- Open Access
Cancer anti-angiogenesis vaccines: Is the tumor vasculature antigenically unique?
- Samuel C. Wagner†1,
- Thomas E. Ichim†1Email author,
- Hong Ma1,
- Julia Szymanski1,
- Jesus A. Perez2,
- Javier Lopez2,
- Vladimir Bogin1,
- Amit N. Patel3,
- Francisco M. Marincola4 and
- Santosh Kesari5
© Wagner et al. 2015
- Received: 13 September 2015
- Accepted: 3 October 2015
- Published: 29 October 2015
Angiogenesis is essential for the growth and metastasis of solid tumors. The tumor endothelium exists in a state of chronic activation and proliferation, fueled by the tumor milieu where angiogenic mediators are aberrantly over-expressed. Uncontrolled tumor growth, immune evasion, and therapeutic resistance are all driven by the dysregulated and constitutive angiogenesis occurring in the vasculature. Accordingly, great efforts have been dedicated toward identifying molecular signatures of this pathological angiogenesis in order to devise selective tumor endothelium targeting therapies while minimizing potential autoimmunity against physiologically normal endothelium. Vaccination with angiogenic antigens to generate cellular and/or humoral immunity against the tumor endothelium has proven to be a promising strategy for inhibiting or normalizing tumor angiogenesis and reducing cancer growth. Here we review tumor endothelium vaccines developed to date including active immunization strategies using specific tumor endothelium-associated antigens and whole endothelial cell-based vaccines designed to elicit immune responses against diverse target antigens. Among the novel therapeutic options, we describe a placenta-derived endothelial cell vaccine, ValloVax™, a polyvalent vaccine that is antigenically similar to proliferating tumor endothelium and is supported by pre-clinical studies to be safe and efficacious against several tumor types.
- Vascular Endothelial Growth Factor
- Human Umbilical Vein Endothelial Cell
- Tumor Vasculature
- Peptide Vaccine
- Tumor Endothelial Cell
Angiogenesis, the outgrowth of new blood vessels from pre-existing capillaries and post-capillary venules, occurs during embryonic development, in the uterus during the menstrual cycle, in the process of wound healing, and in pathological conditions . In healthy adults, endothelial cells can maintain a quiescent state for years, whereas they proliferate and migrate to form new vessels in response to inflammatory conditions and during tumor growth. Several studies have estimated that tumor associated endothelial cells proliferate 30–40 times faster relative to endothelial cells found in healthy vasculature [2–4]. Based on estimates that tumors fail to grow beyond 1–2 mm in the absence of new capillary growth, Dr. Judah Folkman put forth the central hypothesis that tumors release diffusible factors that stimulate endothelial cell proliferation in host capillary blood vessels . Indeed, it has been estimated that eradication of one endothelial cell is capable of neutralizing of up to 100–300 tumor cells . Since the immune system is in direct contact with the tumor vasculature, vaccination against tumor endothelium is theoretically very promising for breaching the immunological barriers created by the tumor microenvironment.
The goal in vaccination strategies is to raise immunity against antigens present in tumor endothelium while avoiding antigens that cross-react with healthy vasculature, thereby preventing deleterious autoimmune reactions. Since the landmark publication by Dr. Folkman, a catalog of molecular players involved in the process of tumor angiogenesis have been identified and characterized. However, clinical outcomes of traditional anti-angiogenic therapies such as monoclonal antibodies have improved patient survival rates only modestly . Vaccination against endothelial cells is poised to overcome the existing problems of drug resistance and adverse side effects associated with other approaches. This report reviews vaccination strategies against the tumor endothelium that have been tested to date, including DNA, protein and peptide vaccines of tumor-endothelium-associated antigens, as well as polyvalent vaccines comprised of whole endothelial cells. Very encouraging data point toward the efficacy of vaccination in raising humoral and cell-mediated immunity against angiogenesis-associated antigens in cancer. In this discussion, we also highlight our novel approach wherein placenta-derived endothelial cell lysates (ValloVax™) are used as a source of antigen for vaccinating against proliferating tumor endothelial cells.
Endothelium is a dynamic and heterogeneous structure influenced by environmental factors such as shear stress, oxygen content of the blood, chemokines, cytokines, and changes in the content of the extracellular matrix . Whereas resting endothelium serves to maintain blood fluidity, regulate blood flow, control vessel wall permeability, and quiesce circulating lymphocytes , environmental cues activate endothelial cells to proliferate, migrate, and form new branches (sprouting). In the tumor milieu, aberrantly elevated and chronic production of angiogenic factors leads to endothelial activation, vascular irregularities, and immune suppression, which are among the well-recognized hallmarks of cancer proliferation . Whereas the structure of normal vascular endothelium is hierarchical and organized, the activated endothelium in cancer consists of dilated and tortuous blood vessels that are chaotically interconnected, leading to heterogeneous vessel density within the tumor, erratic blood flow, and focal regions of hypoxia [11, 12]. At the cellular level, tumor endothelial cells display a disorganized morphology, being loosely connected and exhibiting increased vascular permeability . These conditions cause impaired oxygen and nutrient delivery, conditions which in turn trigger angiogenesis, thereby further promoting tumor endothelial cell activation and vascular growth to meet the metabolic demands of the tumor. In this manner, cancer cells and endothelial cells are involved in a positive feedback loop stimulating each other’s growth. At the same time, the vascular malformations in the tumor milieu serve as obstacles to effective penetration of anti-tumor lymphocytes and chemotherapy drugs into the tumor mass.
Although tumor endothelial cells are considered to be genetically stable as compared to tumor cells, angiogenic factors can fuel genetic aberrations of tumor endothelium. Indeed, resistance of tumor endothelial cells in diverse cancers has been described; for example, tumor endothelial cells in renal carcinoma are reportedly resistant to serum starvation , breast tumor endothelial cells are resistant to vincristine-induced apoptosis  and hepatocellular carcinoma endothelial cells are resistant to 5-fluorouracil and doxorubicin . One study demonstrated aneuploid chromosomes and abnormal centrosomes in tumor endothelial cells . Signaling in response to the dysregulated over-production of angiogenic factors, including vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF), in absence of endothelial stability promoting factors such as PDGF-BB was found to contribute to aneuploidy and centrosome duplication in tumor endothelial cells . Moreover, tumor endothelial cells with excess centrosomes exhibit apoptosis resistance and formation of aberrant spindle projections, likely attributable to gain or loss of genes involved in proliferation, survival, and adhesion . At the mechanistic level, VEGF secreted from tumors affects tumor endothelial cells by upregulating MDR1, a gene encoding a transmembrane glycoprotein P-gp that is known as a multidrug transporter that potentiates resistance to several anti-cancer agents . For example, paclitaxel, which is used to treat several types of cancer together with anti-angiogenic drugs, is transported by P-gp. The VEGFR kinase inhibitor, Ki8751, and a phosphatidylinositol 3-kinase–Akt inhibitor, LY294002, effectively block tumor-induced MDR1 up-regulation, suggesting that VEGF in the tumor microenvironment is an underlying factor in acquired drug resistance . Vaccination against angiogenesis-associated antigens therefore could serve as a valuable asset for improving the efficacy of other cancer therapeutics.
