ALBERT

Description: Symmetry, Vol. 10, Pages 266: Cryptanalysis of an Image Encryption Algorithm Based on Combined Chaos for a BAN System, and Improved Scheme Using SHA-512 and Hyperchaos Symmetry doi: 10.3390/sym10070266 Authors: Musheer Ahmad Eesa Al Solami Xing-Yuan Wang M. N. Doja M. M. Sufyan Beg Amer Awad Alzaidi The issues of identity authentication and privacy protection of individuals in body area network (BAN) systems have raised much concern in past few years. To address the challenges of privacy protection in wireless BAN, an image encryption algorithm has been proposed recently by Wang et al. The encryption algorithm utilized two 1D chaotic maps to generate sub-chaotic matrices which are combined to perform encryption. The algorithm has good statistical encryption performance. However, a cautious inquiry finds that it has some underlying security defects. This paper evaluates the security of the Wang et al. encryption algorithm to show that it is totally breakable under proposed cryptanalysis and hence infeasible for privacy protection in BAN. It has been shown that the plain-image data can be recovered without any prior knowledge of secret key and plain-text. Furthermore, this paper also suggests an improved encryption scheme using secure hash algorithm SHA-512 for one-time keys and a 4D hyperchaotic system to subdue the security insufficiencies of the algorithm under study. The simulation results and analysis demonstrate that the improved image encryption scheme has excellent encryption quality, plain-image sensitivity, and resistance to possible cryptanalytic attacks.

Description: Results to date argue compellingly that disruption of FA/BRCA gene expression plays a pivotal role in human somatic carcinogenesis. Melphalan, a DNA cross-linker, is one of the most widely used and effective drugs in the treatment of multiple myeloma (MM). Although most patients respond to standard and high dose melphalan, eventually patients acquire resistance and develop progressive disease. In 1991, our laboratory reported that acquired resistance in a human myeloma cell line was associated with reduced DNA crosslinks, elevated glutathione levels, and increased radiation survival (Cancer Res. 5:993; 1991). Most recently, we reported that the melphalan-resistant myeloma cell lines, 8226/LR5 and U266/LR6, showed a significant increase in several FA/BRCA genes compared to drug-sensitive cells, and that enhanced interstrand crosslink (ICL) repair via this signaling pathway contributes to acquired drug resistance in melphalan resistant cell lines (Blood 10:698; 2005). Here, we report that IKKa is constitutively phosphorylated in unstimulated 8226/LR5 cells, but not in melphalan-sensitive control cells. The specific phosphorylation of IKKa leads to an increase in basal NF-kB DNA binding activity, and 8226/LR5 cells are found to be markedly sensitive to BMS-345541 (a highly selective inhibitor of IkB) relative to control cells. Importantly, a cytotoxic dose of BMS-345541 induces a dramatic decrease in FA/BRCA gene expression, and a concomitant inhibition of NF-kB DNA binding activity in both 8226/S and 8226/LR5 cells. Furthermore, we show that 8226/LR5 cells experience the highest degree of direct binding between FANCD2 promoter and NF-kB/Rel family members, which, in turn, leads to an increase in basal FANCD2-specific NF-kB activity. Small-interfering RNA (siRNA)-mediated depletion of RelB and p50, but not other NF-kB subunits, in 8226 cells results in impaired NF-kB binding activity, and visible decrease in FANCD2 protein expression. Studies designed to dissect the role of NF-kB in acquired melphalan resistance are in progress, and the results will be presented. Our findings suggest that NF-kB functions as a regulator of FA/BRCA expression, and that this pathway represents a new target for preventing acquired drug resistance in myeloma patients.

