively regulating IFN production.

Conditionally replicating adenovirus expressing TIMP2 for ovarian cancer therapy.

PURPOSE: Current treatments for ovarian cancer have limited therapeutic outcomes due to advanced stage of the disease at diagnosis. Among new therapies, conditionally replicating adenoviruses (CRAds), designed to selectively lyse cancer cells, hold promise. In clinical trials, CRAds exhibited limited efficacy thus far. Second-generation CRAds are being developed to express a therapeutic protein to enhance antitumor efficacy. One attractive target in the tumor microenvironment is the matrix metalloproteinases (MMPs) that degrade the extracellular matrix, and are upregulated in ovarian cancer. Tissue inhibitor of metalloproteinase 2 (TIMP2) is an endogenous inhibitor of MMPs. The present study developed and evaluated a novel CRAd (Ad5/3-CXCR4-TIMP2) for ovarian cancer therapy. EXPERIMENTAL DESIGN: A targeted CRAd, Ad5/3-CXCR4-TIMP2 was developed using the CXCR4 promoter for enhanced replication, and expressing the TIMP2 transgene. The efficacy of this armed CRAd was determined in both established human ovarian cancer cell lines and in primary ovarian tumor samples. RESULTS: Ad5/3-CXCR4-TIMP2 mediated expression of functional TIMP2, as demonstrated by the inhibition of MMP activity. In addition, arming with TIMP2 did not inhibit viral replication or oncolytic potency, as the TIMP2-armed viruses showed enhanced killing of cancer cells when compared to the unarmed viruses. We also examined viral replication in primary ovarian cancer tissues obtained from patients with stage III and IV ovarian cancer. In four of the five tumor samples, Ad5/3-CXCR4-TIMP2 revealed a 21- to 89-fold increase in replication when compared to the Ad5/3 virus. CONCLUSION: Results support the translational potential of Ad5/3-CXCR4-TIMP2 for treatment of patients with advanced ovarian cancer.