Clinical Experience and Recent Advances in the Development of Listeria-Based Tumor Immunotherapies.

The promise of tumor immunotherapy to significantly improve survival in patients who are refractory to long-standing therapies, such as chemotherapy and radiation, is now being realized. While immune checkpoint inhibitors that target PD-1 and CTLA-4 are leading the charge in clinical efficacy, there are a number of other promising tumor immunotherapies in advanced development such as Listeria-based vaccines. Due to its unique life cycle and ability to induce robust CTL responses, attenuated strains of Listeria monocytogenes (Lm) have been utilized as vaccine vectors targeting both infectious disease and cancer. In fact, preclinical studies in a multitude of cancer types have found Listeria-based vaccines to be highly effective at activating anti-tumor immunity and eradicating tumors. Several clinical trials have now recently reported their results, demonstrating promising efficacy against some cancers, and unique challenges. Development of the Lm-based immunotherapies continues with discovery of improved methods of attenuation, novel uses, and more effective combinatorial regimens. In this review, we provide a brief background of Listeria monocytogenes as a vaccine vector, discuss recent clinical experience with Listeria-based immunotherapies, and detail the advancements in development of improved Listeria-based vaccine platforms and in their utilization.

Complement as Prognostic Biomarker and Potential Therapeutic Target in Renal Cell Carcinoma.

Preclinical studies demonstrated that complement promotes tumor growth. Therefore, we sought to determine the best target for complement-based therapy among common human malignancies. High expression of 11 complement genes was linked to unfavorable prognosis in renal cell carcinoma. Complement protein expression or deposition was observed mainly in stroma, leukocytes, and tumor vasculature, corresponding to a role of complement in regulating the tumor microenvironment. Complement abundance in tumors correlated with a high nuclear grade. Complement genes clustered within an aggressive inflammatory subtype of renal cancer characterized by poor prognosis, markers of T cell dysfunction, and alternatively activated macrophages. Plasma levels of complement proteins correlated with response to immune checkpoint inhibitors. Corroborating human data, complement deficiencies and blockade reduced tumor growth by enhancing antitumor immunity and seemingly reducing angiogenesis in a mouse model of kidney cancer resistant to PD-1 blockade. Overall, this study implicates complement in the immune landscape of renal cell carcinoma, and notwithstanding cohort size and preclinical model limitations, the data suggest that tumors resistant to immune checkpoint inhibitors might be suitable targets for complement-based therapy.

Therapeutic Targeting of Vasculature in the Premetastatic and Metastatic Niches Reduces Lung Metastasis.

In the metastasis-targeted organs, angiogenesis is essential for the progression of dormant micrometastases to rapidly growing and clinically overt lesions. However, we observed changes suggesting angiogenic switching in the mouse lungs prior to arrival of tumor cells (i.e., in the premetastatic niche) in the models of breast carcinoma. This angiogenic switching appears to be caused by myeloid-derived suppressor cells recruited to the premetastatic lungs through complement C5a receptor 1 signaling. These myeloid cells are known to secrete several proangiogenic factors in tumors, including IL-1ß and matrix metalloproteinase-9, and we found upregulation of these genes in the premetastatic lungs. Blockade of C5a receptor 1 synergized with antiangiogenic Listeria monocytogenes-based vaccines to decrease the lung metastatic burden by reducing vascular density and improving antitumor immunity in the lungs. This was mediated even when growth of primary breast tumors was not affected by these treatments. This work provides initial evidence that angiogenesis contributes to the premetastatic niche in rapidly progressing cancers and that inhibiting this process through immunotherapy is beneficial for reducing or even preventing metastasis.

The Ribosomal Protein S19 Suppresses Antitumor Immune Responses via the Complement C5a Receptor 1.

