Experimental vaccine strategies for cancer immunotherapy

Recently, cancer immunotherapy has emerged as a therapeutic option for the management of cancer patients. This is based on the fact that our immune system, once activated, is capable of developing specific immunity against neoplastic but not normal cells. Increasing evidence suggests that cell-mediated immunity, particularly T-cell-mediated immunity, is important for the control of tumor cells. Several experimental vaccine strategies have been developed to enhance cell-mediated immunity against tumors. Some of these tumor vaccines have generated promising results in murine tumor systems. In addition, several phase I/II clinical trials using these vaccine strategies have shown extremely encouraging results in patients. In this review, we will discuss many of these promising cancer vaccine strategies. We will pay particular attention to the strategies employing dendritic cells, the central player for tumor vaccine development.     

Temperature-assisted high hydrostatic pressure inactivation of Staphylococcus aureus in a ham model system: evaluation in selective and nonselective medium

Aims: The purpose of this study was to investigate the inactivation kinetics of Staphylococcus aureus in a ham model system by high hydrostatic pressure at ambient (25°C) and selected temperatures (45, 55°C). Selective [Baird Parker (BP) agar] and nonselective [brain heart infusion (BHI) agar] growth media were used for enumeration in order to count viable and sublethally injured cells. Methods and Results: The micro‐organism was exposed to a range of pressures (450, 500, 550, 600 MPa) at ambient temperature (25°C) for up to 45 min. Additionally, the behaviour of the micro‐organism was evaluated at mild temperatures in combination with high pressure treatment, namely: (i) 350, 400 and 450 MPa at 45°C; and (ii) 350 and 400 MPa at 55°C, for up to 12 min. Inactivation kinetics were calculated in terms of D_p and z_p values. Survival curves of S. aureus at ambient temperature were mostly linear, whereas when temperature was applied, tailing was observed in most survival curves. The estimated D_p values and therefore the number of surviving cells, were substantially higher on the selective BP agar in the whole range of pressures applied, indicating that S. aureus showed greater recovery in the selective BP agar than the nonselective BHI agar. Samples pressurized at ambient temperature needed higher pressures (over 500 MPa) to achieve a reduction of the population of the pathogen more than 5 log CFU ml^?1. The same level of inactivation was achieved at lower pressure levels when mild heating was simultaneously applied. Indeed, more than 6 log CFU ml^?1 reductions were obtained at 400 MPa and 55°C within the first 7 min of the process in BHI medium. Conclusion: Elevated temperatures allowed lower pressure levels and shorter processing times of pathogen inactivation than at room temperature. Greater recovery of the pathogen was observed in the selective (BP agar) medium, regardless of pressure and temperature applied. Significance and Impact of the Study: The obtained kinetics could be employed by the industry in selecting optimum pressure/temperature processing conditions. Attention must be given to the selection of the enumeration medium, as the use of an inappropriate medium would lead to underestimation of the surviving cells, thus imposing a risk in the microbiological safety of the product.     

中性体系中超高压－Nisin协同作用下的细菌致死机理

摘要：【目的】为保证超高压中性食品的杀菌强度,可以??????????通过添加Nisin等细菌素协同杀菌以达到商业无菌要求。本文从分子水平和超微结构揭示二者协同作用下的细胞致死机理,为超高压杀菌在中性食品中的应用奠定理论基础。【方法】采用pH7.0的环境体系,100－500 MPa的超高压处理,Nisin浓度为200 IU/mL。通过荧光染色法和紫外吸收法检测细胞膜通透性,傅里叶转换红外光谱法检测细菌细胞壁、蛋白以及核酸的变化,透射电镜观察细菌在协同作用下的形态变化。【结果】结果发现：中性条件下,超高压与     

Current approaches in dendritic cell generation and future implications for cancer immunotherapy

The discovery of tumor-associated antigens, which are either selectively or preferentially expressed by tumors, together with an improved insight in dendritic cell biology illustrating their key function in the immune system, have provided a rationale to initiate dendritic cell-based cancer immunotherapy trials. Nevertheless, dendritic cell vaccination is in an early stage, as methods for preparing tumor antigen presenting dendritic cells and improving their immunostimulatory function are continuously being optimized. In addition, recent improvements in immunomonitoring have emphasized the need for careful design of this part of the trials. Still, valuable proofs-of-principle have been obtained, which favor the use of dendritic cells in subsequent, more standardized clinical trials. Here, we review the recent developments in clinical DC generation, antigen loading methods and immunomonitoring approaches for DC-based trials.     

