Identification Of Tumor-associated Antigens Research Articles

Systems Biology Approaches for the Improvement of Oncolytic Virus-Based Immunotherapies.

Oncolytic virus (OV)-based immunotherapy is mainly dependent on establishing an efficient cell-mediated antitumor immunity. OV-mediated antitumor immunity elicits a renewed antitumor reactivity, stimulating a T-cell response against tumor-associated antigens (TAAs) and recruiting natural killer cells within the tumor microenvironment (TME). Despite the fact that OVs are unspecific cancer vaccine platforms, to further enhance antitumor immunity, it is crucial to identify the potentially immunogenic T-cell restricted TAAs, the main key orchestrators in evoking a specific and durable cytotoxic T-cell response. Today, innovative approaches derived from systems biology are exploited to improve target discovery in several types of cancer and to identify the MHC-I and II restricted peptide repertoire recognized by T-cells. Using specific computation pipelines, it is possible to select the best tumor peptide candidates that can be efficiently vectorized and delivered by numerous OV-based platforms, in order to reinforce anticancer immune responses. Beyond the identification of TAAs, system biology can also support the engineering of OVs with improved oncotropism to reduce toxicity and maintain a sufficient portion of the wild-type virus virulence. Finally, these technologies can also pave the way towards a more rational design of armed OVs where a transgene of interest can be delivered to TME to develop an intratumoral gene therapy to enhance specific immune stimuli.

Identification of tumor-associated antigens and immune subtypes of lower-grade glioma and glioblastoma for mRNA vaccine development

BackgroundmRNA became a promising therapeutic approach in many diseases. This study aimed to identify the tumor antigens specifically expressed in tumor cells for lower-grade glioma (LGG) and glioblastoma (GBM) patients.MethodsIn this work, the mRNA microarray expression profile and clinical data were obtained from 301 samples in the Chinese Glioma Genome Atlas (CGGA) database, the mRNA sequencing data and clinical data of 701 samples were downloaded from The Cancer Genome Atlas (TCGA) database. Genetic alterations profiles were extracted from CGGA and cBioPortal datasets. R language and GraphPad Prism software were applied for the statistical analysis and graph work.ResultsPTBP1 and SLC39A1, which were overexpressed and indicated poor prognosis in LGG patients, were selected as tumor-specific antigens for LGG patients. Meanwhile, MMP9 and SLC16A3, the negative prognostic factors overexpressed in GBM, were identified as tumor-specific antigens for GBM patients. Besides, three immune subtypes (LGG1-LGG3) and eight WGCNA modules were identified in LGG patients. Meanwhile, two immune subtypes (GBM1–GBM2) and 10 WGCNA modules were selected in GBM. The immune characteristics and potential functions between different subtypes were diversity. LGG2 and GBM1 immune subtype were associated with longer overall survival than other subtypes.ConclusionIn this study, PTBP1 and SLC39A1 are promising antigens for mRNA vaccines development in LGG, and MMP9 and SLC16A3 were potential antigens in GBM. Our analyses indicated that mRNA vaccine immunotherapy was more suitable for LGG2 and GBM1 subtypes. This study was helpful for the development of glioma immunotherapies.

Identification Of Liver Mitochondria Tumor Associated Antigens Of The Mice Exposed To N, Nitrosodibutylamine: A Promising Strategy In Development Of Anticancer Treatment

Background: N-Nitrosodibutylamine possess the ability to induce bladder tumor as well as in the liver, and oesophagus when it is administered in the body.Exposure to N-Nitrosodibutylamine can happen by different modes such as by ingestion,inhalation and dermal contact.Methods:In the present investigation an attempt has been done to identify ,isolate as well to purify the tumor associated antigens (TAA) from the liver mitochondria of the mice when exposed to N-Nitrosodibutylamine.Results:It was found that mitochondrial membrane protein of N-Nitrosodibutylamine-exposed animals exhibited differential expression of protein when compared with the control animals. A low molecular weight (~14 kDa) protein was found to be over expressed significantly on liver mitochondrial membrane upon N-Nitrosodibutylamine exposure in mice as compared with the normal control and identified as TAA,showing the sign that some of the proteins could be used as TAA for further study.Conclusion: These identification of TAAs will provide the basis for the use of mitochondria as markers for the early detection for the treatment of liver cancer.