Relatively little is known about factors that initiate immunosuppression in tumors and act at the interface between tumor cells and host cells. In this article, we report novel immunosuppressive properties of the ribosomal protein S19 (RPS19), which is upregulated in human breast and ovarian cancer cells and released from apoptotic tumor cells, whereupon it interacts with the complement C5a receptor 1 expressed on tumor infiltrating myeloid-derived suppressor cells. This interaction promotes tumor growth by facilitating recruitment of these cells to tumors. RPS19 also induces the production of immunosuppressive cytokines, including TGF-ß, by myeloid-derived suppressor cells in tumor-draining lymph nodes, leading to T cell responses skewed toward Th2 phenotypes. RPS19 promotes generation of regulatory T cells while reducing infiltration of CD8+ T cells into tumors. Reducing RPS19 in tumor cells or blocking the C5a receptor 1-RPS19 interaction decreases RPS19-mediated immunosuppression, impairs tumor growth, and delays the development of tumors in a transgenic model of breast cancer. This work provides initial preclinical evidence for targeting RPS19 for anticancer therapy enhancing antitumor T cell responses.

A One Health overview, facilitating advances in comparative medicine and translational research.

TABLE OF CONTENTS: A1 One health advances and successes in comparative medicine and translational researchCheryl StroudA2 Dendritic cell-targeted gorilla adenoviral vector for cancer vaccination for canine melanomaIgor Dmitriev, Elena Kashentseva, Jeffrey N. Bryan, David T. CurielA3 Viroimmunotherapy for malignant melanoma in the companion dog modelJeffrey N. Bryan, David Curiel, Igor Dmitriev, Elena Kashentseva, Hans Rindt, Carol Reinero, Carolyn J. HenryA4 Of mice and men (and dogs!): development of a commercially licensed xenogeneic DNA vaccine for companion animals with malignant melanomaPhilip J. BergmanA5 Successful immunotherapy with a recombinant HER2-expressing Listeria monocytogenes in dogs with spontaneous osteosarcoma paves the way for advances in pediatric osteosarcomaNicola J. Mason, Josephine S. Gnanandarajah, Julie B. Engiles, Falon Gray, Danielle Laughlin, Anita Gaurnier-Hausser, Anu Wallecha, Margie Huebner, Yvonne PatersonA6 Human clinical development of ADXS-HER2Daniel O'ConnorA7 Leveraging use of data for both human and veterinary benefitLaura S. TremlA8 Biologic replacement of the knee: innovations and early clinical resultsJames P. StannardA9 Mizzou BioJoint Center: a translational success storyJames L. CookA10 University and industry translational partnership: from the lab to commercializationMarc JacobsA11 Beyond docking: an evolutionarily guided OneHealth approach to drug discoveryGerald J. Wyckoff, Lee Likins, Ubadah Sabbagh, Andrew SkaffA12 Challenges and opportunities for data applications in animal health: from precision medicine to precision husbandryAmado S. GuloyA13 A cloud-based programmable platform for healthHarlen D. HaysA14 Comparative oncology: One Health in actionAmy K. LeBlancA15 Companion animal diseases bridge the translational gap for human neurodegenerative diseaseJoan R. Coates, Martin L. Katz, Leslie A. Lyons, Gayle C. Johnson, Gary S. Johnson, Dennis P. O'BrienA16 Duchenne muscular dystrophy gene therapyDongsheng DuanA17 Polycystic kidney disease: cellular mechanisms to emerging therapiesJames P. CalvetA18 The domestic cat as a large animal model for polycystic kidney diseaseLeslie A. Lyons, Barbara GandolfiA19 The support of basic and clinical research by the Polycystic Kidney Disease FoundationDavid A. BaronA20 Using naturally occurring large animal models of human disease to enable clinical translation: treatment of arthritis using autologous stromal vascular fraction in dogsMark L. WeissA21 Regulatory requirements regarding clinical use of human cells, tissues, and tissue-based productsDebra A. WebsterA22 Regenerative medicine approaches to Type 1 diabetes treatmentFrancis N. KaranuA23 The zoobiquity of canine diabetes mellitus, man's best friend is a friend indeed-islet transplantationEdward J. RobbA24 One Medicine: a development model for cellular therapy of diabetesRobert J. Harman.

Immunotherapy with a HER2-Targeting Listeria Induces HER2-Specific Immunity and Demonstrates Potential Therapeutic Effects in a Phase I Trial in Canine Osteosarcoma.

