Poison to Promise: The Resurgence of Organophosphorus Fluoride Chemistry.

Organophosphorus(V) fluorides have a long and tumultuous history, with early applications as toxins and nerve agents reflecting their poisonous past. Behind these very real safety considerations, there is also growing potential in a wide range of fields, from chemical biology to drug development. The recent inclusion of organophosphorus(V) fluorides in click chemistry exemplifies the promise these compounds possess and brings these molecules to the brink of a resurgence. In this Perspective, we delve into the history of P(V)-F compounds, discuss the precautions needed to work with them safely, and explore recent advancements in their synthesis and application. We conclude by discussing how this field can continue on a path toward innovation.

Thionyl Fluoride-Mediated One-Pot Substitutions and Reductions of Carboxylic Acids.

Thionyl fluoride (SOF2) is an underutilized reagent that is yet to be extensively studied for its synthetic applications. We previously reported that it is a powerful reagent for both the rapid syntheses of acyl fluorides and for one-pot peptide couplings, but the full scope of these nucleophilic acyl substitutions had not been explored. Herein, we report one-pot thionyl fluoride-mediated syntheses of peptides and amides (35 examples, 45-99% yields) that were not explored in our previous study. The scope of thionyl fluoride-mediated nucleophilic acyl substitutions was also expanded to encompass esters (24 examples, 64-99% yields) and thioesters (11 examples, 24-96% yields). In addition, we demonstrate that the scope of thionyl fluoride-mediated one-pot reactions can be extended beyond nucleophilic acyl substitutions to mild reductions of carboxylic acids using NaBH4 (13 examples, 33-80% yields).

Therapeutic resistance in breast cancer cells can result from deregulated EGFR signaling.

The epidermal growth factor receptor (EGFR) interacts with various downstream molecules including phospholipase C (PLC)/protein kinase C (PKC), Ras/Raf/MEK/ERK, PI3K/PTEN/Akt/GSK-3, Jak/STAT and others. Often these pathways are deregulated in human malignancies such as breast cancer. Various therapeutic approaches to inhibit the activity of EGFR family members including small molecule inhibitors and monoclonal antibodies (MoAb) have been developed. A common problem with cancer treatments is the development of drug-resistance. We examined the effects of a conditionally-activated EGFR (v-Erb-B:ER) on the resistance of breast cancer cells to commonly used chemotherapeutic drugs such as doxorubicin, daunorubicin, paclitaxel, cisplatin and 5-flurouracil as well as ionizing radiation (IR). v-Erb-B is similar to the EGFR-variant EGFRvIII, which is expressed in various cancers including breast, brain, prostate. Both v-Erb-B and EGFRvIII encode the EGFR kinase domain but lack key components present in the extracellular domain of EGFR which normally regulate its activity and ligand-dependence. The v-Erb-B oncogene was ligated to the hormone binding domain of the estrogen receptor (ER) which results in regulation of the activity of the v-Erb-ER construct by addition of either estrogen (E2) or 4-hydroxytamoxifen (4HT) to the culture media. Introduction of the v-Erb-B:ER construct into the MCF-7 breast cancer cell line increased the resistance to the cells to various chemotherapeutic drugs, hormonal-based therapeutics and IR. These results point to the important effects that aberrant expression of EGFR kinase domain can have on therapeutic resistance.

Influences of TP53 and the anti-aging DDR1 receptor in controlling Raf/MEK/ERK and PI3K/Akt expression and chemotherapeutic drug sensitivity in prostate cancer cell lines.