Identification of tumor-associated antigens with diagnostic ability of colorectal cancer by in-depth immunomic and seroproteomic analysis

Colorectal cancer (CRC) is the third most common cancer and the second leading cause of cancer related death worldwide. Its diagnosis at early stages would significantly improve the survival of CRC patients. The humoral immune response has been demonstrated useful for cancer diagnosis, predating clinical symptoms up to 3 years. Here, we employed an in-depth seroproteomic approach to identify proteins that elicit a humoral immune response in CRC patients. The seroproteomic approach relied on the immunoprecipitation with patient-derived autoantibodies of proteins from CRC cell lines with different metastatic properties followed by LC-MS/MS. After bioinformatics, we focused on 31 targets of CRC autoantibodies. After WB and IHC validation, ERP44 and TALDO1 showed potential to discriminate disease-free and metastatic CRC patients, and time to recurrence of CRC patients in stage II. Using plasma samples of 30 healthy individuals, 28 premalignant individuals, and 32 CRC patients, nine out of 13 selected targets for seroreactive analysis showed significant diagnostic ability to discriminate either CRC patients or premalignant subjects from controls. Our results suggest that the here defined panel of CRC autoantibodies and their target proteins should be included in CRC blood-based biomarker panels to get a clinically useful blood-based diagnostic signature for CRC detection. SignificanceColorectal cancer is one of the deadliest cancer types mainly due to its late diagnosis. Its early diagnosis, therefore, is of great importance since it would significantly improve the survival of CRC patients. In our work, the in-depth seroproteomic analysis of colorectal cancer using isolated IgGs from colorectal cancer patients and controls and protein extract of colorectal cancer cells provide the identification of valuable biomarkers with diagnostic and prognostic ability of the disease.

Discovering novel lung cancer associated antigens and the utilization of their autoantibodies in detection of lung cancer.

The identification of tumor-associated antigens (TAAs) and their corresponding autoantibodies in lung cancer (LC) may expand our vision of cancer immunity. This study aims to screen novel TAAs to distinguish LC from the healthy population. In our previous study, 35 genes encoding LC-associated TAAs were identified from the serological analysis of recombinant cDNA expression libraries (SEREX), and Oncomine database was further used to identify potential genes in cancer progression. Autoantibody to TAAs were tested by enzyme-linked immunosorbent assay (ELISA) in sera from 1379 participants in validation set and verification set. Based on analysis of three independent microarrays in Oncomine, ten genes were consistently dysregulated in LC. The sera level and positive frequency of the anti-TOP2A, anti-ACTR3, anti-RPS6KA5 and anti-PSIP1 from LC patients were higher than normal control in validation set. The area under curve (AUC) of anti-TOP2A, anti-ACTR3, anti-RPS6KA5 and anti-PSIP1 was respectively 0.758, 0.787, 0.707, 0.668. The sensitivity of these four autoantibodies for LC detection ranged from 26.63 % to 32.07 % with the specificity over 90 %. Data from the verification set confirmed the results. Except that, the frequency of serum autoantibody against TOP2A (43.3 %) and ACTR3 (50.0 %) was significantly higher in early stage LC than late stage (23.6 % and 22.3 %, respectively). TOP2A, ACTR3, RPS6KA5 and PSIP1 can elicit humoral immune response in LC and their autoantibodies have relationship with the tumorigenesis of LC. Anti-TOP2A and anti-ACTR3 have the potential to serve as a serological biomarkers in early stage LC.