PURPOSE: Recombinant Listeria vaccines induce tumor-specific T-cell responses that eliminate established tumors and prevent metastatic disease in murine cancer models. We used dogs with HER2/neu(+) appendicular osteosarcoma, a well-recognized spontaneous model for pediatric osteosarcoma, to determine whether a highly attenuated, recombinant Listeria monocytogenes expressing a chimeric human HER2/neu fusion protein (ADXS31-164) could safely induce HER2/neu-specific immunity and prevent metastatic disease. EXPERIMENTAL DESIGN: Eighteen dogs that underwent limb amputation or salvage surgery and adjuvant chemotherapy were enrolled in a phase I dose escalation clinical trial and received either 2 × 10(8), 5 × 10(8), 1 × 10(9), or 3.3 × 10(9) CFU of ADXS31-164 intravenously every 3 weeks for 3 administrations. RESULTS: Only low-grade, transient toxicities were observed. ADXS31-164 broke peripheral tolerance and induced antigen-specific IFNγ responses against the intracellular domain of HER2/neu in 15 of 18 dogs within 6 months of treatment. Furthermore, ADXS31-164 reduced the incidence of metastatic disease and significantly increased duration of survival time and 1-, 2-, and 3-year survival rates when compared with a historical control group with HER2/neu(+) appendicular osteosarcoma treated with amputation and chemotherapy alone. CONCLUSIONS: These findings demonstrate that ADXS31-164 administered in the setting of minimal residual disease can induce HER2/neu-specific immunity and may reduce the incidence of metastatic disease and prolong overall survival in a clinically relevant, spontaneous, large animal model of cancer. These findings, therefore, have important translational relevance for children with osteosarcoma and adults with other HER2/neu(+) cancers. Clin Cancer Res; 22(17); 4380-90. ©2016 AACR.

Schistosoma mansoni soluble egg antigens enhance T cell responses to a newly identified HIV-1 Gag H-2b epitope.

Schistosome infection induces significant T helper type 2 (Th2) and anti-inflammatory immune responses and has been shown to negatively impact vaccine efficacy. Our goal was to determine if the administration of schistosome soluble egg antigens (SEA) would negatively influence the induction of cytotoxic T lymphocyte (CTL) and Th1-type T cell responses to an HIV candidate vaccine in the Th1-biased C57BL/6 mouse strain. Initial experiments failed, as we were unable to detect any response to the defined class I epitope for HIV-1 IIIB Gag. Therefore, we initiated an epitope mapping study to identify C57BL/6 (H-2(b)) T cell epitopes in HIV-1 IIIB Gag in order to perform the experiments. This analysis defined two previously unreported minimal class I H-2(b) and class II I-A(b) epitopes for HIV-1 IIIB Gag. The newly defined HIV-1 IIIB Gag epitopes were used to evaluate the influence of SEA on the generation of CTL and Th1-type HIV-1 IIIB Gag responses. Surprisingly, in contrast to our hypothesis, we observed that the coadministration of SEA with a Listeria monocytogenes vector expressing HIV-1 IIIB Gag (Lm-Gag) led to a significantly increased frequency of gamma interferon (IFN-γ)-producing CD8(+) and CD4(+) T cells in C57BL/6 mice compared to mice immunized with Lm-Gag only. These observations suggest that SEA contains, in addition to Th2-type and immune-suppressive molecules, substances that can act with the Lm-Gag vaccine to increase CTL and Th1-type vaccine-specific immune responses.

Schistosoma mansoni soluble egg antigens enhance Listeria monocytogenes vector HIV-1 vaccine induction of cytotoxic T cells.

Vaccines are an important public health measure for prevention and treatment of diseases. In addition to the vaccine immunogen, many vaccines incorporate adjuvants to stimulate the recipient's immune system and enhance vaccine-specific responses. While vaccine development has advanced from attenuated organism to recombinant protein or use of plasmid DNA, the development of new adjuvants that safely increase immune responses has not kept pace. Previous studies have shown that the complex mixture of molecules that comprise saline soluble egg antigens (SEA) from Schistosoma mansoni eggs functions to promote CD4(+) T helper 2 (Th2) responses. Therefore, we hypothesized that coadministration of SEA with a Listeria vector HIV-1 Gag (Lm-Gag) vaccine would suppress host cytotoxic T lymphocyte (CTL) and T helper 1 (Th1) responses to HIV-1 Gag epitopes. Surprisingly, instead of driving HIV-1 Gag-specific responses toward Th2 type, we found that coadministration of SEA with Lm-Gag vaccine significantly increased the frequency of gamma interferon (IFN-γ)-producing Gag-specific Th1 and CTL responses over that seen in mice administered Lm-Gag only. Analysis of the functionality and durability of vaccine responses suggested that SEA not only enlarged different memory T cell compartments but induced functional and long-lasting vaccine-specific responses as well. These results suggest there are components in SEA that can synergize with potent inducers of strong and durable Th1-type responses such as those to Listeria. We hypothesize that SEA contains moieties that, if defined, can be used to expand type 1 proinflammatory responses for use in vaccines.