BACKGROUND: TP53 plays critical roles in sensitivity to chemotherapy, and aging. Collagen is very important in aging. The molecular structure and biochemical properties of collagen changes during aging. The discoidin domain receptor (DDR1) is regulated in part by collagen. Elucidating the links between TP53 and DDR1 in chemosensitivity and aging could improve therapies against cancer and aging. RESULTS: Restoration of WT-TP53 activity resulted in increased sensitivity to chemotherapeutic drugs and elevated expression of key components of the Raf/MEK/ERK, PI3K/Akt and DDR1 pathways. DDR1 could modulate the levels of Raf/MEK/ERK and PI3K/Akt pathways as well as sensitize the cells to chemotherapeutic drugs. In contrast, suppression of WT TP53 with a dominant negative (DN) TP53 gene, suppressed DDR1 protein levels and increased their chemoresistance. CONCLUSION: Restoration of WT TP53 activity or increased expression of the anti-aging DDR1 collagen receptor can result in enhanced sensitivity to chemotherapeutic drugs. Our innovative studies indicate the important links between WT TP53 and DDR1 which can modulate Raf/MEK/ERK and PI3K/Akt signaling as well as chemosensitivity and aging. METHODS: We investigated the roles of wild type (WT) and mutant TP53 on drug sensitivity of prostate cancer cells and the induction of Raf/MEK/ERK, PI3K/Akt and DDR1 expression and chemosensitivity.

Roles of p53, NF-κB and the androgen receptor in controlling NGAL expression in prostate cancer cell lines.

Neutrophil gelatinase-associated lipocalin (NGAL a.k.a lipocalin 2, lnc2) is a secreted protein which can form a complex with matrix metalloproteinase-9 (MMP9). This MMP9/NGAL complex has been associated with metastasis. MMP9 and NGAL are detected in the urine of patients afflicted with many different types of cancer, including prostate cancer. The effects of p53, NF-κB and the androgen receptor (AR) on the expression of NGAL was examined in four prostate cancer cell lines. Prostate cancer cell lines that are AR negative and expressed either mutant or no p53 (DU145 and PC3) displayed higher levels of NGAL expression compared to the prostate cancer cell lines (LNCaP and 22Rv-1) which are AR positive and express wild type (WT) p53. Introduction of WT-p53 into the PC3 prostate cancer cell line, resulted in reduction of the levels of NGAL expression. Conversely, introduction of dominant negative (DN) p53 or a retroviral construct expressing NF-κB into LNCaP cells increased NGAL expression. NGAL expression had functional effects on the ability of the cells to form colonies in soft agar. Whereas suppression of WT-53 in LNCaP cells increased NGAL expression, the introduction of WT-p53 suppressed NGAL transcription activity in PC3 prostate cells which normally express high level of NGAL. NF-κB and p53 were determined to regulate NGAL expression by positive and negative mechanisms, respectively. Our data indicate that prostate cancer growth, progression and sensitivity to chemotherapeutic drugs are regulated in part by NGAL and may involve complex interactions between NGAL, MMP9, NF-κB and p53.

Efforts of a Unit Practice Council to implement practice change utilizing alcohol impregnated port protectors in a burn ICU.

BACKGROUND: Burn patients are an especially high-risk population for development of central line associated bloodstream infections (CLABSI) due to open wounds, extended length of intensive care unit stay, frequent use of central venous catheters, and generally immunocompromised state. Implementing evidence-based practices to prevent these infections is a 2014 National Patient Safety Goal per The Joint Commission. OBJECTIVES: The purpose of this project was introduction of a commercially available alcohol impregnated central venous line port protector to reduce the incidence of CLABSI in the burn unit. METHODS: The Iowa Model for Implementing Evidenced-Based Practice was used to guide this intervention conducted by the Unit Practice Council. A pre- and post-intervention design compared rates of CLABSI before and after introduction of the port protectors. RESULTS: CLABSI infection rates decreased following the intervention from baseline of 7.3 per 1000 line days to an average of 3.04 per 1000 line days during calendar year 2013. CONCLUSIONS: Introduction of an alcohol impregnated central venous line port protector can reduce the incidence of CLABSI in a burn unit.

Novel roles of androgen receptor, epidermal growth factor receptor, TP53, regulatory RNAs, NF-kappa-B, chromosomal translocations, neutrophil associated gelatinase, and matrix metalloproteinase-9 in prostate cancer and prostate cancer stem cells.