Enhanced VISTA Expression in a Subset of Patients with Acute Myeloid Leukemia

Introduction: Identification of tumor associated antigens (TAA) in acute myeloid leukemia (AML) will facilitate the use of targeted immunotherapies such as chimeric antigen receptor T cell therapy and bispecific antibodies. The characteristics of an ideal TAA include tumor specificity, high expression level, large percentage of positive tumor cells, and evidence for expression on putative cancer stem cells. V-domain Ig Suppressor of T cell Activation (VISTA) is a cell surface protein previously shown to be present on hematologic cells and inhibitory towards T cells. VISTA's role and presence within AML has yet to be defined. We performed detailed studies to characterize VISTA expression on tumor cells and mononuclear cell subsets in patients with AML.Methods: Immunophenotyping was performed on mononuclear cells from primary AML patient specimens using mass cytometry (CyTOF) by staining samples with a panel containing 34 antibodies to surface markers and cytokines (Table 1). This antibody panel was used to identify blast cells (variable immunophenotype), leukemic stem cells (LSCs) (CD34+, CD38-), T cells (CD3+), monocytes (CD33+, CD14+, CD11b+, HLA-DR+), myeloid derived suppressor cells (MDSCs) (CD33+, CD11b+, HLA-DR-), and M2 macrophages (CD11b+, CD163+). Raw median VISTA expression and percentage of cells expressing VISTA were calculated for all of these cell types. Samples were split into 2 categories based on blast VISTA expression. A sample was considered VISTA high if >10% of the blasts expressed VISTA, and VISTA low if <10% of the blasts expressed VISTA. Similar data was generated from a pool of healthy donors for comparison.Results: Fourteen bone marrow samples from patients with newly diagnosed AML and 10 from healthy donors were obtained. The average percentage of blasts expressing VISTA was 68% (range 28% - 81%) in the VISTA high group and 4.2% (range 0.3% - 6.8%) in the VISTA low group (p<0.05). VISTA expression was higher in the VISTA high group than in the VISTA low group (17.7 vs -0.1, p<0.05). The VISTA high group had a lower blast percentage in the bone marrow compared to the VISTA low group (56% vs 91%, p<0.05). The VISTA high and low groups had similar percentages of LSCs (2.5% vs 0.6%, p=0.11), but the LSCs in the VISTA high group maintained higher VISTA expression than did those in the VISTA low group (6.81 vs -0.11, p<0.05). The LSCs within the VISTA high group had similar VISTA expression to the blast population (6.81 vs 17.7, p=0.15). Within the non-blast population, VISTA expression was higher on the myeloid cell subsets than on the T cells for both the AML samples (11.5 vs 0.17, p<0.05) and the healthy donor samples (8.07 vs 0.24, p<0.05). VISTA expression was similar between the VISTA high and low groups on T cells (0.36 vs 0.02, p=0.08), monocytes (15.7 vs 19, p=0.7), MDSCs (6.3 vs 4, p=0.26), and M2 macrophages (7.4 vs 16.6, p =0.25). VISTA expression was similar between AML and healthy donor samples for T cells (0.17 vs 0.24, p=0.6), monocytes (15.7 vs 14, p=0.41), MDSCs (6.3 vs 3.6, p=0.22), and M2 macrophages (7.4 vs 6.6, p=0.19). Monocytes had the highest expression of VISTA within the healthy donor samples (p<0.05) and this difference was maintained in the AML samples.Conclusion: A subset of patients with AML has a large percentage of blasts, including LSCs, with high VISTA expression on their surface. VISTA is expressed on normal myeloid cells in the bone marrow of patients with AML, but at low levels and to a similar degree as in the marrow of healthy donors. [Display omitted] [Display omitted] DisclosuresHuang:Janssen Research & Development, LLC: Employment, Other: I am an employee of Janssen and a stock owner . Sasser:Janssen Research & Development, LLC: Employment. Adams:Janssen Research & Development, LLC: Employment. Druker:Agios: Honoraria; Ambit BioSciences: Consultancy; ARIAD: Patents & Royalties, Research Funding; Array: Patents & Royalties; AstraZeneca: Consultancy; Blueprint Medicines: Consultancy, Equity Ownership, Other: travel, accommodations, expenses ; BMS: Research Funding; CTI: Equity Ownership; Curis: Patents & Royalties; Cylene: Consultancy, Equity Ownership; D3 Oncology Solutions: Consultancy; Gilead Sciences: Consultancy, Other: travel, accommodations, expenses ; Lorus: Consultancy, Equity Ownership; MolecularMD: Consultancy, Equity Ownership, Patents & Royalties; Novartis: Research Funding; Oncotide Pharmaceuticals: Research Funding; Pfizer: Patents & Royalties; Roche: Consultancy. Lind:Janssen: Research Funding; Fluidigm: Honoraria.

