Potential Target Antigens for a Universal Vaccine in Epithelial Ovarian Cancer

The prognosis of epithelial ovarian cancer (EOC), the primary cause of death from gynaecological malignancies, has only modestly improved over the last decades. Immunotherapeutic treatment using a cocktail of antigens has been proposed as a "universal" vaccine strategy. We determined the expression of tumor antigens in the context of MHC class I expression in 270 primary tumor samples using tissue microarray. Expression of tumor antigens p53, SP17, survivin, WT1, and NY-ESO-1 was observed in 120 (48.0%), 173 (68.9%), 208 (90.0%), 129 (56.3%), and 27 (11.0%) of 270 tumor specimens, respectively. In 93.2% of EOC, at least one of the investigated tumor antigens was (over)expressed. Expression of MHC class I was observed in 78.1% of EOC. In 3 out 4 primary tumors, (over)expression of a tumor antigen combined with MHC class I was observed. These results indicate that a multiepitope vaccine, comprising these antigens, could serve as a universal therapeutic vaccine for the vast majority of ovarian cancer patients

Immunotherapy of ovarian cancer

In de eerste 3 hoofdstukken van het proefschrift beschrijven we het gebruik van een kankervaccin dat is gebaseerd op het p53-eiwit. P53 is een eiwit dat tot expressie komt in normale cellen. Het eiwit is echter in sterk verhoogde mate aanwezig in de kankercel van ruim de helft van de patienten met eierstokkanker. Omdat kankercellen veel meer p53 bevatten dan normale cellen kan het p53 eiwit een tumorantigeen genoemd worden. In een eerdere klinische studie hebben we een vaccin getest dat bestaat uit lange stukjes (peptiden) van het p53-eiwit. Dit synthetische lange peptide (SLP) vaccin, p53-SLP-vaccin, is getest in patienten met eierstokkanker. Het vaccin is veilig, wordt goed verdragen en kan een p53-specifieke afweerreactie teweeg brengen. De patienten waren eerder behandeld voor eierstokkanker, maar hadden op het moment van deelname aan de studie opnieuw kanker (een recidief) zonder klachten. Wanneer patienten met een recidief van eierstokkanker wei klachten krijgen worden ze opnieuw met chemotherapie behandeld. We waren benieuwd of de chemotherapie het lange termijn geheugen van de p53-specifieke afweerreactie van onze studiepatienten lOU be·invloeden. Ook vroegen we ons af of de gevoeligheid van de kankercellen voor chemotherapie vergroot zou zijn door de aanwezigheid van p53-specifieke T-cellen. In hoofdstuk 2 laten we zien dat er nog steeds p53-specifieke T-cellen te meten zijn, ook nadat patienten opnieuw met chemotherapie zijn behandeld. De respons van de kankercellen op de chemotherapie was echter niet verbeterd door de aanwezigheid van een p53-specifieke afweerreactie. Kennelijk is het p53-SLP-vaccin goed in staat om zowel op de korte als op de lange termijn een p53-specifieke afweerreactie te bewerkstelligen. Zoals gezegd is het p53-SLP-vaccin in staat een p53-specifieke afweerreactie te realiseren; het vaccin bleek echter niet klinisch effectief. Een mogelijke verklaring is de aanwezigheid van regulatoire T-cellen. Deze cellen hebben een remmend effect op het immuunsysteem. Er zijn aanwijzingen dat regulatoire T-cellen specifiek geremd kunnen worden door een lage dosis chemotherapie (bijvoorbeeld cyclofosfamide). In hoofdstuk 3 worden de resultaten beschreven van een klinische studie waarbij we het p53-SLP-vaccin in combinatie met cyclofosfamide testten in patienten met een recidief eierstokkanker. De toevoeging van cyclofosfamide resulteerde niet in een verandering van het aantal en de functie van regulatoire T-cellen. Het resulteerde echter wei in een zeer sterke toename van het aantal p53-specifieke T-helper-cellen in vergelijking met de eerste studie waarbij patienten aileen met het vaccin waren behandeld. Dit resultaat bevestigt dat de toevoeging van cyclofosfamide de werking