Approximately one in six men will be diagnosed with some form of prostate cancer in their lifetime. Over 250,000 men worldwide die annually due to complications from prostate cancer. While advancements in prostate cancer screening and therapies have helped in lowering this statistic, better tests and more effective therapies are still needed. This review will summarize the novel roles of the androgen receptor (AR), epidermal growth factor receptor (EGFR), the EGFRvIII variant, TP53, long-non-coding RNAs (lncRNAs), microRNAs (miRs), NF-kappa-B, chromosomal translocations, neutrophil associated gelatinase, (NGAL), matrix metalloproteinase-9 (MMP-9), the tumor microenvironment and cancer stem cells (CSC) have on the diagnosis, development and treatment of prostate cancer.

Homologous Recombination Repair Factors Rad51 and BRCA1 Are Necessary for Productive Replication of Human Papillomavirus 31.

UNLABELLED: High-risk human papillomavirus 31 (HPV31)-positive cells exhibit constitutive activation of the ATM-dependent DNA damage response (DDR), which is necessary for productive viral replication. In response to DNA double-strand breaks (DSBs), ATM activation leads to DNA repair through homologous recombination (HR), which requires the principal recombinase protein Rad51, as well as BRCA1. Previous studies from our lab demonstrated that Rad51 and BRCA1 are expressed at high levels in HPV31-positive cells and localize to sites of viral replication. These results suggest that HPV may utilize ATM activity to increase HR activity as a means to facilitate viral replication. In this study, we demonstrate that high-risk HPV E7 expression alone is sufficient for the increase in Rad51 and BRCA1 protein levels. We have found that this increase occurs, at least in part, at the level of transcription. Studies analyzing protein stability indicate that HPV may also protect Rad51 and BRCA1 from turnover, contributing to the overall increase in cellular levels. We also demonstrate that Rad51 is bound to HPV31 genomes, with binding increasing per viral genome upon productive replication. We have found that depletion of Rad51 and BRCA1, as well as inhibition of Rad51's recombinase activity, abrogates productive viral replication upon differentiation. Overall, these results indicate that Rad51 and BRCA1 are required for the process of HPV31 genome amplification and suggest that productive replication occurs in a manner dependent upon recombination. IMPORTANCE: Productive replication of HPV31 requires activation of an ATM-dependent DNA damage response, though how ATM activity contributes to replication is unclear. Rad51 and BRCA1 play essential roles in repair of double-strand breaks, as well as the restart of stalled replication forks through homologous recombination (HR). Given that ATM activity is required to initiate HR repair, coupled with the requirement of Rad51 and BRCA1 for productive viral replication, our findings suggest that HPV may utilize ATM activity to ensure localization of recombination factors to productively replicating viral genomes. The finding that E7 increases the levels of Rad51 and BRCA1 suggests that E7 contributes to productive replication by providing DNA repair factors required for viral DNA synthesis. Our studies not only imply a role for recombination in the regulation of productive HPV replication but provide further insight into how HPV manipulates the DDR to facilitate the productive phase of the viral life cycle.

Productive replication of human papillomavirus 31 requires DNA repair factor Nbs1.

UNLABELLED: Activation of the ATM (ataxia telangiectasia-mutated kinase)-dependent DNA damage response (DDR) is necessary for productive replication of human papillomavirus 31 (HPV31). We previously found that DNA repair and homologous recombination (HR) factors localize to sites of HPV replication, suggesting that ATM activity is required to recruit factors to viral genomes that can productively replicate viral DNA in a recombination-dependent manner. The Mre11-Rad50-Nbs1 (MRN) complex is an essential component of the DDR that is necessary for ATM-mediated HR repair and localizes to HPV DNA foci. In this study, we demonstrate that the HPV E7 protein is sufficient to increase levels of the MRN complex and also interacts with MRN components. We have found that Nbs1 depletion blocks productive viral replication and results in decreased localization of Mre11, Rad50, and the principal HR factor Rad51 to HPV DNA foci upon differentiation. Nbs1 contributes to the DDR by acting as an upstream activator of ATM in response to double-strand DNA breaks (DSBs) and as a downstream effector of ATM activity in the intra-S-phase checkpoint. We have found that phosphorylation of ATM and its downstream target Chk2, as well as SMC1 (structural maintenance of chromosome 1), is maintained upon Nbs1 knockdown in differentiating cells. Given that ATM and Chk2 are required for productive replication, our results suggest that Nbs1 contributes to viral replication outside its role as an ATM activator, potentially through ensuring localization of DNA repair factors to viral genomes that are necessary for efficient productive replication. IMPORTANCE: The mechanisms that regulate human papillomavirus (HPV) replication during the viral life cycle are not well understood. Our finding that Nbs1 is necessary for productive replication even in the presence of ATM (ataxia telangiectasia-mutated kinase) and Chk2 phosphorylation offers evidence that Nbs1 contributes to viral replication downstream of facilitating ATM activation. Nbs1 is required for the recruitment of Mre11 and Rad50 to viral genomes, suggesting that the MRN complex plays a direct role in facilitating productive viral replication, potentially through the processing of substrates that are recognized by the key homologous recombination (HR) factor Rad51. The discovery that E7 increases levels of MRN components, and MRN complex formation, identifies a novel role for E7 in facilitating productive replication. Our study not only identifies DNA repair factors necessary for HPV replication but also provides a deeper understanding of how HPV utilizes the DNA damage response to regulate viral replication.