Serum Protein Markers for the Early Detection of Lung Cancer: A Focus on Autoantibodies

Lung cancer has the highest mortality rate among cancer patients in the world, in particular because most patients are only diagnosed at an advanced and noncurable stage. Computed tomography (CT) screening on high-risk individuals has shown that early detection could reduce the mortality rate. However, the still high false-positive rate of CT screening may harm healthy individuals because of unnecessary follow-up scans and invasive follow-up procedures. Alternatively, false-negative and indeterminate results may harm patients due to the delayed diagnosis and treatment of lung cancer. Noninvasive biomarkers, complementary to CT screening, could lower the false-positive and false-negative rate of CT screening at baseline and thereby reduce the number of patients that need follow-up and diagnose patients at an earlier stage of lung cancer. Lung cancer tissue generates lung cancer-associated proteins to which the immune system might produce high-affinity autoantibodies. This autoantibody response to tumor-associated antigens starts during early stage lung cancer and may endure over years. Identification of tumor-associated antigens or the corresponding autoantibodies in body fluids as potential noninvasive biomarkers could thus be an effective approach for early detection and monitoring of lung cancer. We provide an overview of differentially expressed protein, antigen, and autoantibody biomarkers that combined with CT imaging might be of clinical use for early detection of lung cancer.

Novel cancer antigens for personalized immunotherapies: latest evidence and clinical potential.

The clinical success of monoclonal antibody immune checkpoint modulators such as ipilimumab, which targets cytotoxic T lymphocyte-associated antigen 4 (CTLA-4), and the recently approved agents nivolumab and pembrolizumab, which target programmed cell death receptor 1 (PD-1), has stimulated renewed enthusiasm for anticancer immunotherapy, which was heralded by Science as 'Breakthrough of the Year' in 2013. As the potential of cancer immunotherapy has been recognized since the 1890s when William Coley showed that bacterial products could be beneficial in cancer patients, leveraging the immune system in the treatment of cancer is certainly not a new concept; however, earlier attempts to develop effective therapeutic vaccines and antibodies against solid tumors, for example, melanoma, frequently met with failure due in part to self-tolerance and the development of an immunosuppressive tumor microenvironment. Increased knowledge of the mechanisms through which cancer evades the immune system and the identification of tumor-associated antigens (TAAs) and negative immune checkpoint regulators have led to the development of vaccines and monoclonal antibodies targeting specific tumor antigens and immune checkpoints such as CTLA-4 and PD-1. This review first discusses the established targets of currently approved cancer immunotherapies and then focuses on investigational cancer antigens and their clinical potential. Because of the highly heterogeneous nature of tumors, effective anticancer immunotherapy-based treatment regimens will likely require a personalized combination of therapeutic vaccines, antibodies and chemotherapy that fit the specific biology of a patient's disease.

Vaccine based immunotherapy as a strategy to bypass drug resistance in multiple myeloma

Clonal evolution under therapy leads to drug resistance and relapse in multiple myeloma (MM), requiring new treatment mo- dalities to control disease. Immunotherapy and specifically vacci- nation to induce anti-tumor CD8+ cytotoxic T-cells (CTLs) provides one such strategy, but susceptibility of drug resistant MM cells to CTL attack is as yet not defined. To examine this, a first requirement is identification of tumour-associated antigens in drug refractory MM. To this end, we had previously specifically evaluated cancer testis antigens (CTAs) in MM for retention at relapse post- therapy, and identified SPAG9 and MAGEC1 as frontline antigen targets for vaccines. Of these, SPAG9 is generic to relapse MM (expressed in w100% of cases) and MAGEC1 is essentially tumour-specific (expressed w60% of cases). An optimal strategy for vaccination as therapy is in disease remission to attack re-emerging MM cells that escape drug therapy. A second requirement is to model drug resistance in MM, and accordingly we have now generated derivatives of the XG2 MM cell line that are resistant to melphalan (XG2 M) or Bortezomib (XG2 600R3); the RPMI8226-derivative R5 line with multiple drug resistance was also available. A third requirement is to show effective lysis of drug resistant MM cells by antigen specific CTLs. To generate antigen-specific CTLs, we utilized pDOM-CTAepitope DNA vaccine delivery in HLA-A2 transgenic HHD mice. We examined two epitopes in respective vaccines, pDOM-SPAG9p1 and pDOM-MAGEC1p2. Both vac- cines elicited epitope specific T-cell responses in HHD measured by IFN release assays following prime and boost with electroporation. Significantly, using MM cell lines made chimeric for HHD MHC class I as targets, both vaccines induced CTLs in HHD mice that were able to effectively lyse primary (XG2 and others) and drug resistant MM cell lines (XG2M, XG2 600R3, R5) effectively. These data demonstrate that drug resistant MM cells are susceptible to CTL killing, substantiating a role for vaccination in MM to over- come drug resistance and hence prevent relapse.

