Switch to checkpoint inhibition after targeted therapy at time of progression or during ongoing response: A retrospective single‐centre experience in patients with BRAF‐mutated melanoma

BRAF + MEK inhibition is preferentially applied as first‐line therapy in BRAF V600‐mutated melanoma patients with unfavourable prognostic features, due to the ability of targeted therapy (TT) to induce rapid symptom control, decrease tumour burden and normalize lactate dehydrogenase (LDH) levels. In addition, short‐term TT transiently increases tumour antigen presentation and tumour influx of T cells. Therefore, it might be favourable to switch TT to checkpoint inhibition (CPI) before progression (PD). We retrospectively analysed melanoma patients treated first line with TT (TT1) and who subsequently switched to CPI during response to TT (sDR group) or at progression upon TT (sPD group). We identified 74 patients (n = 37 sDR group and n = 37 sPD group). ORR to CPI was 27.0% in the sDR group versus 24.3% in the sPD group (p = .790). Median was PFS 2.5 months versus 1.2 months (p = .145), and median OS was 30.6 versus 14.1 months (p = .007). After adjusting for baseline differences and known prognostic factors, hazard ratios (HRs) favouring sDR were 0.89 for PFS upon CPI (p = .956) and 0.48 for OS (p = .055). Thus, patients switching to CPI during ongoing clinical benefit from TT do not have an inferior outcome. Due to baseline imbalances and small patient population, a favourable trend for the sDR group can be hypothesized only.     

Resistance to the proapoptotic effects of interferon-gamma on melanoma cells used in patient-specific dendritic cell immunotherapy is associated with improved overall survival

The use of whole cell tumor vaccines and various means of loading antigen onto dendritic cells have been under investigation for over a decade. Induction of apoptosis and the exposure of immune-stimulating proteins are thought to be beneficial for the use in immunotherapy protocols, but conclusive evidence in the clinical setting has been lacking. Incubation of melanoma cell lines with interferon-gamma (IFN-γ) increased phosphatidylserine and calreticulin exposure, but not in the IFN-γ-resistant cell line Lu-1205. Short-term autologous melanoma cell lines used for loading dendritic cells for immunotherapy showed differential response to the pro-apoptotic effects of IFN-γ. These IFN-γ-treated tumor cells (TCs) were irradiated and used for loading antigen for dendritic cell therapy. A log-rank comparison of survival for patients whose TCs were found to be either sensitive (upregulated phosphatidylserine and calreticulin) or insensitive to IFN-γ revealed a strongly significant correlation to progression-free (p = 0.003) and overall survival (p = 0.002) favorably in those patients whose cell lines were resistant to the proapoptotic effect of IFN-γ. These results suggest that the use of IFN-γ in anti-melanoma dendritic cell-based immunotherapy may only be beneficial when the cells do not undergo apoptosis in response to IFN-γ and support the contention that the use of some apoptotic cells in vaccines may be detrimental.     

Effect of early adverse events on response and survival outcomes of advanced melanoma patients treated with vemurafenib or vemurafenib plus cobimetinib: A pooled analysis of clinical trial data

This study aimed to evaluate the impact of early adverse events on overall survival (OS), progression‐free survival (PFS) and objective response within a pooled secondary analysis of participants treated with first‐line vemurafenib or vemurafenib plus cobimetinib in the clinical trials BRIM3 and coBRIM. The study included 583 participants who received vemurafenib monotherapy and 247 who received vemurafenib plus cobimetinib. Adverse events requiring vemurafenib/cobimetinib dose adjustment within the first 28 days of therapy were significantly associated with OS (hazard ratio (HR) [95% CI]: dose reduced/interrupted = 0.79 [0.65–0.96]; drug withdrawn = 1.18 [0.71–1.96]; p = 0.032), PFS (HR [95% CI]: dose reduced/interrupted = 0.82 [0.67–0.99]; drug withdrawn = 1.58 [0.97–2.58]; p = 0.017) and objective response (odds ratio (OR) [95% CI]: dose reduced/interrupted = 1.35 [0.99–1.85]; drug withdrawn = 0.17 [0.06–0.43]; p = <0.001). Arthralgia occurring within the first 28 days of vemurafenib or vemurafenib plus cobimetinib therapy was also significantly associated with favourable OS (p = 0.026), PFS (p = 0.042) and objective response (p = 0.047).     

