Successful ex vivo expansion of tumor infiltrating lymphocytes with systemic chemotherapy prior to surgical resection.

Tumor infiltrating lymphocytes (TIL) have demonstrated efficacious clinical outcomes for many patients with various types of solid cancers, including melanoma, gastrointestinal cancer, lung cancer, and head and neck cancer. Currently, the majority of clinical trials require that patients did not receive systemic therapy right before tumor tissue resection to avoid the interference of chemotherapy in the ex vivo TIL expansion. The primary disadvantage of this strategy is limiting the accessibility of TIL therapy for many eligible cancer patients. Over the past decade, substantial progress has been made for ex vivo expansion technologies in T cells. In this study, we investigated the possibility of enrolling patients who underwent chemotherapy prior to surgical resection. We collected seventeen tumor tissues from treatment naive cases, and five from cases that underwent chemotherapies. Cancer indications enrolled in this study were colorectal and lung cancers from both primary and metastatic sites, such as liver and brain. TILs from these tumors were expanded ex vivo to 2.1E8 (total viable lymphocytes counts) on average, with an overall success rate of 90.9%. Subsequently, TIL phenotypes and cytokine production were analyzed using flow cytometry and ELISA, respectively. We demonstrated functional TIL expansion from tumor tissues despite chemotherapy prior to surgical resection. We observed no significant phenotypic or functional differences between groups with and without chemotherapy. TIL expansion rate and characteristics were similar regardless of chemotherapy prior to resection, thereby providing a possibility to recruit patients with the most recent chemotherapy history in TIL therapy trials.

NLRX1 negatively regulates TLR-induced NF-κB signaling by targeting TRAF6 and IKK.

Tight regulation of NF-κB signaling is essential for innate and adaptive immune responses, yet the molecular mechanisms responsible for its negative regulation are not completely understood. Here, we report that NLRX1, a NOD-like receptor family member, negatively regulates Toll-like receptor-mediated NF-κB activation. NLRX1 interacts with TRAF6 or IκB kinase (IKK) in an activation signal-dependent fashion. Upon LPS stimulation, NLRX1 is rapidly ubiquitinated, disassociates from TRAF6, and then binds to the IKK complex, resulting in inhibition of IKKα and IKKß phosphorylation and NF-κB activation. Knockdown of NLRX1 in various cell types markedly enhances IKK phosphorylation and the production of NF-κB-responsive cytokines after LPS stimulation. We further provide in vivo evidence that NLRX1 knockdown in mice markedly enhances susceptibility to LPS-induced septic shock and plasma IL-6 level. Our study identifies a previously unrecognized role for NLRX1 in the negative regulation of TLR-induced NF-κB activation by dynamically interacting with TRAF6 and the IKK complex.

Measurement and Quantitative Characterization of Whole-Body Pharmacokinetics of Exogenously Administered T Cells in Mice.

Here we have investigated whole-body pharmacokinetics (PK) of exogenously administered T cells in a mouse model of melanoma and have developed a physiologically based pharmacokinetic (PBPK) model to quantitatively characterize the data. T cells were isolated from the spleen of tumor-bearing mice, activated, and labeled with chromium-51 to facilitate the quantification. Labeled T cells were injected in the tumor-bearing mice, and PK was measured in 19 different tissues. It was found that T cells disappear from the blood rapidly after administration and accumulate in the tissues to various extents. Spleen, liver, lung, kidney, bone, and lymph nodes accounted for more than 90% of T cells in the body. The distribution of T cells in solid tumors was found to be very low, hovering below 1%ID/g (percent of injected dose per gram of tissue) during the entire study. However, this observation may differ for targeted TCR-T and CAR-T cells. Observed PK profiles also suggest that T-cell-based therapies may be more successful in treating cancers of the lymphatic system and bone marrow metastases compared to solid tumors. A PBPK model was developed to characterize the whole-body PK of T cells, which incorporated key processes such as extravasation, elimination, and recirculation of T cells via lymph flow. Retention factors were incorporated into the spleen, liver, and kidney compartment to adequately capture the PK profiles. The model was able to characterize observed PK profiles reasonably well, and parameters were estimated with good confidence. The PK data and PBPK model presented here provide unprecedented insight into the biodistribution of exogenously administered T cells.