Description: Abstract 1898 Protein kinase C theta (PKCθ), a T cell signaling molecule, has been implicated as a therapeutic target for several autoimmune diseases as well as graft-versus-host disease (GVHD). PKCθ plays a vital role in stabilization of the immunologic synapse between T effector cells and antigen presenting cells (APC), but has been shown to be excluded from the immunologic synapse in T regulatory cells (T reg). PKCθ inhibition reduces the alloreactivity of donor T cells responsible for induction of GVHD while preserving graft-versus-leukemia (GVL) responses. The roles of PKCθ and the potential compensatory alpha isoform (PKCα) are not clearly defined with regard to alloresponses or T cell mediated responses in GVHD. In this context, we measured PKCθ and PKCα/θ gene deficient T cell activation upon TCR-ligation in vitro using [3H]-TdR incorporation and CSFE labeling assays. T cells from PKCθ and PKCα/θ gene deficient donor mice were utilized in vivo in a pre-clinical allogenic murine model of myeloablative bone marrow transplantation (BMT). The development of GVHD was monitored in recipient mice with or without injection of A20-luciferase cells to observe the progression of GVL in vivo. Combined blockade of PKCα and PKCθ causes a significant decrease in T cell proliferation compared to blocking PKCθ alone in vitro. Deficiency in PKCα and PKCθ had no effect on immune reconstitution following irradiation and BMT in vivo. Even with a high transplant load of 5×106 CD4+ and CD8+ T cells, PKCα/θ deficient (PKCα/θ−/−) T cells failed to induce acute GVHD. Our data suggest that the ability of double deficient T cells to induce GVHD was further reduced than PKCθ-deficient T cells. Additionally, a greater number and percentage of B220+ B cells and FoxP3+ T regs were isolated from the spleens of PKCα/θ−/− T cell recipient mice 120 after BMT than were isolated from wild type (WT) or PKCθ−/− T cell recipients. Fewer CD4+ or CD8+ T effector cells were isolated from the spleens of PKCα/θ−/− T cell recipient mice 120 after BMT than were isolated from wild type or PKCθ−/− T cell recipients. Importantly, the activity of B cells isolated from PKCα/θ−/− T cell recipient mice 120 after BMT was greater on a per cell basis, while the activity of T effector cells isolated from these mice was greatly reduced compared to WT or PKCθ−/− T cell recipients. While not absent, GVL was reduced in PKCα/θ−/− T cell recipient mice when compared to WT or PKCθ−/− T cell recipients. This work demonstrates the requirement of PKCα and θ for optimal activation and function of T cells in vitro. These experiments highlight a potential compensatory role for PKCα in the absence of PKCθ in T cell signaling and activation. Combined deficiency of PKCα and θ prevents induction of acute GVHD while improving the maintenance of splenic cellularity in PKCα/θ T cell recipient mice. Additionally, PKCα/θ dual deficient T cell transplant shifts the splenic balance toward a greater number and percentage of T reg and B cells and away from T effector cells following BMT. The reduced and sub-optimally active T effector cells isolated from PKCα/θ−/− T cell recipient mice in combination with reduced GVL stresses the importance of PKCα and θ molecules and their roles in T cell activity in the context of both GVHD and GVL. Dual deficiency of PKCα/θ is associated with a decline of T effector function that is optimal for the amelioration of GVHD, but is perhaps too reduced to substantially maintain effective GVL. Modulation of PKCα and θ signaling presents a valid avenue of investigation as a therapeutic option for GVHD. Disclosures: No relevant conflicts of interest to declare.

Description: Abstract 3000 In T effector cells (Teff), protein kinase C theta (PKCθ) is a stabilizer of the immunologic synapse with antigen presenting cells (APCs). Signal propagation through PKCθ leads to activation of inflammatory NF-kB, thus implicating this molecule as a therapeutic target for T-cell mediated disorders as well as GVHD. PKCθ deficiency impairs donor T-cell mediated alloresponse and GVHD while leaving GVL responses intact to a certain extent. Given PKCθ is known to cooperate with its alpha isoform (PKCa) to optimize T-cell signaling and function, we have studied the role of PKCθ and PKCa in T-cell mediated alloresponse after allogeneic bone marrow transplantation (BMT) by using gene-deficient mice as well as a small molecular inhibitor of PKCa and θ. T cells from PKCa, θ and a/θ deficient (−/−) donor mice were tested in their ability to induce GVHD using an allogeneic murine model of myeloablative BMT. GVHD progression was monitored with or without A20-luciferase cells to observe GVL in vivo. To target PKC pharmacologically the small molecule inhibitor R524 (Rigel, San Francisco, CA), specific for PKCa/θ, was tested on GVHD progression in twice daily oral doses of 40 mg/kg for 6 weeks. Efficacy of R524 was assessed in an MHC-mismatched BMT model with and without the addition of A20-luciferase cells and in a minor antigen-mismatched BMT model with or without the addition of C1498-luciferase-DsRed cells to track GVHD progression and tumor relapse. Using a genetic approach, PKCa−/− donor T-cell alloreactivity is comparable to wild type (WT) T cell responses. WT T cells induced robust GVHD at a dose of 1×106/mouse. The same number of PKCθ−/− and PKCa/θ−/− T cells failed to generate even mild GVHD in recipient mice. At a high T cell dose, 5×106/mouse, PKCa/θ−/− T cells failed to induce severe acute GVHD and increased survival (P 〈 0.0001). The ability of PKCa/θ−/− T cells to induce GVHD was further reduced from PKCθ-/- T cells as evidenced by greater splenic cell number (P 〈 0.05) and B220+ B cell reconstitution in PKCa/θ−/− T-cell recipients. B cell activity in PKCa/θ−/− T-cell recipients was greater on a per cell basis; while the activity of T cells isolated from these mice was greatly reduced compared to WT or PKCθ−/− T-cell recipients. Microscopic tissue damage was also reduced (P 〈 0.05) in GVHD target organs in PKCa/θ−/− T-cell recipients. While not absent, GVL activity was compromised in PKCa/θ−/− T-cell recipients when compared to WT or PKCθ−/− T-cell recipients. Using a pharmacologic approach, the PKCa/θ small molecule inhibitor R524 reduced (P 〈 0.05) CD4+ and CD8+ T-cell proliferation (by 44% and 57% respectively) 5 days post allogeneic BMT. Long-term GVHD efficacy studies in an MHC-mismatched BMT model illustrate R524 mediated weight retention (P 〈 0.002) and decreased mortality (P 〈 0.01) compared to vehicle treatment. These observations were supported by a reduction in donor T cell expansion (P 〈 0.05), increased splenic retention of CD4+, CD8+, and B220+ cells and fewer donor T cells found in GVHD target organs of the recipients treated with R524 (P 〈 0.05). Additionally, CD4+ and CD8+ T cells isolated from spleens, livers and lungs of R524-treated mice produced less CXCR3, CCR6, TNFa and IFNg than their vehicle-treated counterparts. Similar results were obtained from studies using a minor antigen-mismatched BMT model, in which R524 increased recipient weight retention (P 〈 0.01) and reduced mortality by 50% (P 〈 0.002). Crucially, GVL activity was largely preserved in R524-treated mice compared to vehicle controls and mortality rates associated with tumor burden were minimal in both MHC-matched and mismatched models of BMT. In conclusion, we clearly show that PKCa and θ contribute to T cell activity with over-lapping functions in the induction of GVHD and GVL effects. Small molecule inhibition of PKCa/θ signaling with pharmacologic inhibitors presents a valid therapeutic option for GVHD prevention with simultaneous maintenance of GVL activity for clinical consideration. Disclosures: No relevant conflicts of interest to declare.