Oral immunogenicity and protective efficacy in mice of transgenic rice plants producing a vaccine candidate antigen (As16) of Ascaris suum fused with cholera toxin B subunit

Cereal crops such as maize and rice are considered attractive for vaccine production and oral delivery. Here, we evaluated the rice Oryza sativa for production of As16—an antigen protective against the roundworm Ascaris suum. The antigen was produced as a chimeric protein fused with cholera toxin B subunit (CTB), and its expression level in the endosperm reached 50 μg/g seed. Feeding the transgenic (Tg) rice seeds to mice elicited an As16-specific serum antibody response when administered in combination with cholera toxin (CT) as the mucosal adjuvant. Although omitting the adjuvant from the vaccine formulation resulted in failure to develop the specific immune response, subcutaneous booster immunization with bacterially expressed As16 induced the antibody response, indicating priming capability of the Tg rice. Tg rice/CT-fed mice orally administered A. suum eggs had a lower lung worm burden than control mice. This suggests that the rice-delivered antigen functions as a prophylactic edible vaccine for controlling parasitic infection in animals.     

Promoter methylation as biomarkers for diagnosis of melanoma: A systematic review and meta-analysis

Melanoma is one of the most common skin cancer that is characterized by rapid growth, early metastasis, high malignant, and mortality. Accumulating evidence demonstrated that promoter methylation of tumor-suppressor genes is implicated in the pathogenesis of melanoma. In the current study, we performed a meta-analysis to identify promising methylation biomarkers in the diagnosis of melanoma. We carried out a systematic literature search using Pubmed, Embase, and ISI web knowledge database and found that gene promoter methylation of 50 genes was reported to be associated with the risk of melanoma. Meta-analysis revealed that hypermethylation of claudin 11 (CLDN11; odds ratio [OR], 16.82; 95% confidence interval [CI], 1.97–143.29; p = 0.010), O-6-methylguanine-DNA methyltransferase (MGMT; OR, 5.59; 95% CI, 2.51–12.47; p < 0.0001), cyclin-dependent kinase inhibitor 2A (p16; OR, 6.57; 95% CI, 2.19–19.75; p = 0.0008), retinoic acid receptor β (RAR-β2; OR, 24.31; 95% CI, 4.58–129.01; p = 0.0002), and Ras association domain family member (RASSF1A; OR, 9.35; 95% CI, 4.73–18.45; p < 0.00001) was significantly higher in melanoma patients compared with controls. CLDN11 (OR, 14.52; 95% CI, 1.84–114.55; p = 0.01), MGMT (OR, 8.08; 95% CI, 1.84–35.46; p = 0.006), p16 (OR, 9.44; 95% CI, 2.68–33.29; p = 0.0005), and RASSF1A (OR, 7.72; 95% CI, 1.05–56.50; p = 0.04) hypermethylation was significantly increased in primary melanoma compared with controls. Methylation frequency of CLDN11 (OR, 25.56; 95% CI, 2.32–281.66; p = 0.008), MGMT (OR, 4.64; 95% CI, 1.98–10.90; p = 0.0004), p16 (OR, 4.31; 95% CI, 1.33–13.96; p = 0.01), and RASSF1A (OR, 10.10; 95% CI, 2.87–35.54; p = 0.0003) was significantly higher in metastasis melanoma compared with controls. These findings indicated that CLDN11, MGMT, p16, RAR-β2, and RASSF1A hypermethylation is a risk factor and a potential biomarker for melanoma. CLDN11, MGMT, p16, and RASSF1A promoter methylation may take part in the development of melanoma and become useful biomarkers in the early diagnosis of the disease.     