Cellular Immunotherapy in the Treatment of Hematopoietic Malignancies.

Cancer immunotherapy has been shown to be an efficacious therapeutic approach in the treatment of cancers including hematopoietic malignancies. Induction of T cell cytotoxicity against tumors by adoptive cell therapies (ACT), cancer vaccines, gene therapies, and monoclonal antibody therapies has been intensively studied. In particular, immune checkpoint blockade and chimeric antigen receptor T (CAR-T) cell therapies are the recent clinical successes in cancer immunotherapy. This article introduces the main concepts and addresses the most relevant clinical modalities of cellular immunotherapies for hematological malignancies: antigen non-specific T cell therapy, genetically modified T cell receptor (TCR) T cell therapy, chimeric antigen receptor (CAR) T cell therapy, and CAR-T cell clinical trials in leukemia, lymphoma, and multiple myeloma. Clinical trials have shown encouraging results, but future studies may need to incorporate novel CAR constructs or targets with enhanced safety and efficacy to ensure long-term benefits.

Antitumor activity of antibody against cytotoxic T lymphocyte epitope peptide of lymphocyte-specific protein tyrosine kinase.

Although humoral responses against CTL epitope peptides from lymphocyte-specific protein tyrosine kinase (Lck) antigen have been observed in the majority of healthy donors and cancer patients, the biological activity of the antibody has not been determined. We investigated the biological activity of mAb against CTL epitope peptide of Lck antigen at positions 486-494 (anti-Lck-486 mAb). This mAb induced dendritic cell maturation from murine bone marrow cells by the immune complex form in vitro, and inhibited tumor growth in association with a suppression of tumor-infiltrating T cells, including T regulatory cells in a murine model using female BALB/cCrlCrlj mice (H-2Kd ). More potent tumor inhibition was observed when this mAb was given prior to peptide vaccination. These results may help to unveil the biological activity of anti-Lck peptide antibodies against CTL epitope peptides.

Feasibility study of personalized peptide vaccination for hepatocellular carcinoma patients refractory to locoregional therapies.

Overall survival of patients with hepatocellular carcinoma (HCC) refractory to locoregional therapy is dismal, even following treatment with sorafenib, a multikinase inhibitor. To develop a more efficacious treatment, we undertook a feasibility study of personalized peptide vaccination (PPV) for HCC, in which the peptides were selected from 31 peptide candidates based on the pre-existing immunity. Twenty-six HCC patients refractory to locoregional therapies (cohort 1) and 30 patients refractory to both locoregional and systemic therapies (cohort 2) were entered into the study. There were no severe adverse events related to PPV except for one injection site reaction. At the end of the first cycle of six vaccinations, successful CTL or IgG boosting was observed in 57% or 46% of patients in cohort 1 and in 54% or 52% of patients in cohort 2, respectively. Successful IgG boosting at the end of the second cycle was observed in the majority of patients tested. Median overall survival was 18.7 months (95% confidence interval, 12.2-22.5 months) in cohort 1, and 8.5 months (95% confidence interval, 5.9-12.2 months) in cohort 2. Based on the higher rates of immune boosting and the safety profile of PPV, further clinical studies of PPV would be warranted for patients with HCC refractory to not only locoregional therapy but also both locoregional and systemic therapies. The protocol of this study was registered with the UMIN Clinical Trials Registry (UMIN000001882 and UMIN000003590).

A randomized phase II trial of personalized peptide vaccine with low dose cyclophosphamide in biliary tract cancer.

