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Activated human T cells express alternative mRNA transcripts encoding a secreted form of RANKL

Mon, 02/23/2015 - 2:50pm

Receptor activator of nuclear factor-kappaB-ligand (RANKL), encoded by the gene TNFSF11, is required for osteoclastogenesis, and its expression is upregulated in pathologic bone loss. Transcript variants of TNFSF11 messenger RNA (mRNA) have been described that encode a membrane-bound and a putative secreted form of RANKL. We identify a TNFSF11 transcript variant that extends the originally identified transcript encoding secreted RANKL. We demonstrate that this TNFSF11 transcript variant is expressed by the human osteosarcoma cell line, Saos-2, and by both primary human T cells and Jurkat T cells. Of relevance to the production of RANKL in pathologic bone loss, expression of this secreted TNFSF11 transcript is upregulated in Jurkat T cells and primary human T cells upon activation. Furthermore, this transcript can be translated and secreted in Jurkat T cells in vitro and is able to support osteoclast differentiation. Our data highlight the complexity of the TNFSF11 genomic locus, and demonstrate the potential for the expression of alternate mRNA transcripts encoding membrane-bound and secreted forms of RANKL. Implications of alternate mRNA transcripts encoding different RANKL protein isoforms should be carefully considered and specifically examined in future studies, particularly those implicating RANKL in pathologic bone loss.

Osteoblast function is compromised at sites of focal bone erosion in inflammatory arthritis

Mon, 02/23/2015 - 2:50pm

In rheumatoid arthritis (RA), synovial inflammation results in focal erosion of articular bone. Despite treatment attenuating inflammation, repair of erosions with adequate formation of new bone is uncommon in RA, suggesting that bone formation may be compromised at these sites. Dynamic bone histomorphometry was used in a murine model of RA to determine the impact of inflammation on osteoblast function within eroded arthritic bone. Bone formation rates at bone surfaces adjacent to inflammation were similar to those observed in nonarthritic bone; therefore, osteoblast activity is unlikely to compensate for the increased bone resorption at these sites. Within arthritic bone, the extent of actively mineralizing surface was reduced at bone surfaces adjacent to inflammation compared with bone surfaces adjacent to normal marrow. Consistent with the reduction in mineralized bone formation, there was a notable paucity of cells expressing the mid- to late stage osteoblast lineage marker alkaline phosphatase, despite a clear presence of cells expressing the early osteoblast lineage marker Runx2. In addition, several members of the Dickkopf and secreted Frizzled-related protein families of Wnt signaling antagonists were upregulated in arthritic synovial tissues, suggesting that inhibition of Wnt signaling could be one mechanism contributing to impaired osteoblast function within arthritic bone. Together, these data indicate that the presence of inflammation within arthritic bone impairs osteoblast capacity to form adequate mineralized bone, thus contributing to the net loss of bone and failure of bone repair at sites of focal bone erosion in RA.

RANKL protein is expressed at the pannus-bone interface at sites of articular bone erosion in rheumatoid arthritis

Mon, 02/23/2015 - 2:50pm

OBJECTIVES: Receptor activator of NF-kappaB ligand (RANKL) and osteoprotegerin (OPG) have been demonstrated to be critical regulators of osteoclast generation and activity. In addition, RANKL has been implicated as an important mediator of bone erosion in rheumatoid arthritis (RA). However, the expression of RANKL and OPG at sites of pannus invasion into bone has not been examined. The present study was undertaken to further elucidate the contribution of this cytokine system to osteoclastogenesis and subsequent bone erosion in RA by examining the pattern of protein expression for RANKL, OPG and the receptor activator of NF-kappaB (RANK) in RA at sites of articular bone erosion.

METHODS: Tissues from 20 surgical procedures from 17 patients with RA were collected as discarded materials. Six samples contained only synovium or tenosynovium remote from bone, four samples contained pannus-bone interface with adjacent synovium and 10 samples contained both synovium remote from bone and pannus-bone interface with adjacent synovium. Immunohistochemistry was used to characterize the cellular pattern of RANKL, RANK and OPG protein expression immediately adjacent to and remote from sites of bone erosion.

