Exploratory study on the efficacy of bortezomib combining mitoxantrone or CD22-CAR T therapy targeting CD19-negative relapse after CD19-CAR T cell therapy with a simpler cell-line-based model.

Target-negative relapse after CD19 chimeric antigen receptor engineered (CAR) T cell therapy for patients with B lineage acute lymphoblastic leukemia (B-ALL) presents limited treatment options with dismal outcomes. Although CD22-CAR T cells mediate similarly potent antineoplastic effects in patients with CD19dim or even CD19-negative relapse following CD19-directed immunotherapy, a high rate of relapse associated with diminished CD22 cell surface expression has also been observed. Therefore, it is unclear whether any other therapeutic options are available. Mitoxantrone has shown significant antineoplastic activity in patients with relapsed or refractory leukemia over the past decades, and in some cases, the addition of bortezomib to conventional chemotherapeutic agents has demonstrated improved response rates. However, whether this mitoxantrone and bortezomib combination therapy is effective for those patients who have relapsed B-ALL after receiving CD19-CAR T cell therapy remains to be elucidated. In this study, we established a cellular model system using a CD19-positive B-ALL cell line Nalm-6 to investigate the treatment options for CD19-negative relapsed B-ALL after CD19-CAR T cell therapy. In addition to CD22-CAR T therapy, we observed that the combination of bortezomib and mitoxantrone exhibited effective anti-leukemia activity in the CD19-negative Nalm-6 cell line by downregulating p-AKT and p-mTOR. These results suggest that this combination therapy is a possible option for target-negative refractory leukemia cells after CAR-T cell treatment.

Prognostic significance of the tumor suppressor protein p53 gene in childhood acute lymphoblastic leukemia.

The tumor suppressor protein p53 (TP53) gene is associated with various types of cancer; however, little is known about TP53 expression in patients with childhood acute lymphoblastic leukemia (ALL). The aim of the present study was to investigate the prognostic value of TP53 expression in childhood ALL. To achieve this, TP53 mRNA levels of 146 children with ALL and 23 child donors with idiopathic thrombocytopenic purpura were determined by reverse transcription-quantitative PCR. Relapse-free survival (RFS) and overall survival (OS) were analyzed using the Kaplan-Meier method. The results demonstrated that TP53 mRNA level in patients with ALL was higher compared with that in the ITP donors (P=0.019). Patients with highly-expressed TP53 exhibited lower percentages of peripheral blood blast, higher platelet counts and inferior complete remission rates compared with patients with low expression of TP53. Survival analyses revealed that high TP53 expression was associated with poor OS and RFS in childhood ALL (P=0.018 and P=0.028, respectively) and was an independent prognostic factor in multivariate analysis for poor RFS (P<0.001) and OS (P<0.001). In conclusion, high TP53 expression is associated with poor outcomes and may be used as a molecular prognostic marker to be incorporated into an improved risk classification system for childhood ALL.

Low expression of TET2 gene in pediatric acute lymphoblastic leukemia is associated with poor clinical outcome.

INTRODUCTION: TET2, a member of the Ten-Eleven translocation gene family, catalyzes the conversion of 5-methylcytosine to 5-hydroxymethylcytosine in DNA. Low expression of TET2 has been reported as a prognostic factor for several types of malignancies in adult patients. However, there have been few data on the effect of TET2 mRNA level on the prognosis of children with ALL so far. METHODS: In this study, TET2 expression of samples cryopreserved in the liquid nitrogen from January 1, 2007 through December 31, 2011 was retrospectively analyzed in 136 newly diagnosed ALL patients by real-time polymerase chain reaction (PCR) assay. The patients' samples were divided into two groups by the median value of patients group and divided into TET2 low and TET2 high groups. RESULTS: A total of 136 childhood ALL patients demonstrated lower TET2 expression than control group (P = .038). TET2 mRNA expression levels were correlated with the disease status. In addition, patients with low TET2 expression had lower platelet counts and lower CR rates. Survival analysis showed that low TET2 expression in children with ALL was associated with lower 5-year overall survival (OS) (63% vs 88%, P = .011) and event-free survival (EFS) (60% vs 85%, P = .003). Multivariate analysis revealed that low TET2 expression was an independent poor prognostic factor of OS and EFS. CONCLUSION: Low expression of TET2 in children with ALL is associated with poor prognosis and can be used as a molecular prognostic marker for risk group stratification.