Since the prognosis of advanced biliary tract cancer (aBTC) still remains very poor, new therapeutic approaches, including immunotherapies, need to be developed. In the current study, we conducted an open-label randomized phase II study to test whether low dose cyclophosphamide (CPA) could improve antigen-specific immune responses and clinical efficacy of personalized peptide vaccination (PPV) in 49 previously treated aBTC patients. Patients with aBTC refractory to at least one regimen of chemotherapies were randomly assigned to receive PPV with low dose CPA (100 mg/day for 7 days before vaccination) (PPV/CPA, n = 24) or PPV alone (n = 25). A maximum of four HLA-matched peptides were selected based on the pre-existing peptide-specific IgG responses, followed by subcutaneous administration. T cell responses to the vaccinated peptides in the PPV/CPA arm tended to be greater than those in the PPV alone arm. The PPV/CPA arm showed significantly better progression-free survival (median time: 6.1 vs 2.9 months; hazard ratio (HR): 0.427; P = 0.008) and overall survival (median time: 12.1 vs 5.9 months; HR: 0.376; P = 0.004), compared to the PPV alone arm. The PPV alone arm, but not the PPV/CPA arm, showed significant increase in plasma IL-6 after vaccinations, which might be associated with inhibition of antigen-specific T cell responses. These results suggested that combined treatment with low dose CPA could provide clinical benefits in aBTC patients under PPV, possibly through prevention of IL-6-mediated immune suppression. Further clinical studies would be recommended to clarify the clinical efficacy of PPV/CPA in aBTC patients.

Immunological evaluation of peptide vaccination for cancer patients with the HLA -A11+ or -A33+ allele.

The HLA-A11 or -A33 allele is found in approximately 18% or 10% of the Asian population, respectively, but each of which is a minor allele worldwide, and therefore no clinical trials were previously conducted. To develop a therapeutic peptide vaccine for each of them, we investigated immunological responses of advanced cancer patients with the HLA-A11+ /A11+ (n = 18) or -A33+ /A33+ (n = 13) allele to personalized peptide vaccine (PPV) regimens. The primary sites of HLA-A11+/A11+ or -A33+/A33+ patients were the colon (n = 4 or 2), stomach (2 or 3), breast (3 or 2), lung and pancreas (2 or 2), and so on. For PPV, a maximum of four peptides were selected from nine different peptides capable of binding to HLA-A11 and -A33 molecules based on the pre-existing peptide-specific IgG responses. There were no severe adverse events related to PPV. At the end of the first cycle, peptide-specific CTL responses were augmented in 4/12 or 2/9 of HLA-A11+ /A11+ or -A33+ /A33+ patients, while peptide-specific IgG responses were augmented in 6/14 or 4/10 patients, respectively. Clinical responses consisted of four stable diseases and 14 progressive diseases in HLA-A11+ /A11+ patients, versus seven and six in -A33+ /A33+ patients, respectively. Further clinical study of PPV could be recommended because of the safety and positive immunological responses.

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.

A randomized phase II clinical trial of personalized peptide vaccination with metronomic low-dose cyclophosphamide in patients with metastatic castration-resistant prostate cancer.

This study investigated the effect of metronomic cyclophosphamide (CPA) in combination with personalized peptide vaccination (PPV) on regulatory T cells (Treg) and myeloid-derived suppressor cells (MDSC), and whether it could improve the antitumor effect of PPV. Seventy patients with metastatic castration-resistant prostate cancer were randomly assigned (1:1) to receive PPV plus oral low-dose CPA (50 mg/day), or PPV alone. PPV treatment used a maximum of four peptides chosen from 31 pooled peptides according to human leukocyte antigen types and antigen-specific humoral immune responses before PPV, for 8 subcutaneous weekly injections. Peptide-specific cytotoxic T lymphocyte (CTL) and immunoglobulin G responses were measured before and after PPV. The incidence of grade 3 or 4 hematologic adverse events was higher in the PPV plus CPA arm than in the PPV alone arm. Decrease in Treg and increase in MDSC were more pronounced in PPV plus CPA treatment than in PPV alone (p = 0.036 and p = 0.048, respectively). There was no correlation between the changes in Treg or MDSC and CTL response. There was no difference in positive immune responses between the two arms, although overall survival in patients with positive immune responses was longer than in those with negative immune responses (p = 0.001). Significant differences in neither progression-free survival nor overall survival were observed between the two arms. Low-dose CPA showed no change in the antitumor effect of PPV, possibly due to the simultaneous decrease in Treg and increase in MDSC, in patients under PPV.

Identification of novel Lck-derived T helper epitope long peptides applicable for HLA-A2(+) cancer patients as cancer vaccine.