Heterogeneity among tumor endothelial cells, as choreographed by the over-activity of certain angiogenic pathways, also shields tumors by affecting their visibility to the immune system. Production of angiogenic molecules by tumors inhibits the expression of adhesion molecules involved in leukocyte interactions with blood vessel walls, including intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), E-selectin and CD34 . These features of tumor endothelium prevent adhesion and extravasation of effector T cells into the tumor. Additionally, tumor endothelial cells have been described as anergic, marked by unresponsiveness to inflammatory cytokines that would normally induce adhesion molecule expression but instead allow the endothelium to escape immune surveillance [20, 21]. The anergic phenotype of tumor endothelial cells can be reversed by anti-angiogenesis therapy, which upregulates the expression of endothelial adhesion molecules in the tumor vasculature [22, 23]. The tumor endothelium is also prohibitive to entry of tumor-reactive T lymphocytes by suppression or direct killing of effector T cells via molecules such as Fas ligand (FasL) . Notably, VEGF-A, interleukin 10 (IL-10) and prostaglandin E2 (PGE2) are involved in eliciting FasL expression in endothelial cells, as evidenced by observations that pharmacological inhibition of these molecules leads to a marked influx of tumor-rejecting CD8+ cells and tumor growth suppression in mice. Hence, anti-angiogenesis vaccination can break down the immunological firewall that the tumor endothelium imposes to protect the cancer.
Cancer vaccines consist of tumor-associated antigens delivered in a pro-inflammatory context to generate potent antitumor immune responses and overcome the cancer’s varied immunosuppressive mechanisms. Vaccination strategies directed against tumor-associated endothelium are designed to take advantage of both quantitative and qualitative differences between tumor endothelial cells and non-malignant endothelial cells. Ideal vaccine candidates include receptors/proteins that are upregulated in tumor endothelium, owing to its activated and proliferating state, but are sparsely expressed during physiological angiogenesis in healthy adult tissues. Notably, angiogenesis-associated molecules can be over-expressed on endothelial cells in an organ- or tumor-specific manner, and can sometimes be expressed on resting endothelial cells . Thus, an attractive tumor endothelial vaccine should be capable of eliciting anti-angiogenesis and anti-cancer immunity against diverse tumor types, while avoiding autoimmune reactions against physiological angiogenesis, such that occur in the female reproductive tract and during wound healing, or against quiescent endothelium.
Whether immunity and/or autoimmunity ensue as a result of vaccination is dependent not only on the target tumor endothelial antigen but also on the vaccination protocol. Candidate vaccines have employed numerous delivery approaches, including proteins, peptides, dendritic cells pulsed with the antigen(s), naked DNA or recombinant DNA delivered by carriers, and mRNA vaccines, as well as whole cell vaccines or endothelial membrane components. To develop rationales for clinical vaccine design, pre-clinical studies have been conducted for many angiogenesis-associated molecules using various delivery systems. In some cases, differences in safety and efficacy between vaccines hinge on a plethora of secondary variables including the use of different delivery vectors, the choice of adjuvants and the varying routes of vaccine administration.
Two general approaches to anti-angiogenesis vaccination have yielded promising results for inducing specific and robust immunity against tumor endothelium and reducing tumor growth and metastasis: (1) vaccines expressing defined angiogenesis-associated antigens; and, (2) vaccines comprised of whole endothelial cells and/or mixtures of endothelial antigens. In the latter category are vaccines consisting of placenta-derived endothelial antigens, an approach currently being advanced by Batu Biologics, Inc. (ValloVax™ vaccine).
Evidence has shown the feasibility of targeting molecules that are expressed by angiogenic endothelium, and the data also suggest that anti-angiogenic vaccination can be applied synergistically with tumor immunotherapy and/or chemotherapy to invoke anti-tumor immunity. Theoretically, the approach of targeting certain defined angiogenesis-associated molecules on tumor endothelium has the risk to evoke only a transient decrease in cancer progression, owing to the fact that angiogenesis can proceed via compensatory pathways. Despite this caveat, success at achieving anti-angiogenic immunity has been demonstrated by vaccinating against the following antigens.
VEGF-A and VEGFR
VEGF/VEGFR is perhaps the best-studied angiogenic pathway involved in the growth and survival of tumor endothelium. Clinical therapies addressing tumor angiogenesis have therefore largely focused on inhibiting VEGF and its cognate receptor. VEGF-A exists in several pro-angiogenic variants that are secreted by both tumor cells and endothelial cells and activate VEGFR1 and VEGFR2. Numerous VEGF-expressing vectors have been administered for vaccinating against VEGF. A Xenopus VEGF DNA vaccine that was protective and therapeutic in several tumor models in mice, an effect that was mediated by anti-tumor activity of CD4+ lymphocytes . A phase I clinical study has investigated CIGB-247, a vaccine comprising a human VEGF variant molecule in combination with a bacterial adjuvant, in 30 patients with advanced solid tumors . After eight consecutive weeks of subcutaneous immunization and re-vaccination on week 12, this vaccine was safe at three dose levels and also demonstrated immunogenicity in three sequential analyses of patients’ blood samples. Despite the critical importance of VEGF in hematopoiesis [28–30], no abnormalities in leukocytes or megakaryocytes were found, and additionally, serum biochemistry parameters were not altered in patients received the VEGF vaccination. Based on this clinical report, VEGF is a highly promising vaccine target as a cancer treatment strategy. The implications of this body of research are that immunity can be induced to self-antigens, without triggering a fulminant autoimmune response.
VEGFR2 (also FLK-1 and KDR), which is highly expressed on proliferating endothelial cells of the tumor vasculature, has also been the focus of numerous pre-clinical studies. Animal models using VEGFR2 DNA, protein, and peptide vaccines have demonstrated their ability to elicit potent humoral and cellular immunity, suppression of angiogenesis, tumor necrosis and/or suppression of metastastatic progression [31–38]. Interestingly, none of the aforementioned studies have reported hematopoietic or other abnormalities.
Since VEGFR2 is also expressed at lower levels in normal vascular endothelium, vaccination could theoretically cause side effects or an autoimmune response. Neithammer et al. reported that in murine models a DNA vaccine against VEGFR2 suppressed angiogenesis in the tumor vasculature as verified by in vitro inhibition of endothelial cell proliferation, deposition of antibodies in tumor vasculature, a reduction of microvessel density and antitumor activity in vivo without impairment of fertility, neuromuscular performance or hematopoiesis; however, a slight delay in wound healing in immunized mice was noted . In another report, there was no delay in wound healing observed in response to a dendritic cell-based VEGFR2 vaccine although reduced litter sizes and fetal loss were found in vaccinated animals . However, in other studies where VEGFR2 protein vaccines were found to generate potent immunity against tumor endothelium, no effects on wound healing, reproduction or other organ toxicities were observed [35, 41].
Immunogenic epitopes of VEGFR2 have been identified and peptide vaccines have been optimized for binding to MHC class I molecules in order to elicit activity of cytotoxic T lymphocytes (CTL) specific for tumor endothelium. In a clinical study of pancreatic cancer, administration of immunogenic VEGFR2-169 peptide vaccine in combination with gemcitabine was well tolerated with no severe adverse events and peptide-specific CTL were induced . Other than injection site reactions, other peptide vaccines tested for targeting VEGFR were also deemed to have manageable toxicities [43–45].
bFGF and FGF-R
Basic fibroblast growth factor (bFGF or FGF-2) and its receptor (FGFR-1, CD331) have been targeted in pre-clinical mouse models to evaluate their anti-angiogenic and anti-tumor effects. FGFR-1 is highly expressed on angiogenic endothelium as well as on endothelial progenitor cells [46, 47]. In one such study, vaccination with xenogeneic FGFR-1 plasmid DNA inhibited tumor endothelial cell proliferation and produced anti-cancer immunity in three murine tumor models . Studies utilizing recombinant protein vaccines targeting FGFR-1 demonstrated significantly decreased tumor volume compared to controls, as well as decreased microvessel density in tumors without any observable overt toxicity . Peptide bFGF vaccines combined with adjuvants also induced antigen-specific antibody and cell-mediated responses [50, 51]. Importantly, in a study where a liposome-based peptide bFGF vaccine was administered in murine cancer models, immunity against tumor endothelium was elicited while physiological angiogenesis was unperturbed, as evidenced by normal wound healing times and no impairments in the reproductive ability of vaccinated animals or in the viability or health of the offspring .
