Novel microtubule-targeting agents, pyrrolo-1,5-benzoxazepines, induce apoptosis in multi-drug-resistant cancer cells

PURPOSE: The development of multi-drug resistance (MDR) due to the expression of members of the ATP binding cassette (ABC) transporter family is a major obstacle in cancer treatment. The broad range of substrate specificities associated with these transporters leads to the efflux of many anti-cancer drugs from tumour cells. Therefore, the development of new chemotherapeutic agents that are not substrates of these transporters is important. We have recently demonstrated that some members of a novel series of pyrrolo-1,5-benzoxazepine (PBOX) compounds are microtubule-depolymerising agents that potently induce apoptosis in several cancer cell lines and impair growth of mouse breast tumours. The aim of this current study was to establish whether PBOXs were capable of inducing apoptosis in cancer cells expressing either P-glycoprotein or breast cancer resistance protein (BCRP), two of the main ABC transporters associated with MDR.

METHODS: We performed in vitro studies to assess the effects of PBOXs on cell proliferation, cell cycle and apoptosis in human cancer cell lines and their drug-resistant substrains expressing either P-glycoprotein or BCRP. In addition, we performed a preliminary molecular docking study to examine interactions between PBOXs and P-glycoprotein.

RESULTS: We established that three representative PBOXs, PBOX-6, -15 and -16 were capable of inducing apoptosis in drug-resistant HL60-MDR1 cells (expressing P-glycoprotein) and HL60-ABCG2 cells (expressing BCRP) with similar potencies as in parental human promyelocytic leukaemia HL60 cells. Likewise, resistance to PBOX-6 and -16 was not evident in P-glycoprotein-expressing A2780-ADR cells in comparison with parent human ovarian carcinoma A2780 cells. Finally, we deduced by molecular docking that PBOX-6 is not likely to form favourable interactions with the substrate binding site of P-glycoprotein.

CONCLUSION: Our results suggest that pro-apoptotic PBOX compounds may be potential candidates for the treatment of P-glycoprotein- or BCRP-associated MDR cancers.

BubR1 is required for a sustained mitotic spindle checkpoint arrest in human cancer cells treated with tubulin-targeting pyrrolo-1,5-benzoxazepines

Intrinsic or acquired resistance to chemotherapy is a major clinical problem that has evoked the need to develop innovative approaches to predict and ultimately reverse drug resistance. A prolonged G(2)M arrest has been associated with apoptotic resistance to various microtubule-targeting agents (MTAs). In this study, we describe the functional significance of the mitotic spindle checkpoint proteins, BubR1 and Bub3, in maintaining a mitotic arrest after microtubule disruption by nocodazole and a novel series of MTAs, the pyrrolo-1,5-benzoxazepines (PBOXs), in human cancer cells. Cells expressing high levels of BubR1 and Bub3 (K562, MDA-MB-231, and HeLa) display a prolonged G(2)M arrest after exposure to MTAs. On the other hand, cells with low endogenous levels of mitotic spindle checkpoint proteins (SK-BR-3 and HL-60) transiently arrest in mitosis and undergo increased apoptosis. The phosphorylation of BubR1 correlated with PBOX-induced G(2)M arrest in four cell lines tested, indicating an active mitotic spindle checkpoint. Gene silencing of BubR1 by small interfering RNA interference reduced PBOX-induced G(2)M arrest without enhancing apoptotic efficacy. Further analysis demonstrated that PBOX-treated BubR1-depleted cells were both mononucleated and multinucleated with a polyploid DNA content, suggesting a requirement for BubR1 in cytokinesis. Taken together, these results suggest that BubR1 contributes to the mitotic checkpoint induced by the PBOXs.

Identification of tubulin as the molecular target of proapoptotic pyrrolo-1,5-benzoxazepines