Abstract 1324: A translational platform to design antibodies targeting triple negative breast cancer-specific antigens for cancer immunotherapy

Abstract Triple negative breast cancers (TNBCs) are characterised by the lack of estrogen receptor, progesterone receptor and human epidermal growth factor receptor 2 (HER2) rendering them insensitive to current endocrine and anti-HER2 therapies, therefore novel treatment options are required. Breast cancers have been shown to be immunogenic and patient tumors feature infiltrating immune cells. Since monoclonal antibodies are an important therapeutic modality in the clinical management of many cancers, engineering antibodies against triple negative breast cancer specific antigens that may activate immune effector cells within tumors and in the circulation may lead to tumour clearance and may provide new treatment strategies for this patient group. Here we describe a platform for selection of TNBC-associated antigens, engineering of novel antibodies against these antigens and functional studies to elucidate antibody mechanisms of action and therapeutic potential in vitro and vivo. Our approach involves identification of tumour-associated antigens from patient tumor specimens using a transcriptomics approach and validation and evaluation of biological functions in breast cancer cells using cell culture models. Antibody heavy (VH) and light (VL) sequences are cloned in a dual expression vector system for recombinant expression in HEK293 cells to rapidly produce antibodies with reactivity to specific tumor antigens. The system is optimised for the seamless expression of V genes of any species including human sequences, along with human constant (C) genes of different antibody classes and subclasses. The latter permits the study of different class/subclass antibodies in order to identify those most effective in triggering immune cell activation and tumor clearance. Engineered monoclonal antibodies are subsequently tested for their antigen binding properties and capacity to activate immune effector cells against cancer cells in vitro. The therapeutic potential of selected candidates is evaluated in vivo using xenograft TNBC mouse models engrafted with components of the human immune system using adoptive transfer of healthy volunteer and patient derived peripheral blood cells. Xenograft models featuring human immune components serve to evaluate the therapeutic potential of monoclonal antibodies in the context of their capacity to redirect immune effector functions against cancer cells. Our findings demonstrate a translational pathway designed to facilitate the discovery and evaluation of novel antibody therapeutic options for triple negative breast cancer immunotherapy. Citation Format: Kristina M. Ilieva, Rebecca Marlow, Anthony Cheung, Erika Francesch, Panagiotis Karagiannis, Matthew Fittall, Silvia Crescioli, Andrew Tutt, Sophia Karagiannis. A translational platform to design antibodies targeting triple negative breast cancer-specific antigens for cancer immunotherapy. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1324. doi:10.1158/1538-7445.AM2015-1324

