Phase I study of a new cancer vaccine of ten mixed peptides for advanced cancer patients

A phase I study of a new cancer vaccine (KRM‐10), consisting of a mixture of 10 different short peptides, was conducted for patients with advanced gastrointestinal cancers. Primary or secondary endpoints included the dose‐limiting toxicity (DLT), or safety and immune responses, respectively. Peptide‐specific cytotoxic T lymphocytes (CTL) and immunoglobulin G (IgG), together with soluble inflammatory factors, were measured before and after vaccination. Twenty‐one patients were vaccinated with KRM‐10 at dose levels of 10 (n = 6), 20 (n = 8) or 30 mg (n = 7) of peptides every week for 6 weeks. No DLT were observed in the dose range evaluated. Common treatment‐related adverse events were a grade 1 injection site reaction in 15 patients, and fever in three patients (grade 1 in two patients and grade 2 in one patient). CTL activity to at least one peptide at the time of the third and sixth vaccination increased in 2 and 3 of 6 (10 mg), 2 of 8 and 4 of 6 (20 mg), or 2 and 1 of 6 (30 mg) patients, respectively. IgG levels, at the third and sixth vaccination, were also increased in 1 and 1 of 6 (10 mg), 2 of 8 and 4 of 6 (20 mg), or 1 and 3 of 6 (30 mg) patients, respectively. The KRM‐10 vaccine consisting of 20 mg of peptides was determined as the optimal dose for a coming phase II trial because of its safety, and also for demonstrating the most potent activity for augmenting the immune response of the three doses tested. This trial was registered at the UMIN Clinical Trials Registry as UMIN000008820.     

Efficacy of the NCCV Cocktail‐1 vaccine for refractory pediatric solid tumors: A phase I clinical trial

Pediatric refractory solid tumors are aggressive malignant diseases, resulting in an extremely poor prognosis. KOC1, FOXM1, and KIF20A are cancer antigens that could be ideal targets for anticancer immunotherapy against pediatric refractory solid tumors with positive expression for these antigens. This nonrandomized, open‐label, phase I clinical trial evaluated the safety and efficacy of the NCCV Cocktail‐1 vaccine, which is a cocktail of cancer peptides derived from KOC1, FOXM1, and KIF20A, in patients with pediatric refractory solid tumors. Twelve patients with refractory pediatric solid tumors underwent NCCV Cocktail‐1 vaccination weekly by intradermal injections. The primary endpoint was the safety of the NCCV Cocktail‐1 vaccination, and the secondary endpoints were the immune response, as measured by interferon‐r enzyme‐linked immunospot assay, and the clinical outcomes including tumor response and progression‐free survival. The NCCV Cocktail‐1 vaccine was well tolerated. The clinical response of this trial showed that 4 patients had stable disease after 8 weeks and 2 patients maintained remission for >11 months. In 4, 8, and 5 patients, the NCCV Cocktail‐1 vaccine induced the sufficient number of peptide‐specific CTLs for KOC1, FOXM1, and KIF20A, respectively. Patients with high peptide‐specific CTL frequencies for KOC1, FOXM1, and KIF20A had better progression‐free survival than those with low frequencies. The findings of this clinical trial showed that the NCCV Cocktail‐1 vaccine could be a novel therapeutic strategy, with adequate effects against pediatric refractory solid tumors. Future large‐scale trials should evaluate the efficacy of the NCCV Cocktail‐1 vaccination.     

Heat shock cognate protein 70 encodes antigenic epitopes recognised by HLA-B4601-restricted cytotoxic T lymphocytes from cancer patients

Heat shock cognate protein 70 (HSC70), a highly conserved protein and a member of the family of molecular chaperones, has the ability to induce cytotoxic T lymphocyte (CTL) responses through binding and carrying antigenic peptides. We demonstrated in this study that the HSC70 gene encodes two antigenic peptides recognised by HLA-B46-restricted and tumour-reactive CTLs established from tumour-infiltrating lymphocytes of a colon cancer. These HSC70-derived peptides, at amino-acid positions 106-114 and 233-241, had the ability to induce HLA-B46-restricted and peptide-specific CTLs, which are reactive to tumour cells, from peripheral blood mononuclear cells of the majority of epithelial cancer patients tested. These results, along with those from the previous studies, indicate the two ways of HSC70 involvement in the immune responses to tumours: chaperones and antigens, and thus may provide a new insight for the development of HSC70-directed cancer-specific immunotherapy.     