The expression of several cell adhesion molecules, most notable integrin alpha(v)beta(3), has been associated with tumor angiogenesis .
Integrin αvβ3 plays a role in several physiological processes including angiogenesis, pathological neovascularization, and tumor metastasis . An αvβ3 ligand vaccine induced a humoral response associated with significant antitumor activity without recourse of adjuvant therapy. The immune response was driven by antibody-dependent cellular toxicity and complement-directed cytotoxicity to the tumor-associated endothelial cells . The effectiveness of utilizing antibodies blocking αvβ3 as a method of inhibiting angiogenesis, has led to the clinical development of monoclonal anti-αvβ3 antibodies for the treatment of advanced solid tumors. MEDI-522, a second generation humanized anti-αvβ3, successfully completed a Phase I study in patients with a variety of metastatic solid tumors, and a maximum tolerated dose was not identified by traditional dose-limiting toxicities .
Another target of anti-angiogenic vaccine strategies is the angiostain binding protein angiomotin, a membrane-associated pro-angiogenic protein present on endothelial cells in angiogenic tissues . Holmgren et al. used DNA vaccination with a xenogeneic (human) angiomotin DNA construct to increase immunogenicity of the vaccine, and injected the DNA intramuscularly followed by electroporation . This vaccine protected mice from tumor challenge and acted synergistically with a vaccine targeting the Her-2 oncogene in a transgenic breast cancer model system. No evidence for autoimmunity in normal vasculature of the retinas was noted at 16 or 70 weeks post-treatment. In another report, an angiomotin DNA vaccine was shown to alter blood vessel architecture to increase permeability and thus improve the efficacy of chemotherapy in an animal model .
This transmembrane glycoprotein is primarily expressed in proliferating endothelial cells and is upregulated by hypoxia; therefore, endoglin is strongly expressed in tumor endothelial cells . The functional role for endoglin in hematopoiesis is also exemplified by CD105 knockout studies where embryonic lethality occurred as a result of impaired angiogenesis in the yolk sac and heart defects. CD105 expression on tumor vessels is a prognostic factor correlated with poor overall and disease-free survival, tumor recurrence, and metastasis of various cancers [60, 61]. CD105 vaccination approaches employing bacterial surface display of protein  and orally administered DNA vaccines  effectively targeted the vasculature and inhibited tumor growth in the absence of observable effects on healthy tissues.
This is an intracellular tumor-associated antigen that is upregulated in cancers and tumor endothelial cells but is not expressed in healthy differentiated tissues. Survivin is a member of the inhibitor of apoptosis protein family, and serves to inhibit programmed cell death, promote cellular proliferation and enhance angiogenesis . Survivin vaccines elicit apoptosis of tumor cells and its vasculature with variable efficacy; while intradermal electroporation was found to be very effective in animal models, naked DNA administration is less effective in the prophylactic or therapeutic settings [65, 66].
Robo4 is a member of the Roundabout family proteins and is a transmembrane cell adhesion molecule that is specifically expressed in endothelial cells and hematopoietic stem cells and progenitor cells . Robo4 guides the formation of the vasculature by controlling endothelial cell migration and coordinating blood vessel sprouting . Tonic Robo4 signaling in healthy tissues maintains vascular integrity by inhibiting VEGF-induced endothelial cell migration and vascular permeability . Robo4 is pathologically over-expressed in the tumor endothelium , it promotes atypical vascular patterning that reduces blood-tumor barrier permeability, thereby likely impeding the access of chemotherapy drugs to tumors . In one study, a protein vaccine against this target was developed, comprised of the extracellular domain of mouse Robo4, fused to the Fc domain of human immunoglobulin within an adjuvant. Vaccinated mice had a strong antibody response to Robo4, impaired fibrovascular invasion in vitro, and reduced growth of implanted Lewis lung carcinoma . Further studies will be needed to determine how breaking tolerance to Robo4 affects physiological angiogenesis, where it appears to counteract angiogenic signaling .
Tie-2 is an endothelium-specific receptor tyrosine kinase that binds to angiopoietin and is upregulated on diseased endothelium in diverse conditions including cancer, atherosclerosis, and macular degeneration. A protein vaccine based on chicken Tie-2 reduced growth of hepatomas and melanomas in mice, whereas a murine Tie-2 vaccine was ineffective, indicating the need for further enhancement of this protein’s immunogenicity . The chicken Tie-2 vaccine exhibited apoptotic and anti-angiogenic effects on endothelial cells. The ligand for Tie-2, angiopoietin-2 is also over-expressed during neoangiogenesis in tumors and acts synergistically with VEGF in promoting tumor vascularization .
This polypeptide is expressed as a marker for neonatal lung vasculature and adult pathological angiogenesis. HP59 protein has been identified in the vasculature of many types of tumors including lung, colon, ovary, and breast cancers of different stages, but not in the corresponding normal tissues; therefore, this protein could be an ideal target antigen for vaccinating against tumor endothelium . Vaccination of mice with immunogenic HP59 peptides attenuated the growth of Lewis lung tumors and inhibited pathological angiogenesis, as evidenced by an absence of HP59-expressing blood vessels. HP59 possesses a 41-amino acid sequence at the NH2 terminal that has no homology with any known protein; therefore, this antigen could be used for raising specific immunity against the pathological vasculature in human cancer .
This tyrosine kinase receptor is activated by members of the platelet-derived growth factor family, and is broadly involved in regulating cellular proliferation and differentiation in physiological and pathological contexts. PDGF-B, the ligand, produced by tumor cells binds to PDGFR on stromal and perivascular cells to promote tumor growth and angiogenesis, as well as eliciting other pathways of tumor nourishment [83, 84]. PDGFRβ supports angiogenesis in two distinct ways: (1) by activating pericytes, which support endothelial cell proliferation and elaboration of the tumor vasculature; and (2) by upregulating expression of the proangiogeneic factor FGF-2 .
Anti-PDGF drugs have been widely used for treating various cancers; however, their mechanisms of action at a molecular level are not strictly anti-angiogeneic. Paradoxically, PDGF ligand inhibition can increase tumor angiogenesis, which supports increased tumor growth but, in doing so, normalizes blood vessel walls to allow for more efficient delivery of chemotherapy drugs [85, 86]. On this basis, inhibiting the PDGF pathway can serve as an effective adjunct to cytotoxic drugs and yield a net gain in eradicating tumors .
To assess the effects of vaccinating against PDGFRβ, a DNA vaccine delivered via Salmonella typhimurium was given to mice orally in murine colon, breast and lung carcinoma model systems . Significantly, in a therapeutic setting where tumors were implanted 20 days before vaccination, the size of tumors in the PDGFRβ-immunized group was one-fourth the size of the control group. Angiogenic markers within tumors were reduced and robust cytotoxic T cell responses against PDGFR-β were detectable, indicating that angiogenesis was effectively targeted.