Optimized dendritic cell-based immunotherapy for melanoma: the TriMix-formula

Since decades, the main goal of tumor immunologists has been to increase the capacity of the immune system to mediate tumor regression. In this regard, one of the major focuses of cancer immunotherapy has been the design of vaccines promoting strong tumor-specific cytotoxic T lymphocyte responses in cancer patients. Here, dendritic cells (DCs) play a pivotal role as they are regarded as nature's adjuvant and as such have become the natural agents for antigen delivery in order to finally elicit strong T cell responses (Villadangos and Schnorrer in Nat Rev Immunol 7:543-555, 2007; Melief in Immunity 29:372-383, 2008; Palucka and Banchereau in Nat Rev Cancer 12:265-277, 2012; Vacchelli et al. in Oncoimmunology 2:e25771, 2013; Galluzzi et al. in Oncoimmunology 1:1111-1134, 2012). Therefore, many investigators are actively pursuing the use of DCs as an efficient way of inducing anticancer immune responses. Nowadays, DCs can be generated at a large scale in closed systems, yielding sufficient numbers of cells for clinical application. In addition, with the identification of tumor-associated antigens, which are either selectively or preferentially expressed by tumors, a whole range of strategies using DCs for immunotherapy have been designed and tested in clinical studies. Despite the evidence that DCs loaded with tumor-associated antigens can elicit immune responses in vivo, clinical responses remained disappointingly low. Therefore, optimization of the cellular product and route of administration was urgently needed. Here, we review the path we have followed in the development of TriMixDC-MEL, a potent DC-based cellular therapy, discussing its development as well as further modifications and applications.

Glycoprotein non-metastatic b (GPNMB): A metastatic mediator and emerging therapeutic target in cancer

Molecularly targeted therapies are rapidly growing with respect to their clinical development and impact on cancer treatment due to their highly selective anti-tumor action. However, many aggressive cancers such as triple-negative breast cancer (TNBC) currently lack well-defined therapeutic targets against which such agents can be developed. The identification of tumor-associated antigens and the generation of antibody drug-conjugates represent an emerging area of intense interest and growth in the field of cancer therapeutics. Glycoprotein non-metastatic b (GPNMB) has recently been identified as a gene that is over-expressed in numerous cancers, including TNBC, and often correlates with the metastatic phenotype. In breast cancer, GPNMB expression in the tumor epithelium is associated with a reduction in disease-free and overall survival. Based on these findings, glembatumumab vedotin (CDX-011), an antibody-drug conjugate that selectively targets GPNMB, is currently being investigated in clinical trials for patients with metastatic breast cancer and unresectable melanoma. This review discusses the physiological and potential pathological roles of GPNMB in normal and cancer tissues, respectively, and details the clinical advances and challenges in targeting GPNMB-expressing malignancies.

Screening of Multiple Myeloma by Polyclonal Rabbit Anti-Human Plasmacytoma Cell Immunoglobulin

Antibody-based immunotherapy has been effectively used for tumor treatment. However, to date, only a few tumor-associated antigens (TAAs) or therapeutic targets have been identified. Identification of more immunogenic antigens is essential for improvements in multiple myeloma (MM) diagnosis and therapy. In this study, we synthesized a polyclonal antibody (PAb) by immunizing rabbits with whole human plasmacytoma ARH-77 cells and identified MM-associated antigens, including enlonase, adipophilin, and HSP90s, among others, via proteomic technologies. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay showed that 200 µg/mL PAb inhibits the proliferation of ARH-77 cells by over 50% within 48 h. Flow cytometric assay indicated that PAb treatment significantly increases the number of apoptotic cells compared with other treatments (52.1% vs. NS, 7.3% or control rabbit IgG, 9.9%). In vivo, PAb delayed tumor growth and prolonged the lifespan of mice. Terminal deoxynucleotidyl transferase dUTP nick end labeling assay showed that PAb also induces statistically significant changes in apoptosis compared with other treatments (P<0.05). We therefore conclude that PAb could be used for the effective screening and identification of TAA. PAb may have certain anti-tumor functions in vitro and in vivo. As such, its combination with proteomic technologies could be a promising approach for sieving TAA for the diagnosis and therapy of MM.