RESULTS: Cellular expression of RANKL protein was relatively restricted in the bone microenvironment; staining was focal and confined largely to sites of osteoclast-mediated erosion at the pannus-bone interface and at sites of subchondral bone erosion. RANK-expressing osteoclast precursor cells were also present in these sites. OPG protein expression was observed in numerous cells in synovium remote from bone but was more limited at sites of bone erosion, especially in regions associated with RANKL expression.

CONCLUSIONS: The pattern of RANKL and OPG expression and the presence of RANK-expressing osteoclast precursor cells at sites of bone erosion in RA contributes to the generation of a local microenvironment that favours osteoclast differentiation and activity. These data provide further evidence implicating RANKL in the pathogenesis of arthritis-induced joint destruction.

TRANCE/RANKL knockout mice are protected from bone erosion in a serum transfer model of arthritis

Mon, 02/23/2015 - 2:50pm

There is considerable evidence that osteoclasts are involved in the pathogenesis of focal bone erosion in rheumatoid arthritis. Tumor necrosis factor-related activation-induced cytokine, also known as receptor activator of nuclear factor-kappaB ligand (TRANCE/RANKL) is an essential factor for osteoclast differentiation. In addition to its role in osteoclast differentiation and activation, TRANCE/RANKL also functions to augment T-cell dendritic cell cooperative interactions. To further evaluate the role of osteoclasts in focal bone erosion in arthritis, we generated inflammatory arthritis in the TRANCE/RANKL knockout mouse using a serum transfer model that bypasses the requirement for T-cell activation. These animals exhibit an osteopetrotic phenotype characterized by the absence of osteoclasts. Inflammation, measured by clinical signs of arthritis and histopathological scoring, was comparable in wild-type and TRANCE/RANKL knockout mice. Microcomputed tomography and histopathological analysis demonstrated that the degree of bone erosion in TRANCE/RANKL knockout mice was dramatically reduced compared to that seen in control littermate mice. In contrast, cartilage erosion was present in both control littermate and TRANCE/RANKL knockout mice. These results confirm the central role of osteoclasts in the pathogenesis of bone erosion in arthritis and demonstrate distinct mechanisms of cartilage destruction and bone erosion in this animal model of arthritis.

A longitudinal study of multicultural curriculum in medical education

Thu, 02/19/2015 - 8:50pm

OBJECTIVE: To evaluate impact a multicultural interclerkship had on students' perception of knowledge, interview skills, and empathy towards serving culturally diverse populations and role student demographics played in learning.

METHODS: Data extracted from students' self-reported course evaluations and pre/post questionnaires during multiculturalism interclerkship across 11 academic years. Inquired students' opinion about four areas: effectiveness, small group leaders, usefulness, and overall experience. Subscale and item ratings were compared using trend tests including multivariate analyses.

RESULTS: During studied years, 883 students completed course evaluation with high overall mean rating of 3.08 (S = 0.45) and subscale mean scores ranging from 3.03 to 3.30. Trends in three of four subscales demonstrated clear uptrend (p < 0.0001). Positive correlations between ratings of leaders and "usefulness" were observed (p < 0.0001). Pre/post matched dataset (n = 967) indicated majority of items (19/23) had statistically significant higher post interclerkship ratings compared to pre scores with nine of 19 having statistically significant magnitudes of change. Questionnaire had high overall reliability (Cronbach alpha = 0.8), and item-to-group correlations ranged from 0.40 to 0.68 (p < 0.0001).

CONCLUSIONS: By increasing students' exposure and interaction with diverse patients, their knowledge, attitude, and skills were increased and expanded in positive manner. These findings might inform those who are interested in enhancing this important competence. This is especially true given increasing scrutiny this global topic is receiving within and across healthcare professions around the world.

A survey study of evidence-based medicine training in US and Canadian medical schools

Thu, 02/19/2015 - 8:50pm

PURPOSE: The authors conducted a survey examining (1) the current state of evidence-based medicine (EBM) curricula in US and Canadian medical schools and corresponding learning objectives, (2) medical educators' and librarians' participation in EBM training, and (3) barriers to EBM training.