Tumor endothelial marker 1, also known as endosialin or CD248, is a type 1 membrane protein involved in developmental, physiological and pathological angiogenesis. TEM1 is overexpressed in the vasculature of carcinomas, brain tumors and sarcomas [88–91] and is present on endothelial cells, pericytes, and fibroblasts [92, 93]. A DNA vaccine expressing full-length mouse Tem1 cDNA fused to tetanus toxoid adjuvant inhibited tumor growth and progression without affecting reproduction or wound healing . Splenocytes from TEM1 vaccinated mice secreted IFN-γ and lysed TEM1-expressing endothelial cells in vitro. These data support the use of TEM1 protein vaccine in conjunction with an adjuvant to break tolerance to tumor endothelium for the treatment of solid tumors.
Designing a tumor endothelial vaccine based on TEM8 is a very enticing possibility since this molecule is selectively upregulated on tumor endothelial cells and, unlike the majority of other putative vaccine targets, TEM8 is undetectable in endothelium during physiological angiogenesis . The first TEM8 DNA vaccine was a syngeneic vaccine that was tested in a rat model of breast cancer and murine melanoma . Although the vaccine had no activity as a single agent, TEM8 DNA significantly enhanced anti-tumor immunity when administered with a rat HER2 DNA vaccine for breast cancer, and also in combination with a tyrosine-related protein-1 DNA vaccine in melanoma. Ruan et al. developed a xenogeneic TEM8 DNA vaccine carried by xenogeneic Salmonella typhimurium that was capable of generating TEM8-specific CD8+ cytotoxic T cells and protected mice from lethal tumor challenge . Another vaccination approach consisted of dendritic cells transduced with recombinant adenovirus encoding TEM8, which also effectively protected mice from lethal challenges against hepatocellular carcinoma . Lastly, a DNA vaccine encoding syngeneic TEM8 and murine beta-defensin 2, which activates dendritic cells to stimulate potent immunity, was used to inhibit tumor growth in a murine colon cancer model . This vaccine was also highly effective, causing the collapse of tumor vessels by evoking an antigen specific CD8+ T cell response. Further studies are warranted to determine whether TEM8-based vaccines can be safe and efficacious for clinical use.
In summary, several therapeutic approaches have been explored targeting single antigen expressed on the tumor endothelium, demonstrating the feasibility of targeting the tumor vasculature by vaccine therapy without inducing systemic autoimmunity to quiescent endothelium in vivo. Signals of efficacy have been observed in preclinical studies around these targets, and several tumor endothelium-targeting vaccines have made it into the clinical setting for the treatment of cancer. As such, in an effort to reduce the development of treatment resistance and to potentiate the strength of an immune response and resultant survival benefit, the authors would like to explore the concept of a polyvalent vaccine targeting several angiogenesis associated antigens.
Although great strides have been made in advancing anti-angiogenesis tumor vaccines against specific targets, it is clear that the presence of many interrelated and compensatory pathways, as well as genetic instability of tumor endothelium could theoretically overcome targeted inhibition in a clinical setting. Accordingly, polyvalent vaccination approaches are being developed using whole endothelial cells or isolated proteins from endothelial cell membranes. Another advantage of such polyvalent vaccines expressing numerous angiogenic antigens is to allow antigen-presenting cells to process and present immunodominant epitopes for generating anti-angiogenic immunity.
Preclinical studies of endothelial cell vaccines
Human umbilical vein endothelial cells (HUVEC) have been the standard for cell-based models of tumor angiogenesis, having the ability to proliferate extensively and expressing a number of pro-angiogenic molecules that mimic tumor neovasculature. Specifically, the aforementioned antigens such as VEGFR2, αvβ3 and endoglin, which are common biomarkers associated with tumor angiogenesis, are expressed in the primary culture of HUVECs . In a report by Wei et al. , paraformaldehyde fixed xenogeneic whole HUVEC used as a vaccine markedly inhibited tumor growth in prophylactic and therapeutic murine cancer models. The anti-angiogenic effect of this vaccine depended on CD4+ T cells eliciting endothelial-specific antibody responses. Using a syngeneic vaccine consisting of fixed hepatic sinusoidal endothelial cells, Okaji et al. also demonstrated potent preventative and therapeutic anti-tumor immunity in a lung metastasis model of murine colon cancer . Both antibody and cytotoxic T cell responses against endothelial cells were detected in response to this vaccine. Similar studies performed in animal models have reported potent angiogenesis inhibition and tumor targeting when using syngeneic endothelial cell vaccines [102, 103], xenogeneic endothelial cells , and xenogeneic endothelial proteins . Although the experimental design differences between these studies preclude any definitive conclusions concerning the optimal vaccine candidate, these data collectively provide a compelling proof of concept that tolerance to tumor endothelium can be broken using whole endothelial cells as vaccines.
Clinical progress with endothelial cell vaccines
HUVECs have been used in pilot studies to test the anti-angiogenic effects of vaccination in patients with malignant brain tumors and metastatic colorectal cancer [106, 107]. Vaccinations were performed using 5 × 10(7) HUVECs given intradermally on a weekly basis for the first month and the every 2 weeks from the second month onward. In a published report where a total of 230 vaccinations were administered to 6 patients with recurrent malignant brain tumors and 3 patients with metastatic colorectal cancer, MRI results showed partial or complete responses lasting for a minimum of 9 months in 3 of the patients with brain tumors . Moreover, antibodies directed against HUVEC antigens were detected in eight out of nine patients and HUVEC-specific CTLs were detected in 6 of 7 tested patients. No adverse events were reported with the exception of skin reactions at the vaccine injection site.
In a related study, 17 patients with recurrent glioblastoma were treated with the HUVEC vaccine using the same protocol of intradermal delivery . These patients had been previously treated with surgery and chemoradiotherapy, and were also undergoing salvage treatments including concomitant and adjuvant chemotherapy during the course of the study. The results showed that HUVEC vaccine therapy is feasible for recurrent glioblastoma based on significant prolongations of the tumor doubling times and reduced tumor growth rates at 3- and 6-month follow ups. Despite the fact that 352 vaccinations were performed, no adverse events were observed with the exception of skin reactions at the injection sites. For comparative purposes, the investigators point out that bevacizumab, a humanized monoclonal antibody to VEGF, is associated with grade 3 adverse events in 46.4 % of patients when used alone and in 65.8 % of patients when used in combination with chemotherapy , demonstrating that a HUVEC vaccine appears to be much safer than conventional anti-angiogenesis drug therapies . The authors conclude that, for invasive and large tumors, HUVEC vaccination is feasible for use in combination with other treatment modalities and similar trials should be conducted for other types of cancer.
Among the vaccine candidates for targeting tumor endothelium, the placenta is a source of significant numbers of proliferating and angiogenic cells owing to its immunological status at the maternal-fetal interface. In the placenta, trophoblast cells invade into the maternal uterine wall where they remodel the spiral arteries and supply angiogenic factors for de novo blood vessel formation and expansion of the pre-existing vascular network . These processes must meet the high demands for blood supply and nutrient exchange of the growing fetus, analogous to the elaborate vascular networks required for tumor growth.
The idea of using placenta as a source of antigens for vaccination against tumors originated in the 1970s from Dr. Valentin Govallo who noted the immunological similarities between pregnancy and cancer and demonstrated that immunity to placental trophoblast cells conferred radiological tumor reductions in lung cancer patients (reviewed in ). Indeed, a substantial body of literature validates numerous parallels in proliferation, angiogenesis, invasion, and immune suppression between cancer and pregnancy that is attributable to shared characteristics of fetal-derived trophoblast cells of the placenta and tumor cells . Common angiogenesis-associated molecules that have been identified as highly expressed in tumor cells and in placenta include VEGF family members , FGF/FGFR, survivin , calreticulin , angiomotin , ROBO4 , and PDGFB/PDGFR . Placenta is also a practical source for purification of endothelial cells for therapeutic purposes with yields reported from 2 to 10 billion primary endothelial cells per placenta [118–120]. In contrast, the theoretical yield of total cells from umbilical cord (including mesenchymal and endothelial cells) is 500 million cells with reported yields of less than half of this amount following isolation and processing . Placental endothelial cells are thus a feasible alternative to HUVECs as vaccines for targeting tumor vasculature.