We have demonstrated previously that certain members of a series of novel pyrrolo-1,5-benzoxazepine (PBOX) compounds potently induce apoptosis in a variety of human chemotherapy-resistant cancer cell lines and in primary ex vivo material derived from cancer patients. A better understanding of the molecular mechanisms underlying the apoptotic effects of these PBOX compounds is essential to their development as antineoplastic therapeutic agents. This study sought to test the hypothesis that proapoptotic PBOX compounds target the microtubules. We show that a representative proapoptotic PBOX compound, PBOX-6, induces apoptosis in both the MCF-7 and K562 cell lines. An accumulation of cells in G2/M precedes apoptosis in response to PBOX-6. PBOX-6 induces prometaphase arrest and causes an accumulation of cyclin B1 levels and activation of cyclin B1/CDK1 kinase in a manner similar to that of two representative antimicrotubule agents, nocodazole and paclitaxel. Indirect immunofluorescence demonstrates that both PBOX-6 and another pro-apoptotic PBOX compound, PBOX-15, cause microtubule depolymerization in MCF-7 cells. They also inhibit the assembly of purified tubulin in vitro, whereas a nonapoptotic PBOX compound (PBOX-21) has no effect on either the cellular microtubule network or on the assembly of purified tubulin. This suggests that the molecular target of the pro-apoptotic PBOX compounds is tubulin. PBOX-6 does not bind to either the vinblastine or the colchicine binding site on tubulin, suggesting that it binds to an as-yet-uncharacterised novel site on tubulin. The ability of PBOX-6 to bind tubulin and cause microtubule depolymerization confirms it as a novel candidate for antineoplastic therapy.

Selective induction of apoptosis by the pyrrolo-1,5-benzoxazepine 7-[[dimethylcarbamoyl]oxy]-6-(2-naphthyl)pyrrolo-[2,1-d] (1,5)-benzoxazepine (PBOX-6) in Leukemia cells occurs via the c-Jun NH2-terminal kinase-dependent phosphorylation and inactivation of Bcl-2 and Bcl-XL

Overexpression of the Bcl-2 proto-oncogene in tumor cells confers resistance against chemotherapeutic drugs. In this study, we describe how the novel pyrrolo-1,5-benzoxazepine compound 7-[[dimethylcarbamoyl]oxy]-6-(2-naphthyl)pyrrolo-[2,1-d] (1,5)-benzoxazepine (PBOX-6) selectively induces apoptosis in Bcl-2-overexpressing cancer cells, whereas it shows no cytotoxic effect on normal peripheral blood mononuclear cells. PBOX-6 overcomes Bcl-2-mediated resistance to apoptosis in chronic myelogenous leukemia (CML) K562 cells by the time- and dose-dependent phosphorylation and inactivation of antiapoptotic Bcl-2 family members Bcl-2 and Bcl-XL. PBOX-6 also induces Bcl-2 phosphorylation and apoptosis in wild-type T leukemia CEM cells and cells overexpressing Bcl-2. This is in contrast to chemotherapeutic agents such as etoposide, actinomycin D, and ultraviolet irradiation, whereby overexpression of Bcl-2 confers resistance against apoptosis. In addition, PBOX-6 induces Bcl-2 phosphorylation and apoptosis in wild-type Jurkat acute lymphoblastic leukemia cells and cells overexpressing Bcl-2. However, Jurkat cells containing a Bcl-2 triple mutant, whereby the principal Bcl-2 phosphorylation sites are mutated to alanine, demonstrate resistance against Bcl-2 phosphorylation and apoptosis. PBOX-6 also induces the early and transient activation of c-Jun NH2-terminal kinase (JNK) in CEM cells. Inhibition of JNK activity prevents Bcl-2 phosphorylation and apoptosis, implicating JNK in the upstream signaling pathway leading to Bcl-2 phosphorylation. Collectively, these findings identify Bcl-2 phosphorylation and inactivation as a critical step in the apoptotic pathway induced by PBOX-6 and highlight its potential as an effective antileukemic agent.

Transcriptional response of T cells to IFN-alpha:changes induced in IFN-alpha-sensitive and resistant cutaneous T cell lymphoma

Interferon-alpha (IFN-alpha) therapy is commonly used in the treatment of neoplastic and autoimmune diseases, including cutaneous T cell lymphoma (CTCL). However, the IFN-alpha response is unpredictable, and the IFN-alpha cell targets and pathways are only partially understood. To delineate the molecular mechanisms of IFN-alpha activity, gene expression profiling was performed in a time-course experiment of both IFN-alpha sensitive and IFN-alpha-resistant variants of a CTCL cell line. These experiments revealed that IFN-alpha is responsible for the regulation of hundreds of genes in both variants and predominantly involves genes implicated in signal transduction, cell cycle control, apoptosis, and transcription regulation. Specifically, the IFN-alpha response of tumoral T cells is due to a combination of induction of apoptosis in which TNFSF10 and HSXIAPAF1 may play an important role and cell cycle arrest achieved by downregulation of CDK4 and CCNG2 and upregulation of CDKN2C and tumor suppressor genes (TSGs). Resistance to IFN-alpha appears to be associated with failure to induce IRF1 and IRF7 and deregulation of the apoptotic signals of HSXIAPAF1, TRADD, BAD, and BNIP3. Additionally, cell cycle progression is heralded by upregulation of CDC25A and CDC42. A critical role of NF-kappaB in promoting cell survival in IFN-alpha-resistant cells is indicated by the upregulation of RELB and LTB.