Melanocortin 1 receptor (MC1R) polymorphisms’ influence on size and dermoscopic features of nevi

The melanocortin 1 receptor (MC1R) is a highly polymorphic gene. The loss‐of‐function MC1R variants (“R”) have been strongly associated with red hair color phenotype and an increased melanoma risk. We sequenced the MC1R gene in 175 healthy individuals to assess the influence of MC1R on nevus phenotype. We identified that MC1R variant carriers had larger nevi both on the back [p‐value = .016, adjusted for multiple parameters (adj. p‐value)] and on the upper limbs (adj. p‐value = .007). Specifically, we identified a positive association between the “R” MC1R variants and visible vessels in nevi [p‐value = .033, corrected using the FDR method for multiple comparisons (corrected p‐value)], dots and globules in nevi (corrected p‐value = .033), nevi with eccentric hyperpigmentation (corrected p‐value = .033), a high degree of freckling (adj. p‐value = .019), and an associative trend with presence of blue nevi (corrected p‐value = .120). In conclusion, the MC1R gene appears to influence the nevus phenotype.     

Ten-year survival analysis for renal carcinoma patients treated with an autologous tumour lysate vaccine in an adjuvant setting

About 30% of renal cell carcinomas (RCC) will develop recurrence after surgery. Despite evidence for a significantly improved survival by autologous tumour cell vaccination therapy, the procedure has not become standard. Between August 1993 and December 1996, 1,267 RCC patients undergoing radical nephrectomy in 84 German hospitals were subsequently treated by autologous tumour cell vaccination therapy. The study group comprised 692 patients with complete follow-up (stages pT2-3, pNx-2, M0 based on the TNM classification, 4th edition). Subsequent propensity-score matching according to 7 defined criteria with 861 control patients undergoing nephrectomy alone without adjuvant treatment at the Carl-Thiem-Hospital Cottbus, resulted in 495 matched pairs. Overall and stage-specific survival rates were analysed after a median follow-up of 131 months. The 5- and 10-year overall survival (OS) rates were 80.6 and 68.9% in the vaccine group and 79.2 and 62.1% in the control group (p = 0.066). Patients with pT3 stage RCC revealed 5- and 10-year OS rates of 71.3 and 53.6% in the study group and 65.4 and 36.2% in the control group (p = 0.022). In multivariable analysis, patients in the vaccine group showed a significantly improved survival both in the whole study group (HR = 1.28, p = 0.030) and in the subgroup presenting with pT3 stage tumours (HR = 1.67, p = 0.011). Adjuvant treatment with autologous vaccination therapy resulted in a significantly improved overall survival in pT3 stage RCC patients, suggesting benefit especially in this subgroup. However, controlled clinical trials integrating the recent TNM classification and further risk constellations are required to define additional patient groups that may derive benefit from this treatment.     

NK cell activation by dendritic cell vaccine: a mechanism of action for clinical activity

Recent reports revealed that dendritic cell (DC)–natural killer (NK) cell interaction plays an important role in tumor immunity, but few DC vaccine studies have attempted to evaluate the non-specific, yet potentially clinically relevant, NK response to immunization. In this study, we first analyzed in vitro activation of NK cells by DCs similar to those used in clinical trials. Subsequently, NK cell responses were analyzed in a phase I clinical trial of a vaccine consisting of autologous DCs loaded with a fowlpox vector encoding CEA. The data were compared with the clinical outcome of the patients. DC enhances NK activity in vitro, partly by sustaining NK cell survival and by enhancing the expression of NK-activating receptors, including NKp46 and NKG2D. Among nine patients in our clinical trial, NK cytolytic activity increased in four (range 2.5–5 times greater lytic activity) including three who had increased NK cell frequency, was stable in two and decreased in three. NKp46 and NKG2D expression showed a good correlation with the patients’ NK activity. When patients were grouped by clinical activity (stable disease/no evidence of disease (stable/NE, n=5) vs progressive disease (N=4) at 3 months), the majority in the stable/NE group had increases in NK activity (P=0.016). Anti-CEA T cell response was enhanced in all the nine patients analyzed, but was not significantly different between the two groups (P=0.14). Thus, NK responses following DC vaccination may correlate more closely with clinical outcome than do T cell responses. Monitoring of NK response during vaccine studies should be routinely performed.     