Batu Biologics is currently advancing ValloVax™, a vaccine consisting of placenta-derived endothelial cells pretreated with interferon gamma to enhance immunogenicity. Pre-clinical studies revealed that ValloVax™ potently inhibits tumor growth in 3 histologically distinct animal models and also suppresses pulmonary metastasis subsequent to intravenous tumor administration . This new approach awaits clinical validation and may serve as a novel anti-angiogenesis vaccine that can be deployed against a variety of tumor types.
Blocking tumor-induced angiogenesis has become a focal point for the development of new cancer therapeutic drugs. Vaccines represent a promising approach for overcoming tolerance to the angiogenic factors that support cancer growth and metastasis. Antigens that are highly expressed on proliferating tumor endothelium, while comparatively downregulated or absent on quiescent endothelium in healthy tissues and during normal physiological angiogenesis are ideal candidates for vaccination strategies targeting the tumor vasculature. Two major types of vaccines are being developed; namely, vaccines against defined angiogenesis-associated antigens, and whole endothelial cell vaccines. The data available thus far support the feasibility of these approaches for generating humoral and cell-mediated immunity against the tumor vasculature and the safety of endothelial cell-based vaccines.
SCW, TEI, HM, JS, JAP, JL, VN, ANP, FMM, and SK all contributed to conceptualization, writing, and review of the manuscript. All authors read and approved the final manuscript.
The authors thank John Peck for his unwavering support in advancing our work and clinical development.
SCW, HM, are board members and officers of Batu Biologics Inc. TI is board member of Batu Biologics Inc. SK is member of scientific advisory board for Batu Biologics Inc.
Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
- Stockmann C, et al. The impact of the immune system on tumor: angiogenesis and vascular remodeling. Front Oncol. 2014;4:69.PubMed CentralPubMedView ArticleGoogle Scholar
- Hirst DG, Denekamp J, Hobson B. Proliferation kinetics of endothelial and tumour cells in three mouse mammary carcinomas. Cell Tissue Kinet. 1982;15(3):251–61.PubMedGoogle Scholar
- Denekamp J, Hobson B. Endothelial-cell proliferation in experimental tumours. Br J Cancer. 1982;46(5):711–20.PubMed CentralPubMedView ArticleGoogle Scholar
- Hobson B, Denekamp J. Endothelial proliferation in tumours and normal tissues: continuous labelling studies. Br J Cancer. 1984;49(4):405–13.PubMed CentralPubMedView ArticleGoogle Scholar
- Folkman J. Tumor angiogenesis: therapeutic implications. N Engl J Med. 1971;285(21):1182–6.PubMedView ArticleGoogle Scholar
- Folkman J. Fighting cancer by attacking its blood supply. Sci Am. 1996;275(3):150–4.PubMedView ArticleGoogle Scholar
- Ellis LM, Fidler IJ. Finding the tumor copycat. Therapy fails, patients don’t. Nat Med. 2010;16(9):974–5.PubMedView ArticleGoogle Scholar
- Dudley AC. Tumor endothelial cells. Cold Spring Harb Perspect Med. 2012;2(3):a006536.PubMed CentralPubMedView ArticleGoogle Scholar
- Cines DB, et al. Endothelial cells in physiology and in the pathophysiology of vascular disorders. Blood. 1998;91(10):3527–61.PubMedGoogle Scholar
- Nagy JA, Dvorak AM, Dvorak HF. VEGF-A and the induction of pathological angiogenesis. Annu Rev Pathol. 2007;2:251–75.PubMedView ArticleGoogle Scholar
- Jain RK. Determinants of tumor blood flow: a review. Cancer Res. 1988;48(10):2641–58.PubMedGoogle Scholar
- Less JR, et al. Microvascular architecture in a mammary carcinoma: branching patterns and vessel dimensions. Cancer Res. 1991;51(1):265–73.PubMedGoogle Scholar
- Goel S, et al. Normalization of the vasculature for treatment of cancer and other diseases. Physiol Rev. 2011;91(3):1071–121.PubMed CentralPubMedView ArticleGoogle Scholar
- Bussolati B, et al. Altered angiogenesis and survival in human tumor-derived endothelial cells. FASEB J. 2003;17(9):1159–61.PubMedGoogle Scholar
- Grange C, et al. Isolation and characterization of human breast tumor-derived endothelial cells. Oncol Rep. 2006;15(2):381–6.PubMedGoogle Scholar
- Xiong YQ, et al. Human hepatocellular carcinoma tumor-derived endothelial cells manifest increased angiogenesis capability and drug resistance compared with normal endothelial cells. Clin Cancer Res. 2009;15(15):4838–46.PubMedView ArticleGoogle Scholar
- Hida K, Klagsbrun M. A new perspective on tumor endothelial cells: unexpected chromosome and centrosome abnormalities. Cancer Res. 2005;65(7):2507–10.PubMedView ArticleGoogle Scholar
- Taylor SM, et al. Angiogenic factor signaling regulates centrosome duplication in endothelial cells of developing blood vessels. Blood. 2010;116(16):3108–17.PubMed CentralPubMedView ArticleGoogle Scholar
- Akiyama K, et al. Tumor endothelial cells acquire drug resistance by MDR1 up-regulation via VEGF signaling in tumor microenvironment. Am J Pathol. 2012;180(3):1283–93.PubMedView ArticleGoogle Scholar
- Griffioen AW. Anti-angiogenesis: making the tumor vulnerable to the immune system. Cancer Immunol Immunother. 2008;57(10):1553–8.PubMed CentralPubMedView ArticleGoogle Scholar
- Griffioen AW, et al. Tumor angiogenesis is accompanied by a decreased inflammatory response of tumor-associated endothelium. Blood. 1996;88(2):667–73.PubMedGoogle Scholar
- Griffioen AW, et al. Angiogenesis inhibitors overcome tumor induced endothelial cell anergy. Int J Cancer. 1999;80(2):315–9.PubMedView ArticleGoogle Scholar
- Dirkx AE, et al. Anti-angiogenesis therapy can overcome endothelial cell anergy and promote leukocyte-endothelium interactions and infiltration in tumors. FASEB J. 2006;20(6):621–30.PubMedView ArticleGoogle Scholar
- Motz GT, et al. Tumor endothelium FasL establishes a selective immune barrier promoting tolerance in tumors. Nat Med. 2014;20(6):607–15.PubMed CentralPubMedView ArticleGoogle Scholar
- Seaman S, et al. Genes that distinguish physiological and pathological angiogenesis. Cancer Cell. 2007;11(6):539–54.PubMed CentralPubMedView ArticleGoogle Scholar
- Wei YQ, et al. Immunogene therapy of tumors with vaccine based on Xenopus homologous vascular endothelial growth factor as a model antigen. Proc Natl Acad Sci USA. 2001;98(20):11545–50.PubMed CentralPubMedView ArticleGoogle Scholar
- Gavilondo JV, et al. Specific active immunotherapy with a VEGF vaccine in patients with advanced solid tumors. Results of the CENTAURO antigen dose escalation phase I clinical trial. Vaccine. 2014;32(19):2241–50.PubMedView ArticleGoogle Scholar