The present study attempted to identify T helper epitope long peptides capable of inducing cytotoxic T lymphocytes (CTL) from Lck antigen (p56(Lck) ), the src family tyrosine kinase, which is known to be aberrantly expressed in metastatic cancers cells, in order to develop a long peptide-based cancer vaccine for HLA-A2(+) cancer patients. Based on the biding motif to the HLA-DR and HLA-A2 alleles, 94 peptides were prepared from the Lck antigen. These peptides were screened for their reactivity to immunoglobulin G (IgG) from plasma of cancer patients, followed by testing of their ability to induce both CD4(+) and CD8(+) T lymphocytes showing not only peptide-specific IFN-γ production but cytotoxicity against HLA-A2(+) cancer cells from peripheral blood mononuclear cells (PBMC) of HLA-A2(+) cancer patients. Among 94 peptides tested, the three T helper epitope long peptides and their inner CTL epitope short peptides with HLA-A2 binding motifs were frequently recognized by IgG of cancer patients, and efficiently induced both CD4(+) IFN-γ(+) and CD8(+) IFN-γ(+) T lymphocytes. Patients' PBMC stimulated with these long peptides showed cytotoxicity against HLA-A2(+) Lck(+) cancer cells in HLA-class I and HLA-class II dependent manners. These three peptides might be useful for long peptide-based vaccines for HLA-A2(+) cancer patients with Lck(+) tumor cells.

Personalized peptide vaccination for cervical cancer patients who have received prior platinum-based chemotherapy.

A feasibility study was performed to evaluate the immunological efficacy and safety of a personalized peptide vaccine (PPV) for cervical cancer patients who have received platinum-based chemotherapy. A total of 24 patients with standard chemotherapy-resistant cervical cancer, including 18 recurrent cases, were enrolled in this study and received a maximum of 4 peptides based on HLA-A types and IgG levels to the vaccine candidate peptides in pre-vaccination plasma. The parental protein expression of most of the vaccine peptides was confirmed in the cervical cancer tissues. No vaccine-related systemic grade 3 or 4 adverse events were observed in any patients. Due to disease progression, 2 patients failed to complete the first cycle of vaccinations (sixth vaccination). Cytotoxic T-lymphocyte (CTL) or IgG responses specific for the peptides used for vaccination were augmented in half of cases after the first cycle. The median overall survival was 8.3 months. The clinical responses of the evaluable 18 cases consisted of 1 case with a partial response and 17 cases with disease progression; the remaining 6 cases were not evaluable. Performance status, injection site skin reaction and circulating PD-1(+) CD4(+) T-cells were significantly prognostic of overall survival, and multivariate analysis also indicated that the performance status and circulating PD-1(+) CD4(+) T-cells were prognostic. Because of the safety and immunological efficacy of PPV and the possible prolongation of overall survival, further clinical trials of PPV at a larger scale in advanced or recurrent cervical cancer patients who have received prior platinum-based chemotherapy are recommended.

Immunological evaluation of peptide vaccination for cancer patients with the HLA-A26 allele.

To develop a peptide vaccine for cancer patients with the HLA-A26 allele, which is a minor population worldwide, we investigated the immunological responses of HLA-A26(+) /A26(+) cancer patients to four different CTL epitope peptides under personalized peptide vaccine regimens. In personalized peptide vaccine regimens, two to four peptides showing positive peptide-specific IgG responses in pre-vaccination plasma were selected from the four peptide candidates applicable for HLA-A26(+) /A26(+) cancer patients and administered s.c. Peptide-specific CTL and IgG responses along with cytokine levels were measured before and after vaccination. Cell surface markers in PBMCs and plasma cytokine levels were also measured. In this study, 21 advanced cancer patients, including seven lung, three breast, two pancreas, and two colon cancer patients, were enrolled. Their HLA-A26 genotypes were HLA-A26:01 (n = 24), HLA-A26:03 (n = 10), and HLA-A26:02 (n = 8). One, 14, and 6 patients received two, three, and four peptides, respectively. Grade 1 or 2 skin reactions at the injection sites were observed in the majority of patients, but no severe adverse events related to the vaccination were observed. Peptide-specific CTL responses were augmented in 39% or 22% of patients after one or two cycles of vaccination, respectively. Notably, peptide-specific IgG were augmented in 63% or 100% of patients after one or two cycles of vaccination, respectively. Personalized peptide vaccines with these four CTL epitope peptides could be feasible for HLA-A26(+) advanced cancer patients because of their safety and higher rates of immunological responses.