Heat shock protein 90 targets a chaperoned peptide to the static early endosome for efficient cross‐presentation by human dendritic cells

The presentation of an exogenous antigen in a major histocompatibility complex class‐I‐ restricted fashion to CD8⁺ T cells is called cross‐presentation. Heat shock proteins (HSPs) such as Hsp70, gp96, and Hsp90 have been shown to elicit efficient CTL responses by cross‐presentation through an as‐yet entirely unknown mechanism. Hsp90 is the most abundant cytosolic HSP and is known to act as a molecular chaperone. We have shown that a tumor antigen peptide complexed with Hsp90 could be cross‐presented by dendritic cells (DCs) through an endosomal pathway in a murine system. However, it has not been determined whether human DCs also cross‐present an Hsp90–peptide complex and induce peptide‐specific CTLs. In this study, we found that an Hsp90–cancer antigen peptide complex was efficiently cross‐presented by human monocyte‐derived DCs and induced peptide‐specific CTLs. Furthermore, we observed that the internalized Hsp90–peptide complex was strictly sorted to the Rab5⁺, EEA1⁺ static early endosome and the Hsp90‐chaperoned peptide was processed and bound to MHC class I molecules through an endosome‐recycling pathway. Our data indicate that targeting of the antigen to a “static” early endosome by Hsp90 is essential for efficient cross‐presentation.     

Immunization with a P53 synthetic long peptide vaccine induces P53‐specific immune responses in ovarian cancer patients,a phase II trial

The prognosis of ovarian cancer, the primary cause of death from gynecological malignancies, has only modestly improved over the last decades. Immunotherapy is one of the new treatment modalities explored for this disease. To investigate safety, tolerability, immunogenicity and obtain an impression of clinical activity of a p53 synthetic long peptide (p53‐SLP) vaccine, twenty patients with recurrent elevation of CA‐125 were included, eighteen of whom were immunized 4 times with 10 overlapping p53‐SLP in Montanide ISA51. The first 5 patients were extensively monitored for toxicity, but showed no ≥ grade 3 toxicity, thus accrual was continued. Overall, toxicity was limited to grade 1 and 2, mostly locoregional, inflammatory reactions. IFN‐γ producing p53‐specific T‐cell responses were induced in all patients who received all 4 immunizations as measured by IFN‐γ ELISPOT. An IFN‐γ secretion assay showed that vaccine‐induced p53‐specific T‐cells were CD4⁺, produced both Th1 and Th2 cytokines as analyzed by cytokine bead array. Notably, Th2 cytokines dominated the p53‐specific response. P53‐specific T‐cells were present in a biopsy of the last immunization site of at least 9/17 (53%) patients, reflecting the migratory capacity of p53‐specific T‐cells. As best clinical response, stable disease evaluated by CA‐125 levels and CT‐scans, was observed in 2/20 (10%) patients, but no relationship was found with vaccine‐induced immunity. This study shows that the p53‐SLP vaccine is safe, well tolerated and induces p53‐specific T‐cell responses in ovarian cancer patients. Upcoming trials will focus on improving T helper‐1 polarization and clinical efficacy. © 2009 UICC     

Inhibition of mitogen‐activated protein kinase pathway can induce upregulation of human leukocyte antigen class I without PD‐L1‐upregulation in contrast to interferon‐γ treatment

Recently, we reported that human leukocyte antigen (HLA) class I expression is predominantly regulated by the mitogen‐activated protein kinase (MAPK) pathway as one of the oncogenic regulations of HLA class I expression. In the present study, we examined mechanisms of how HLA class I and PD‐L1 are regulated by MAPK inhibitors and interferon‐γ (IFN‐γ). Furthermore, we evaluated the expression of major signal transduction molecules by Western blot and anti‐tumor CTL activity by a cytotoxic assay when HLA class I and PD‐L1 were modulated by MAPK inhibitors and/or IFN‐γ. As a result, we confirmed, as a more general phenomenon, that the inhibition of MAPK could upregulate HLA class I expression in a panel of human solid tumors (n = 26). Of note, we showed that MAPK inhibitors act on the upregulation of HLA class I expression through a different pathway from IFN‐γ; there was an additive effect in the upregulation of HLA class I when treated with the combination of MAPK inhibitors and IFN‐γ, and there was no overlapping activation of JAK2/STAT1 and Erk1/2 molecules when treated with either IFN‐γ or MAPK inhibitors. Furthermore, we showed that IFN‐γ–treatment impaired the tumor‐specific CTL activity due to the upregulation of PD‐L1 in spite of the upregulation of HLA class I, while MAPK inhibitors can augment the tumor‐specific CTL activity due to the upregulated HLA class I without PD‐L1 alterations. In conclusion, in addition to the original anti‐proliferative activity, MAPK inhibitors may work toward the enhancement of T‐cell‐mediated anti‐tumor immunity through the upregulation of HLA class I without the upregulation of PD‐L1.     

