We prospectively analyzed p15 and p16 promoter methylation patterns using methylation-specific polymerase chain reaction (PCR) in patients with adult and childhood acute leukemias and studied the association of methylation patterns with chromosomal abnormalities and prognostic variables. In nearly all French-American-British leukemia subtypes, we found p15methylation in bone marrow or peripheral blood cells from 58% (46/79) of patients with acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), or acute biphenotypic leukemia (ABL). An identical alteration was detected in blood plasma from 11 of 12 of these patients (92%). We also demonstrated for the first time concomitant p16and p15 methylation in 22% (8/37) of adults with AML or ALL, exclusively in those with M2, M4, or L2 subtypes. According to cytogenetic data from 35 patients with ALL, AML, or ABL, 82% (14/17) of those with unmethylated p15 alleles had normal karyotypes or hyperdiploidies associated with a favorable prognosis. Conversely, 44% (8/18) of patients with p15 methylation had chromosomal translocations, inversions, or deletions, suggesting an interplay of these abnormalities with p15 methylation. As a prognostic marker for disease monitoring, p15 methylation appears to be more widely applicable than BCR-ABL, AF4-MLL, andAML1-ETO transcripts, which were detectable in only 8% (4/48) of patients by reverse transcriptase-PCR. Thirty-nine of 43 blood samples (91%) sequentially collected from 12 patients with AML, ALL, or ABL showed p15 methylation status in excellent concordance with morphologic disease stage. Early detection of p15methylation at apparent remission or its acquisition during follow-up may prove valuable for predicting relapse. Overall survival of patients with p15 methylation was notably shortened among 38 adults with AML and 12 adults with ALL. Aberrant p15 methylation may have important prognostic implications for clinical monitoring and risk assessment.

Severe acute respiratory syndrome (SARS) is a public health concern worldwide. By studying the human leukocyte antigen (HLA) types A, B, DR, and DQ al

Nasopharyngeal carcinoma (NPC) is an Epstein–Barr virus-associated disease with high prevalence in Southern Chinese. Using multiparametric flow cytometry, we identified significant expansions of circulating naïve and memory CD4+CD25^high T cells in 56 NPC patients compared with healthy age- and sex-matched controls. These were regulatory T cells (Treg), as they overexpressed Foxp3 and GITR, and demonstrated enhanced suppressive activities against autologous CD4+CD25− T-cell proliferation in functional studies on five patients. Abundant intraepithelial infiltrations of Treg with very high levels of Foxp3 expression and absence of CCR7 expression were also detected in five primary tumours. Our current study is the first to demonstrate an expansion of functional Treg in the circulation of NPC patients and the presence of infiltrating Treg in the tumour microenvironment. As Treg may play an important role in suppressing antitumour immunity, our findings provide critical insights for clinical management of NPC.

Platelet factor 4 (PF4) is an angiostatic chemokine that suppresses tumor growth and metastasis. We previously revealed frequent transcriptional silencing of PF4 in multiple myeloma, but the functional roles of this chemokine are still unknown. We studied the apoptotic effects of PF4 on myeloma cell lines and primary myeloma in vitro, and investigated the involved signaling pathway. The in vivo effects were also studied using a mouse model. PF4 not only suppressed myeloma-associated angiogenesis, but also inhibited growth and induced apoptosis in myeloma cells. We found that PF4 negatively regulated STAT3 and concordantly inhibited constitutive and interleukin-6-induced phosphorylation of STAT3, and down-regulated the expression of STAT3 target genes (Mcl-1, survivin and VEGF). Overexpression of constitutively activated STAT3 could rescue PF4-induced apoptotic effects. Furthermore, we found that PF4 induced the expression of SOCS3, a STAT3 inhibitor, and gene silencing of SOCS3 abolished its ability to inhibit STAT3 activation, suggesting a critical role of SOCS3 in PF4-induced STAT3 inhibition. Knockdown of LRP1, a putative PF4 receptor, could also abolish PF4-induced apoptosis and STAT3 inhibition. Finally, the tumor growth inhibitory effect of PF4 was confirmed by in vivo mouse models. Immunostaining of rabbit bone xenografts from PF4-treated mice showed induction of apoptosis of myeloma cells and inhibition of angiogenesis, which was associated with suppression of STAT3 activity. Together, our preclinical data indicate that PF4 may be a potential new targeting agent for the treatment of myeloma.

Detection of chromosomal translocation is a key component in diagnosis and management of acute myeloid leukemia (AML). Targeted RNA next-generation sequencing (NGS) is emerging as a powerful and clinically practical tool, but it depends on expression of RNA transcript from the underlying DNA translocation. Here, we show the clinical utility of nanopore long-read sequencing in rapidly detecting DNA translocation with exact breakpoints. In a newly diagnosed patient with AML, conventional karyotyping showed translocation t(10;12)(q22;p13) but RNA NGS detected NUP98-NSD1 fusion transcripts from a known cryptic translocation t(5;11)(q35;p15). Rapid PCR-free nanopore whole-genome sequencing yielded a 26,194 bp sequencing read and revealed the t(10;12) breakpoint to be DUSP13 and GRIN2B in head-to-head configuration. This translocation was then classified as a passenger structural variant. The sequencing also yielded a 20,709 bp sequencing read and revealed the t(5;11) breakpoint of the driver NUP98-NSD1 fusion. The identified DNA breakpoints also served as markers for molecular monitoring, in addition to fusion transcript expression by digital PCR and sequence mutations by NGS. We illustrate that third-generation nanopore sequencing is a simple and low-cost workflow for DNA translocation detection.