Phase I trial of a cancer vaccine consisting of 20 mixed peptides in patients with castration-resistant prostate cancer: dose-related immune boosting and suppression.

The heterogeneity expression of tumor-associated antigens (TAA) and variability of human T cell repertoire suggest that effective cancer vaccine requires induction of a wide breadth of cytotoxic T lymphocyte (CTL) specificities. This can be achieved with vaccines targeting multiple TAA. We evaluated the safety and immune dynamics of a cancer vaccine consisting of 20 mixed peptides (KRM-20) designed to induce CTLs against 12 different TAA in patients with castration-resistant prostate cancer (CRPC). Patients received each of three different randomly assigned doses of KRM-20 (6, 20, or 60 mg) once a week for 6 weeks. KRM-20 was applicable for patients with positive human leukocyte antigen (HLA) A2, A3, A11, A24, A26, A31 or A33 alleles, which cover the majority of the global population. To evaluate the minimum immunological effective dose (MIED), peptide-specific CTL and immunoglobulin G (IgG) responses, and immune suppressive subsets were evaluated during the vaccination. Total of 17 patients was enrolled. No serious adverse drug reactions were encountered. The MIED of KRM-20 in CTL or IgG response calculated by logistic regression model was set as 16 or 1.6 mg, respectively. The frequency of immune suppressive subsets was fewer in the 20 mg cohort than that in 6 or 60 mg cohort. Clinical responses determined by prostate-specific antigen levels were two partial responses (from the 20 mg cohort), five no changes and ten progressive diseases. Twenty milligrams of KRM-20 could be recommended for further studies because of the safety and ability to augment CTL activity.

[Pretreatment and Posttreatment Evaluations of Circulating Tumor Cells（CTC）in Patients with Metastatic Breast Cancer].

Recently, circulating tumor cells (CTCs) of cancer patients have been analyzed as a biomarker of disease progression and as a new prognostic tool.We conducted a pilot study to determine whether CTCs present before and after chemotherapy or peptide vaccination predicted outcome in patients with metastatic breast cancer (MBC). CTCs were isolated from the peripheral blood of MBC patients and evaluated using the CellSearch®System (Janssen Diagnostics). The expression of the HLA classⅠ molecule in the CTCs was analyzed simultaneously by using a flow cytometric system. A CTC-positive test result was defined as five or more CTCs/7.5 mL blood. Nine patients (median [range] age, 64.2 years [44-80 years]) were included. Four (57.1%) of 7 MBC patients with Luminal A subtype tested positive for CTC.Of 6 MBC patients, 5 (83.3%) had a decreased mean number of CTCs, from 35.8/7.5 to 0.4/7.5 mL, after 3 courses of systemic chemotherapy. Furthermore, CTC-HLA class Ⅰexpression level was decreased in 2 (66.7%) of 3 patients after cytotoxic chemotherapy but increased in 4 (80.0%) of 5 patients after 6 vaccinations with tumor-specific peptide vaccines. The number of CTCs and CTC-HLA class Ⅰ expression level in the patients with MBC changed after the treatment with systemic chemotherapy or peptide-vaccine therapy.

Feasibility study of personalized peptide vaccination for metastatic recurrent triple-negative breast cancer patients.