A phase I trial of DNA vaccination with a plasmid expressing prostate-specific antigen in patients with hormone-refractory prostate cancer

Prostate-specific antigen (PSA) is a serine protease secreted at low levels by normal luminal epithelial cells of the prostate and in significantly higher levels by prostate cancer cells. Therefore, PSA is a potential target for various immunotherapeutical approaches against prostate cancer. DNA vaccination has been investigated as immunotherapy for infectious diseases in patients and for specific treatment of cancer in certain animal models. In animal studies, we have demonstrated that vaccination with plasmid vector pVAX/PSA results in PSA-specific cellular response and protection against tumour challenge. The purpose of the trial was to evaluate the safety, feasibility and biological efficacy of pVAX/PSA vaccine in the clinic. A phase I trial of pVAX/PSA, together with cytokine granulocyte/macrophage-colony stimulating factor (GM-CSF) (Molgramostim) and IL-2 (Aldesleukin) as vaccine adjuvants, was carried out in patients with hormone-refractory prostate cancer. To evaluate the biologically active dose, the vaccine was administered during five cycles in doses of 100, 300 and 900 microg, with three patients in each cohort. Eight patients were evaluable. A PSA-specific cellular immune response, measured by IFN-gamma production against recombinant PSA protein, and a rise in anti-PSA IgG were detected in two of three patients after vaccination in the highest dose cohort. A decrease in the slope of PSA was observed in the two patients exhibiting IFN-gamma production to PSA. No adverse effects (WHO grade >2) were observed in any dose cohort. We demonstrate that DNA vaccination with a PSA-coding plasmid vector, given with GM-CSF and IL-2 to patients with prostate cancer, is safe and in doses of 900 microg the vaccine can induce cellular and humoral immune responses against PSA protein.     

Proteomics analysis of differential protein expression identifies heat shock protein 47 as a predictive marker for lymph node metastasis in patients with colorectal cancer

The discovery of biomarkers to predict the potential for lymph node (LN) metastasis in patients with colorectal cancer (CRC) is essential for developing improved strategies for treating CRC. In the present study, they used isobaric tags for relative and absolute quantitation to conduct a proteomic analysis designed to identify novel biomarkers for predicting LN metastasis in patients with CRC. They identified 60 differentially expressed proteins specifically associated with LN metastasis in CRC patients and classified the molecular and functional characteristics of these proteins by bioinformatic approaches. A literature search led them to select heat shock protein 47 (HSP47) as the most suitable candidate biomarker for predicting LN metastasis. Validation analysis by immunohistochemistry showed that HSP47 expression in patients with CRC and the number of HSP47‐positive spindle cells in the tumor stroma were significantly higher compared with those in adjacent normal colonic mucosa, and the number of the latter cells increased with tumor progression. Further, the number of HSP47‐positive spindle cells in stroma was a more informative marker for identifying LN metastasis than HSP47expression. Multivariate analysis identified spindle cells that expressed elevated levels of HSP47 as an independent predictive biomarker for CRC with LN metastasis. Moreover, these cells served as an independent marker of disease‐free and overall survival of patients with CRC. Their data indicate that the number of HSP47‐positive spindle cells in the stroma of CRC may serve as a novel predictive biomarker of LN metastasis, early recurrence and poor prognosis.     