Activation of the c-Jun N-terminal kinase (JNK) signaling pathway is essential during PBOX-6-induced apoptosis in chronic myelogenous leukemia (CML) cells

The mitogen-activated protein (MAP) kinase family is activated in response to a wide variety of external stress signals such as UV irradiation, heat shock, and many chemotherapeutic drugs and leads to the induction of apoptosis. A novel series of pyrrolo-1,5-benzoxazepines have been shown to potently induce apoptosis in chronic myelogenous leukemia (CML) cells, which are resistant to many chemotherapeutic agents. In this study we have delineated part of the mechanism by which a representative compound known as PBOX-6 induces apoptosis. We have investigated whether PBOX-6 induces activation of MAP kinase signaling pathways in CML cells. Treatment of K562 cells with PBOX-6 resulted in the transient activation of two JNK isoforms, JNK1 and JNK2. In contrast, PBOX-6 did not activate the extracellular signal-regulated kinase (ERK) or p38. Apoptosis was found to occur independently of the small GTPases Ras, Rac, and Cdc42 but involved phosphorylation of the JNK substrates, c-Jun and ATF-2. Pretreatment of K562 cells with the JNK inhibitor, dicoumarol, abolished PBOX-6-induced phosphorylation of c-Jun and ATF-2 and inhibited the induced apoptosis, suggesting that JNK activation is an essential component of the apoptotic pathway induced by PBOX-6. Consistent with this finding, transfection of K562 cells with the JNK scaffold protein, JIP-1, inhibited JNK activity and apoptosis induced by PBOX-6. JIP-1 specifically scaffolds JNK, MKK7, and members of the mixed-lineage kinase (MLK) family, implicating these kinases upstream of JNK in the apoptotic pathway induced by PBOX-6 in K562 cells.

The emergence of DNA methylation as a key modulator of aberrant cell death in prostate cancer

It is now well established that cancer cells exhibit a number of genetic defects in the machinery that governs programmed cell death and that sabotage of apoptosis is one of the principal factors aiding in the evolution of the carcinogenic phenotype. A number of studies have implicated aberrant DNA methylation as a key survival mechanism in cancer, whereby promoter hypermethylation silences genes essential for many processes including apoptosis. To date, studies on the methylation profile of apoptotic genes have largely focused on cancers of the breast, colon and stomach, with only limited data available on prostate cancer. Here we discuss the major developments in the field of DNA methylation and its role in the regulation of aberrant apoptosis in prostate cancer. The most significant advances have involved the discovery of apoptotic gene targets of methylation, including XAF1, (fragile histidine triad (FHIT ), cellular retinol binding protein 1 (CRBP1), decoy receptor 1(DCR1), decoy receptor 2 (DCR2 ), target of methylation-induced silenceing 1 (TMS1), TNF receptor superfamily, member 6 (FAS), Reprimo (RPRM) and GLI pathogenesis-related 1 (GLIPR1). These genes are reported to be hypermethylated in prostate cancer and some offer potential as diagnostic and prognostic markers. We also introduce the concept of an 'apoptotic methylation signature' for prostate cancer and evaluate its potential in a diagnostic, prognostic and therapeutic setting.

Abl expression, tumour grade, and apoptosis in chondrosarcoma

AIMS: To determine whether Abl immunoreactivity correlates with grade and cell kinetics (apoptosis and mitosis) in chondrosarcoma.

METHODS: Sections from 16 chondrosarcomas were stained immunohistochemically using a polyclonal antibody to the c-Abl/Bcr-Abl oncoprotein. Apoptotic indices and mitotic indices were assessed in all tumours. Sections from 24 paraffin wax blocks of human fetal rib (gestational ages, 15-42 weeks) were also stained to determine whether the Abl protein is synthesised consistently throughout endochondral ossification.

RESULTS: Abl staining in immature fetal rib chondrocytes at all stages of development was predominantly nuclear, and 70% of cells showed moderate to strong staining. Abl immunoreactivity was minimal or absent in hypertrophic chondrocytes about to undergo apoptosis at the growth plate. There was strong Abl staining in grade 1 and grade 2 chondrosarcomas but staining was greatly reduced or absent in grade 3 chondrosarcomas. There was a very significant linear correlation between apoptotic index (mean, 0.68%; range, 0-3.2%) and mitotic index (mean, 0.23%; range, 0-0.9%), and both indices were significantly lower in grade 1 than in grade 2 and grade 3 chondrosarcomas.