Effect of temperature on gelatinization and retrogradation in high hydrostatic pressure treatment of potato starch-water mixtures

The effect of temperature (20-70 °C) on the gelatinization and retrogradation of potato starch-water mixtures (10-70%, w/w) treated with high hydrostatic pressure (HHP) (400-1000 MPa) was investigated. Gelatinization enthalpy change (ΔH_gel) and re-gelatinization enthalpy change of retrograded crystalline part (ΔH_retro) of the HHP-treated starch were evaluated using differential scanning calorimetry. The value of ΔH_gel of 10-20% (w/w) mixtures decreased with increased pressure and temperature, while ΔH_gel of 30-50% (w/w) mixtures decreased to certain values with increased pressure and the values depended on treatment temperature. With higher temperature and pressure conditions, ΔH_gel of 10-40% (w/w) mixtures reached zero, but ΔH_gel of 50-70% (w/w) mixtures did not. Retrogradation was observed with HHP-treated 20-60% (w/w) mixtures and the value of ΔH_retro depended on the starch content, pressure, and temperature. The value of ΔH_retro trended to increase with increase in starch content. In addition, retrogradation was promoted by HHP treatment at low temperature. Gelatinizaiton and retrogradation behaviors of HHP-treated (400-1000 MPa) potato starch-water mixtures (10-70%, w/w) at 20-70 °C were summerized in a series of state diagrams.     

Intermittent hydrostatic pressure maintains and enhances the chondrogenic differentiation of cartilage progenitor cells cultivated in alginate beads

The objective of this study was to explore the effects of intermittent hydrostatic pressure (IHP) on the chondrogenic differentiation of cartilage progenitor cells (CPCs) cultivated in alginate beads. CPCs were isolated from the knee joint cartilage of rabbits, and infrapatellar fat pad‐derived stem cells (FPSCs) and chondrocytes (CCs) were included as the control cell types. Cells embedded in alginate beads were treated with IHP at 5 Mpa and 0.5 Hz for 4 h/day for 1, 2, or 4 weeks. The cells' migratory and proliferative capacities were evaluated using the scratch and Live/Dead assays, respectively. Hematoxylin and eosin staining, safranin O staining, and immunohistochemical staining were performed to determine the effects of IHP on the synthesis of extracellular matrix (ECM) proteins. Real‐time polymerase chain reaction analysis was performed to measure the expression of genes related to chondrogenesis. The scratch and Live/Dead assays revealed that IHP significantly promoted the migration and proliferation of FPSCs and CPCs to different extents. The staining experiments showed greater production of cartilage ECM components (glycosaminoglycans and collagen II) by cells exposed to IHP, and the gene expression analysis demonstrated that IHP stimulated the expression of chondrocyte‐related genes. Importantly, these effects of IHP were more prominent in CPCs than in FPSCs and CCs. Considering all of our experimental results combined, we conclude that CPCs demonstrated a stronger chondrogenic differentiation capacity than the FPSCs and CCs under stimulation with IHP. Thus, the use of CPCs, combined with mechanical stimulation, may represent a valuable strategy for cartilage tissue engineering.     

Oscillatory shear stress and hydrostatic pressure modulate cell-matrix attachment proteins in cultured endothelial cells

Summary Endothelial cells (ECs) may behave as hemodynamic sensors, translating mechanical information from the blood flow into biochemical signals, which may then be transmitted to underlying smooth muscle cells. The extracellular matrix (ECM), which provides adherence and integrity for the endothelium, may serve an important signaling function in vascular diseases such as atherogenesis, which has been shown to be promoted by low and oscillating shear stresses. In this study, confluent bovine aortic ECs (BAECs) were exposed to an oscillatory shear stress or to a hydrostatic pressure of 40 mmHg for time periods of 12 to 48 h. Parallel control cultures were maintained in static condition. Although ECs exposed to hydrostatic pressure or to oscillatory flow had a polygonal morphology similar to that of control cultures, these cells possessed more numerous central stress fibers and exhibited a partial loss of peripheral bands of actin, in comparison to static cells. In EC cultures exposed to oscillatory flow or hydrostatic pressure, extracellular fibronectin (Fn) fibrils were more numerous than in static cultures. Concomitantly, a dramatic clustering ofα ₅β₁ Fn receptors and of the focal contact-associated proteins vinculin and talin occurred. Laminin (Ln) and collagen type IV formed a network of thin fibrils in static cultures, which condensed into thicker fibers when BAECs were exposed to oscillatory shear stress or hydrostatic pressure. The ECM-associated levels of Fn and Ln were found to be from 1.5-to 5-fold greater in cultures exposed to oscillatory shear stress or pressure for 12 and 48 h, than in static cultures. The changes in the organization and composition of ECM and focal contacts reported here suggest that ECs exposed to oscillatory shear stress or hydrostatic pressure may have different functional characteristics from cells in static culture, even though ECs in either environment exhibit a similar morphology.     