The common Scandinavian human leucocyte antigen ancestral haplotype 62.1 as prognostic factor in patients with advanced malignant melanoma

Purpose  We have previously demonstrated an association of the human leukocyte antigen (HLA), HLA-A2 allele with ovarian and prostate cancer mortality as well as a segregation of the ancestral HLA haplotype (AHH) 62.1 [(A2) B15 Cw3 DRB1*04] in patients with stage III–IV serous ovarian cancer. The objective of the present study was to determine the role of the HLA phenotype on the prognosis in stage III–IV malignant melanoma patients. Patients and methods  A cohort of metastatic malignant melanoma patients (n = 91), in stage III (n = 26) or IV (n = 65) were analysed for HLA-A, -B, -Cw and -DRB1 types by PCR/sequence-specific primer method. The frequencies of HLA alleles in the patients were compared to that of healthy Swedish bone marrow donors. The effect of HLA types on prognosis was defined by Kaplan–Meier and Cox analysis. Results  The presence of the AHH 62.1 in clinical stage IV patients was significantly and independently associated with the worst survival rate recorded from the appearance of metastasis (HR = 2.14; CI = 1.02–4.4; P = 0.04). In contrast, the period from the primary diagnosis to metastasis was the longest in patients with this haplotype (HR = 0.40; CI = 0.17–0.90; P = 0.02). Conclusions  Melanoma patients in our cohort with 62.1 AHH which is associated with autoimmune diseases have an initial strong anti-tumour control with longer metastasis-free period. These patients have rapid progression after the appearance of metastasis, responding poorly to chemo- or/and immunotherapy. This apparently paradoxical clinical process could be due to the interplay between tumour clones escape and immune surveillance ending up with a rapid disease progression. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Prognostic significance of tumor iNOS and COX-2 in stage III malignant cutaneous melanoma

Purpose  New prognostic markers are needed for malignant melanoma. Inducible nitric oxide synthase (iNOS) and cyclooxygenase type 2 (COX-2) have been described to correlate with progression of melanoma. Moreover, activating mutations in BRAF/NRAS oncogenes are often detected in melanoma. The BRAF/NRAS mutation status and expression of COX-2 and iNOS were examined to compare their prognostic value for overall survival (OS) in stage III malignant cutaneous melanoma. Experimental design  The expression of iNOS and COX-2 in metastatic lymph nodes from 21 rapidly progressing (OS from date of diagnosis of stage III disease ≤14 months) and 17 slowly progressing (OS ≥60 months) stage III cutaneous melanoma patients was examined by immunohistochemistry. The presence of BRAF/NRAS mutations was analyzed using direct DNA sequencing. χ² exact trend test and logistic regression analysis were used for statistical analysis. Results  Both iNOS (P = 0.002) and COX-2 (P = 0.048) alone significantly predicted OS. The BRAF/NRAS mutation status did not significantly differ between patient groups, although iNOS significantly (P = 0.013) correlated with BRAF mutation frequency. Furthermore, the odds ratio (OR) with respect to OS of iNOS (OR = 10.4) was higher than that of COX-2 (OR = 5.6) and was stable in the multivariate analysis of OS together with disease stage IIIB/C, ulceration, number of metastatic lymph nodes, and Breslow tumor thickness. Conclusion  Our data show that iNOS is an independent and stronger prognostic factor for OS in stage III malignant cutaneous melanoma than COX-2.     