METHODS: A survey instrument with thirty-four closed and open-ended questions was sent to curricular deans at US and Canadian medical schools. The survey sought information on enrollment and class size; EBM learning objectives, curricular activities, and assessment approaches by year of training; EBM faculty; EBM tools; barriers to implementing EBM curricula and possible ways to overcome them; and innovative approaches to EBM education. Both qualitative and quantitative methods were used for data analysis. Measurable learning objectives were categorized using Bloom's taxonomy.

RESULTS: One hundred fifteen medical schools (77.2%) responded. Over half (53%) of the 900 reported learning objectives were measurable. Knowledge application was the predominant category from Bloom's categories. Most schools integrated EBM into other curricular activities; activities and formal assessment decreased significantly with advanced training. EBM faculty consisted primarily of clinicians, followed by basic scientists and librarians. Various EBM tools were used, with PubMed and the Cochrane database most frequently cited. Lack of time in curricula was rated the most significant barrier. National agreement on required EBM competencies was an extremely helpful factor. Few schools shared innovative approaches.

CONCLUSIONS: Schools need help in overcoming barriers related to EBM curriculum development, implementation, and assessment.

IMPLICATIONS: Findings can provide a starting point for discussion to develop a standardized competency framework.

UMCCTS Newsletter, February 2015

Wed, 02/11/2015 - 2:36pm

This is the February 2015 issue of the UMass Center for Clinical and Translational Science Newsletter containing news and events of interest.

Glioblastoma multiforme therapy and mechanisms of resistance

Wed, 02/11/2015 - 11:39am

Glioblastoma multiforme (GBM) is a grade IV brain tumor characterized by a heterogeneous population of cells that are highly infiltrative, angiogenic and resistant to chemotherapy. The current standard of care, comprised of surgical resection followed by radiation and the chemotherapeutic agent temozolomide, only provides patients with a 12-14 month survival period post-diagnosis. Long-term survival for GBM patients remains uncommon as cells with intrinsic or acquired resistance to treatment repopulate the tumor. In this review we will describe the mechanisms of resistance, and how they may be overcome to improve the survival of GBM patients by implementing novel chemotherapy drugs, new drug combinations and new approaches relating to DNA damage, angiogenesis and autophagy.

Evaluation of novel imidazotetrazine analogues designed to overcome temozolomide resistance and glioblastoma regrowth

Wed, 02/11/2015 - 11:39am

The cellular responses to two new temozolomide (TMZ) analogues, DP68 and DP86, acting against glioblastoma multiforme (GBM) cell lines and primary culture models are reported. Dose-response analysis of cultured GBM cells revealed that DP68 is more potent than DP86 and TMZ and that DP68 was effective even in cell lines resistant to TMZ. On the basis of a serial neurosphere assay, DP68 inhibits repopulation of these cultures at low concentrations. The efficacy of these compounds was independent of MGMT and MMR functions. DP68-induced interstrand DNA cross-links were demonstrated with H2O2-treated cells. Furthermore, DP68 induced a distinct cell-cycle arrest with accumulation of cells in S phase that is not observed for TMZ. Consistent with this biologic response, DP68 induces a strong DNA damage response, including phosphorylation of ATM, Chk1 and Chk2 kinases, KAP1, and histone variant H2AX. Suppression of FANCD2 expression or ATR expression/kinase activity enhanced antiglioblastoma effects of DP68. Initial pharmacokinetic analysis revealed rapid elimination of these drugs from serum. Collectively, these data demonstrate that DP68 is a novel and potent antiglioblastoma compound that circumvents TMZ resistance, likely as a result of its independence from MGMT and mismatch repair and its capacity to cross-link strands of DNA. Mol Cancer Ther; 14(1); 111-9. ©2014 AACR.

Evaluation of [In]-Labeled Zinc-Dipicolylamine Tracers for SPECT Imaging of Bacterial Infection

Tue, 02/10/2015 - 10:11pm

PURPOSE: This study prepared three structurally related zinc-dipicolylamine (ZnDPA) tracers with [111In] labels and conducted biodistribution and single-photon emission computed tomography/computed tomography (SPECT/CT) imaging studies of a mouse leg infection model.