Molecular Mimics of the Tumour Antigen MUC1

A key requirement for the development of cancer immunotherapy is the identification of tumour-associated antigens that are differentially or exclusively expressed on the tumour and recognized by the host immune system. However, immune responses to such antigens are often muted or lacking due to the antigens being recognized as “self”, and further complicated by the tumour environment and regulation of immune cells within. In an effort to circumvent the lack of immune responses to tumour antigens, we have devised a strategy to develop potential synthetic immunogens. The strategy, termed mirror image phage display, is based on the concept of molecular mimicry as demonstrated by the idiotype/anti-idiotype paradigm in the immune system. Here as ‘proof of principle’ we have selected molecular mimics of the well-characterised tumour associated antigen, the human mucin1 protein (MUC1) from two different peptide phage display libraries. The putative mimics were compared in structure and function to that of the native antigen. Our results demonstrate that several of the mimic peptides display T-cell stimulation activity in vitro when presented by matured dendritic cells. The mimic peptides and the native MUC1 antigenic epitopes can cross-stimulate T-cells. The data also indicate that sequence homology and/or chemical properties to the original epitope are not the sole determining factors for the observed immunostimulatory activity of the mimic peptides.

Immuno-PET of Cancer: A Revival of Antibody Imaging

See page [1173][1] With the identification of tumor-associated antigens more than 30 y ago and the development of hybridoma technology in 1975, investigators realized that Paul Ehrlich's vision to effectively target drugs to diseased tissues could be tested. It was shown that monoclonal antibodies

Conditions of tumor-associated antigens as a proper target for therapeutic antibodies against solid cancers

Since the success of rituximab and trastuzumab for treatment of non-Hodgkin's lymphoma and breast cancer, respectively, a huge therapeutic potential of monoclonal antibodies (mAbs) was realized and development of therapeutic mAbs has been widely tried against various cancers. However, the successful examples are still limited and therapeutic mAbs are not yet available for the majority of human cancers. We established a procedure for comprehensive identification of tumor-associated antigens (TAAs) through the extensive isolation of human mAbs that may become therapeutic. Thirty-twoTAAs have been identified and 555 mAbs that bound to one of the TAAs have been isolated to date. Now we are trying to select TAAs as proper targets for therapy and candidate mAbs as drugs from among them. The immunohistochemical analysis using many fresh lung cancer specimens suggested probabilities of proper targets, and moreover, presence of cancer-specific epitopes that could be distinguished from normal epitopes on the same molecules by mAbs. For Abs to efficiently kill the cancer cells they should have the ability to induce immunological cytotoxicity such as ADCC and/or CDC. They should also be able to inhibit the function mediated by the target Ags. For clinical point of view, the continuous presence of the target molecule on the cell surface until cell death might be essential for successful treatment. Therefore, it will be required for targets TAAs to play essential roles in tumorigenesis. Otherwise the cancer cells that do not express them could selectively survive during treatment and finally become dominant. It was also suggested that even the same molecules could play different roles in tumorigenesis quite often in different patients. Therefore when we develop therapeutic Abs, we should obtain information about the conditions of patients including genetic background to whom the treatment will be effective. I will discuss how we can accomplish this purpose.

Identification of HLA-A2-restricted CTL epitopes of a novel tumour-associated antigen, KIF20A, overexpressed in pancreatic cancer

Background:Identification of tumour-associated antigens (TAAs) that induce cytotoxic T lymphocytes (CTLs) specific to cancer cells is critical for the development of anticancer immunotherapy. In this study, we aimed at identifying a novel TAA of pancreatic cancer for immunotherapy.Methods:On the basis of the genome-wide cDNA microarray analysis, we focused on KIF20A (also known as RAB6KIFL/MKlp2) as a candidate TAA in pancreatic cancer cells. The HLA-A2 (A*02:01)-restricted CTL epitopes of KIF20A were identified using HLA-A2 transgenic mice (Tgm) and the peptides were examined to check whether they could generate human CTLs exhibiting cytotoxic responses against KIF20A+, HLA-A2+ tumour cells in vitro.Results:KIF20A was overexpressed in pancreatic cancer and in some other malignancies, but not in their non-cancerous counterparts and many normal adult tissues. We found that KIF20A-2 (p12–20, LLSDDDVVV), KIF20A-8 (p809–817, CIAEQYHTV), and KIF20A-28 (p284–293, AQPDTAPLPV) peptides could induce HLA-A2-restricted CTLs in HLA-A2 Tgm without causing autoimmunity. Peptide-reactive human CTLs were generated from peripheral blood mononuclear cells of HLA-A2+ healthy donors by in vitro stimulation with the three peptides, and those CTLs successfully exhibited cytotoxic responses to cancer cells expressing both KIF20A and HLA-A2.Conclusion:KIF20A is a novel promising candidate for anticancer immunotherapeutic target for pancreatic cancers.