Assessment of immunologic response and recurrence patterns among patients with clinical recurrence after vaccination with a preventive HER2/neu peptide vaccine: from US Military Cancer Institute Clinical Trials Group Study I-01 and I-02

Background  E75, a HER2/neu immunogenic peptide, is expressed in breast cancer (BCa). We have performed clinical trials of E75 + GM-CSF vaccine in disease-free, node-positive and node-negative BCa patients at high recurrence risk and recurrences were noted in both control and vaccine groups. Methods  Among the 186 BCa patients enrolled, 177 completed the study. Patients were HLA typed; the HLA-A2⁺/A3⁺ patients were vaccinated; HLA-A2⁻/A3⁻ patients were followed as controls. Standard clinicopathological factors, immunologic response to the vaccine, and recurrences were collected and assessed. Results  The control group recurrence rate was 14.8 and 8.3% in the vaccinated group (P = 0.17). Comparing the 8 vaccinated recurrences (V-R) to the 88 vaccinated nonrecurrent patients (V-NR), the V-R group had higher nodal stage (≥N2: 75 vs. 5%, P = 0.0001) and higher grade tumors (%grade 3: 88 vs. 31%, P = 0.003). The V-R group did not fail to respond immunologically as noted by equivalent dimer responses and post-DTH responses. Compared to control recurrent patients (C-R), V-R patients trended toward higher-grade tumors and hormone-receptor negativity. C-R patients had 50% bone-only recurrences, compared to V-R patients with no bone-only recurrences (P = 0.05). Lastly, V-R mortality rate was 12.5% compared with 41.7% for the C-R group (P = 0.3). Conclusions  The vaccinated patients who recurred had more aggressive disease compared to V-NR patients. V-R patients had no difference in immune response to the vaccine either in vitro or in vivo. V-R patients, when compared to C-R patients, trended towards more aggressive disease, decreased recurrence rates, decreased mortality, and no bone-only recurrences. Supported by the United States Military Cancer Institute and the Department of Clinical Investigation at Walter Reed Army Medical Center. Funded primarily by the Clinical Breast Care Project. The opinions or assertions contained herein are the private views of the authors and are not to be construed as official or reflecting the views of the Department of the Army, the Department of the Navy, or the Department of Defense. This work represents original research that has not been submitted elsewhere for publication.     

Protective roles of vitamin C and 5-aminosalicylic acid on reproduction in acrylamide intoxicated male mice

ContextSerious health risks have been connected to ongoing, escalating exposure to environmental toxins and one of them is acrylamide (ACR), an organic compound. Although there are many published reports on ACR toxicity, limited information is available regarding the use of two potential antioxidants against ACR-instigated reproductive toxicity.AimsThe study focused on investigating the protective effects of vitamin C and 5-ASA against ACR-incited reproductive toxicity.MethodsA total of 50 male mice aged 4 weeks old were treated for 90 days with different concentrations either of ACR or ACR and vitamin C or ACR and 5- ASA or ACR, vitamin C, and 5- ASA.Key resultsACR significantly reduced serum testosterone level (p = 0.0037), sperm concentration (p = 0.0004), and percentage of sperm motility (p = 0.003), as well as increased sperm abnormality; head (p = 0.0058), tail (p = 0.001), and midpiece (p = 0.0339). Besides, the weight (p = 0.0006) and length (p = 0.0105) of testes, as well as weight (p = 0.0001) and length (p = 0.0021) of epididymis were decreased along with atrophy of seminiferous tubules of the testis, and disintegration of the tubular epithelium of epididymis on ACR exposed mice which were improved by vitamin C and 5-ASA administration.ConclusionsVitamin C and 5-ASA can potentially mitigate the negative effects of ACR on male reproduction; however, combined application is recommended for better performance.ImplicationsIn Bangladesh, this work is anticipated to address the health benefits of vitamin C and 5-ASA, particularly in improving the reproductive health of males against ACR toxicity.     