Description: Key Points PKCα and PKCθ cooperate in T-cell alloresponses, which contribute to GVHD. Pharmacologic inhibition of PKCα and PKCθ prevents GVHD and largely preserves GVL responses.

Description: Abstract 2458 Poster Board II-435 Introduction: When used as therapy for hematopoietic malignances, allogeneic hematopoietic stem cell transplantation (HCT) relies on the graft-versus-leukemia (GVL) effect to eradicate residual tumor cells through immunologic mechanisms. However, graft-versus-host-disease (GVHD) is a potentially lethal complication of allogeneic BMT. Thus, inhibition of GVHD, while preserving GVL and protective responses against infectious agents, can enhance the therapeutic potential of BMT. GVHD is initiated by alloreactive donor T cells that recognize mismatched major and/or minor histocompatibility antigens and cause severe damage to hematopoietic and epithelial tissues. Protein kinase C theta isoform (PKCθ) is crucial in TCR signaling and regulates the nature of lymphocyte-specific effector responses. The aim of this study was to investigate the significance of PKCθ in donor T-cell-mediated GVHD, anti-leukemia, and antiviral infection. Methods: Our results were validated using clinically relevant murine bone marrow transplantation (BMT) models. GVHD was measured by recipient survival and clinical signs such as body weight loss. Leukemia/lymphoma was induced by intravenous injection of a luciferase expressing A20 clone (a B lymphoma cell line derived from BALB/c mice). Tumor invasion was quantitated using the Xenogen IVIS-200 bioluminescence imaging system. Murine cytomegalovirus (MCMV) infection was induced by intraperitoneally injecting MCMV in the recipients after GVHD was established. Results: Using 3 distinct mouse models of allogeneic BMT, we found that T cells lacking PKCθ are unable to undergo robust expansion and cause damage to recipient hematopoietic or epithelial tissues, demonstrating that an essential requirement for PKCθ in alloreactivity and GVHD development. In contrast, PKCθ-/- T cells retain the ability to respond to virus infection post-BMT. Mechanistically, absence of PKCθ raises the threshold for T cell activation that selectively impacts alloresponses, but T cell responses triggered by infection or following administration of antigen plus microbial adjuvant are relatively well preserved in the absence of PKCθ Furthermore, we evaluated the role of PKCθ in GVHD and GVL responses, and found that PKCθ-/- T cells have at least a 10-fold reduction in the ability to induce GVHD but only a ∼2.5-fold reduction in GVL compared to WT T cells. Thus, absence of PKCθ impacts GVHD responses more significantly than GVL responses. Conclusion: These findings validate PKCθ as a potentially unique therapeutic target that is required for GVHD induction but not for GVL or protective responses to infectious agents in allogeneic BMT. Disclosures: No relevant conflicts of interest to declare.