PROCEDURES: Two monovalent tracers, ZnDPA-[111In]DTPA and ZnDPA-[111In]DOTA, each with a single zinc-dipicolylamine targeting unit, and a divalent tracer, Bis(ZnDPA)-[111In]DTPA, with two zinc-dipicolylamine units were prepared. Organ biodistribution and SPECT and CT imaging studies were performed on living mice with a leg infection created by injection of clinically relevant Gram positive Streptococcus pyogenes. Fluorescent and luminescent Eu3+-labeled versions of these tracers were also prepared and used to measure relative affinity for the exterior membrane surface of bacterial cells and mimics of healthy mammalian cells.

RESULTS: All three 111In-labeled radiotracers were prepared with a radiopurity of greater than 90%. The biodistribution studies showed that the two monovalent tracers were cleared from the body through the liver and kidney, with retained percentage injected dose for all organs of < 8% at 20h and infected leg target to non-target ratio (T/NT) ratio of greater than or equal to 3.0. Clearance of the divalent tracer from the bloodstream was slower and primarily through the liver, with a retained percentage injected dose for all organs greater than 37% at 20h and T/NT ratio rising to 6.2 after 20 h. The SPECT/CT imaging indicated the same large difference in tracer pharmacokinetics and higher accumulation of the divalent tracer at the site of infection.

CONCLUSIONS: All three [111In]-ZnDPA tracers selectively targeted the site of a clinically relevant mouse infection model that could not be discerned by visual external inspection of the living animal. The highest target selectivity, observed with a divalent tracer equipped with two zinc-dipicolylamine targeting units, compares quite favorably with the imaging selectivities previously reported for other nuclear tracers that target bacterial cell surfaces. The tracer pharmacokinetics depended heavily on tracer molecular structure suggesting that it may be possible to rapidly fine tune the structural properties for optimized in vivo imaging performance and clinical translation.

Her2/neu small interfering RNA delivered in culture by a streptavidin nanoparticle

Tue, 02/10/2015 - 10:11pm

A three-component nanoparticle consisting of biotinylated Trastuzumab antiHer2 antibody, tat transferring peptide and radiolabeled antisense oligomer, linked together through streptavidin, have shown promise in the delivery to Her2+ tumor in mice following intravenous administration and with evidence of radiotherapeutic efficacy. These results have encouraged us to consider the nanoparticle as a delivery vehicle for RNA interference therapy in which the radiolabeled antisense oligomer is replaced with an unlabeled siRNA duplex. The siRNA stability within the nanoparticle was first confirmed by incubation with RNase A. The interferon responses, that indicate off-target cytotoxicity, were evaluated by quantitative real-time RT-PCR in BT-474 (Her2+) human breast cancer cells by measuring the mRNA expression of 2', 5'-oligoadenylate synthetase (OAS1) and Stat-1, two key interferon-responsive genes. Thereafter the cytotoxicity induced by the siRNA nanoparticle was evaluated by a clonogenic survival assay in BT-474 cells while the Her2 expression of these target cells was evaluated for evidence of specific gene silencing. The siRNA within the three-component anti- Her2/neu siRNA nanoparticle was largely protected from RNase-dependent degradation and did not activate an interferon response. The nanoparticle effectively and significantly inhibited colony formation of the target cells and silenced the Her2 gene expression at 5 nM compared with the identical nanoparticle with a scrambled siRNA. Our delivery nanoparticle, with tumor targeting provided by the antibody and its accumulation without entrapment, possibly due to the transfecting peptide, delivered an siRNA duplex to the proper subcellular localization for specific and effective gene silencing in culture by what appears to be an siRNA mechanism.