- Katoh O, et al. Expression of the vascular endothelial growth factor (VEGF) receptor gene, KDR, in hematopoietic cells and inhibitory effect of VEGF on apoptotic cell death caused by ionizing radiation. Cancer Res. 1995;55(23):5687–92.PubMedGoogle Scholar
- Hattori K, et al. Vascular endothelial growth factor and angiopoietin-1 stimulate postnatal hematopoiesis by recruitment of vasculogenic and hematopoietic stem cells. J Exp Med. 2001;193(9):1005–14.PubMed CentralPubMedView ArticleGoogle Scholar
- Larrivee B, et al. Vascular endothelial growth factor receptor-2 induces survival of hematopoietic progenitor cells. J Biol Chem. 2003;278(24):22006–13.PubMedView ArticleGoogle Scholar
- Chen R, et al. Anti-metastatic effects of DNA vaccine encoding single-chain trimer composed of MHC I and vascular endothelial growth factor receptor 2 peptide. Oncol Rep. 2015;33(5):2269–76.PubMedGoogle Scholar
- McKinney KA, et al. Effect of a novel DNA vaccine on angiogenesis and tumor growth in vivo. Arch Otolaryngol Head Neck Surg. 2010;136(9):859–64.PubMedView ArticleGoogle Scholar
- Liang PH, et al. Construction of a DNA vaccine encoding Flk-1 extracellular domain and C3d fusion gene and investigation of its suppressing effect on tumor growth. Cancer Immunol Immunother. 2010;59(1):93–101.PubMedView ArticleGoogle Scholar
- Xie K, et al. Anti-tumor effects of a human VEGFR-2-based DNA vaccine in mouse models. Genet Vaccines Ther. 2009;7:10.PubMed CentralPubMedView ArticleGoogle Scholar
- Liu JY, et al. Immunotherapy of tumors with vaccine based on quail homologous vascular endothelial growth factor receptor-2. Blood. 2003;102(5):1815–23.PubMedView ArticleGoogle Scholar
- Zuo SG, et al. Orally administered DNA vaccine delivery by attenuated Salmonella typhimurium targeting fetal liver kinase 1 inhibits murine Lewis lung carcinoma growth and metastasis. Biol Pharm Bull. 2010;33(2):174–82.PubMedView ArticleGoogle Scholar
- Wada S, et al. Rationale for antiangiogenic cancer therapy with vaccination using epitope peptides derived from human vascular endothelial growth factor receptor 2. Cancer Res. 2005;65(11):4939–46.PubMedView ArticleGoogle Scholar
- Ishizaki H, et al. Inhibition of tumor growth with antiangiogenic cancer vaccine using epitope peptides derived from human vascular endothelial growth factor receptor 1. Clin Cancer Res. 2006;12(19):5841–9.PubMedView ArticleGoogle Scholar
- Niethammer AG, et al. A DNA vaccine against VEGF receptor 2 prevents effective angiogenesis and inhibits tumor growth. Nat Med. 2002;8(12):1369–75.PubMedView ArticleGoogle Scholar
- Li Y, et al. Active immunization against the vascular endothelial growth factor receptor flk1 inhibits tumor angiogenesis and metastasis. J Exp Med. 2002;195(12):1575–84.PubMed CentralPubMedView ArticleGoogle Scholar
- Seavey MM, et al. An anti-vascular endothelial growth factor receptor 2/fetal liver kinase-1 Listeria monocytogenes anti-angiogenesis cancer vaccine for the treatment of primary and metastatic Her-2/neu+ breast tumors in a mouse model. J Immunol. 2009;182(9):5537–46.PubMed CentralPubMedView ArticleGoogle Scholar
- Miyazawa M, et al. Phase I clinical trial using peptide vaccine for human vascular endothelial growth factor receptor 2 in combination with gemcitabine for patients with advanced pancreatic cancer. Cancer Sci. 2010;101(2):433–9.PubMedView ArticleGoogle Scholar
- Okamoto I, et al. Clinical phase I study of elpamotide, a peptide vaccine for vascular endothelial growth factor receptor 2, in patients with advanced solid tumors. Cancer Sci. 2012;103(12):2135–8.PubMedView ArticleGoogle Scholar
- Hayashi H, et al. Phase I trial of OTS11101, an anti-angiogenic vaccine targeting vascular endothelial growth factor receptor 1 in solid tumor. Cancer Sci. 2013;104(1):98–104.PubMedView ArticleGoogle Scholar
- Suzuki H, et al. Multiple therapeutic peptide vaccines consisting of combined novel cancer testis antigens and anti-angiogenic peptides for patients with non-small cell lung cancer. J Transl Med. 2013;11:97.PubMed CentralPubMedView ArticleGoogle Scholar
- Magnusson PU, et al. Fibroblast growth factor receptor-1 expression is required for hematopoietic but not endothelial cell development. Arterioscler Thromb Vasc Biol. 2005;25(5):944–9.PubMedView ArticleGoogle Scholar
- Burger PE, et al. Fibroblast growth factor receptor-1 is expressed by endothelial progenitor cells. Blood. 2002;100(10):3527–35.PubMedView ArticleGoogle Scholar
- He QM, et al. Inhibition of tumor growth with a vaccine based on xenogeneic homologous fibroblast growth factor receptor-1 in mice. J Biol Chem. 2003;278(24):21831–6.PubMedView ArticleGoogle Scholar
- Zheng SJ, et al. Synergistic anti-tumor effect of recombinant chicken fibroblast growth factor receptor-1-mediated anti-angiogenesis and low-dose gemcitabine in a mouse colon adenocarcinoma model. World J Gastroenterol. 2007;13(17):2484–9.PubMed CentralPubMedView ArticleGoogle Scholar
- Li M, et al. bFGF peptide combined with the pVAX-8CpG plasmid as adjuvant is a novel anticancer vaccine inducing effective immune responses against Lewis lung carcinoma. Mol Med Rep. 2012;5(3):625–30.PubMedGoogle Scholar
- Plum SM, et al. Administration of a liposomal FGF-2 peptide vaccine leads to abrogation of FGF-2-mediated angiogenesis and tumor development. Vaccine. 2000;19(9–10):1294–303.PubMedView ArticleGoogle Scholar
- Plum SM, et al. Generation of a specific immunological response to FGF-2 does not affect wound healing or reproduction. Immunopharmacol Immunotoxicol. 2004;26(1):29–41.PubMedView ArticleGoogle Scholar
- Liu Z, Wang F, Chen X. Integrin alpha(v)beta(3)-Targeted Cancer Therapy. Drug Dev Res. 2008;69(6):329–39.PubMed CentralPubMedView ArticleGoogle Scholar
- Wilder RL. Integrin alpha V beta 3 as a target for treatment of rheumatoid arthritis and related rheumatic diseases. Ann Rheum Dis. 2002; 61(Suppl 2):ii96–ii99.Google Scholar
- McNeel DG, et al. Phase I trial of a monoclonal antibody specific for alphavbeta3 integrin (MEDI-522) in patients with advanced malignancies, including an assessment of effect on tumor perfusion. Clin Cancer Res. 2005;11(21):7851–60.PubMedView ArticleGoogle Scholar
- Bratt A, et al. Angiomotin belongs to a novel protein family with conserved coiled-coil and PDZ binding domains. Gene. 2002;298(1):69–77.PubMedView ArticleGoogle Scholar