Inhibition of GSK-3ß activity can result in drug and hormonal resistance and alter sensitivity to targeted therapy in MCF-7 breast cancer cells.

The PI3K/Akt/mTORC1 pathway plays prominent roles in malignant transformation, prevention of apoptosis, drug resistance, and metastasis. One molecule regulated by this pathway is GSK-3ß. GSK-3ß is phosphorylated by Akt on S9, which leads to its inactivation; however, GSK-3ß also can regulate the activity of the PI3K/Akt/mTORC1 pathway by phosphorylating molecules such as PTEN, TSC2, p70S6K, and 4E-BP1. To further elucidate the roles of GSK-3ß in chemotherapeutic drug and hormonal resistance of MCF-7 breast cancer cells, we transfected MCF-7 breast cancer cells with wild-type (WT), kinase-dead (KD), and constitutively activated (A9) forms of GSK-3ß. MCF-7/GSK-3ß(KD) cells were more resistant to doxorubicin and tamoxifen compared with either MCF-7/GSK-3ß(WT) or MCF-7/GSK-3ß(A9) cells. In the presence and absence of doxorubicin, the MCF-7/GSK-3ß(KD) cells formed more colonies in soft agar compared with MCF-7/GSK-3ß(WT) or MCF-7/GSK-3ß(A9) cells. In contrast, MCF-7/GSK-3ß(KD) cells displayed an elevated sensitivity to the mTORC1 blocker rapamycin compared with MCF-7/GSK-3ß(WT) or MCF-7/GSK-3ß(A9) cells, while no differences between the 3 cell types were observed upon treatment with a MEK inhibitor by itself. However, resistance to doxorubicin and tamoxifen were alleviated in MCF-7/GSK-3ß(KD) cells upon co-treatment with an MEK inhibitor, indicating regulation of this resistance by the Raf/MEK/ERK pathway. Treatment of MCF-7 and MCF-7/GSK-3ß(WT) cells with doxorubicin eliminated the detection of S9-phosphorylated GSK-3ß, while total GSK-3ß was still detected. In contrast, S9-phosphorylated GSK-3ß was still detected in MCF-7/GSK-3ß(KD) and MCF-7/GSK-3ß(A9) cells, indicating that one of the effects of doxorubicin on MCF-7 cells was suppression of S9-phosphorylated GSK-3ß, which could result in increased GSK-3ß activity. Taken together, these results demonstrate that introduction of GSK-3ß(KD) into MCF-7 breast cancer cells promotes resistance to doxorubicin and tamoxifen, but sensitizes the cells to mTORC1 blockade by rapamycin. Therefore GSK-3ß is a key regulatory molecule in sensitivity of breast cancer cells to chemo-, hormonal, and targeted therapy.

Signaling intermediates (MAPK and PI3K) as therapeutic targets in NSCLC.

