Harm Westdorp

Effective Clinical Responses in Metastatic Melanoma Patients after Vaccination with Primary Myeloid Dendritic Cells

Purpose: Thus far, dendritic cell (DC)-based immunotherapy of cancer was primarily based on in vitro–generated monocyte-derived DCs, which require extensive in vitro manipulation. Here, we report on a clinical study exploiting primary CD1c+ myeloid DCs, naturally circulating in the blood. Experimental Design: Fourteen stage IV melanoma patients, without previous systemic treatment for metastatic disease, received autologous CD1c+ myeloid DCs, activated by only brief (16 hours) ex vivo culture and loaded with tumor-associated antigens of tyrosinase and gp100. Results: Our results show that therapeutic vaccination against melanoma with small amounts (3–10 × 106) of myeloid DCs is feasible and without substantial toxicity. Four of 14 patients showed long-term progression-free survival (12–35 months), which directly correlated with the development of multifunctional CD8+ T-cell responses in three of these patients. In particular, high CD107a expression, indicative for cytolytic activity, and IFNγ as well as TNFα and CCL4 production was observed. Apparently, these T-cell responses are essential to induce tumor regression and promote long-term survival by stalling tumor growth. Conclusions: We show that vaccination of metastatic melanoma patients with primary myeloid DCs is feasible and safe and results in induction of effective antitumor immune responses that coincide with improved progression-free survival. Clin Cancer Res; 22(9); 2155–66. ©2015 AACR.

Opportunities for immunotherapy in microsatellite instable colorectal cancer

Microsatellite instability (MSI), the somatic accumulation of length variations in repetitive DNA sequences called microsatellites, is frequently observed in both hereditary and sporadic colorectal cancer (CRC). It has been established that defects in the DNA mismatch repair (MMR) pathway underlie the development of MSI in CRC. After the inactivation of the DNA MMR pathway, misincorporations, insertions and deletions introduced by DNA polymerase slippage are not properly recognized and corrected. Specific genomic regions, including microsatellites, are more prone for DNA polymerase slippage and, therefore, more susceptible for the introduction of these mutations if the DNA MMR capacity is lost. Some of these susceptible genomic regions are located within the coding regions of genes. Insertions and deletions in these regions may alter their reading frame, potentially resulting in the transcription and translation of frameshift peptides with c-terminally altered amino acid sequences. These frameshift peptides are called neoantigens and are highly immunogenic, which explains the enhanced immunogenicity of MSI CRC. Neoantigens contribute to increased infiltration of tumor tissue with activated neoantigen-specific cytotoxic T lymphocytes, a hallmark of MSI tumors. Currently, neoantigen-based vaccination is being studied in a clinical trial for Lynch syndrome and in a trial for sporadic MSI CRC of advanced stage. In this Focussed Research Review, we summarize current knowledge on molecular mechanisms and address immunological features of tumors with MSI. Finally, we describe their implications for immunotherapeutic approaches and provide an outlook on next-generation immunotherapy involving neoantigens and combinatorial therapies in the setting of MSI CRC.

Immunotherapy for prostate cancer: lessons from responses to tumor-associated antigens.

Prostate cancer (PCa) is the most common cancer in men and the second most common cause of cancer-related death in men. In recent years, novel therapeutic options for PCa have been developed and studied extensively in clinical trials. Sipuleucel-T is the first cell-based immunotherapeutic vaccine for treatment of cancer. This vaccine consists of autologous mononuclear cells stimulated and loaded with an immunostimulatory fusion protein containing the prostate tumor antigen prostate acid posphatase. The choice of antigen might be key for the efficiency of cell-based immunotherapy. Depending on the treatment strategy, target antigens should be immunogenic, abundantly expressed by tumor cells, and preferably functionally important for the tumor to prevent loss of antigen expression. Autoimmune responses have been reported against several antigens expressed in the prostate, indicating that PCa is a suitable target for immunotherapy. In this review, we will discuss PCa antigens that exhibit immunogenic features and/or have been targeted in immunotherapeutic settings with promising results, and we highlight the hurdles and opportunities for cancer immunotherapy.

Adjuvant dendritic cell vaccination induces tumor-specific immune responses in the majority of stage III melanoma patients.