Attenuated Listeria monocytogenes: a powerful and versatile vector for the future of tumor immunotherapy.

For over a century, inactivated or attenuated bacteria have been employed in the clinic as immunotherapies to treat cancer, starting with the Coley's vaccines in the 19th century and leading to the currently approved bacillus Calmette-Guérin vaccine for bladder cancer. While effective, the inflammation induced by these therapies is transient and not designed to induce long-lasting tumor-specific cytolytic T lymphocyte (CTL) responses that have proven so adept at eradicating tumors. Therefore, in order to maintain the benefits of bacteria-induced acute inflammation but gain long-lasting anti-tumor immunity, many groups have constructed recombinant bacteria expressing tumor-associated antigens (TAAs) for the purpose of activating tumor-specific CTLs. One bacterium has proven particularly adept at inducing powerful anti-tumor immunity, Listeria monocytogenes (Lm). Lm is a gram-positive bacterium that selectively infects antigen-presenting cells wherein it is able to efficiently deliver tumor antigens to both the MHC Class I and II antigen presentation pathways for activation of tumor-targeting CTL-mediated immunity. Lm is a versatile bacterial vector as evidenced by its ability to induce therapeutic immunity against a wide-array of TAAs and specifically infect and kill tumor cells directly. It is for these reasons, among others, that Lm-based immunotherapies have delivered impressive therapeutic efficacy in preclinical models of cancer for two decades and are now showing promise clinically. In this review, we will provide an overview of the history leading up to the development of current Lm-based immunotherapies, the advantages and mechanisms of Lm as a therapeutic vaccine vector, the preclinical experience with Lm-based immunotherapies targeting a number of malignancies, and the recent findings from clinical trials along with concluding remarks on the future of Lm-based tumor immunotherapies.

HIV-1 vaccine-specific responses induced by Listeria vector vaccines are maintained in mice subsequently infected with a model helminth parasite, Schistosoma mansoni.

In areas co-endemic for helminth parasites and HIV/AIDS, infants are often administered vaccines prior to infection with immune modulatory helminth parasites. Systemic Th2 biasing and immune suppression caused by helminth infection reduces cell-mediated responses to vaccines such as tetanus toxoid and BCG. Therefore, we asked if infection with helminthes post-vaccination, alters already established vaccine induced immune responses. In our model, mice are vaccinated against HIV-1 Gag using a Listeria vaccine vector (Lm-Gag) in a prime-boost manner, then infected with the human helminth parasite Schistosoma mansoni. This allows us to determine if established vaccine responses are maintained or altered after helminth infection. Our second objective asked if helminth infection post-vaccination alters the recipient's ability to respond to a second boost. Here we compared responses between uninfected mice, schistosome infected mice, and infected mice that were given an anthelminthic, which occurred coincident with the boost or four weeks prior, as well as comparing to un-boosted mice. We report that HIV-1 vaccine-specific responses generated by Listeria vector HIV-1 vaccines are maintained following subsequent chronic schistosome infection, providing further evidence that Listeria vector vaccines induce potent vaccine-specific responses that can withstand helminth infection. We also were able to demonstrate that administration of a second Listeria boost, which markedly enhanced the immune response, was minimally impacted by schistosome infection, or anthelminthic therapy. Surprisingly, we also observed enhanced antibody responses to HIV Gag in vaccinated mice subsequently infected with schistosomes.

Interferon γ-induced intratumoral expression of CXCL9 alters the local distribution of T cells following immunotherapy with Listeria monocytogenes.