B-CLL患者瘤细胞膜表面Id-ScFv和人Hsp70的制备及其抗瘤效应

背景与目的:恶性B型淋巴细胞表面免疫球蛋白(surface membrane immunoglobulin,SmIg)表达的独特型决定簇(Idiotypic determinant,Id)不仅是该类肿瘤特异性标记,也是该类肿瘤的特异性抗原,可诱导机体对其产生特异性免疫应答,但由于Id是自体成分,分子量小,免疫原性较弱。人热休克蛋白70(heat shock protein,Hsp70)是一类重要的抗原提呈分子,能有效加强抗原肽的免疫原性。本研究通过制备慢性B型淋巴细胞性白血病(B-cell chronic lymphatic leukemia,B-CLL)患者瘤细胞膜表面独特型单链抗体(Idiotypic determinant single-chain antibody,Id-ScFv)和Hsp70两种蛋白,在体外研究两者联合抗瘤作用并初步探讨其机制。方法:分别在大肠杆菌中表达Id-ScFv和Hsp70两种蛋白,表达产物经SDS-PAGE电泳(sodium dodecyl sulfate polyacrylamide gel electropheresis)及ELISA(enzyme-linked immunosorbentassay)检测鉴定后,分别用金属螯合层析和离子交换层析纯化并在体外将这两种蛋白结合成复合物(Hsp70-Id)。用MTT法及检测Id-ScFv组、人Hsp70组及Hsp70-Id组刺激外周血单个核细胞(peripheral blood mononuclear cell,PBMC)的增殖作用,以ELISA法检测各组培养上清中IL-12和TNF-α水平,并用流式细胞术检测各组PBMC亚群的变化。用活细胞计数法检测各组被激活的PBMC对慢性B细胞白血病细胞株Daudi、慢性髓性白血病细胞株K562和肝癌细胞株HepG2的杀伤作用。结果:经SDS-PAGE电泳分析纯化后表达产物的分子量大约为30ku(Id-ScFv蛋白)和70ku(Hsp70蛋白),分别与其理论预期值相符。PBMC的增殖作用、培养上清中IL-12和TNF-琢水平,Hsp70-Id组明显强于Id-ScFv组和人Hsp70组(P<0.05),而Id-ScFv组和人Hsp70组明显强于阴性对照组(P<0.05)。流式细胞术检测显示在Hsp70-Id组、Id-ScFv组和人Hsp70组PBMC中CD8 T细胞亚群的百分率均有增加,其中Hsp70-Id组最明显。在Id-ScFv组和Hsp70-Id组激活的PBMC对Daudi细胞的杀伤作用较K562、HepG2细胞强(P<0.05),且Hsp70-Id组激活的PBMC对Daudi细胞的杀伤作用明显强于Id-ScFv组(P<0.001)。结论:成功获取B-CLL患者瘤细胞膜表面Id-ScFv和人Hsp70两种纯化蛋白;Hsp70-Id在体外可增强PBMC的特异性杀瘤作用,其机制可能与促进PBMC的增殖、活化CD8 T细胞并诱导具有抗肿瘤作用的Th1型细胞因子的分泌有关。     

上皮性卵巢癌患者血清可溶性白细胞介素-2受体、人附睾蛋白4、T淋巴细胞亚群水平及临床意义

目的探讨上皮性卵巢癌患者的血清可溶性白细胞介素-2受体(SIL-2R)、人附睾蛋白4(HE4)、T淋巴细胞亚群水平及临床意义。方法选择100例上皮性卵巢癌患者和100例卵巢良性肿瘤患者,分别作为观察组和对照组,比较两组患者的血清SIL-2R、HE4及T淋巴细胞亚群(CD3⁺、CD4⁺、CD8⁺和CD4⁺/CD8⁺)水平,分析不同临床特征上皮性卵巢癌患者的血清SIL-2R、HE4及T淋巴细胞亚群水平。结果观察组患者的血清SIL-2R、HE4水平均明显高于对照组,差异均有统计学意义(P﹤0.01)。观察组患者的血清CD3⁺、CD4⁺水平及CD4⁺/CD8⁺均明显低于对照组,CD8⁺水平明显高于对照组,差异均有统计学意义(P﹤0.01)。临床分期为Ⅲ~Ⅳ期、中/低分化、有腹腔积液的上皮性卵巢癌患者的血清SIL-2R、HE4水平分别高于临床分期为Ⅰ~Ⅱ期、高分化、无腹腔积液的患者,差异均有统计学意义(P﹤0.01)。中/低分化、有淋巴结转移、有腹腔积液上皮性卵巢癌患者的血清CD3⁺、CD4⁺水平分别低于高分化、无淋巴结转移、无腹腔积液的患者,CD8⁺水平分别高于高分化、无淋巴结转移、无腹腔积液的患者,差异均有统计学意义(P﹤0.05)。上皮性卵巢癌患者的血清SIL-2R、HE4水平与CD3⁺、CD4⁺、CD4⁺/CD8⁺均呈负相关(P﹤0.05),与CD8⁺呈正相关(P﹤0.05)。结论上皮性卵巢癌患者血清中SIL-2R、HE4水平均较高,T淋巴细胞亚群CD3⁺、CD4⁺水平及CD4⁺/CD8⁺均较低,SIL-2R、HE4水平与T淋巴细胞亚群之间具有相关性。     