Activin-like kinase 3 is important for kidney regeneration and reversal of fibrosis

Tue, 02/10/2015 - 10:11pm

Molecules associated with the transforming growth factor beta (TGF-beta) superfamily, such as bone morphogenic proteins (BMPs) and TGF-beta, are key regulators of inflammation, apoptosis and cellular transitions. Here we show that the BMP receptor activin-like kinase 3 (Alk3) is elevated early in diseased kidneys after injury. We also found that its deletion in the tubular epithelium leads to enhanced TGF-beta1-Smad family member 3 (Smad3) signaling, epithelial damage and fibrosis, suggesting a protective role for Alk3-mediated signaling in the kidney. A structure-function analysis of the BMP-Alk3-BMP receptor, type 2 (BMPR2) ligand-receptor complex, along with synthetic organic chemistry, led us to construct a library of small peptide agonists of BMP signaling that function through the Alk3 receptor. One such peptide agonist, THR-123, suppressed inflammation, apoptosis and the epithelial-to-mesenchymal transition program and reversed established fibrosis in five mouse models of acute and chronic renal injury. THR-123 acts specifically through Alk3 signaling, as mice with a targeted deletion for Alk3 in their tubular epithelium did not respond to therapy with THR-123. Combining THR-123 and the angiotensin-converting enzyme inhibitor captopril had an additive therapeutic benefit in controlling renal fibrosis. Our studies show that BMP signaling agonists constitute a new line of therapeutic agents with potential utility in the clinic to induce regeneration, repair and reverse established fibrosis.

Radiolabeled Zn-DPA as a potential infection imaging agent

Tue, 02/10/2015 - 10:11pm

INTRODUCTION: A zinc-dipicolylamine analog (Zn-DPA) conjugated with a fluorophore (PSVue(R)794) has been shown to image bacterial infections in mice. However, radiolabeled Zn-DPA has not previously been considered for nuclear imaging of infection.

METHODS: Both 111In-labeled DOTA-biotin and Zn-DPA-biotin were combined using streptavidin (SA) as a noncovalent linker. Mice injected intramuscularly with Streptococcus pyogenes (infection model) or with lipopolysaccharide (LPS) (inflammation model) were coinjected intravenously with 6 mug of DPA as PSVue794 and as 111In-DOTA-biotin/SA/biotin-Zn-DPA. Periodic fluorescent and SPECT (single photon emission computed tomography)/CT (computed tomography) images were acquired, and biodistributions were obtained at 22 h.

RESULTS: Histological examination confirmed the validity of both the infection and inflammation animal models. Both the whole-body optical and nuclear images showed obvious accumulations in the target thigh in both models at all time points. At 22 h, the average target thigh accumulation of 111In was 1.66%ID/g (S.D. 0.15) in the infection mice compared to 0.58%ID/g (S.D. 0.07) in the inflammation mice (P < .01), and the 111In target/normal thigh ratio was 2.8 fold higher in the infection animals compared to the inflammation animals.

CONCLUSIONS: These preliminary results show that Zn-DPA within streptavidin targets S. pyogenes-infected mice similarly to its free fluorescent analogue. The significantly higher accumulation in the live bacterial infection thigh compared to that of the LPS-induced inflammation thigh suggests that Zn-DPA may be a promising imaging agent to distinguish between bacterial infections and sterile inflammations.

Assessing antibody pharmacokinetics in mice with in vivo imaging

Tue, 02/10/2015 - 10:11pm

Recent advances in small-animal molecular imaging instrumentation combined with well characterized antibody-labeling chemistry have enabled detailed in vivo measurements of antibody distribution in mouse models. This article reviews the strengths and limitations of in vivo antibody imaging methods with a focus on positron emission tomography and single-photon emission computed tomography and a brief discussion of the role of optical imaging in this application. A description of the basic principles behind the imaging techniques is provided along with a discussion of radiolabeling methods relevant to antibodies. Practical considerations of study design and execution are presented through a discussion of sensitivity and resolution tradeoffs for these techniques as defined by modality, signaling probe (isotope or fluorophore) selection, labeling method, and radiation dosimetry. Images and analysis results from a case study are presented with a discussion of output data content and relevant informatics gained with this approach to studying antibody pharmacokinetics.