van een kankervaccin kan verbeteren. Hoe dat kan, moet nog verder uitgezocht worden. In de afgelopen 10 jaar hebben meerdere onderzoeksgroepen een p53-vaccin getest in kankerpatienten. In hoofdstuk 4 wordt een overzicht gegeven van aile klinische studies die tot dusver zijn uitgevoerd met een p53-vaccin. Dit overzicht bestaat niet aileen uit studies uitgevoerd in patienten met eierstokkanker, maar ook in patienten met andere soorten kanker. Uit deze studies blijkt dat behandeling met een p53-vaccin weliswaar een p53-specifieke afweerreactie kan veroorzaken in kankerpatienten, maar nog geen klinisch effect heeft. Epithelial ovarian cancer (EOC) is the most common cause of death in gynaecologic malignancies. Due to a lack of specific symptoms most EOC patients are diagnosed with advanced stage of disease. The majority of patients with advanced stage EOC achieve a complete clinical response due to the current therapy of aggressive cytoreductive surgery and platinum-taxane based chemotherapy. Unfortunately, most patients develop tumor recurrence, resulting in five-year survival rates 0' only 30%. Therefore, new therapeutic strategies are urgently required. Since the prognosis of EOC patients is influenced by tumor infiltrating immune effector cells [1·3], an immunotherapeutic approach to treat ovarian cancer seems promising. Immunization strategies targeting various single tumor-associated antigens have been developed and tested in EOC patients. In most studies these vaccines induce antigen-specific immunity, but substantial clinical efficacy has not been observed. This thesis describes the results of two clinical trials on an antigen-specific vaccine targeting tumor· antigen p53 in ovarian cancer patients with recurrent disease. Next, we studied intratumoral parameters in EOC in an effort to increase our knowledge on how to improve the clinical efficacy of antigen-specific vaccines in EOC patients. A short introduction on cancer immunotherapy is provided in chapter 1. This chapter highlights the background of ovarian cancer and tumor immunology, and illustrates the possibilities for clini· cal implementation of (antigen-specific) immunotherapy. In the following three chapters the use of p53 as a target for antigen-specific vaccines in cancer patients is described. P53, a tumor-suppressor protein, is a potential target for cancer immuno· therapy as mutations in the p53 gene are frequently seen in human oncogenesis. Persistent over· expression of p53 is present in 50-60% of ovarian cancers. Therefore, the majority of ovarian cancer patients might benefit from p53-directed immunotherapy. Previously, a phase 1/11 clinical trial has been performed to study the immunogenicity and clinical efficacy of a p53-specific synthetic long peptide (SLP) vaccine in recurrent ovarian cancer patients [4]. Induction of p53-specific T-cells, but limited clinical efficacy was observed in the study cohort. We were interested if treatment with secondary chemotherapy after treatment with the p53-SLP vaccine influences the long-term memory of the p53-specific immune response. Therefore, we con ducted a follow-up study in 8 recurrent ovarian cancer patients initially treated with the p53-SLP vaccine in our phase 11/1 clinical trial. described in chapter 2. Antonia et al. reported that vaccine induced p53-specific immune responses were associated with improved response to secondary chemotherapy in small cell lung cancer patients [5]. Treatment with the p53-SLP vaccine in oU' study cohort of ovarian cancer patients, did not affect responses to secondary chemotherapy or survival. However, p53-specific T-cells could still be retrieved despite the use of chemotherapy. From our previous phase 1/11 clinical trial and the current follow-up study we can conclude that the p53-SLP vaccine is able to induce both short-and long-term immunity.

