SEC31A-ALK Fusion Gene in Lung Adenocarcinoma.

Anaplastic lymphoma kinase (ALK) fusion is a common mechanism underlying pathogenesis of non-small cell lung carcinoma (NSCLC) where these rearrangements represent important diagnostic and therapeutic targets. In this study, we found a new ALK fusion gene, SEC31A-ALK, in lung carcinoma from a 53-year-old Korean man. The conjoined region in the fusion transcript was generated by the fusion of SEC31A exon 21 and ALK exon 20 by genomic rearrangement, which contributed to generation of an intact, in-frame open reading frame. SEC31A-ALK encodes a predicted fusion protein of 1,438 amino acids comprising the WD40 domain of SEC31A at the N-terminus and ALK kinase domain at the C-terminus. Fluorescence in situ hybridization studies suggested that SEC31A-ALK was generated by an unbalanced genomic rearrangement associated with loss of the 3'-end of SEC31A. This is the first report of SEC31A-ALK fusion transcript in clinical NSCLC, which could be a novel diagnostic and therapeutic target for patients with NSCLC.

A whole recombinant yeast-based therapeutic vaccine elicits HBV X, S and Core specific T cells in mice and activates human T cells recognizing epitopes linked to viral clearance.

Chronic hepatitis B infection (CHB) is characterized by sub-optimal T cell responses to viral antigens. A therapeutic vaccine capable of restoring these immune responses could potentially improve HBsAg seroconversion rates in the setting of direct acting antiviral therapies. A yeast-based immunotherapy (Tarmogen) platform was used to make a vaccine candidate expressing hepatitis B virus (HBV) X, surface (S), and Core antigens (X-S-Core). Murine and human immunogenicity models were used to evaluate the type and magnitude of HBV-Ag specific T cell responses elicited by the vaccine. C57BL/6J, BALB/c, and HLA-A*0201 transgenic mice immunized with yeast expressing X-S-Core showed T cell responses to X, S and Core when evaluated by lymphocyte proliferation assay, ELISpot, intracellular cytokine staining (ICS), or tumor challenge assays. Both CD4+ and CD8+ T cell responses were observed. Human T cells transduced with HBc18-27 and HBs183-91 specific T cell receptors (TCRs) produced interferon gamma (IFNγ following incubation with X-S-Core-pulsed dendritic cells (DCs). Furthermore, stimulation of peripheral blood mononuclear cells (PBMCs) isolated from CHB patients or from HBV vaccine recipients with autologous DCs pulsed with X-S-Core or a related product (S-Core) resulted in pronounced expansions of HBV Ag-specific T cells possessing a cytolytic phenotype. These data indicate that X-S-Core-expressing yeast elicit functional adaptive immune responses and supports the ongoing evaluation of this therapeutic vaccine in patients with CHB to enhance the induction of HBV-specific T cell responses.

A novel fusion of TPR and ALK in lung adenocarcinoma.

INTRODUCTION: Anaplastic lymphoma kinase (ALK) fusion is the most common mechanism for overexpression and activation in non-small-cell lung carcinoma. Several fusion partners of ALK have been reported, including echinoderm microtubule-associated protein-like 4, TRK-fused gene, kinesin family member 5B, kinesin light chain 1 (KLC1), protein tyrosine phosphatase and nonreceptor type 3, and huntingtin interacting protein 1 (HIP1). METHODS AND RESULTS: A 60-year-old Korean man had a lung mass which was a poorly differentiated adenocarcinoma with ALK overexpression. By using an Anchored Multiplex polymerase chain reaction assay and sequencing, we found that tumor had a novel translocated promoter region (TPR)-ALK fusion. The fusion transcript was generated from an intact, in-frame fusion of TPR exon 15 and ALK exon 20 (t(1;2)(q31.1;p23)). The TPR-ALK fusion encodes a predicted protein of 1192 amino acids with a coiled-coil domain encoded by the 5'-2 of the TPR and juxtamembrane and kinase domains encoded by the 3'-end of the ALK. CONCLUSIONS: The novel fusion gene and its protein TRP-ALK, harboring coiled-coil and kinase domains, could possess transforming potential and responses to treatment with ALK inhibitors. This case is the first report of TPR-ALK fusion transcript in clinical tumor samples and could provide a novel diagnostic and therapeutic candidate target for patients with cancer, including non-small-cell lung carcinoma.