ABSTRACT Purpose: To determine the effectiveness of adjuvant dendritic cell (DC) vaccination to induce tumor-specific immunological responses in stage III melanoma patients. Experimental design: Retrospective analysis of stage III melanoma patients, vaccinated with autologous monocyte-derived DC loaded with tumor-associated antigens (TAA) gp100 and tyrosinase after radical lymph node dissection. Skin-test infiltrating lymphocytes (SKILs) obtained from delayed-type hypersensitivity skin-test biopsies were analyzed for the presence of TAA-specific CD8+ T cells by tetrameric MHC-peptide complexes and by functional TAA-specific T cell assays, defined by peptide-recognition (T2 cells) and/or tumor-recognition (BLM and/or MEL624) with specific production of Th1 cytokines and no Th2 cytokines. Results: Ninety-seven patients were analyzed: 21 with stage IIIA, 34 with stage IIIB, and 42 had stage IIIC disease. Tetramer-positive CD8+ T cells were present in 68 patients (70%), and 24 of them showed a response against all 3 epitopes tested (gp100:154–162, gp100:280–288, and tyrosinase:369–377) at any point during vaccinations. A functional T cell response was found in 62 patients (64%). Rates of peptide-recognition of gp100:154–162, gp100:280–288, and tyrosinase:369–377 were 40%, 29%, and 45%, respectively. Median recurrence-free survival and distant metastasis-free survival of the whole study population were 23.0 mo and 36.8 mo, respectively. Conclusions: DC vaccination induces a functional TAA-specific T cell response in the majority of stage III melanoma patients, indicating it is more effective in stage III than in stage IV melanoma patients. Furthermore, performing multiple cycles of vaccinations enhances the chance of a broader immune response.

Immune-related Adverse Events of Dendritic Cell Vaccination Correlate With Immunologic and Clinical Outcome in Stage III and IV Melanoma Patients.

The purpose of this study was to determine the toxicity profile of dendritic cell (DC) vaccination in stage III and IV melanoma patients, and to evaluate whether there is a correlation between side effects and immunologic and clinical outcome. This is a retrospective analysis of 82 stage III and 137 stage IV melanoma patients, vaccinated with monocyte-derived or naturally circulating autologous DCs loaded with tumor-associated antigens gp100 and tyrosinase. Median follow-up time was 54.3 months in stage III patients and 12.9 months in stage IV patients. Treatment-related adverse events occurred in 84% of patients; grade 3 toxicity was present in 3% of patients. Most common adverse events were flu-like symptoms (67%) and injection site reactions (50%), and both correlated with the presence of tetramer-positive CD8+ T cells (both P<0.001). In stage III melanoma patients experiencing flu-like symptoms, median overall survival (OS) was not reached versus 32.3 months in patients without flu-like symptoms (P=0.009); median OS in patients with an injection site reaction was not reached versus 53.7 months in patients without an injection site reaction (P<0.05). In stage IV melanoma patients (primary uveal and mucosal melanomas excluded), median OS in patients with or without flu-like symptoms was 13.1 versus 8.9 months, respectively (P=0.03); median OS in patients with an injection site reaction was 15.7 months versus 9.8 months in patients without an injection site reaction (P=0.003). In conclusion, DC vaccination is safe and tolerable and the occurrence of the immune-related side effects, such as flu-like symptoms and injection site reactions, correlates with immunologic and clinical outcome.

Metastatic melanoma mimicking solitary fibrous tumor: report of two cases

Malignant melanomas are known for their remarkable morphological variation and aberrant immunophenotype with loss of lineage-specific markers, especially in recurrences and metastases. Hot spot mutations in BRAF, NRAS, GNAQ, and GNA11 and mutations in KIT are oncogenic events in melanomas. Therefore, genotyping can be a useful ancillary diagnostic tool. We present one case each of recurrent and metastatic melanoma, both showing histological and immunohistochemical features of solitary fibrous tumor (SFT). Mutational analysis detected BRAF and NRAS mutations in the primary and secondary lesions, respectively. This result confirmed the diagnosis of recurrent/metastastic melanoma.

Survival of metastatic melanoma patients after dendritic cell vaccination correlates with expression of leukocyte phosphatidylethanolamine-binding protein 1/Raf kinase inhibitory protein

Immunotherapy for metastatic melanoma offers great promise but, to date, only a subset of patients have responded. There is an urgent need to identify ways of allocating patients to the most beneficial therapy, to increase survival and decrease therapy-associated morbidity and costs. Blood-based biomarkers are of particular interest because of their straightforward implementation in routine clinical care. We sought to identify markers for dendritic cell (DC) vaccine-based immunotherapy against metastatic melanoma through gene expression analysis of peripheral blood mononuclear cells. A large-scale microarray analysis of 74 samples from two treatment centers, taken directly after the first round of DC vaccination, was performed. We found that phosphatidylethanolamine binding protein 1 (PEBP1)/Raf Kinase inhibitory protein (RKIP) expression can be used to identify a significant proportion of patients who performed poorly after DC vaccination. This result was validated by q-PCR analysis on blood samples from a second cohort of 95 patients treated with DC vaccination in four different centers. We conclude that low PEBP1 expression correlates with poor overall survival after DC vaccination. Intriguingly, this was only the case for expression of PEBP1 after, but not prior to, DC vaccination. Moreover, the change in PEBP1 expression upon vaccination correlated well with survival. Further analyses revealed that PEBP1 expression positively correlated with genes involved in T cell responses but inversely correlated with genes associated with myeloid cells and aberrant inflammation including STAT3, NOTCH1, and MAPK1. Concordantly, PEBP1 inversely correlated with the myeloid/lymphoid-ratio and was suppressed in patients suffering from chronic inflammatory disease.Immunotherapy for metastatic melanoma offers great promise but, to date, only a subset of patients have responded. There is an urgent need to identify ways of allocating patients to the most beneficial therapy, to increase survival and decrease therapy-associated morbidity and costs. Blood-based biomarkers are of particular interest because of their straightforward implementation in routine clinical care. We sought to identify markers for dendritic cell (DC) vaccine-based immunotherapy against metastatic melanoma through gene expression analysis of peripheral blood mononuclear cells. A large-scale microarray analysis of 74 samples from two treatment centers, taken directly after the first round of DC vaccination, was performed. We found that phosphatidylethanolamine binding protein 1 (PEBP1)/ Raf Kinase inhibitory protein (RKIP) expression can be used to identify a significant proportion of patients who performed poorly after DC vaccination. This result was validated by q-PCR analysis on blood samples from a second cohort of 95 patients treated with DC vaccination in four different centers. We conclude that low PEBP1 expression correlates with poor overall survival after DC vaccination. Intriguingly, this was only the case for expression of PEBP1 after, but not prior to, DC vaccination. Moreover, the change in PEBP1 expression upon vaccination correlated well with survival. Further analyses revealed that PEBP1 expression positively correlated with genes involved in T cell responses but inversely correlated with genes associated with myeloid cells and aberrant inflammation including STAT3, NOTCH1, and MAPK1. Concordantly, PEBP1 inversely correlated with the myeloid/ lymphoid-ratio and was suppressed in patients suffering from chronic inflammatory disease.