The ability of Listeria monocytogenes-based anticancer vaccines to induce tumor regression depends on the responsiveness of malignant cells to interferon γ (IFNγ). Inhibition of IFNγ limits the recruitment of T cells to the tumors of vaccinated mice. We hypothesized that vaccination with immunotherapeutic L. monocytogenes induces the IFNγ-dependent production of chemokines that regulate the migration of tumor-infiltrating T cells. To gain further insights into this issue, we examined the chemokine responses of a transplantable, human papillomavirus (HPV)-immortalized murine tumor model (TC-1) following the administration of a L. monocytogenes-based immunotherapeutic agent that expresses E7 from HPV-16. Here, we report that the administration of L. monocytogenes-based anticancer vaccines increases the secretion of chemokine (C-X-C motif) ligand 9 (CXCL9), and CXCL10 by tumors, hence favoring the recruitment of T cells bearing the cognate chemokine (C-X-C motif) receptor 3 (CXCR3). Furthermore, the expression of CXCL9, but not CXCL10, in TC-1 tumors was significantly reduced upon anti-IFNγ antibody treatment. CXCL9 was highly expressed by TC-1 cells following the administration of IFNγ and tumor necrosis factor α (TNFα), in vitro. Moreover, the inhibition of CXCL9 in TC-1 cells reduced the proportion of CD8+ T cells infiltrating tumors in vaccinated mice, while increasing that of CD4+ T cells, thus altering T-cell subset distribution. We conclude that the administration of L. monocytogenes-based anticancer vaccines regulates TH1 chemokine responses and that malignant cells are an important source of these chemokines.

Live-attenuated Listeria-based immunotherapy.

For decades Listeria monocytogenes has been used as a model of host-disease immunology, and a considerable body of knowledge has been amassed regarding the complex immune response to L. monocytogenes. Attenuated strains of L. monocytogenes are currently being assessed as therapeutic bacterial vectors to present tumor-associated antigens to the immune system for the clinical treatment of cancer. L. monocytogenes immunotherapy utilizes many synchronous and disparate action mechanisms that stimulate innate and cell-mediated adaptive immunity while reducing immunosuppressive influences in the tumor microenvironment. Other effects not typically associated with immunotherapy include the stimulation of myeloid hematopoiesis and vascular changes that enable chemotaxis. Preliminary clinical results using L. monocytogenes bearing the HPV oncogene E7 indicate good tolerability and a strong efficacy signal, warranting further development. This article reviews the current status of L. monocytogenes as a cancer immunotherapeutic and the complex immune responses that underlie L. monocytogenes immunotherapy.

Successful vaccination of immune suppressed recipients using Listeria vector HIV-1 vaccines in helminth infected mice.

Vaccines for HIV, malaria and TB remain high priorities, especially for sub-Saharan populations. The question is: will vaccines currently in development for these diseases function in populations that have a high prevalence of helminth infection? Infection with helminth parasites causes immune suppression and a CD4+ Th2 skewing of the immune system, thereby impairing Th1-type vaccine efficacy. In this study, we conduct HIV vaccine trials in mice with and without chronic helminth infection to mimic the human vaccine recipient populations in Sub-Saharan Africa and other helminth parasite endemic regions of the world, as there is large overlap in global prevalence for HIV and helminth infection. Here, we demonstrate that Listeria monocytogenes functions as a vaccine vector to drive robust and functional HIV-specific cellular immune responses, irrespective of chronic helminth infection. This observation represents a significant advance in the field of vaccine research and underscores the concept that vaccines in the developmental pipeline should be effective in the target populations.

Listeria monocytogenes-derived listeriolysin O has pathogen-associated molecular pattern-like properties independent of its hemolytic ability.

There is a constant need for improved adjuvants to augment the induction of immune responses against tumor-associated antigens (TAA) during immunotherapy. Previous studies have established that listeriolysin O (LLO), a cholesterol-dependent cytolysin derived from Listeria monocytogenes, exhibits multifaceted effects to boost the stimulation of immune responses to a variety of antigens. However, the direct ability of LLO as an adjuvant and whether it acts as a pathogen-associated molecular pattern (PAMP) have not been demonstrated. In this paper, we show that a detoxified, nonhemolytic form of LLO (dtLLO) is an effective adjuvant in tumor immunotherapy and may activate innate and cellular immune responses by acting as a PAMP. Our investigation of the adjuvant activity demonstrates that dtLLO, either fused to or administered as a mixture with a human papillomavirus type 16 (HPV-16) E7 recombinant protein, can augment antitumor immune responses and facilitate tumor eradication. Further mechanistic studies using bone marrow-derived dendritic cells suggest that dtLLO acts as a PAMP by stimulating production of proinflammatory cytokines and inducing maturation of antigen-presenting cells (APC). We propose that dtLLO is an effective adjuvant for tumor immunotherapy, and likely for other therapeutic settings.