CONCLUSIONS: These data suggest that abl gene expression is associated with differentiation and apoptosis inhibition in fetal and neoplastic chondrocytes. However, these putative effects cannot be ascribed solely to the Abl protein, because several additional factors contribute to the regulation of both differentiation and apoptosis.

A miR-433 mediated mechanism of chemoresistance in high grade serous ovarian cancer (HGSOC).

Higher expression of the miR-433 microRNA (miRNA) is associated with poorer survival outcomes in patients with HGSOC that may be overcome by a greater understanding of the functional role of this miRNA. We previously described miR-433 as a critical cell cycle regulator and mediator of cellular senescence. Downregulation of the mitotic arrest deficiency 2 (MAD2) protein by miR-433 led to increased cellular resistance to paclitaxel in epithelial ovarian cancer cells (EOC). Furthermore immunohistochemical (IHC) analysis of MAD2 expression in patients with HGSOC showed that loss of MAD2 was significantly associated with poorer patient survival. Higher miR-433 expression is also associated with an increased resistance to the platins which is unrelated to loss of MAD2 expression. In silico analysis of the miR-433 target proteins in the TCGA database identified the association between a number of miR-433 targets and poorer patient survival. IHC analysis of the miR-433 target, histone deacetylase 6 (HDAC6), confirmed that its expression was significantly associated with a decrease in patient overall survival. The knock-down of HDAC6 by siRNA in EOC cells did not attenuate apoptotic responses to paclitaxel or platin although lower endogenous HDAC6 expression was associated with more resistant EOC cell lines. In vitro analysis revealed that EOC cells which survived chemotherapeutic kill with high doses of paclitaxel expressed higher miR-433 and concomitant decreased expression of the miR-433 targets. These cells were more chemoresistant compared to the parental cell line and repopulated as 3d organoid cultures in non-adherent stem cell selective conditions; thus indicating that the cells which survive chemotherapy were viable, capable of regrowth and had an increased potential for pluripotency. In conclusion, our data suggests that chemotherapy is not driving the transcriptional upregulation of miR-433 but rather selecting a population of cells with high miR-433 expression that may contribute to chemoresistant disease and tumour recurrence.

Tumor necrosis factor receptor expression and signaling in renal cell carcinoma

Clear cell renal cell carcinoma (ccRCC), a tubular epithelial cell (TEC) malignancy, frequently secretes tumor necrosis factor (TNF). TNF signals via two distinct receptors (TNFRs). TNFR1, expressed in normal kidney primarily on endothelial cells, activates apoptotic signaling kinase 1 and nuclear factor-kappaB (NF-kappaB) and induces cell death, whereas TNFR2, inducibly expressed on endothelial cells and on TECs by injury, activates endothelial/epithelial tyrosine kinase (Etk), which trans-activates vascular endothelial growth factor receptor 2 (VEGFR2) to promote cell proliferation. We investigated TNFR expression in clinical samples and function in short-term organ cultures of ccRCC tissue treated with wild-type TNF or specific muteins selective for TNFR1 (R1-TNF) or TNFR2 (R2-TNF). There is a significant increase in TNFR2 but not TNFR1 expression on malignant TECs that correlates with increasing malignant grade. In ccRCC organ cultures, R1-TNF increases TNFR1, activates apoptotic signaling kinase and NF-kappaB, and promotes apoptosis in malignant TECs. R2-TNF increases TNFR2, activates NF-kappaB, Etk, and VEGFR2 and increases entry into the cell cycle. Wild-type TNF induces both sets of responses. R2-TNF actions are blocked by pretreatment with a VEGFR2 kinase inhibitor. We conclude that TNF, acting through TNFR2, is an autocrine growth factor for ccRCC acting via Etk-VEGFR2 cross-talk, insights that may provide a more effective therapeutic approach to this disease.