Feasibility of monitoring advanced melanoma patients using cell‐free DNA from plasma

To determine the feasibility of liquid biopsy for monitoring of patients with advanced melanoma, cell‐free DNA was extracted from plasma for 25 Stage III/IV patients, most (84.0%) having received previous therapy. DNA concentrations ranged from 0.6 to 390.0 ng/ml (median = 7.8 ng/ml) and were positively correlated with tumor burden as measured by imaging (Spearman rho = 0.5435, p = .0363). Using ultra‐deep sequencing for a 61‐gene panel, one or more mutations were detected in 12 of 25 samples (48.0%), and this proportion did not vary significantly for patients on or off therapy at the time of blood draw (52.9% and 37.5% respectively; p = .673). Sixteen mutations were detected in eight different genes, with the most frequent mutations detected in BRAF, NRAS, and KIT. Allele fractions ranged from 1.1% to 63.2% (median = 29.1%). Among patients with tissue next‐generation sequencing, nine of 11 plasma mutations were also detected in matched tissue, for a concordance of 81.8%.     

Spatial distribution and effects of Lantana camara invasion on soil properties and native vegetation community composition in Tugwi-Mukosi Recreational Park,Masvingo, Zimbabwe

This study investigated spatial distribution and effects of Lantana camara invasion on soil properties and vegetation composition in Tugwi-Mukosi Recreational Park. Supervised classification with the Random Forest classifier was used to map L. camara in the park. The study area was stratified into two categories based on soil type and the extent of L. camara invasion. Stratified random sampling was used in data collection to assess differences in species diversity and soil properties in the park. Results revealed that L. camara covered an area of 1772 ha. Using Random Forest classification, the study obtained an accuracy score of 92.1% and an F1 score of 86.25%. Two-way ANOVA showed significant effect of L. camara invasion on soil moisture (F = 28.143, p = 0.000), organic matter (F = 13.377, p = 0.003), pH (F = 1272.369, p = 0.000), total nitrogen (F = 51.762, p = 0.000), total phosphorus (F = 5.000, p = 0.045), woody species density (F = 4.987, p = 0.027), basal area (F = 10.393, p = 0.001), grass species richness (F = 196.258, p = 0.000) and grass cover (F = 3.637, p = 0.042). These results suggest that L. camara was modifying the soil and vegetation properties of the ecosystem which has implications on biodiversity.     

Periostin accelerates human malignant melanoma progression by modifying the melanoma microenvironment

Given no reliable therapy for advanced malignant melanoma, it is important to elucidate the molecular mechanisms underlying the disease progression. Using a quantitative proteomics approach, the ‘isobaric tags for relative and absolute quantitation (iTRAQ)’ method, we identified that the extracellular matrix protein, periostin (POSTN), was highly expressed in invasive melanoma compared with normal skin. An immunohistochemical analysis showed that POSTN was expressed in all invasive melanoma (n = 20) and metastatic lymph node (n = 5) tissue samples, notably restricted in their stroma. In terms of the intercellular regulation of POSTN, we found that there was upregulation of POSTN when melanoma cells and normal human dermal fibroblasts (NHDFs) were cocultured, with restricted expression of TGF‐β1 and TGF‐β3. In a functional analyses, recombinant and NHDF‐derived POSTN significantly accelerated melanoma cell proliferation via the integrin/mitogen‐activated protein kinase (MAPK) signaling pathway in vitro. The size of implanted melanoma tumors was significantly suppressed in POSTN/Rag2 double knockout mice compared with Rag2 knock‐out mice. These results indicate that NHDF‐derived POSTN accelerates melanoma progression and might be a promising therapeutic target for malignant melanoma.     

Lentiviral vector expression of tumour antigens in dendritic cells as an immunotherapeutic strategy

