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 HLAA26 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.This study was supported in part by the Japan Agency for Medical Research and development, AMED, a research program of the Regional Innovation Cluster Program of the Ministry of Education, Culture, Sports, Science and Technology of Japan, and a grant from the Sendai Kousei Hospital

Aldehyde dehydrogenase 1 identifies cells with cancer stem cell-like properties in a human renal cell carcinoma cell line.

Cancer stem cells (CSC) or cancer stem cell-like cells (CSC-LCs) have been identified in many malignant tumors. CSCs are proposed to be related with drug resistance, tumor recurrence, and metastasis and are considered as a new target for cancer treatment; however, there are only a few reports on CSCs or CSC-LCs in renal cell carcinoma (RCC). Different approaches have been reported for CSC identification, but there are no universal markers for CSC. We used two different approaches, the traditional side population (SP) approach, and the enzymatic (aldehyde dehydrogenase 1 (ALDH1)) approach to identify CSC-LC population in two RCC cell lines, ACHN and KRC/Y. We found that ACHN and KRC/Y contain 1.4% and 1.7% SP cells, respectively. ACHN SP cells showed a higher sphere forming ability, drug resistance, and a slightly higher tumorigenic ability in NOD/SCID mice than Non-SP (NSP) cells, suggesting that cells with CSC-LC properties are included in ACHN SP cells. KRC/Y SP and NSP cells showed no difference in such properties. ALDH1 activity analysis revealed that ACHN SP cells expressed a higher level of activity than NSP cells (SP vs. NSP: 32.7% vs 14.6%). Analysis of ALDH1-positive ACHN cells revealed that they have a higher sphere forming ability, self-renewal ability, tumorigenicity and express higher mRNA levels of CSC-LC property-related genes (e.g., ABC transporter genes, self-replication genes, anti-apoptosis genes, and so forth) than ALDH1-negative cells. Drug treatment or exposure to hypoxic condition induced a 2- to 3-fold increase in number of ALDH1-positive cells. In conclusion, the results suggest that the ALDH1-positive cell population rather than SP cells show CSC-LC properties in a RCC cell line, ACHN

Immunological evaluation of peptide vaccination for cancer patients with the HLA

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

Tumorigenicity of aldehyde dehydrogenase 1 (ALDH1)-positive and ALDH1-negative cells in ACHN.

<p>Tumorigenicity of aldehyde dehydrogenase 1 (ALDH1)-positive and ALDH1-negative cells in ACHN.</p

Quantification of mRNA expression of CSC-LC property-related genes in ALDH1-positive and ALDH1-negative ACHN cells by real-time PCR.

<p>ALDH1-positive cells showed significantly higher mRNA expression of ALDH1A1, transporter-related genes (ABCB1 and ABCG2), self-replication genes (BMI-1 and c-MYC), anti-apoptosis genes (BCL2 and CFLAR), hypoxia-related genes (HIF1α and VEGFA) and EMT-related genes (Twist) than ALDH1-negative cells in ACHN. However, there was no significant difference in mRNA expression of Snail between ALDH1-positive and ALDH1-negative cells. The experiments were repeated at least four times, and almost identical results were obtained.</p

A self-renewal capacity between ALDH1-positive and ALDH1-negative ACHN cells.

<p>The sphere forming ability of ALDH1-positive cells in ACHN was maintained during the second and third passages (* <i>P</i><0.0001).</p

SP cells analysis and expression of CSC markers in SP and NSP cells.

<p>(A) ACHN and KRC/Y were labeled with Hoechst 33342, and then analyzed by FCM. The SP cell rates in ACHN and KRC/Y were 1.4% (A–a) and 1.7% (A–c), respectively, which decreased significantly in the presence of reserpine (A–b, A–d). The experiment was repeated at least three times for each cell line and almost identical results were obtained. A representative figure of our experiments is shown. (B) There was no apparent difference in CD90 and EpCAM expression between SP and NSP cells in ACHN. In the KRC/Y cell line, although there was no difference in EpCAM expression, SP cells expressed a higher CD105-positive cell rate than NSP cells (SP vs NSP : 24.6% vs 4.6%). The experiments were repeated twice, and almost identical results were obtained. A representative figure of our experiments is shown.</p