The RAS/RAF/MEK/ ERK and the PI3K/AKT/mTOR pathways govern fundamental physiological processes, such as cell proliferation, differentiation, metabolism, cytoskeleton reorganization and cell death and survival. Constitutive activation of these signal transduction pathways is a required hallmark of cancer and dysregulation, on either genetic or epigenetic grounds, of these pathways has been implicated in the initiation, progression and metastastic spread of lung cances. Targeting components of the MAPK and PI3K cascades is thus an attractive strategy in the development of novel therapeutic approaches to treat lung cancer, although the use of single pathway inhibitors has met with limited clinical success so far. Indeed, the presence of intra- and inter-pathway compensatory loops that re-activate the very same cascade, either upstream or downstream the point of pharmacological blockade, or activate the alternate pathway following the blockade of one signaling cascade has been demonstrated, potentially driving preclinical (and possibly clinical) resistance. Therefore, the blockade of both pathways with combinations of signaling inhibitors might result in a more efficient anti-tumor effect, and thus potentially overcome and/or delay clinical resistance, as compared with single agent. The current review aims at summarizing the current status of preclinical and clinical research with regard to pathway crosstalks between the MAPK and PI3K cascades in NSCLC and the rationale for combined therapeutic pathway targeting.

p53 expression controls prostate cancer sensitivity to chemotherapy and the MDM2 inhibitor Nutlin-3.

Prostate cancer is the second most commonly diagnosed cancer in men, and approximately one-third of those diagnosed succumb to the disease. The development of prostate cancer from small regions of hyperplasia to invasive tumors requires genetic and epigenetic alterations of critical cellular components to aid in the development of cells more adapted for aberrant growth. The p53 transcription factor is a critical element in the cell's ability to regulate the cell cycle and its response to DNA damage. Mutations within the DNA-binding domain of p53 are common and allow the formation of tetramers; however, these alterations prevent this protein complex from associating with target gene promoters. In the present study, we examined the effects of p53 functionality in prostate cancer cells that harbored wild-type (WT) or mutant forms of the protein in response to commonly used chemotherapeutic drugs. The androgen receptor positive 22Rv-1 and LNCaP prostate cancer cell lines carry WT p53 and were demonstrated to have a decrease in chemotherapeutic drug sensitivity when transfected with a dominant-negative (DN) p53. Conversely, expression of the WT p53 in the p53-mutated and more advanced DU145 prostate cancer cell line significantly increased its overall sensitivity to anti-neoplastic drugs. Furthermore, analysis of colony formation in soft agar revealed that the functional status of p53 in each cell line altered the cell's ability to proliferate in an anchorage-independent fashion. Prostate cancer colony growth was more prevalent when p53 transcriptional activity was decreased, whereas growth was more limited in the presence of functional p53. These results demonstrate that the functional status of the tumor suppressor p53 is important in the progression of prostate cancer and dictates the overall effectiveness a given drug would have on disease treatment.

Ectopic NGAL expression can alter sensitivity of breast cancer cells to EGFR, Bcl-2, CaM-K inhibitors and the plant natural product berberine.

Neutrophil gelatinase-associated lipocalin (NGAL, a.k.a Lnc2) is a member of the lipocalin family and has diverse roles. NGAL can stabilize matrix metalloproteinase-9 from autodegradation. NGAL is considered as a siderocalin that is important in the transport of iron. NGAL expression has also been associated with certain neoplasias and is implicated in the metastasis of breast cancer. In a previous study, we examined whether ectopic NGAL expression would alter the sensitivity of breast epithelial, breast and colorectal cancer cells to the effects of the chemotherapeutic drug doxorubicin. While abundant NGAL expression was detected in all the cells infected with a retrovirus encoding NGAL, this expression did not alter the sensitivity of these cells to doxorubicin as compared with empty vector-transduced cells. We were also interested in determining the effects of ectopic NGAL expression on the sensitivity to small-molecule inhibitors targeting key signaling molecules. Ectopic NGAL expression increased the sensitivity of MCF-7 breast cancer cells to EGFR, Bcl-2 and calmodulin kinase inhibitors as well as the natural plant product berberine. Furthermore, when suboptimal concentrations of certain inhibitors were combined with doxorubicin, a reduction in the doxorubicin IC 50 was frequently observed. An exception was observed when doxorubicin was combined with rapamycin, as doxorubicin suppressed the sensitivity of the NGAL-transduced MCF-7 cells to rapamycin when compared with the empty vector controls. In contrast, changes in the sensitivities of the NGAL-transduced HT-29 colorectal cancer cell line and the breast epithelial MCF-10A cell line were not detected compared with empty vector-transduced cells. Doxorubicin-resistant MCF-7/Dox (R) cells were examined in these experiments as a control drug-resistant line; it displayed increased sensitivity to EGFR and Bcl-2 inhibitors compared with empty vector transduced MCF-7 cells. These results indicate that NGAL expression can alter the sensitivity of certain cancer cells to small-molecule inhibitors, suggesting that patients whose tumors exhibit elevated NGAL expression or have become drug-resistant may display altered responses to certain small-molecule inhibitors.