Generation of a Novel Dendritic-cell Vaccine Using Melanoma and Squamous Cancer Stem Cells

We identified cancer stem cell (CSC)-enriched populations from murine melanoma D5 syngeneic to C57BL/6 mice and the squamous cancer SCC7 syngeneic to C3H mice using ALDEFLUOR/ALDH as a marker, and tested their immunogenicity using the cell lysate as a source of antigens to pulse dendritic cells (DCs). DCs pulsed with ALDH^high CSC lysates induced significantly higher protective antitumor immunity than DCs pulsed with the lysates of unsorted whole tumor cell lysates in both models and in a lung metastasis setting and a s.c. tumor growth setting, respectively. This phenomenon was due to CSC vaccine-induced humoral as well as cellular anti-CSC responses. In particular, splenocytes isolated from the host subjected to CSC-DC vaccine produced significantly higher amount of IFNγ and GM-CSF than splenocytes isolated from the host subjected to unsorted tumor cell lysate pulsed-DC vaccine. These results support the efforts to develop an autologous CSC-based therapeutic vaccine for clinical use in an adjuvant setting.     

High pressure homogenization inactivation of Escherichia coli and Staphylococcus aureus in phosphate buffered saline,milk and apple juice

High pressure homogenization (HPH) offers new opportunities for food pasteurization/sterilization. Escherichia coli and Staphylococcus aureus suspended in phosphate buffered saline (PBS) buffer, milk and apple juice at initial concentration of ~10⁶ log₁₀ CFU per ml were subjected to HPH treatments up to 200 MPa with inlet temperatures at 4–40°C. After HPH at 200 MPa with the inlet temperature at 40°C, the count of E. coli suspended in PBS, milk and apple juice reduced by 3·42, 3·67 and 3·19 log₁₀ CFU per ml respectively while the count of S. aureus decreased by 2·21, 1·02 and 2·33 log₁₀ CFU per ml respectively suggesting that S. aureus was more resistant. The inactivation data were well fitted by the polynomial equation. Milk could provide a protective effect for S. aureus against HPH. After HPH at 200 MPa with the inlet temperature at 20°C, the cell structure of E. coli was destroyed, while no obvious damages were found for S. aureus.     

Modelling the effect of high pressure on the inactivation kinetics of a pressure-resistant strain of Pediococcus damnosus in phosphate buffer and gilt-head seabream (Sparus aurata)

AIMS: The aim of this research was to: (i) determine the inactivation pattern of a pressure-resistant strain of Pediococcus damnosus by high hydrostatic pressure in phosphate buffer (pH 6.7) and gilt-head seabream using the linear, biphasic and Weibull models; and (ii) validate the applicability of the Weibull model to predict survival curves at other experimental pressure levels. METHODS AND RESULTS: A pressure-resistant strain of P. damnosus was exposed to a range of pressures (500, 550, 600 and 650 MPa) in phosphate buffer (pH 6.7) and gilt-head seabream for up to 8 min at ambient temperature (23 degrees C). Inactivation kinetics were described by the linear, biphasic and Weibull models. Increasing the magnitude of the pressure applied resulted in increasing levels of inactivation. Pronounced tailing effect was observed at pressures over 600 MPa. The Weibull and biphasic models consistently produced better fit than the linear model as inferred by the values of the root mean squared error, coefficient of determination (R2) and accuracy factor (A(f)). The scale factor (b) of the Weibull model was linearly correlated with pressure (P) treatment in the whole pressure range. Substituting the b parameter in the initial Weibull function and calculating the shape factor (n) by linear interpolation, high pressure (P) was directly incorporated into the model providing reasonable predictions of the survival curves at 570 and 630 MPa. Comparison between the survival curves in phosphate buffer and gilt-head seabream showed a clear protective effect of the food matrix on the resistance of the micro-organism, especially at 500 and 550 MPa. CONCLUSIONS: The Weibull and biphasic models were more flexible to describe the survival curves of P. damnosus in the experimental pressure range, taking also into account the tailing effect that could not be included in the linear model. The Weibull model could also give reasonable predictions of the survival curves at other experimental pressures in both pressure menstrua. As the food matrix has a protective effect in microbial inactivation, the development of accurate mathematical models should be done directly on real food to avoid under- or over-processing times. SIGNIFICANCE AND IMPACT OF THE STUDY: The development of accurate models to describe the survival curves of micro-organisms under high hydrostatic pressure treatment would be very important to the food industry for process optimisation, food safety and extension of the applicability of high pressure processing.     