Vaccination with immunotherapeutic Listeria monocytogenes induces IL-17(+) γδ T cells in a murine model for HPV associated cancer.

Interleukin 17 (IL-17) is produced during infection with Listeria monocytogenes and is also an important regulator of tumor development with both pro- and anti-tumorigenic effects. αß T cells and γδ T cells are among the principle producers of IL-17 in response to infection and other proinflammatory conditions. Listeria-based cancer immunotherapies induce IFNγ directed Th1 dependent tumor regression; however, the role of IL-17 in Listeria based immunotherapy has not been addressed. Therefore, we investigated the ability of attenuated Listeria-based immunotherapy to induce IL-17 producing cells in a model of cervical cancer and the potential impact that these cells have on anti-tumor vaccine efficacy. Here we show that vaccination of tumor bearing mice with Listeria vaccines resulted in elevated levels of intratumoral IL-17 and increased IL-17 production by γδ TCR+ cells, exclusively. IL-17 producing cells were lacking in tumors of γδ T-cell-deficient mice; however, the absence of γδ T cells, including IL-17+ γδ T cells, did not alter tumor progression or abrogate the efficacy of the Listeria-based vaccine indicating that αß T cells are key for clearance of the tumor. Th1 responses, known to be responsible for anti-tumor Listeria-based vaccine efficacy, appear to be sufficient for tumor regression in γδ T-cell-deficient mice. We conclude that the efficacy of Listeria-based vaccine does not rely on γδ T cells (or IL-17 produced by them) in a TC.1 tumor model; however, Listeria-based immunotherapy can be used to induce IL-17+ γδ T cells that are important for regression observed in alternative cancer models.

Listeria monocytogenes and its products as agents for cancer immunotherapy.

This review covers the use of Listeria monocytogenes and its virulence factors as cancer immunotherapeutics. We describe their development as vectors to carry protein tumor antigen and eukaryotic DNA plasmids to antigen-presenting cells and efforts to harness their tumor-homing properties. We also describe their use as vectors of angiogenic molecules to induce an immune response that will destroy tumor vasculature. The background knowledge necessary to understand the biology behind the rationale to develop Listeria as a vaccine vector for tumor immunotherapy is included as well as a brief summary of the major therapies that have used this approach thus far.

The ubiquitin-like protein, ISG15, is a novel tumor-associated antigen for cancer immunotherapy.

The recent announcement of the first FDA-approved therapeutic vaccine for prostate cancer, Sipuleucel-T, is a watershed moment for the field of tumor immunotherapy. However, while Sipuleucel-T provides a powerful tool to clinicians for the most prevalent form of cancer in men, there remains an unmet need for a similar therapeutic strategy against breast cancer, the most prevalent cancer in women. While current breast cancer vaccines in development target several antigens, the most prevalent is the tumor-associated antigen, HER2. Initial results with HER2 vaccines appear promising in terms of efficacy; however, the lack of HER2 overexpression by a majority of breast tumors and the safety concerns associated with current HER2-targeted immunotherapy suggest that additional therapeutic strategies would be beneficial. Recently, several studies have identified ISG15 as a molecule highly expressed in numerous malignancies. ISG15 is a small ubiquitin-like protein regulated by type-I interferon and classically associated with viral defense. Elevated ISG15 expression in breast cancer is especially well documented and is independent of HER2, progesterone receptor, and estrogen receptor status. Additionally, high ISG15 expression in breast cancer correlates with an unfavorable prognosis and poor responses to traditional treatment strategies such as chemotherapy and radiation. To overcome these challenges, we employ a novel strategy to specifically target tumor-associated ISG15 expression with immunotherapy. We demonstrate that vaccination against ISG15 results in significant CD8-mediated reductions in both primary and metastatic mammary tumor burden. These results validate ISG15 as a tumor-associated antigen for cancer immunotherapy.

Active immunotherapy combined with blockade of a coinhibitory pathway achieves regression of large tumor masses in cancer-prone mice.