Effects of ectopic expression of NGAL on doxorubicin sensitivity.

Neutrophil gelatinase-associated lipocalin (NGAL, a.k.a Lnc2) is a member of the lipocalin family which has diverse roles including stabilizing matrix metalloproteinase-9 from auto-degradation and as siderocalins which are important in the transport of iron. NGAL also has important biological functions involved in immunity and inflammation as well as responses to kidney damage. NGAL expression has also been associated with certain neoplasia and is important in the metastasis of breast cancer. Many advanced cancer patients have elevated levels of NGAL in their urine and it has been proposed that NGAL may be a prognostic indicator for certain cancers (e.g. breast, brain, and others). NGAL expression is detected in response to various chemotherapeutic drugs including doxorubicin and docetaxel. We were interested in the roles of NGAL expression in cancer and whether it is associated with chemotherapeutic drug resistance. In the present study, we investigated whether increased NGAL expression led to resistance to the chemotherapeutic drug doxorubicin in normal breast epithelial cells (MCF-10A), breast cancer cells (MCF-7), and colorectal cancer cells (HT-29). We infected the various cell lines with a retrovirus encoding NGAL which we constructed. Increased NGAL expression was readily detected in the NGAL-infected cells but not the empty vector-infected cells. However, increased NGAL expression did not alter the sensitivity of the cells to the chemotherapeutic drug doxorubicin. Thus, although NGAL expression is often detected after chemotherapeutic drug treatment, it by itself, does not lead to doxorubicin resistance.

Ras/Raf/MEK/ERK and PI3K/PTEN/Akt/mTOR cascade inhibitors: how mutations can result in therapy resistance and how to overcome resistance.

The Ras/Raf/MEK/ERK and PI3K/PTEN/Akt/mTOR cascades are often activated by genetic alterations in upstream signaling molecules such as receptor tyrosine kinases (RTK). Targeting these pathways is often complex and can result in pathway activation depending on the presence of upstream mutations (e.g., Raf inhibitors induce Raf activation in cells with wild type (WT) RAF in the presence of mutant, activated RAS) and rapamycin can induce Akt activation. Targeting with inhibitors directed at two constituents of the same pathway or two different signaling pathways may be a more effective approach. This review will first evaluate potential uses of Raf, MEK, PI3K, Akt and mTOR inhibitors that have been investigated in pre-clinical and clinical investigations and then discuss how cancers can become insensitive to various inhibitors and potential strategies to overcome this resistance.

Mutations and deregulation of Ras/Raf/MEK/ERK and PI3K/PTEN/Akt/mTOR cascades which alter therapy response.

The Ras/Raf/MEK/ERK and PI3K/PTEN/Akt/mTOR cascades are often activated by genetic alterations in upstream signaling molecules such as receptor tyrosine kinases (RTK). Certain components of these pathways, RAS, NF1, BRAF, MEK1, DUSP5, PP2A, PIK3CA, PIK3R1, PIK3R4, PIK3R5, IRS4, AKT, NFKB1, MTOR, PTEN, TSC1, and TSC2 may also be activated/inactivated by mutations or epigenetic silencing. Upstream mutations in one signaling pathway or even in downstream components of the same pathway can alter the sensitivity of the cells to certain small molecule inhibitors. These pathways have profound effects on proliferative, apoptotic and differentiation pathways. Dysregulation of components of these cascades can contribute to: resistance to other pathway inhibitors, chemotherapeutic drug resistance, premature aging as well as other diseases. This review will first describe these pathways and discuss how genetic mutations and epigenetic alterations can result in resistance to various inhibitors.