Systematic analysis of HSP gene expression and effects on cell growth and survival at high hydrostatic pressure in Saccharomyces cerevisiae

We systematically investigated the role of HSP genes in the growth and survival of Saccharomyces cerevisiae under high hydrostatic pressure together with analysis of pressure-regulated gene expression. Cells of strain BY4742 were capable of growth at moderate pressure of 25 MPa. When pressure of 25 MPa was applied to the cells, the expression of HSP78, HSP104, and HSP10 was upregulated by about 3- to 4-fold, and that of HSP32, HSP42, and HSP82 was upregulated by about 2- to 2.6-fold. However, the loss of one of the six genes did not markedly affect growth at 25 MPa, while the loss of HSP31 impaired high-pressure growth. These results suggest that Hsp31 plays a role in high-pressure growth but that the six upregulated genes do not. Extremely high pressure of 125 MPa decreased the viability of the wild-type cells to 1% of the control level. Notably, the loss of HSP genes other than HSP31 enhanced the survival rate by about fivefold at 125 MPa, suggesting that the cellular defensive system against high pressure could be strengthened upon the loss of the HSP genes. In this paper, we describe the requirement for and significance of a subset of HSP genes in yeast cell growth at moderate pressure and survival at extremely high pressure.     

Reformation in chimeric antigen receptor based cancer immunotherapy: Redirecting natural killer cell

Natural killer (NK) cells are an important subset of lymphocytes which play a critical role in host immunity against cancers. With MHC-independent recognition, short lifespan and potent cytotoxicity, NK cells make a promising candidate for chimeric antigen receptor (CAR)-engineered cancer immunotherapy. Due to innate biological properties of NK cells, CAR-NK may outperform CAR-T therapy in terms of less side effects and more universal access, which may become a great reformation in CAR-based cancer immunotherapy. The CARs used in peripheral blood (PB) NK cells as well as NK cell line like NK-92 are the most important outfits defining antigenic specificity. The constructs of CARs used in NK cells from different sources vary, which all undergo generational optimization. The anti-tumor effects of CAR-NK have been validated in numerous preclinical trials for cancers, including hematologic malignancies and many solid tumors, which provide evidence for potential clinical application of CAR-NK. Additionally, this review concludes the challenges faced in the application of CAR-NK. Although CAR-NK is considered as one of the most possible “off-the-shelf” products, the improvement for the efficiency of expansion and transduction as well as the solution for underlying safety issues is still needed. Possible coping strategies for challenges and upgrades in techniques are also highlighted for future development in CAR-NK cancer immunotherapy.     

Immune checkpoint blockade and its combination therapy with small-molecule inhibitors for cancer treatment

Initially understood for its physiological maintenance of self-tolerance, the immune checkpoint molecule has recently been recognized as a promising anti-cancer target. There has been considerable interest in the biology and the action mechanism of the immune checkpoint therapy, and their incorporation with other therapeutic regimens. Recently the small-molecule inhibitor (SMI) has been identified as an attractive combination partner for immune checkpoint inhibitors (ICIs) and is becoming a novel direction for the field of combination drug design. In this review, we provide a systematic discussion of the biology and function of major immune checkpoint molecules, and their interactions with corresponding targeting agents. With both preclinical studies and clinical trials, we especially highlight the ICI + SMI combination, with its recent advances as well as its application challenges.     

Challenges in cancer research and multifaceted approaches for cancer biomarker quest

Recent advances in cancer biology have subsequently led to the development of new molecularly targeted anti-cancer agents that can effectively hit cancer-related proteins and pathways. Despite better insight into genomic aberrations and diversity of cancer phenotypes, it is apparent that proteomics too deserves attention in cancer research. Currently, a wide range of proteomic technologies are being used in quest for new cancer biomarkers with effective use. These, together with newer technologies such as multiplex assays could significantly contribute to the discovery and development of selective and specific cancer biomarkers with diagnostic or prognostic values for monitoring the disease state. This review attempts to illustrate recent advances in the field of cancer biomarkers and multifaceted approaches undertaken in combating cancer.     

Potential roles and targeted therapy of the CXCLs/CXCR2 axis in cancer and inflammatory diseases

The chemokine receptor CXCR2 and its ligands are implicated in the progression of tumours and various inflammatory diseases. Activation of the CXCLs/CXCR2 axis activates multiple signalling pathways, including the PI3K, p38/ERK, and JAK pathways, and regulates cell survival and migration. The CXCLs/CXCR2 axis plays a vital role in the tumour microenvironment and in recruiting neutrophils to inflammatory sites. Extensive infiltration of neutrophils during chronic inflammation is one of the most important pathogenic factors in various inflammatory diseases. Chronic inflammation is considered to be closely correlated with initiation of cancer. In addition, immunosuppressive effects of myeloid-derived suppressor cells (MDSCs) against T cells attenuate the anti-tumour effects of T cells and promote tumour invasion and metastasis. Over the last several decades, many therapeutic strategies targeting CXCR2 have shown promising results and entered clinical trials. In this review, we focus on the features and functions of the CXCLs/CXCR2 axis and highlight its role in cancer and inflammatory diseases. We also discuss its potential use in targeted therapies.