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Purpose: Transcription factors are commonly deregulated in cancer, and they have been widely considered as difficult to target due to their nonenzymatic mechanism of action. Altered expression levels of members of the ETS-transcription factors are often observed in many different tumors, including lymphomas. Here, we characterized two small molecules, YK-4-279 and its clinical derivative, TK-216, targeting ETS factors via blocking the protein-protein interaction with RNA helicases, for their antilymphoma activity., Experimental Design: The study included preclinical in vitro activity screening on a large panel of cell lines, both as single agent and in combination; validation experiments on in vivo models; and transcriptome and coimmunoprecipitation experiments., Results: YK-4-279 and TK-216 demonstrated an antitumor activity across several lymphoma cell lines, which we validated in vivo . We observed synergistic activity when YK-4-279 and TK-216 were combined with the BCL2 inhibitor venetoclax and with the immunomodulatory drug lenalidomide. YK-4-279 and TK-216 interfere with protein interactions of ETS family members SPIB, in activated B-cell-like type diffuse large B-cell lymphomas, and SPI1, in germinal center B-cell-type diffuse large B-cell lymphomas., Conclusions: The ETS inhibitor YK-4-279 and its clinical derivative TK-216 represent a new class of agents with in vitro and in vivo antitumor activity in lymphomas. Although their detailed mechanism of action needs to be fully defined, in DLBCL they might act by targeting subtype-specific essential transcription factors., (©2019 American Association for Cancer Research.)

Tetraspanins are commonly believed to act only as "molecular facilitators," with no direct role in signal transduction. We herein demonstrate that upon ligation, CD37, a tetraspanin molecule expressed on mature normal and transformed B cells, becomes tyrosine phosphorylated, associates with proximal signaling molecules, and initiates a cascade of events leading to apoptosis. Moreover, we have identified two tyrosine residues with opposing regulatory functions: one lies in the N-terminal domain of CD37 in a predicted "ITIM-like" motif and mediates SHP1-dependent death, whereas the second lies in a predicted "ITAM motif" in the C-terminal domain of CD37 and counteracts death signals by mediating phosphatidylinositol 3-kinase-dependent survival., (Copyright © 2012 Elsevier Inc. All rights reserved.)

G protein-coupled receptors mediate their biological responses through the generation of second messengers, such as cAMP. The down-regulation of their activity (desensitization) is carried out, in part, by the family of G protein-coupled receptor kinases, which phosphorylate activated receptors. The Gprk2 gene in Drosophila melanogaster is a putative member of this family. The GPRK2 protein is expressed most abundantly in the ovaries and in the mushroom bodies, the brain region that is implicated in learning and memory in insects. Many of the genes that are involved in learning in Drosophila are members of a cAMP-signaling pathway and are also expressed in the mushroom bodies. These observations suggest that the Gprk2 gene may be involved in a cAMP-mediated pathway. To investigate this possibility, we tested for a genetic interaction between Gprk2 and dunce (which encodes cAMP-specific phosphodiesterase). A mutant allele of Gprk2, called gprk2(6936), has decreased fertility as a result of reduced levels of egg laying and hatching, and developing egg chambers display defects in the formation of anterior structures. Similarly, many alleles of dunce are sterile, with an ovary phenotype that resembles gprk2(6936). Introduction of a single copy of a hypomorphic or null allele of dunce into the gprk2(6936) background suppressed all of these defects to a significant degree. Suppression was also observed when a single copy of gprk2(6936) was introduced into a dunce background. Like mutants of rutabaga (which encodes a calcium/calmodulin-dependent adenylate cyclase), gprk2(6936) has reduced levels of cAMP. Ovaries from gprk2(6936) females contain about one third of the normal amount of cAMP. In addition, in every mutant combination where fertility is increased, cAMP levels are closer to wild type levels. These results suggest that Gprk2 is functioning in a cAMP-signaling pathway and that the underlying basis of the interaction between Gprk2 and dunce is a normalization of cAMP levels., (Copyright 2001 Academic Press.)