INTRODUCTION: Since treatment modalities for metastatic recurrent triple-negative breast cancer (mrTNBC) are limited, a novel treatment approach including immunotherapy is required. We have developed a novel regimen of personalized peptide vaccination (PPV), in which vaccine antigens are individually selected from a pool of different peptide candidates based on the pre-existing host immunity. Herein we conducted a phase II study of PPV for metastatic recurrent breast cancer patients to investigate the feasibility of PPV for mrTNBC. METHODS: Seventy-nine patients with metastatic recurrent breast cancer who had metastases and had failed standard chemotherapy and/or hormonal therapy were enrolled. They were subgrouped as the mrTNBC group (n = 18), the luminal/human epidermal growth factor receptor 2 (HER2)-negative group (n = 41) and the HER2-positive group (n = 18), while the remaining two patients had not been investigated. A maximum of four human leukocyte antigen (HLA)-matched peptides showing higher peptide-specific immunoglobulin G (IgG) responses in pre-vaccination plasma were selected from 31 pooled peptide candidates applicable for the four HLA-IA phenotypes (HLA-A2, -A24, or -A26 types, or HLA-A3 supertypes), and were subcutaneously administered weekly for 6 weeks and bi-weekly thereafter. Measurement of peptide-specific cytotoxic T lymphocyte (CTL) and IgG responses along with other laboratory analyses were conducted before and after vaccination. RESULTS: No severe adverse events associated with PPV were observed in any of the enrolled patients. Boosting of CTL and/or IgG responses was observed in most of the patients after vaccination, irrespective of the breast cancer subtypes. There were three complete response cases (1 mrTNBC and 2 luminal/HER2-negative types) and six partial response cases (1 mrTNBC and 5 luminal/HER2-negative types). The median progression-free survival time and median overall survival time of mrTNBC patients were 7.5 and 11.1 months, while those of luminal/HER2-negative patients were 12.2 and 26.5 months, and those of HER2-positive patients were 4.5 and 14.9 months, respectively. CONCLUSIONS: PPV could be feasible for mrTNBC patients because of the safety, immune responses, and possible clinical benefits. CLINICAL TRIAL REGISTRATION NUMBER: UMIN000001844 (Registration Date: April 5, 2009).

PD-1 expression on peripheral blood T-cell subsets correlates with prognosis in non-small cell lung cancer.

PD-1 expression in peripheral blood T-cells has been reported in several kinds of cancers, including lung cancer. However, the relationship between PD-1 expression in peripheral blood T-cells and prognosis after treatment with a cancer vaccine has not been reported. To elucidate this relationship, we analyzed PD-1 expression in the peripheral blood T-cells of patients with non-small cell lung cancer. The blood samples used in this study were obtained from patients enrolled in phase II clinical trials of a personalized peptide vaccine. Seventy-eight samples obtained before and after a single vaccination cycle (consisting of six or eight doses) were subjected to the analysis. PD-1 was expressed on lymphocytes in the majority of samples. The relative contents of PD1(+) CD4(+) T-cells against total lymphocytes before and after the vaccination cycle correlated with overall survival (OS) with a high degree of statistical significance (P < 0.0001 and P = 0.0014). A decrease in PD-1(+) CD8(+) T-cells after one cycle of vaccination also correlated with longer OS (P = 0.032). The IgG response to the non-vaccinated peptides suggested that the epitope spreading seemed to occur more frequently in high-PD-1(+) CD4(+) T-cell groups. Enrichment of CD45RA(-) CCR7(-) effector-memory phenotype cells in PD-1(+) T-cells in PBMCs was also shown. These results suggest that PD-1 expression on the peripheral blood T-cell subsets can become a new prognostic marker in non-small cell lung cancer patients treated with personalized peptide vaccination.

Human dermal fibroblast migration induced by fibronectin in autocrine and paracrine manners.

Although fibronectin (FN) is known as a chemoattractant for human dermal fibroblasts (HDFs), it is unclear whether HDF migration is stimulated by FN produced by HDFs (autocrine manner) or by keratinocytes (paracrine manner). In this study, we investigated HDF migration by Boyden chamber assay using conditioned media from HDFs and HaCaT cells (keratinocyte cell line). Immunoblotting and enzyme-linked immunosorbent assay revealed that FN existed in both conditioned media. Boyden chamber assay showed both conditioned media stimulated HDF migration, which was inhibited by anti-FN antibody. Antibodies to both integrin ß1and ß3 subunits inhibited HDF migration induced by HDF-conditioned medium almost completely and that by HaCaT cell-conditioned medium with 50-60%. These results suggested that HDF migration was stimulated by FN in both autocrine and paracrine manners. However, the mechanisms of HDF migration by FN, particularly the role of integrin ß1 and ß3 subunits, were slightly different between autocrine and paracrine manners.

Feasibility study of personalized peptide vaccination for recurrent ovarian cancer patients.