Antibody responses to galectin-8, TARP and TRAP1 in prostate cancer patients treated with a GM-CSF-secreting cellular immunotherapy.

A critical factor in clinical development of cancer immunotherapies is the identification of tumor-associated antigens that may be related to immunotherapy potency. In this study, protein microarrays containing >8,000 human proteins were screened with serum from prostate cancer patients (N = 13) before and after treatment with a granulocyte-macrophage colony-stimulating factor (GM-CSF)-secreting whole cell immunotherapy. Thirty-three proteins were identified that displayed significantly elevated (P <or= 0.05) signals in post-treatment samples, including three proteins that have previously been associated with prostate carcinogenesis, galectin-8, T-cell alternative reading frame protein (TARP) and TNF-receptor-associated protein 1 (TRAP1). Expanded analysis of antibody induction in metastatic, castration-resistant prostate cancer (mCRPC) patients (N = 92) from two phase 1/2 trials of prostate cancer immunotherapy, G-9803 and G-0010, indicated a significant (P = 0.03) association of TARP antibody induction and median survival time (MST). Antibody induction to TARP was also significantly correlated (P = 0.036) with an increase in prostate-specific antigen doubling time (PSADT) in patients with a biochemical (PSA) recurrence following prostatectomy or radiation therapy (N = 19) from in a previous phase 1/2 trial of prostate cancer immunotherapy, G-9802. RNA and protein encoding TARP and TRAP1 was up-regulated in prostate cancer tissue compared to matched normal controls. These preliminary findings suggest that antibody induction to TARP may represent a possible biomarker for treatment response to GM-CSF secreting cellular immunotherapy in prostate cancer patients and demonstrates the utility of using protein microarrays for the high-throughput screening of patient-derived antibody responses.

Identification of Tumor-Associated Antigens from Medullary Breast Carcinoma by a Modified SEREX Approach

Medullary breast carcinoma (MBC) is a relatively rare malignancy with heavy lymphocytic infiltration that despite cytologically anaplastic features and high mitotic index has more favorable prognosis than other types of breast cancer. Lymphocytic infiltration of tumors reflects ongoing immune response against tumor antigens which could represent a great interest as potential targets for cancer immunotherapy. The search for MBC antigens by SEREX methodology has not been successful due to a very high titer of false positive clones, representing immunoglobulin genes. Here, we describe a novel approach for generating cDNA expression libraries from MBC tumor samples which are depleted of IgG cDNA clones and, therefore, are suitable for the identification of novel tumor-associated antigens (TAA) by SEREX approach. Modified methodology allowed us to isolate a panel of known and novel TAA which are currently under further investigation.

Immune surveillance of human cancer: if the cytotoxic T‐lymphocytes play the music, does the tumoral system call the tune?

Accumulating evidence indicates that the innate and adaptive immune systems participate in the recognition and destruction of cancer cells by a process known as cancer immunosurveillance. Tumor antigen-specific cytotoxic T-lymphocytes (CTL) are the major effectors in the immune response against tumor cells. The identification of tumor-associated antigen (TAA) recognized primarily by CD 8(+) T-lymphocytes has led to the development of several vaccination strategies that induce or potentiate specific immune responses. However, large established tumors, which are associated with the acquisition of tumor resistance to specific lysis, are usually not fully controlled by the immune system. Recently, it has become clear that the immune system not only protects the host against tumor development but also sculpts the immunogenic phenotype of a developing tumor and can favor the emergence of resistant tumor cell variants. Moreover, it has become obvious that the evasion of immunosurveillance by tumor cells is under the control of the tumor microenvironment complexity and plasticity. In this review, we will focus on some new mechanisms associated with the acquisition of tumor resistance to specific lysis during tumor progression, involving genetic instability, structural changes in cytoskeleton, and hypoxic stress. We will also discuss the interaction between CTLs and tumor endothelial cells, a major component of tumor stroma.