Targeting the cancer initiating cell: the ultimate target for cancer therapy.

An area of therapeutic interest in cancer biology and treatment is targeting the cancer stem cell, more appropriately referred to as the cancer initiating cell (CIC). CICs comprise a subset of hierarchically organized, rare cancer cells with the ability to initiate cancer in xenografts in genetically modified murine models. CICs are thought to be responsible for tumor onset, self-renewal/maintenance, mutation accumulation and metastasis. CICs may lay dormant after various cancer therapies which eliminate the more rapidly proliferating bulk cancer (BC) mass. However, CICs may remerge after therapy is discontinued as they may represent cells which were either intrinsically resistant to the original therapeutic approach or they have acquired mutations which confer resistance to the primary therapy. In experimental mouse tumor transplant models, CICs have the ability to transfer the tumor to immunocompromised mice very efficiently while the BCs are not able to do so as effectively. Often CICs display increased expression of proteins involved in drug resistance and hence they are intrinsically resistant to many chemotherapeutic approaches. Furthermore, the CICs may be in a suspended state of proliferation and not sensitive to common chemotherapeutic and radiological approaches often employed to eliminate the rapidly proliferating BCs. Promising therapeutic approaches include the targeting of certain signal transduction pathways (e.g., RAC, WNT, PI3K, PML) with small molecule inhibitors or targeting specific cell-surface molecules (e.g., CD44), with effective cytotoxic antibodies. The existence of CICs could explain the high frequency of relapse and resistance to many currently used cancer therapies. New approaches should be developed to effectively target the CIC which could vastly improve cancer therapies and outcomes. This review will discuss recent concepts of targeting CICs in certain leukemia models.

Paper spray ionization devices for direct, biomedical analysis using mass spectrometry.

Paper spray ionization has been developed as a direct, fast and low-cost sampling and ionization method for qualitative and quantitative mass spectrometric (MS) analysis of complex mixtures. Analyte ions are generated by applying a high voltage and a small volume (~10 µL) of spray solvent onto a porous substrate. The sample can be preloaded onto the paper or mixed into the spray solution. The geometry of the paper and the method of supplying the necessary internal standard are important factors that affect the ionization efficiency and subsequently the sensitivity and quantitation accuracy of the analytical data. As the cut angle of the paper tip is changed, the spray plume, the total spray current and the electric field intensity at the tip all vary correspondingly, with resulting differences in signal intensity. Sample load is another important factor for obtaining a stable MS signal and accurate quantitative results. The optimal sample load was found to be dependent on the paper size. The dissolution and spray process was also investigated and analyte transfer on paper was shown to be largely associated with bulk solution flow towards the spray tip. The information gathered from these systematic studies provides guidance for the design and optimization of a disposable sample cartridge for paper spray MS, a device which potentially is suitable for fast clinical analysis, especially for point-of-care diagnostics.

Polymer-based miniature flexible capacitive pressure sensor for intraocular pressure (IOP) monitoring inside a mouse eye.

This paper presents an ultra-thin and flexible polymer-based capacitive pressure sensor for intraocular pressure (IOP) monitoring in a mouse eye. Due to the size limitation of the anterior chamber in the mouse eye, a volume of approximately 700 × 700 × 150 µm(3) on a small substrate is available for the MEMS capacitive pressure sensor. Moreover, the sensor would ideally be easily injectable into place. Further complicating the sensing is the need to operate the device on the curved surface of the anterior chamber with minimum damage to the eye tissue. Therefore, a thin and flexible substrate is required. We fabricate sensors by exploiting Parylene as a biocompatible structural material in a suitable form factor and 25 µm thick liquid crystal polymer (LCP) as a soft and flexible host substrate. Using our approach, the flexibility and small form factor necessary for a mouse eye implant is achieved, along with the sensitivity required to monitor IOP fluctuations. This device will allow better study of glaucoma through close monitoring in mice after integration with other components.