Glial remodeling and neural plasticity in human retinal detachment with proliferative vitreoretinopathy.

PURPOSE: To investigate glial remodeling and neuronal plasticity in adult human retinal detachment complicated by proliferative vitreoretinopathy (PVR) and to grade pathologic changes with a severity scoring system. METHODS: Sixteen full-thickness retinectomy specimens obtained at retinal relaxing surgery for PVR were fixed in 4% paraformaldehyde immediately after excision and compared to similarly processed normal donor retinas. Agarose-embedded sections (100-microm-thick) were double labeled for immunohistochemistry by confocal microscopy, with antibodies against rod opsin and GFAP; vimentin and M/L-cone opsin; calbindin D and S-cone opsin; and cytochrome oxidase and synaptophysin. These staining patterns formed the basis of a retinal pathology scoring system, and immunohistochemistry was also used to detect CD68, neurofilaments, protein kinase C, growth-associated protein-43, and a pan-cone-specific enzymatic marker. Morphology was also assessed by light microscopy of resin-embedded semithin sections. RESULTS: Prolonged detachment was characterized by photoreceptor degeneration and intracellular redistribution of opsin proteins to the plasma membrane in the outer nuclear layer (ONL). Remodeling of rod synaptic terminals was characterized by terminal retraction and also by axon extension to the inner retina in most specimens. Rod bipolar cell dendrites extended into the ONL, as did fine, horizontal cell processes. Large ganglion cells showed upregulated neurofilament and GAP-43 expression, with neurites sprouting from somata and axon collaterals. Anti-cytochrome oxidase labeling of surviving inner segments was reduced but detectable in all specimens, as was anti-calbindin D labeling of horizontal and amacrine cells. All specimens demonstrated a marked upregulation of Muller cell and astrocyte expression of GFAP and vimentin. More severe degenerative changes correlated with trauma and prolonged detachment duration when scored according to this system. CONCLUSIONS: The neural and glial components of detached neurosensory retina complicated by PVR exhibit pathology that changes characteristically with increasing detachment severity. Even in advanced degeneration, most of the structural motifs necessary for functional recovery are retained. Evidence of remodeling in the first-, second-, and third-order neurons of detached adult human retina may represent an attempt to re-establish synaptic connectivity.

Characterization of transcriptional regulation during negative selection in vivo.

Negative selection is the process whereby immature thymocytes expressing TCRs with high affinity for self-peptide:MHC complexes are induced to undergo apoptosis. The transcriptional events that occur as a result of TCR signaling during negative selection are not well-characterized. Using oligonucleotide arrays, we have identified 33 genes that exhibit changes in RNA levels in CD4(+)CD8(+) thymocytes during negative selection in vivo. Of 18 genes that have been further characterized, 13 are regulated in response to stimulation with Ag or anti-CD3 and anti-CD28 Abs ex vivo, indicating that these genes are regulated independently of activation of the peripheral immune system. These data also support the idea that anti-CD3/CD28-mediated thymocyte apoptosis is a valid model for negative selection in vivo. A detailed examination of the regulation of many of the identified genes in response to treatment with dexamethasone or gamma-radiation or in response to anti-CD3/anti-CD28 stimulation in the presence of pharmacological inhibitors of mitogen-activated protein kinase kinase kinase 1, p38 mitogen-activated protein kinase, phosphatidylinositol 3-kinase, calcineurin, and cyclin-dependent kinase 2 has facilitated the elucidation of a map of the transcriptional events that occur downstream of the TCR. These studies support a model whereby similar signal transduction pathways are activated by stimuli that induce positive and negative selection and are consistent with the idea that the balance between opposing proapoptotic and antiapoptotic pathways determines cell fate. The data presented in this study also suggest that calcineurin functions to amplify TCR signals by promoting sustained increases in the levels of specific transcripts.