A phase I/II trial of a WT1 (Wilms' tumor gene) peptide vaccine in patients with solid malignancy: safety assessment based on the phase I data

OBJECTIVE: We conducted a phase I study to investigate the safety of a weekly WT1 tumor vaccine therapy in patients with solid tumors that had been refractory to all other anti-cancer therapies. METHODS: Skin-test-negative patients were intradermally injected weekly for 12 weeks with 3.0 mg of an HLA-A*2402-restricted modified 9-mer WT1 peptide emulsified in Montanide ISA51 adjuvant. We estimated the Bayesian posterior probability of the occurrence of grade 3 or 4 toxicity when receiving the weekly WT1 vaccination. This analysis provided the basis for making a decision to terminate the phase I study and switch to phase II. Moreover, we performed an exploratory assessment of the anti-tumor effects of WT1 treatment. RESULTS: Ten patients received 114 vaccinations with WT1 on a weekly schedule. No grade 3 or 4 toxicities were observed. Based on the Bayesian approach, it was highly likely that the probability of grade 3 or 4 toxicity was below 20% (the posterior probability = 0.914). Fifteen grade 2 and two grade 1 toxicities were observed; all of these incidents, however, were determined by the Independent Data and Safety Monitoring Committee to be unrelated to the WT1 treatment. One patient exhibited a partial response; five additional patients had stable disease while receiving weekly WT1 treatment. CONCLUSION: This paper confirms that the potential toxicities of the treatment schedule of weekly WT1 vaccination are acceptable and suggested a potential anti-tumor effect. Consequently, we validated the decision to continue to the phase II trial.     

Results of the first phase 1 clinical trial of the HER‐2/neu peptide (GP2) vaccine in disease‐free breast cancer patients

BACKGROUND:

HER‐2/neu, overexpressed in breast cancer, is a source of immunogenic peptides that include GP2 and E75. Phase 2 testing of E75 as an adjuvant vaccine has suggested a clinical benefit. GP2, derived from the transmembrane portion of HER‐2/neu, has differing binding characteristics and may be more immunogenic than E75. Results of the first phase 1 trial of GP2 peptide vaccine are presented.

METHODS:

Disease‐free, lymph node‐negative, human leukocyte antigen (HLA)‐A2⁺ breast cancer patients were enrolled. This dose escalation trial included 4 groups to determine safety and optimal GP2 peptide/granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) dose. Toxicities were monitored. Immunologic response was assessed ex vivo via the HLA‐A2:immunoglobulin dimer assay to detect GP2‐specific CD8⁺ T cells (and E75‐specific CD8⁺ T cells to assess epitope spreading) and in vivo via delayed type hypersensitivity (DTH) reaction (medians/ranges).

RESULTS:

Eighteen patients were enrolled. All toxicities were grade ≤2. Eight (88.9%) of 9 patients in the first 3 dose groups required GM‐CSF dose reductions for local reactions ≥100 mm or grade ≥2 systemic toxicity. GM‐CSF dose was reduced to 125 μg for the final dose group. All patients responded immunologically ex vivo (GP2‐specific CD8⁺ T cells from prevaccination to maximum, 0.4% [0.0%‐2.0%] to 1.1% [0.4%‐3.6%], P < .001) and in vivo (GP2 pre‐ to postvaccination DTH, 0 mm [0.0‐19.5 mm] to 27.5 mm [0.0‐114.5 mm, P < .001). E75‐specific CD8⁺ T cells also increased in response to GP2 from prevaccination to maximum (0.8% [0.0%‐2.41%] to 1.6% [0.86%‐3.72%], P < .001).

CONCLUSIONS:

The GP2 peptide vaccine appears safe and well tolerated with minimal local/systemic toxicity. GP2 elicited HER‐2/neu–specific immune responses, including epitope spreading, in high‐risk, lymph node‐negative breast cancer patients. These findings support further investigation of the GP2 vaccine for the prevention of breast cancer recurrence. Cancer 2010. © 2010 American Cancer Society.     