Immunotherapy holds the key to cancer treatment and prevention in Constitutional Mismatch Repair Deficiency (CMMRD) syndrome

Monoallelic germline mutations in one of the DNA mismatch repair (MMR) genes cause Lynch syndrome, with a high lifetime risks of colorectal and endometrial cancer at adult age. Less well known, is the constitutional mismatch repair deficiency (CMMRD) syndrome caused by biallelic germline mutations in MMR genes. This syndrome is characterized by the development of childhood cancer. Patients with CMMRD are at extremely high risk of developing multiple cancers including hematological, brain and intestinal tumors. Mutations in MMR genes impair DNA repair and therefore most tumors of patients with CMMRD are hypermutated. These mutations lead to changes in the translational reading frame, which consequently result in neoantigen formation. Neoantigens are recognized as foreign by the immune system and can induce specific immune responses. The growing evidence on the clinical efficacy of immunotherapies, such as immune checkpoint inhibitors, offers the prospect for treatment of patients with CMMRD. Combining neoantigen-based vaccination strategies and immune checkpoint inhibitors could be an effective way to conquer CMMRD-related tumors. Neoantigen-based vaccines might also be a preventive treatment option in healthy biallelic MMR mutation carriers. Future studies need to reveal the safety and efficacy of immunotherapies for patients with CMMRD.

Adjuvant Dendritic Cell Vaccination in High-Risk Uveal Melanoma.

Uveal melanoma (UM) is the most common primary intraocular malignancy in adults with an annual incidence of 4 to 10 per million in the white population. The 5-year overall survival (OS) rate is approximately 70% to 80%. Up to 50% of patients with UM develop metastases, usually after a long disease-free interval (2-5 years). If metastatic disease is present, the prognosis is dismal with a 1-year OS rate of 10% to 40%. Currently, no effective systemic treatment improving OS is available for patients with metastatic UM, nor has any adjuvant treatment shown survival benefit. nOur research group and others have performed several prospective dendritic cell (DC) vaccination studies in patients with cutaneous melanoma showing little toxicity and promising immunologic and clinical results. [...]

Ipilimumab administered to metastatic melanoma patients who progressed after dendritic cell vaccination

ABSTRACT Background: Ipilimumab has proven to be effective in metastatic melanoma patients. The purpose of this study was to determine the efficacy of ipilimumab in advanced melanoma patients who showed progressive disease upon experimental dendritic cell (DC) vaccination. Methods: Retrospective analysis of 48 stage IV melanoma patients treated with ipilimumab after progression upon DC vaccination earlier in their treatment. DC vaccination was given either as adjuvant treatment for stage III disease (n = 18) or for stage IV disease (n = 30). Ipilimumab (3 mg/kg) was administered every 3 weeks for up to 4 cycles. Results: Median time between progression upon DC vaccination and first gift of ipilimumab was 5.4 mo. Progression-free survival (PFS) rates for patients that received ipilimumab after adjuvant DC vaccination, and patients that received DC vaccination for stage IV melanoma, were 35% and 7% at 1 y and 35% and 3% at 2 y, while the median PFS was 2.9 mo and 3.1 mo, respectively. Median overall survival of patients pre-treated with adjuvant DC vaccination for stage III melanoma was not reached versus 8.0 mo (95% CI, 5.2–10.9) in the group pre-treated with DC vaccination for stage IV disease (HR of death, 0.36; p = 0.017). Grade 3 immune-related adverse events occurred in 19% of patients and one death (2%) was related to ipilimumab. Conclusions: Clinical responses to ipilimumab were found in a considerable number of advanced melanoma patients with progression after adjuvant DC vaccination for stage III disease, while the effect was very limited in patients who showed progression after DC vaccination for stage IV disease.