Potential function of CTLA-4 in the tumourigenic capacity of melanoma stem cells.

Extensive clinical evidence supports that cytotoxic T lymphocyte antigen-4 (CTLA-4) is expressed in a variety of human malignant tumour cells in addition to T cells. In certain types of cancer, the overexpression of CTLA-4 is associated with poor patient prognosis. However, few studies have demonstrated the effects of tumour-intrinsic CTLA-4 in cancer stem cells, including melanoma stem cells (MSCs). In the present study, it was demonstrated that melanoma cell-intrinsic CTLA-4 induced tumour cell proliferation in vitro and suppressed tumour cell apoptosis. Furthermore, CTLA-4 was expressed in aldehyde dehydrogenase (ALDH)+ MSCs. CTLA-4 inhibited MSCs proliferation in vitro by blocking antibodies and significantly downregulated ALDH1A1, ALDH1A3 and ALDH2 mRNA expression (P<0.01). Functionally, blocking CTLA-4 in melanoma cell lines suppressed the properties of stem-like cells, including ALDH activity and significantly suppressed the ability of these cells to form spheres in vitro (P<0.05). In addition, the blocking of CTLA-4 in melanoma cells suppressed the properties of stem-like cells in vivo, including the capacity for tumourigenesis. The presence of residual ALDH+ MSCs within the tumour was observed, and the blocking CTLA-4 significantly decreased the number of residual ALDH+ MSCs in vivo (P<0.01). Altogether, these results indicate the identification of a novel mechanism underlying melanoma progression in the present study and that CTLA-4-targeted therapy may benefit candidate CTLA-4-targeted therapy by improving the long-term outcome for patients with advanced stages of melanoma.

Cancer Stem Cell Vaccination With PD-L1 and CTLA-4 Blockades Enhances the Eradication of Melanoma Stem Cells in a Mouse Tumor Model.

Immune checkpoint inhibitors and monoclonal antibodies reinvigorate cancer immunotherapy. However, these immunotherapies only benefit a subset of patients. We previously reported that ALDH tumor cells were highly enriched for cancer stem cells (CSCs), and ALDH CSC lysate-pulsed dendritic cell (CSC-DC) vaccine was shown to induce CSC-specific cytotoxic T lymphocytes. In this study, we investigated the CSC targeting effect of the CSC-DC vaccine combined with a dual blockade of programmed death-ligand 1 and cytotoxic T-lymphocyte-associated protein (CTLA-4) in B16-F10 murine melanoma tumor model. Our data showed that animals treated with the dual blockade of programmed death-ligand 1 and CTLA-4 and CSC-DC vaccine conferred significantly more tumor regression than the CSC-DC vaccine alone. Importantly, the triple combination treatment dramatically eliminated ALDH CSCs in vivo. We observed that CSC-DC vaccine in combination with anti-PD-L1 and anti-CTLA-4 administration resulted in ∼1.7-fold fewer PD-1CD8 T cells and ∼2.5-fold fewer CTLA-4CD8 T cells than the populations observed following the CSC-DC vaccination alone. Moreover, significant antitumor effects and dramatically eliminated ALDH CSCs following the triple combination treatment were accompanied by significantly enhanced T-cell expansion, suppressed transforming growth factor ß secretion, enhanced IFN-γ secretion, and significantly enhanced host specific CD8 T-cell response against CSCs. Collectively, these data showed that administration of a-PD-L1 and a-CTLA-4 combined with CSC-DC vaccine may represent an effective immunotherapeutic strategy for cancer patients in clinical.