CONTEXT: To develop a personalized peptide vaccine (PPV) for recurrent ovarian cancer patients and evaluate its efficacy from the point of view of overall survival (OS), Phase II study of PPV was performed. PATIENTS AND METHODS: Forty-two patients, 17 with platinum-sensitive and 25 with platinum-resistant recurrent ovarian cancer, were enrolled in this study and received a maximum of four peptides based on HLA-A types and IgG responses to the peptides in pre-vaccination plasma. RESULTS: Expression of 13 of the 15 parental tumor-associated antigens encoding the vaccine peptides, with the two prostate-related antigens being the exceptions, was confirmed in the ovarian cancer tissues. No vaccine-related systemic severe adverse events were observed in any patients. Boosting of cytotoxic T lymphocytes or IgG responses specific for the peptides used for vaccination was observed in 18 or 13 of 42 cases at 6th vaccination, and 19 or 29 of 30 cases at 12th vaccination, respectively. The median survival time (MST) values of the platinum-sensitive- and platinum-resistant recurrent cases were 39.3 and 16.2 months, respectively. The MST of PPV monotherapy or PPV in combination with any chemotherapy during the 1st to 12th vaccination of platinum-sensitive cases was 39.3 or 32.2 months, and that of platinum-resistant cases was 16.8 or 16.1 months, respectively. Importantly, lymphocyte frequency and epitope spreading were significantly prognostic of OS. DISCUSSION AND CONCLUSION: Because of the safety and possible prolongation of OS, a clinical trial of PPV without chemotherapy during the 1st to 12th vaccination in recurrent ovarian cancer patients is merited.

Exploring the Potential of Humoral Immune Response to Commensal Bifidobacterium as a Biomarker for Human Health, including Both Malignant and Non-Malignant Diseases: A Perspective on Detection Strategies and Future Directions.

In this comprehensive review, we explore the pivotal role of commensal Bifidobacterium (c-BIF) as potent non-self-antigens through antigenic mimicry, along with exploring the potential of humoral immune responses for both malignant and non-malignant disease. c-BIF, a predominant component of the human gut microbiome encompassing around 90% of the human genome, has emerged as a pivotal player in human biology. Over recent decades, there has been extensive research elucidating the intricate connections between c-BIF and various facets of human health, with particular emphasis on their groundbreaking impact on anti-cancer effects and the management of non-malignant diseases. The multifaceted role of c-BIF is explored, ranging from enhancing anti-tumor immunity to improving the efficacy of anti-cancer and anti-infectious disease strategies, and serving as predictive biomarkers for various diseases. Recent studies highlight not only c-BIF's promotion of anti-tumor immunity but also their role in enhancing the efficacy of immune checkpoint inhibitors. The review emphasizes the promising avenue of manipulating the gut microbiota, particularly c-BIF, for modulating cancer immunotherapy with targeted effects on tumor cells while minimizing harm to normal tissue. In the context of infectious and inflammatory diseases, the crucial role of c-BIFs in the management of COVID-19 symptoms is examined, emphasizing their impact on the severity of and immune response to COVID-19. Furthermore, c-BIF exhibits preventive and therapeutic effects on Human Papillomaviruses (HPV) and shows promise in improving inflammatory bowel diseases. The potential application of c-BIF as a biomarker for immunotherapy is explored, with a specific emphasis on its predictive and prognostic value in cancer. Suggestions are made regarding the use of humoral immune responses to cytotoxic T lymphocyte (CTL) epitope peptides that share motifs with c-BIF, proposing them as potential markers for predicting overall survival in diverse cancer patients. In conclusion, c-BIF emerges as a crucial and multifaceted determinant of human health, across anti-tumor immunity to infectious and inflammatory disease management. The manipulation of c-BIF and gut microbiota presents a promising avenue for advancing therapeutic strategies, particularly in the realm of cancer immunotherapy. Additionally, this review highlights the significance of c-BIF as potent non-self-antigens via antigenic mimicry, emphasizing the importance of robust humoral immune responses against c-BIF for preventing various diseases, including inflammatory conditions. Elevated levels of circulating antibodies against c-BIF in healthy individuals may serve as potential indicators of lower risks for malignant and non-malignant diseases.