- Holmgren L, et al. A DNA vaccine targeting angiomotin inhibits angiogenesis and suppresses tumor growth. Proc Natl Acad Sci USA. 2006;103(24):9208–13.PubMed CentralPubMedView ArticleGoogle Scholar
- Arigoni M, et al. A vaccine targeting angiomotin induces an antibody response which alters tumor vessel permeability and hampers the growth of established tumors. Angiogenesis. 2012;15(2):305–16.PubMed CentralPubMedView ArticleGoogle Scholar
- Duff SE, et al. CD105 is important for angiogenesis: evidence and potential applications. FASEB J. 2003;17(9):984–92.PubMedView ArticleGoogle Scholar
- Basilio-de-Oliveira RP, Nunes VL. Pannain, Prognostic angiogenic markers (endoglin, VEGF, CD31) and tumor cell proliferation (Ki67) for gastrointestinal stromal tumors. World J Gastroenterol. 2015;21(22):6924–30.PubMed CentralPubMedGoogle Scholar
- Svatek RS, et al. Preoperative plasma endoglin levels predict biochemical progression after radical prostatectomy. Clin Cancer Res. 2008;14(11):3362–6.PubMedView ArticleGoogle Scholar
- Huang FY, et al. Bacterial surface display of endoglin by antigen 43 induces antitumor effectiveness via bypassing immunotolerance and inhibition of angiogenesis. Int J Cancer. 2014;134(8):1981–90.PubMedView ArticleGoogle Scholar
- Jarosz M, et al. Therapeutic antitumor potential of endoglin-based DNA vaccine combined with immunomodulatory agents. Gene Ther. 2013;20(3):262–73.PubMedView ArticleGoogle Scholar
- Ryan BM, O’Donovan N, Duffy MJ. Survivin: a new target for anti-cancer therapy. Cancer Treat Rev. 2009;35(7):553–62.PubMedView ArticleGoogle Scholar
- Lladser A, et al. Intradermal DNA electroporation induces survivin-specific CTLs, suppresses angiogenesis and confers protection against mouse melanoma. Cancer Immunol Immunother. 2010;59(1):81–92.PubMedView ArticleGoogle Scholar
- Lladser A, et al. Naked DNA immunization as an approach to target the generic tumor antigen survivin induces humoral and cellular immune responses in mice. Immunobiology. 2006;211(1–2):11–27.PubMedView ArticleGoogle Scholar
- Park KW, et al. Robo4 is a vascular-specific receptor that inhibits endothelial migration. Dev Biol. 2003;261(1):251–67.PubMedView ArticleGoogle Scholar
- Bedell VM, et al. Roundabout4 is essential for angiogenesis in vivo. Proc Natl Acad Sci U S A. 2005;102(18):6373–8.PubMed CentralPubMedView ArticleGoogle Scholar
- Jones CA, et al. Robo4 stabilizes the vascular network by inhibiting pathologic angiogenesis and endothelial hyperpermeability. Nat Med. 2008;14(4):448–53.PubMed CentralPubMedView ArticleGoogle Scholar
- Seth P, et al. Magic roundabout, a tumor endothelial marker: expression and signaling. Biochem Biophys Res Commun. 2005;332(2):533–41.PubMedView ArticleGoogle Scholar
- Cai H, et al. Roundabout 4 regulates blood-tumor barrier permeability through the modulation of ZO-1, Occludin, and Claudin-5 expression. J Neuropathol Exp Neurol. 2015;74(1):25–37.PubMedView ArticleGoogle Scholar
- Zhuang X, et al. Robo4 vaccines induce antibodies that retard tumor growth. Angiogenesis. 2015;18(1):83–95.PubMedView ArticleGoogle Scholar
- Acevedo LM, Weis SM, Cheresh DA. Robo4 counteracts VEGF signaling. Nat Med. 2008;14(4):372–3.PubMedView ArticleGoogle Scholar
- Lou YY, et al. Immunogene therapy of tumors with a vaccine based on the ligand-binding domain of chicken homologous integrin beta3. Immunol Invest. 2002;31(1):51–69.PubMedView ArticleGoogle Scholar
- Daly C, et al. Angiopoietin-2 functions as a Tie2 agonist in tumor models, where it limits the effects of VEGF inhibition. Cancer Res. 2013;73(1):108–18.PubMedView ArticleGoogle Scholar
- Niu G, et al. Constitutive Stat3 activity up-regulates VEGF expression and tumor angiogenesis. Oncogene. 2002;21(13):2000–8.PubMedView ArticleGoogle Scholar
- Arteaga CL. The epidermal growth factor receptor: from mutant oncogene in nonhuman cancers to therapeutic target in human neoplasia. J Clin Oncol. 2001;19(18 Suppl):32S–40S.PubMedGoogle Scholar
- Fontanini G, et al. Evaluation of epidermal growth factor-related growth factors and receptors and of neoangiogenesis in completely resected stage I-IIIA non-small-cell lung cancer: amphiregulin and microvessel count are independent prognostic indicators of survival. Clin Cancer Res. 1998;4(1):241–9.PubMedGoogle Scholar
- Grandis JR, et al. Normalization of EGFR mRNA levels following restoration of wild-type p53 in a head and neck squamous cell carcinoma cell line. Int J Oncol. 1998;13(2):375–8.PubMedGoogle Scholar
- Garcia B, et al. Effective inhibition of the epidermal growth factor/epidermal growth factor receptor binding by anti-epidermal growth factor antibodies is related to better survival in advanced non-small-cell lung cancer patients treated with the epidermal growth factor cancer vaccine. Clin Cancer Res. 2008;14(3):840–6.PubMedView ArticleGoogle Scholar
- Fernandez Lorente A, et al. Effect of blockade of the EGF system on wound healing in patients vaccinated with CIMAvax(R) EGF. World J Surg Oncol. 2013;11:275.PubMed CentralPubMedView ArticleGoogle Scholar
- Fu C, et al. Identification of a novel membrane protein, HP59, with therapeutic potential as a target of tumor angiogenesis. Clin Cancer Res. 2001;7(12):4182–94.PubMedGoogle Scholar
- Xue Y, et al. PDGF-BB modulates hematopoiesis and tumor angiogenesis by inducing erythropoietin production in stromal cells. Nat Med. 2012;18(1):100–10.View ArticleGoogle Scholar
- Pietras K, et al. Functions of paracrine PDGF signaling in the proangiogenic tumor stroma revealed by pharmacological targeting. PLoS Med. 2008;5(1):e19.PubMed CentralPubMedView ArticleGoogle Scholar
- Falcon BL, et al. Increased vascular delivery and efficacy of chemotherapy after inhibition of platelet-derived growth factor-B. Am J Pathol. 2011;178(6):2920–30.PubMed CentralPubMedView ArticleGoogle Scholar
- Hosaka K, et al. Tumour PDGF-BB expression levels determine dual effects of anti-PDGF drugs on vascular remodelling and metastasis. Nat Commun. 2013;4:2129.PubMedView ArticleGoogle Scholar
- Kaplan CD, et al. A novel DNA vaccine encoding PDGFRbeta suppresses growth and dissemination of murine colon, lung and breast carcinoma. Vaccine. 2006;24(47–48):6994–7002.PubMedView ArticleGoogle Scholar
- Brady J, et al. Human endosialin (tumor endothelial marker 1) is abundantly expressed in highly malignant and invasive brain tumors. J Neuropathol Exp Neurol. 2004;63(12):1274–83.PubMedView ArticleGoogle Scholar