Antigen-specific immunotherapy in ovarian cancer and p53 as tumor antigen

This review discusses the results of different immunization strategies, identifies possible drawbacks in study design and provides potential solutions for augmentation of clinical efficacy. A potential target for cancer immunotherapy is p53, as approximately 50% of ovarian cancer cells carry p53 mutations. Therefore we review the immunological and clinical responses observed in ovarian cancer patients vaccinated with p53 targeting vaccines in particular. In most studies antigen-specific vaccine-induced immunological responses were observed. Unfortunately, no clinical responses with significant reduction of tumor-burden have been reported. Based on the currently available results we emphasize the necessity of multimodality treatment of ovarian cancer, combining classical cytoreductive surgery, (neo) adjuvant chemotherapy, immunotherapy and/or targeted therapy

Long-term clinical and immunological effects of p53-SLP (R) vaccine in patients with ovarian cancer

Vaccine-induced p53-specific immune responses were previously reported to be associated with improved response to secondary chemotherapy in patients with small cell lung cancer. We investigated long-term clinical and immunological effects of the p53-synthetic long peptide (p53-SLP (R)) vaccine in patients with recurrent ovarian cancer. Twenty patients were immunized with the p53-SLP (R) vaccine between July 2006 and August 2007. Follow-up information on patients was obtained. Clinical responses to secondary chemotherapy after p53-SLP (R) immunizations were determined by computerized tomography and/or tumor marker levels (CA125). Disease-specific survival was compared to a matched historical control group. Immune responses were analyzed by flow cytometry, proliferation assay, interferon gamma (IFN-gamma) ELISPOT and/or cytokine bead array. Lymphocytes cultured from skin biopsy were analyzed by flow cytometry and proliferation assay. Of 20 patients treated with the p53-SLP (R) vaccine, 17 were subsequently treated with chemotherapy. Eight of these patients volunteered another blood sample. No differences in clinical response rates to secondary chemotherapy or disease-specific survival were observed between immunized patients and historical controls (p = 0.925, resp. p = 0.601). p53-specific proliferative responses were observed in 5/8 patients and IFN-gamma production in 2/7 patients. Lymphocytes cultured from a prior injection site showing inflammation during chemotherapy did not recognize p53-SLP (R). Thus, treatment with the p53-SLP (R) vaccine does not affect responses to secondary chemotherapy or survival, although p53-specific T-cells do survive chemotherapy

Potentiation of a p53-SLP vaccine by cyclophosphamide in ovarian cancer:A single-arm phase II study

The purpose of the current phase II single-arm clinical trial was to evaluate whether pretreatment with low-dose cyclophosphamide improves immunogenicity of a p53-synthetic long peptide (SLP) vaccine in patients with recurrent ovarian cancer. Patients with ovarian cancer with elevated serum levels of CA-125 after primary treatment were immunized four times with the p53-SLP vaccine. Each immunization was preceded by administration of 300 mg/m2 intravenous cyclophosphamide as a means to affect regulatory T cells (Tregs). Vaccine-induced p53-specific interferon-gamma (IFN-?)-producing T cells evaluated by IFN-? ELISPOT were observed in 90% (9/10) and 87.5% (7/8) of evaluable patients after two and four immunizations, respectively. Proliferative p53-specific T cells, observed in 80.0% (8/10) and 62.5% (5/8) of patients, produced both T-helper 1 and T-helper-2 cytokines. Cyclophosphamide induced neither a quantitative reduction of Tregs determined by CD4+FoxP3+ T cell levels nor a demonstrable qualitative difference in Treg function tested in vitro. Nonetheless, the number of vaccine-induced p53-specific IFN-?-producing T cells was higher in our study compared to a study in which a similar patient group was treated with p53-SLP monotherapy (p = 0.012). Furthermore, the strong reduction in the number of circulating p53-specific T cells observed previously after four immunizations was currently absent. Stable disease was observed in 20.0% (2/10) of patients, and the remainder of patients (80.0%) showed clinical, biochemical and/or radiographic evidence of progressive disease. The outcome of this phase II trial warrants new studies on the use of low-dose cyclophosphamide to potentiate the immunogenicity of the p53-SLP vaccine or other antitumor vaccines

Vaccination against Oncoproteins of HPV16 for Noninvasive Vulvar/Vaginal Lesions:Lesion Clearance Is Related to the Strength of the T-Cell Response

Purpose: Therapeutic vaccination with human papillomavirus type 16 (HPV16) E6 and E7 synthetic long peptides (SLP) is effective against HPV16-induced high-grade vulvar and vaginal intraepithelial neoplasia (VIN/VaIN). However, clinical non-responders displayed weak CD8(+) T-cell reactivity. Here, we studied if imiquimod applied at the vaccine site could improve CD8(+) T-cell reactivity, clinical efficacy, and safety of HPV16-SLP (ISA101). Experimental Design: A multicenter open-label, randomized controlled trial was conducted in patients with HPV16(-) high-grade VIN/VaIN. Patients received ISA101 vaccination with or without application of 5% imiquimod at the vaccine site. The primary objective was the induction of a directly ex vivo detectable HPV16-specific CD8(+) T-cell response. The secondary objectives were clinical responses (lesion size, histology, and virology) and their relation with the strength of vaccination-induced immune responses. Results: Forty-three patients were assigned to either ISA101 with imiquimod (n = 21) or ISA101 only (n = 22). Imiquimod did not improve the outcomes of vaccination. However, vaccine-induced clinical responses were observed in 18 of 34 (53%; 95% CI, 35.1-70.2) patients at 3 months and in 15 of 29 (52%; 95% CI, 32.5-70.6) patients, 8 of whom displayed a complete histologic response, at 12 months after the last vaccination. All patients displayed vaccine-induced T-cell responses, which were significantly stronger in patients with complete responses. Importantly, viral clearance occurred in all but one of the patients with complete histologic clearance. Conclusions: This new study confirms that clinical efficacy of ISA101 vaccination is related to the strength of vaccine-induced HPV16-specific T-cell immunity and is an effective therapy for HPV16-induced high-grade VIN/VaIN. (C) 2016 AACR