Hyperthermia on immune regulation: a temperature's story

Over the last decade the linkage between hyperthermia, heat shock proteins and fever with the body's immune system has been well investigated. The immunomodulatory function of hyperthermia has been found to be quite sensitively regulated by temperature, as different levels of heating can bring different modulatory effect on different sensitive targets. Understanding these intrinsic mechanisms could bring new inspirations on the design of clinical trials combining local tumor hyperthermia with immunotherapy in cancer patients. This review will attempt to tell the story about the effect of temperature on immune regulation, with special emphasis on the clinical application of hyperthermia and the feasibility of combining it with immunotherapy in the clinic.     

Heat shock protein 70 as a predictive marker for platinum-based adjuvant chemotherapy in patients with resected non-small cell lung cancer

Objectives

Although adjuvant platinum-based chemotherapy improves survival in completely resected non-small cell lung cancer (NSCLC), its effect is limited. We evaluated whether the expression of heat shock protein 70 (Hsp70) is associated with clinical outcomes in patients with completely resected NSCLC who were treated with or without adjuvant platinum-based chemotherapy.

Patients and methods

Patients who underwent curative resection for NSCLC and diagnosed as stage IIA through IIIA were included. Immunohistochemical staining for Hsp70 was performed on surgical specimens and survival rates were compared by Hsp70 expression and adjuvant platinum-based chemotherapy.

Results

Of 327 enrolled patients, Hsp70 expression was positive in 220 (67.3%). For patients who did not receive adjuvant chemotherapy, Hsp70 expression did not significantly affect survival. However, for patients who received adjuvant chemotherapy, those with Hsp70-positive tumors had a longer disease-free survival outcome than cases with Hsp70-negative tumors (not reached vs. 27.3 months; P = 0.002), although there was no significant difference in overall survival (97.0 vs. 58.9 months, P = 0.080). In the adjuvant chemotherapy group, multivariate modeling showed that patients with Hsp70-postitive tumors had a lower risk of recurrence and death after adjusting for age, sex, performance status, pathologic stage, and histological type (disease-free survival: adjusted hazard ratio, 0.537; 95% CI, 0.362–0.796; P = 0.002; overall survival: adjusted hazard ratio, 0.663; 95% CI, 0.419–1.051; P = 0.080).

Conclusion

Hsp70 is a positive predictive factor in completely resected NSCLC with received platinum-based adjuvant chemotherapy.     

Addition of interferon‐α to the p53‐SLP® vaccine results in increased production of interferon‐γ in vaccinated colorectal cancer patients: A phase I/II clinical trial

We previously established safety and immunogenicity of a p53 synthetic long peptides (p53‐SLP®) vaccine. In the current trial, we investigated whether combination of interferon‐alpha (IFN‐α) with p53‐SLP® is both safe and able to improve the induced p53‐specific IFN‐γ response. Eleven colorectal cancer patients successfully treated for metastatic disease were enrolled in this study. Of these, nine patients completed follow‐up after two injections with p53‐SLP® together with IFN‐α. Safety and p53‐specific immune responses were determined before and after vaccination. Furthermore, cryopreserved PBMCs were compared head‐to‐head to cryopreserved PBMCs obtained in our previous trial with p53‐SLP® only. Toxicity of p53‐SLP® vaccination in combination with IFN‐α was limited to Grade 1 or 2, with predominantly small ongoing swellings at the vaccination site. All patients harbored p53‐specific T cells after vaccination and most patients showed p53‐specific antibodies. Compared to the previous trial, addition of IFN‐α significantly improved the frequency of p53‐specific T cells in IFN‐γ ELISPOT. Moreover, in this trial, p53‐specific T cells were detectable in blood samples of all patients in a direct ex vivo multiparameter flowcytometric assay, opposed to only 2 of 10 patients vaccinated with p53‐SLP® only. Finally, patients in this trial displayed a broader p53‐specific immunoglobulin‐G response, indicating an overall better p53‐specific T‐helper response. Our study shows that p53‐SLP® vaccination combined with IFN‐α injection is safe and capable of inducing p53‐specific immunity. When compared to a similar trial with p53‐SLP® vaccination alone the combination was found to induce significantly more IFN‐γ producing p53‐specific T cells.