Inhibition of dUTPase induces synthetic lethality with thymidylate synthase-targeted therapies in non-small cell lung cancer

Chemotherapies that target thymidylate synthase (TS) continue to see considerable clinical expansion in non-small cell lung cancer (NSCLC). One drawback to TS-targeted therapies is drug resistance and subsequent treatment failure. Novel therapeutic and biomarker-driven strategies are urgently needed. The enzyme deoxyuridine triphosphate nucleotidohydrolase (dUTPase) is reported to protect tumor cells from aberrant misincorporation of uracil during TS inhibition. The goal of this study was to investigate the expression and significance of dUTPase in mediating response to TS-targeted agents in NSCLC. The expression of dUTPase in NSCLC cell lines and clinical specimens was measured by quantitative real-time reverse transcriptase PCR and immunohistochemistry. Using a validated RNA interference approach, dUTPase was effectively silenced in a panel of NSCLC cell lines and response to the fluoropyrimidine fluorodeoxyuridine (FUdR) and the antifolate pemetrexed was analyzed using growth inhibition and clonogenic assays. Apoptosis was analyzed by flow cytometry. Significant variation in the quantity and cellular expression of dUTPase was observed, including clear evidence of overexpression in NSCLC cell line models and tumor specimens at the mRNA and protein level. RNA interference-mediated silencing of dUTPase significantly sensitized NSCLC cells to growth inhibition induced by FUdR and pemetrexed. This sensitization was accompanied by a significant expansion of intracellular dUTP pools and significant decreases in NSCLC cell viability evaluated by clonogenicity and apoptotic analyses. Together, these results strongly suggest that uracil misincorporation is a potent determinant of cytotoxicity to TS inhibition in NSCLC and that inhibition of dUTPase is a mechanism-based therapeutic approach to significantly enhance the efficacy of TS-targeted chemotherapeutic agents.

Alterations in Circulating Immune Cells in Neovascular Age-Related Macular Degeneration

Neovascular age-related macular degeneration (nAMD) is the leading cause of irreversible blindness in developed countries. Recent advances have highlighted the essential role of inflammation in the development of the disease. In addition to local retinal chronic inflammatory response, systemic immune alterations have also been observed in AMD patients. In this study we investigated the association between the frequency of circulating leukocyte populations and the prevalence as well as clinical presentations of nAMD. Leukocyte subsets of 103 nAMD patients (most of them were receiving anti-VEGF therapy prior to enrolment) and 26 controls were analysed by flow cytometry by relative cell size, granularity and surface markers. Circulating CD11b(+) cells and CD16(hi)HLA-DR(-) neutrophils were significantly increased (P = 0.015 and 0.009 respectively) in nAMD when compared to controls. The percentage of circulating CD4(+) T-cells was reduced in nAMD patients without subretinal fibrosis (P = 0.026) compared to patients with subretinal fibrosis. There was no correlation between the percentage of circulating leukocytes and the responsiveness to anti-VEGF therapy in nAMD patients. Our results suggest that higher levels of circulating CD11b(+) cells and neutrophils are associated with nAMD and that reduced levels of CD4(+) T-cells are associated with the absence of subretinal fibrosis in nAMD.

Single nucleotide polymorphisms in the APCS gene influence geographic atrophy secondary to VEGF inhibition therapy in neovascular macular degeneration.

Introduction: Age-related macular degeneration (AMD) is a leading cause of vision loss in the elderly mostly due to the development of neovascular AMD (nAMD) or geographic atrophy (GA). Intravitreal injections of anti-vascular endothelial growth factor (VEGF) agents are an effective therapeutic option for nAMD. Following anti-VEGF treatments, increased atrophy of the retinal pigment epithelium (RPE) and choriocapillaries that resembles GA has been reported. We sought to evaluate the underlying genetic influences that may contribute to this process. Methods: We selected 68 single nucleotide polymorphisms (SNPs) from genes previously identified as susceptibility factors in AMD, along with 43 SNPs from genes encoding the VEGF protein and its cognate receptors as this pathway is targeted by treatment. We enrolled 467 consecutive patients (Feb 2009 to October 2011) with nAMD who received anti-VEGF therapy. The acutely presenting eye was designated as the study eye and retinal tomograms graded for macular atrophy at study exit. Statistical analysis was performed using PLINK to identify SNPs with a P value < 0.01. Logistic regression models with macular atrophy as dependent variable were fitted with age, gender, smoking status, common genetic risk factors and the identified SNPs as explanatory variables. Results: Grading for macular atrophy was available in 304 study eyes and 70% (214) were classified as showing macular atrophy. In the unadjusted analysis we observed significant associations between macular atrophy and two independent SNPs in the APCS gene: rs6695377: odds ratio (OR) = 1.98; 95% confidence intervals (CI): 1.23, 3.19; P = 0.004; rs1446965: OR = 2.49, CI: 1.29, 4.82; P = 0.006 and these associations remained significant after adjustment for covariates. Conclusions: VEGF is a mitogen and growth factor for choroidal blood vessels and the RPE and its inhibition could lead to atrophy of these key tissues. Anti-VEGF treatment can interfere with ocular vascular maintenance and may be associated with RPE and choroidal atrophy. As such, these medications, which block the effects of VEGF, may influence the development of GA. The top associated SNPs are found in the APCS gene, a highly conserved glycoprotein that encodes Serum amyloid P (SAP) which opsonizes apoptotic cells. SAP can bind to and activate complement components via binding to C1q, a mechanism by which SAP may remove cellular debris, affecting regulation of the three complement pathways.