- Rouleau C, et al. Endosialin protein expression and therapeutic target potential in human solid tumors: sarcoma versus carcinoma. Clin Cancer Res. 2008;14(22):7223–36.PubMedView ArticleGoogle Scholar
- Carson-Walter EB, et al. Characterization of TEM1/endosialin in human and murine brain tumors. BMC Cancer. 2009;9:417.PubMed CentralPubMedView ArticleGoogle Scholar
- Bagley RG. Endosialin: from vascular target to biomarker for human sarcomas. Biomark Med. 2009;3(5):589–604.PubMedView ArticleGoogle Scholar
- MacFadyen JR, et al. Endosialin (TEM1, CD248) is a marker of stromal fibroblasts and is not selectively expressed on tumour endothelium. FEBS Lett. 2005;579(12):2569–75.PubMedView ArticleGoogle Scholar
- Christian S, et al. Endosialin (Tem1) is a marker of tumor-associated myofibroblasts and tumor vessel-associated mural cells. Am J Pathol. 2008;172(2):486–94.PubMed CentralPubMedView ArticleGoogle Scholar
- Facciponte JG, et al. Tumor endothelial marker 1-specific DNA vaccination targets tumor vasculature. J Clin Invest. 2014;124(4):1497–511.PubMed CentralPubMedView ArticleGoogle Scholar
- St. Croix B, et al. Genes expressed in human tumor endothelium. Science. 2000;289(5482):1197–202.PubMedView ArticleGoogle Scholar
- Felicetti P, et al. Tumor endothelial marker 8 enhances tumor immunity in conjunction with immnization against differentiation Ag. Cytotherapy. 2007;9(1):23–34.View ArticleGoogle Scholar
- Ruan Z, et al. DNA vaccine against tumor endothelial marker 8 inhibits tumor angiogenesis and growth. J Immunother. 2009;32(5):486–91.PubMedView ArticleGoogle Scholar
- Yang X, Zhu H, Hu Z. Dendritic cells transduced with TEM8 recombinant adenovirus prevents hepatocellular carcinoma angiogenesis and inhibits cells growth. Vaccine. 2010;28(43):7130–5.PubMedView ArticleGoogle Scholar
- Liu P, et al. Anti-tumor angiogenesis effect of genetic fusion vaccine encoding murine beta-defensin 2 and tumor endothelial marker-8 in a CT-26 murine colorectal carcinoma model. Int J Clin Exp Med. 2015;8(3):4744–52.PubMed CentralPubMedGoogle Scholar
- Wei YQ, et al. Immunotherapy of tumors with xenogeneic endothelial cells as a vaccine. Nat Med. 2000;6(10):1160–6.PubMedView ArticleGoogle Scholar
- Okaji Y, et al. Vaccination with autologous endothelium inhibits angiogenesis and metastasis of colon cancer through autoimmunity. Cancer Sci. 2004;95(1):85–90.PubMedView ArticleGoogle Scholar
- Yoshiura K, et al. Inhibition of B16 melanoma growth and metastasis in C57BL mice by vaccination with a syngeneic endothelial cell line. J Exp Clin Cancer Res. 2009;28:13.PubMed CentralPubMedView ArticleGoogle Scholar
- Scappaticci FA, Nolan GP. Induction of anti-tumor immunity in mice using a syngeneic endothelial cell vaccine. Anticancer Res. 2003;23(2B):1165–72.PubMedGoogle Scholar
- Corsini E, et al. Immunotherapy with bovine aortic endothelial cells in subcutaneous and intracerebral glioma models in rats: effects on survival time, tumor growth, and tumor neovascularization. Cancer Immunol Immunother. 2004;53(11):955–62.PubMedView ArticleGoogle Scholar
- Zhang W, Liu JN, Tan XY. Vaccination with xenogeneic tumor endothelial proteins isolated in situ inhibits tumor angiogenesis and spontaneous metastasis. Int J Cancer. 2009;125(1):124–32.PubMedView ArticleGoogle Scholar
- Okaji Y, et al. Pilot study of anti-angiogenic vaccine using fixed whole endothelium in patients with progressive malignancy after failure of conventional therapy. Eur J Cancer. 2008;44(3):383–90.PubMedView ArticleGoogle Scholar
- Tanaka M, et al. Human umbilical vein endothelial cell vaccine therapy in patients with recurrent glioblastoma. Cancer Sci. 2013;104(2):200–5.PubMedView ArticleGoogle Scholar
- Friedman HS, et al. Bevacizumab alone and in combination with irinotecan in recurrent glioblastoma. J Clin Oncol. 2009;27(28):4733–40.PubMedView ArticleGoogle Scholar
- Reynolds LP, Redmer DA. Angiogenesis in the placenta. Biol Reprod. 2001;64(4):1033–40.PubMedView ArticleGoogle Scholar
- Harandi A. Immunoplacental therapy, a potential multi-epitope cancer vaccine. Med Hypotheses. 2006;66(6):1182–7.PubMedView ArticleGoogle Scholar
- Holtan SG, et al. Cancer and pregnancy: parallels in growth, invasion, and immune modulation and implications for cancer therapeutic agents. Mayo Clin Proc. 2009;84(11):985–1000.PubMed CentralPubMedView ArticleGoogle Scholar
- Vonnahme KA, et al. Placental vascularity and growth factor expression in singleton, twin, and triplet pregnancies in the sheep. Endocrine. 2008;33(1):53–61.PubMedView ArticleGoogle Scholar
- Muschol-Steinmetz C, et al. Function of survivin in trophoblastic cells of the placenta. PLoS One. 2013;8(9):e73337.PubMed CentralPubMedView ArticleGoogle Scholar
- Olsen DT, et al. Purification and characterization of a soluble calnexin from human placenta. Protein Expr Purif. 2013;92(1):105–11.PubMedView ArticleGoogle Scholar
- Troyanovsky B, et al. Angiomotin: an angiostatin binding protein that regulates endothelial cell migration and tube formation. J Cell Biol. 2001;152(6):1247–54.PubMed CentralPubMedView ArticleGoogle Scholar
- Liao WX, et al. Human placental expression of SLIT/ROBO signaling cues: effects of preeclampsia and hypoxia. Biol Reprod. 2012;86(4):111.PubMed CentralPubMedView ArticleGoogle Scholar
- Holmgren L, et al. Angiogenesis during human extraembryonic development involves the spatiotemporal control of PDGF ligand and receptor gene expression. Development. 1991;113(3):749–54.PubMedGoogle Scholar
- Moscatelli DA, et al. Purification and biological activities of an angiogenesis factor from human placenta. Anticancer Res. 1986;6(4):861–3.PubMedGoogle Scholar
- Ugele B, Lange F. Isolation of endothelial cells from human placental microvessels: effect of different proteolytic enzymes on releasing endothelial cells from villous tissue. Vitro Cell Dev Biol Anim. 2001;37(7):408–13.View ArticleGoogle Scholar
- Kacemi A, et al. Isolation of villous microvessels from the human placenta. C R Acad Sci III. 1997;320(2):171–7.PubMedView ArticleGoogle Scholar
- Schugar RC, et al. High harvest yield, high expansion, and phenotype stability of CD146 mesenchymal stromal cells from whole primitive human umbilical cord tissue. J Biomed Biotechnol. 2009;2009:789526.PubMed CentralPubMedView ArticleGoogle Scholar
- Ichim TE, et al. Induction of tumor inhibitory anti-angiogenic response through immunization with interferon Gamma primed placental endothelial cells: ValloVax. J Transl Med. 2015;13:90.PubMed CentralPubMedView ArticleGoogle Scholar