Comparison of 18F PET and 99mTc SPECT imaging in phantoms and in tumored mice

Tue, 02/10/2015 - 10:11pm

Our objective was to compare the performance of a micro-single photon emission computed tomography (micro-SPECT) with that of a micro-positron emission tomography (microPET) in a Her2+ tumored mice using an anti-Her2 nanoparticle radiolabeled with (99m)Tc and (18)F. Camera performance was first compared using phantoms; then a tumored mouse administered the (99m)Tc-nanoparticle was imaged on a Bioscan NanoSPECT/CT, while another tumored mouse received the identical nanoparticle, labeled now with (18)F, and was imaged on a Philips Mosaic HP PET camera. The nanoparticle was radiolabeled with (99m)Tc via MAG(3) chelation and with (18)F via SFB as an intermediate. Phantom imaging showed that the resolution of the SPECT camera was clearly superior, but even with 4 heads and multipinhole collimators, detection sensitivity was 15-fold lower. Radiolabeling of the nanoparticle by chelation with (99m)Tc was considerably easier and safer than manual covalent attachment of (18)F. Both cameras provided accurate quantitation of radioactivity over a broad range. In conclusion, when deciding between (99m)Tc vs (18)F, an advantage rests with the chelation of (99m)Tc over covalent attachment of (18)F, achieved manually or otherwise, but with these small animal cameras, this choice also results in trading lower sensitivity for higher resolution.

Auger-mediated cytotoxicity of cancer cells in culture by an 125I-antisense oligomer delivered as a three-component streptavidin nanoparticle

Tue, 02/10/2015 - 10:11pm

We reported recently that a three-component nanoparticle, consisting of a targeting antibody, a transfecting peptide and an 111In-antiRIalpha MORF antisense oligomer, provided Auger electron-mediated, antisense-mediated, cytotoxicity of cells in culture. We have now measured the cytotoxicity of the nanoparticle in culture with the 111In replaced by 125I, another attractive Auger electron emitter. The nanoparticle consisted of streptavidin linking the 125I labeled antiRIalpha mRNA antisense MORF oligomer, the tat transfecting peptide and the anti-Her2 Trastuzumab antibody. Cytotoxicity was evaluated by a clonogenic survival assay in BT-474 (Her2+) human breast cancer cells. In a dose escalation study, as measured by the surviving fraction, the cytotoxicity of tumor cells to the 125I-labeled antisense nanoparticle was significantly higher than that for the identical sense control. When compared with our previous study with 111In as label, a similar level of cytotoxicity was achieved but the observed minimal therapeutic dose for the 125I-labeled nanoparticle in BT-474 cells was lower than that for 111In-labeled nanoparticle in SK-BR-3 cells. Thus, a radiolabeled antisense MORF oligomer delivered into cells by a three-component nanoparticle is an effective vehicle for Auger radiotherapy when radiolabeled with 111In or 125I.

In vivo delivery of antisense MORF oligomer by MORF/carrier streptavidin nanoparticles

Tue, 02/10/2015 - 10:11pm

Tumor targeting by oligomers is largely limited by the pharmacokinetics and cell-membrane transport obstacles. In this article, we describe the use of a delivery nanoparticle, in which streptavidin served as a convenient bridge between a biotinylated oligomer and a biotinylated cell-membrane-penetrating peptide, to improve the delivery of an antisense phosphorodiamidate morpholino (MORF) oligomer in vivo. A biotinylated (99m)Tc-radiolabeled MORF oligomer with a base sequence antisense to the RIalpha mRNA and its sense control were incorporated separately into nanoparticles, along with biotinylated tat or polyarginine carrier. The streptavidin nanoparticles were administrated intravenously to both normal and nude mice bearing SUM149 breast tumor xenografts. The biodistributions showed much higher normal tissue levels for the radiolabeled MORFs, independent of antisense or sense or tat or polyarginine, when administered as the nanoparticles, compared to naked. A statistically significant higher accumulation of both antisense nanoparticles, compared to the respective sense control nanoparticles, was observed, along with much higher tumor accumulations, compared to historical naked controls. This study has provided evidence that the in vivo function of an antisense oligomer within the streptavidin nanoparticle is not impeded, and, as such, the MORF/streptavidin/carrier nanoparticles may be suitable for in vivo tumor delivery of antisense MORF and other oligomers.

Tumor delivery of antisense oligomer using trastuzumab within a streptavidin nanoparticle

Tue, 02/10/2015 - 10:11pm

PURPOSE: Trastuzumab (Herceptin) is often internalized following binding to Her2+ tumor cells. The objective of this study was to investigate whether trastuzumab can be used as a specific carrier to deliver antisense oligomers into Her2+ tumor cells both in vitro and in vivo.