PTEN deficiency promotes macrophage infiltration and hypersensitivity of prostate cancer to IAP antagonist/radiation combination therapy

PTEN loss is prognostic for patient relapse post-radiotherapy in prostate cancer (CaP). Infiltration of tumor-associated macrophages (TAMs) is associated with reduced disease-free survival following radical prostatectomy. However, the association between PTEN loss, TAM infiltration and radiotherapy response of CaP cells remains to be evaluated. Immunohistochemical and molecular analysis of surgically-resected Gleason 7 tumors confirmed that PTEN loss correlated with increased CXCL8 expression and macrophage infiltration. However PTEN status had no discernable correlation with expression of other inflammatory markers by CaP cells, including TNF-α. In vitro, exposure to conditioned media harvested from irradiated PTEN null CaP cells induced chemotaxis of macrophage-like THP-1 cells, a response partially attenuated by CXCL8 inhibition. Co-culture with THP-1 cells resulted in a modest reduction in the radio-sensitivity of DU145 cells. Cytokine profiling revealed constitutive secretion of TNF-α from CaP cells irrespective of PTEN status and IR-induced TNF-α secretion from THP-1 cells. THP-1-derived TNF-α increased NFκB pro-survival activity and elevated expression of anti-apoptotic proteins including cellular inhibitor of apoptosis protein-1 (cIAP-1) in CaP cells, which could be attenuated by pre-treatment with a TNF-α neutralizing antibody. Treatment with a novel IAP antagonist, AT-IAP, decreased basal and TNF-α-induced cIAP-1 expression in CaP cells, switched TNF-α signaling from pro-survival to pro-apoptotic and increased radiation sensitivity of CaP cells in co-culture with THP-1 cells. We conclude that targeting cIAP-1 can overcome apoptosis resistance of CaP cells and is an ideal approach to exploit high TNF-α signals within the TAM-rich microenvironment of PTEN-deficient CaP cells to enhance response to radiotherapy.

Host Defence Peptide LL-37; Effects of truncation on antimicrobial activity

Background: The oral cavity is an ideal environment for colonisation by micro-organisms. A first line of defence against microbial infection is the secretion of broad spectrum host defence peptides (HDPs). In the current climate of antibiotic resistance, exploiting naturally occurring HDPs or synthetic derivatives (mimetics) to combat infection is particularly appealing. The human cathelicidin, LL-37 is one such HDP expressed ubiquitously by epithelial cells and neutrophils. LL-37 exhibits the ability to bind lipopolysaccharide (LPS) and displays broad spectrum activity against a wide range of bacteria. The current study focuses on truncation of LL-37 and defining the antimicrobial and LPS binding activity of the resultant mimetics. Objectives: To assess the antimicrobial and LPS binding activity of LL-37 and three truncated mimetics (KE-18, EF-14 and KR-12). Methods: Peptides were synthesised in-house by Fmoc solid phase peptide synthesis or obtained commercially. Antimicrobial activity was determined using a radial diffusion assay and ability to bind LPS was determined by indirect ELISA. Results: LL-37 and mimetics displayed antimicrobial activity against Streptococcus mutans and Enterococcus Faecalis. KE-18 and KR-12 were shown to possess antimicrobial activity against both pathogens whereas EF-14 was the least antimicrobial. In terms of LPS binding, KE-18 and KR-12 were both effective whereas EF-14 showed the least activity of the three mimetics. Conclusion: Truncation of LL-37 can yield peptides which retain antimicrobial activities and have the ability to bind LPS. Interestingly in some cases the truncation of LL-37 produced mimetics with greater potency than the parent molecule in terms of antimicrobial activity and LPS binding. This work was funded by DEL and the Diabetes Wellness Foundation.