Therapeutic cancer vaccines need to stimulate a refractory immune system to make an effective anti-tumour response. We have explored the use of lentiviral vectors to deliver tumour antigen genes to dendritic cells (DC) as a possible mechanism of immune stimulation. Direct injection of a lentiviral vector encoding the melanoma antigen NY-ESO-1 in HLA-A2 transgenic mice primed NY-ESO-1-specific CD8+ cells that could be expanded by boosting with an NY-ESO-1 vaccinia virus. The expanded cells could kill NY-ESO-1_157–165 peptide-pulsed targets in vivo. In order to examine the priming step directly, we constructed another lentiviral vector expressing the melanoma antigen Melan-A (MART-1). Here we show that Melan-A protein is also efficiently expressed after transduction of human DC cultured from peripheral blood mononuclear cells. When these transduced DC are co-cultured with autologous naïve T cells, they cause the expansion of cells that recognise the HLA-A2 restricted Melan-A_27–35 epitope. The expanded cells are functional in that they release IFN-γ upon antigen stimulation. Melan-A lentiviral vector transduced DC caused a similar level of naïve T-cell expansion to Melan-A_27–35 peptide-pulsed DC in four experiments using different HLA-A2 positive donors. These data suggest that a vaccine based either on DC transduced with a lentiviral vector ex vivo, or on direct lentiviral vector injection, should be assessed in a phase I clinical trial.This article is a symposium paper from the “Robert Baldwin Symposium: 50 years of Cancer Immunotherapy”, held in Nottingham, Great Britain, on 30th June 2005.     

Clinical Application of a Dendritic Cell Vaccine Raised Against Heat-Shocked Glioblastoma

Establishment of a detection platform for glioblastoma-dendritic cell (DC) vaccine preparation and to determine the efficacy of the vaccine in a clinical trial. Autologous glioblastoma-DC vaccine was prepared from a glioblast specimen procured from surgical resection. The specimen was used to enrich the vaccine with peripherally blood-derived DCs after heat-shock induced, glioblastoma apoptosis. The control group received conventional treatment of surgery and radio-chemotherapy post-operation. The therapeutic group received a combination of glioblastoma-DC vaccine and conventional therapy. A comparison of the functional immune parameters, including tumor control, rate live time, Karnofsky scores, and complications occurring in each group were observed and recorded. The proportions of peripheral CD3⁺, CD3⁺CD4⁺, CD4⁺/CD8⁺, and NK cells were significantly higher after DC vaccination than the control group (P < 0.05). Serum levels of IL-2, IL-12, and IFN-γwere significantly higher after DC vaccination than in the control group (P < 0.05). Nine months after vaccination, tumor control rate is significantly improved in the DC group compared with the control group (P < 0.05); survival rate was significantly higher in DC group than in control group (P < 0.05) and the time to relapse was significantly longer in DC group than that in control group (P < 0.05). Karnofsky scores were better in DC vaccination group 6 and 9 months post-treatment compared with the control group (P < 0.05). The combination of glioma DC vaccine and radiotherapy/chemotherapy post-operatively enhances the immune function of patients, increases the tumor control rate, prolongs the survival time and relapse duration, improves the quality of life, and therefore provides a more effective intervention of treating glioblastoma.     

In vitro heat shock of human monocytes results in a proportional increase of inducible Hsp70 expression according to the basal content

Heat shock proteins play an important role as molecular chaperones of the cell. Inducible heat shock protein 70 is rapidly synthesised in response to numerous stressors and monocytes are sensitive to changes in core temperature resulting in a circadian variation of Hsp70 expression. Monocytes were isolated via density centrifugation from nine healthy male volunteers at 5 am, 1 pm and 9 pm, representing the nadir (5 am), peak (9 pm) and intermediate (1 pm) of Hsp70 expression in the 24-h cycle. Analysis of freshly isolated monocytes for Hsp70 expression confirmed Hsp70 levels at the three selected time points. Monocytes were subjected to in vitro heat shock at 40°C (±0.1) for 90 min with a 90 min 37°C (±0.1) exposure acting as a control. A significant increase in Hsp70 was observed at 5 am (p < 0.001) and 1 pm (p = 0.028) at 40°C when compared to 37°C but not at 9 pm (p = 0.19). A significant increase was also observed from the basal levels of Hsp70, measured on freshly isolated monocytes and the levels detected after heat shock at 40°C at 5 am (p < 0.001) and 1 pm (p = 0.001), which was not observed at 9 pm (p = 0.15). Furthermore, a significant correlation was observed in the heat shock response at 40°C and that obtained at 37°C (p < 0.001). In conclusion, the heat shock response in monocytes is directly proportional to the amount of Hsp70 present in the cells and the stress response may be much higher at different times of the day.