METHODS: A biotinylated MORF oligomer antisense to RhoC mRNA and its biotinylated sense control were labeled with either lissamine for fluorescence detection or 99mTc for radioactivity detection and were linked to biotinylated trastuzumab via streptavidin. The nanoparticles were studied in SUM190 (RhoC+, Her2+) study and SUM149 (RhoC+, Her2-) control cells in culture and as xenografts in mice.

RESULTS: As evidence of unimpaired Her2+ binding of trastuzumab within the nanoparticle, accumulations were clearly higher in SUM190 compared to SUM149 cells and, by whole-body imaging, targeting of SUM190 tumor was similar to that expected for a radiolabeled trastuzumab. As evidence of internalization, fluorescence microscopy images of cells grown in culture and obtained from xenografts showed uniform cytoplasm distribution of the lissamine-MORF. An invasion assay showed decreased RhoC expression in SUM190 cells when incubated with the antisense MORF nanoparticles at only 100 nM.

CONCLUSION: Both in cell culture and in animals, the nanoparticle with trastuzumab as specific carrier greatly improved tumor delivery of the antisense oligomer against RhoC mRNA into tumor cells overexpressing Her2 and may be of general utility.

Auger radiation-induced, antisense-mediated cytotoxicity of tumor cells using a 3-component streptavidin-delivery nanoparticle with 111In

Tue, 02/10/2015 - 10:11pm

When antisense oligomers are intracellular, they migrate to and are retained in the nucleus of tumor cells and therefore may be used to carry Auger electron-emitting radionuclides such as (111)In for effective tumor radiotherapy.

METHODS: Our nanoparticle consists of streptavidin that links 3 biotinylated components: the antiHer2 antibody trastuzumab (to improve pharmacokinetics), the tat peptide (to improve cell membrane transport), and the (111)In-labeled antiRIalpha messenger RNA antisense morpholino (MORF) oligomer.

RESULTS: As evidence of unimpaired function, tumor cell and nuclear accumulations were orders of magnitude higher after incubation with (99m)Tc-MORF/tat/trastuzumab than after incubation with free (99m)Tc-MORF and significantly higher with the antisense than with the sense MORF. In mice, tumor and normal-tissue accumulations of the (99m)Tc-MORF/tat/trastuzumab nanoparticle were comparable to those of free (99m)Tc-trastuzumab, confirming the improved pharmacokinetics due to the trastuzumab component. Although kidneys, liver, and other normal tissues also accumulated the nanoparticle, immunohistochemical evaluation of tissue sections in mice receiving the Cy3-MORF/tat/trastuzumab nanoparticle showed evidence of nuclear accumulation only in tumor tissue. In a dose escalation study, as measured by the surviving fraction, the nanoparticle significantly increased the kill of SK-BR-3 breast cancer Her2+/RIalpha+ cells, compared with all controls.

CONCLUSION: Significant radiation-induced antisense-mediated cytotoxicity of tumor cells in vitro was achieved using an Auger electron-emitting antisense MORF oligomer administered as a member of a 3-component streptavidin-delivery nanoparticle.

Community health worker interventions for Latinos with type 2 diabetes: a systematic review of randomized controlled trials

Tue, 02/10/2015 - 5:28pm

This systematic review aimed to synthesize glucose (HbA1c) outcomes of community health worker (CHW)-delivered interventions for Latinos with type 2 diabetes that were tested in randomized controlled trials and to summarize characteristics of the targeted populations and interventions, including the background, training, and supervision of the CHWs. Searches of PubMed and Google Scholar databases and references from selected articles identified 12 studies that met the inclusion criteria. Of these, seven reported statistically significant improvements in HbA1c. Study participants were largely low-income, female, and Spanish-speaking and had uncontrolled diabetes. The CHWs led the interventions alone, in pairs, or as part of a team. Interventions varied considerably in session time, duration, and number. Most met standards for tailored, high-intensity interventions and half were theory-based. Overall, methodological quality was good but there were inconsistencies in the reporting of key information. Future research should report in greater detail CHW background, training, and supervision; examine factors associated with intervention effectiveness; and provide data on cost and cost-effectiveness.