Vaccines that aim to expand tumor-specific CD8(+) T cells have yielded disappointing results in cancer patients although they showed efficacy in transplantable tumor mouse models. Using a system that more faithfully mimics a progressing cancer and its immunoinhibitory microenvironment, we here show that in transgenic mice, which gradually develop adenocarcinomas due to expression of HPV-16 E7 within their thyroid, a highly immunogenic vaccine expressing E7 only induces low E7-specific CD8(+) T-cell responses, which fail to affect the size of the tumors. In contrast, the same type of vaccine expressing E7 fused to herpes simplex virus (HSV)-1 glycoprotein D (gD), an antagonist of the coinhibitory B- and T-lymphocyte attenuator (BTLA)/CD160-herpes virus entry mediator (HVEM) pathways, stimulates potent E7-specific CD8(+) T-cell responses, which can be augmented by repeated vaccination, resulting in initial regression of even large tumor masses in all mice with sustained regression in more than half of them. These results indicate that active immunization concomitantly with blockade of the immunoinhibitory HVEM-BTLA/CD160 pathways through HSV-1 gD may result in sustained tumor regression.

Targeting tumor vasculature with novel Listeria-based vaccines directed against CD105.

The FDA approval of bevacizumab (Avastin®, Genentech/Roche), a monoclonal antibody raised against human VEGF-A, as second-line therapy for colon and lung carcinoma validated the approach of targeting human tumors with angiogenesis inhibitors. While the VEGF/VEGFR pathway is a viable target for anti-angiogenesis tumor therapy, additional targets involved in tumor neovascularization have been identified. One promising target present specifically on tumor vasculature is endoglin (CD105), a member of the TGF-ß receptor complex expressed on vascular endothelium and believed to play a role in angiogenesis. Monoclonal antibody therapy and preventive vaccination against CD105 has met with some success in controlling tumor growth. This report describes the in vivo proof-of-concept studies for two novel therapeutic vaccines, Lm-LLO-CD105A and Lm-LLO-CD105B, directed against CD105 as a strategy to target neovascularization of established tumors. Listeria-based vaccines directed against CD105 lead to therapeutic responses against primary and metastatic tumors in the 4T1-Luc and NT-2 mouse models of breast cancer. In a mouse model for autochthonous Her-2/neu-driven breast cancer, Lm-LLO-CD105A vaccination prevented tumor incidence in 20% of mice by week 58 after birth while all control mice developed tumors by week 40. In comparison with previous Listeria-based vaccines targeting tumor vasculature, Lm-LLO-CD105A and Lm-LLO-CD105B demonstrated equivalent or superior efficacy against two transplantable mouse models of breast cancer. Support is provided for epitope spreading to endogenous tumor antigens and reduction in tumor vascularity after vaccination with Listeria-based CD105 vaccines. Reported here, these CD105 therapeutic vaccines are highly effective in stimulating anti-angiogenesis and anti-tumor immune responses leading to therapeutic efficacy against primary and metastatic breast cancer.

Listeria-derived ActA is an effective adjuvant for primary and metastatic tumor immunotherapy.

Tumor immunotherapy is currently at the cusp of becoming an important aspect of comprehensive cancer treatment in the clinic. However, the need for improved adjuvants to augment immune responses against tumor antigens is always present. In this paper, we characterize the Listeria monocytogenes-derived actin-nucleating protein, ActA, as a novel adjuvant for use in tumor immunotherapy. ActA is a virulence factor that is expressed on the cell surface of L. monocytogenes and facilitates the production of actin tails that propel Listeria throughout the cytosol of an infected host cell. It is believed that this ActA-dependent cytosolic motility allows Listeria to evade adaptive host cell defenses and facilitates its invasion into a proximal uninfected host cell. However, there is evidence that ActA fused to a tumor antigen and delivered by L. monocytogenes can perform a beneficial function in tumor immunotherapy as an adjuvant. Our investigation of this adjuvant activity demonstrates that ActA, either fused to or administered as a mixture with a tumor antigen, can augment anti-tumor immune responses, break immune tolerance and facilitate tumor eradication, which suggests that ActA is not only an effective adjuvant in tumor immunotherapy but can also be applied in a number of therapeutic settings.