Interleukin-12 is proposed to have anti-neoplastic activity on the basis of both its anti-angiogenic and immunologic effects. Gene gun therapy with interleukin-12 cDNA into the peritumoral area of immunocompetent 129/J mice with life-threatening primary vascular tumors reduced tumor volume 7.5-fold and almost tripled the duration of mouse survival, in contrast with luciferase-bombarded control mice. Epidermal expression of mouse interleukin-12 elevated tumoral and serum levels of interferon-gamma and tumor necrosis factor-alpha, increased the tumoral populations of T lymphocyte and natural killer cells, and induced tumor apoptosis. Gene transfer of interleukin-12 had little effect on tumor volumes and survival of tumor-bearing athymic nude mice, emphasizing the requirement for T cell directed cellular immunity. Peritumoral gene gun introduction of interleukin-12 may be a novel, cost-effective approach to limit the growth and associated mortality of life-threatening tumors.

Polysialoganglioside GT1b, a keratinocyte membrane glycosphingolipid, inhibits normal keratinocyte adhesion and migration on a fibronectin matrix. The specificity of the inhibition for cells plated on a fibronectin matrix and competition of GT1b inhibition with peptide RGDS suggest that GT1b abrogates the alpha 5 beta 1/fibronectin interaction. We examined the effects of GT1b on the adhesion and migration of keratinocyte-derived cell lines and correlated GT1b responsiveness and alpha 5 beta 1 integrin expression. GT1b (5 nM) significantly inhibited migration of normal human keratinocytes, immortalized keratinocytes, and squamous cell carcinoma SCC12F2 cells on fibronectin, but not on collagen I. Concentrations as high as 5 microM had no effect on SCC13 or HaCaT cells. Likewise, GT1b inhibited fibronectin-dependent cell adhesion of normal human keratinocytes, immortalized keratinocytes, and SCC12F2 cells, but had no effect on SCC13 or HaCaT cells. Flow cytometric and Western immunoblot analysis of integrin expression showed significantly decreased alpha 5 and beta 1 integrin expression in SCC13 and HaCaT cells compared to normal keratinocytes, immortalized keratinocytes, and SCC12F2 cells. Incubation with TGF-beta 1 increased alpha 5 beta 1 integrin expression and induced responsiveness to GT1b in HaCaT cells. These data imply that GT1b "response" requires sufficient expression of alpha 5 beta 1 and further suggest that the mechanism of the inhibitory effect of GT1b involves GT1b/alpha 5 beta 1 interaction.

Angiostatin inhibits angiogenesis and metastatic tumor growth; however, its usefulness in treating primary nonmetastasizing tumors is less well understood. We now report the effectiveness of human angiostatin administration in a mouse hemangioendothelioma model. Human angiostatin was administered to mice with s.c. hemangioendothelioma and associated disseminated intravascular coagulopathy (Kasabach-Merritt syndrome). Angiostatin significantly reduced tumor volume in comparison to nontreated controls, increased survival, and prevented the profound thrombocytopenia and anemia of Kasabach-Merritt syndrome. Apoptosis of tumor cells was induced by angiostatin, but tumor cell proliferation was not inhibited. These data suggest angiostatin as a novel treatment for nonmetastasizing vascular tumors and for Kasabach-Merritt syndrome.

A novel photoreactive deoxycytidine analog, 4-[N-(p-azidobenzoyl)-2-aminoethyl]-dCTP (ABdCTP), has been synthesized and incorporated at specific sites within the SUP4 tRNA(Tyr) gene. Immobilized single-stranded DNA was annealed to specific oligonucleotides and AB-dCMP incorporated into DNA by primer extension. DNA photoaffinity labeling with AB-dCMP was used to survey protein-DNA contacts in initiation and elongation complexes of RNA polymerase III (Pol III), and compared to DNA photoaffinity labeling using the previously described photoreactive deoxyuridine analog, 5-[N-(pazidobenzoyl)-3-aminoallyl]-dUMP (AB-dUMP) [Bartholomew et al. (1993) Mol. Cell.Biol. 13,942-952]. In contrast to previous studies, we have used a crude protein fraction rather than highly purified preparations of Pol III and transcription factors TFIIIC and TFIIIB to examine if some component of the transcription complex is lost upon purification. Eleven nucleotide positions from bp-17 to bp +17 (+1 being the start site of transcription) on the nontranscribed strand were modified and shown to have little or no effect on transcription complex formation, initiation, or elongation as determined by multiple-round transcription assays. Efficient photoaffinity labeling by DNA containing AB-dCMP gave results comparable to that with AB-dUMP at proximal nucleotide positions and provided new evidence for the placement of the 160 and 31 kDa subunits of Pol III near the 5' end of the transcriptional bubble in an elongation complex. A novel 40 kDa protein was cross-linked at bps -17, -9, and -8 in a TFIIIC-dependent manner that had not been previously detected.