Synergistic effects of nelfinavir and bortezomib on proteotoxic death of NSCLC and multiple myeloma cells.

Exploiting protein homeostasis is a new therapeutic approach in cancer. Nelfinavir (NFV) is an HIV protease inhibitor that induces endoplasmic reticulum (ER) stress in cancer cells. Under conditions of ER stress, misfolded proteins are transported from the ER back to the cytosol for subsequent degradation by the ubiquitin-proteasome system. Bortezomib (BZ) is a proteasome inhibitor and interferes with degradation of misfolded proteins. Here, we show that NFV and BZ enhance proteotoxicity in non-small cell lung cancer (NSCLC) and multiple myeloma (MM) cells. The combination synergistically inhibited cell proliferation and induced cell death. Activating transcription factor (ATF)3 and CCAAT-enhancer binding protein homologous protein (CHOP), markers of ER stress, were rapidly increased, and their siRNA-mediated knockdown inhibited cell death. Knockdown of double-stranded RNA activated protein kinase-like ER kinase, a signal transducer in ER stress, significantly decreased apoptosis. Pretreatment with the protein synthesis inhibitor, cycloheximide, decreased levels of ubiquitinated proteins, ATF3, CHOP, and the overall total cell death, suggesting that inhibition of protein synthesis increases cell survival by relieving proteotoxic stress. The NFV/BZ combination inhibited the growth of NSCLC xenografts, which correlated with the induction of markers of ER stress and apoptosis. Collectively, these data show that NFV and BZ enhance proteotoxicity in NSCLC and MM cells, and suggest that this combination could tip the precarious balance of protein homeostasis in cancer cells for therapeutic gain.

Ceramide mediates nanovesicle shedding and cell death in response to phosphatidylinositol ether lipid analogs and perifosine.

Anticancer phospholipids that inhibit Akt such as the alkylphospholipid perifosine (Per) and phosphatidylinositol ether lipid analogs (PIAs) promote cellular detachment and apoptosis and have a similar cytotoxicity profile against cancer cell lines in the NCI60 panel. While investigating the mechanism of Akt inhibition, we found that short-term incubation with these compounds induced rapid shedding of cellular nanovesicles containing EGFR, IGFR and p-Akt that occurred in vitro and in vivo, while prolonged incubation led to cell detachment and death that depended on sphingomyelinase-mediated generation of ceramide. Pretreatment with sphingomyelinase inhibitors blocked ceramide generation, decreases in phospho-Akt, nanovesicle release and cell detachment in response to alkylphospholipids and PIAs in non-small cell lung cancer cell lines. Similarly, exogenous ceramide also decreased active Akt and induced nanovesicle release. Knockdown of neutral sphingomyelinase decreased, whereas overexpression of neutral or acid sphingomyelinase increased cell detachment and death in response to the compounds. When transferred in vitro, PIA or Per-induced nanovesicles increased ceramide levels and death in recipient cells. These results indicate ceramide generation underlies the Akt inhibition and cytotoxicity of this group of agents, and suggests nanovesicle shedding and uptake might potentially propagate their cytotoxicity in vivo.

Prospective multicenter trial assessing effectiveness, refractive predictability and safety of a new aberration free, bi-aspheric intraocular lens.

PURPOSE: To determine the effectiveness and safety of the Softec HD IOL; and to present refractive outcomes for lenses manufactured at an IOL power tolerance of 0.11 D. METHODS: Three-hundred and ninety adult patients requiring removal of a cataractous lens with implantation of a monofocal IOL in at least one eye were eligible for study participation across eight US investigative sites. Patients were enrolled unilaterally. After routine surgery, subjects were examined for adverse events (AEs), best corrected visual acuity (BCVA) and manifest refraction correction at 12 months postoperatively. RESULTS: Three-hundred and sixty-six (95%) of patients completed the 12-month postoperative visit. The percent of patients achieving best corrected Snellen acuity 20/40 or better was 98.9%, and 81.1% of patients achieved best corrected Snellen acuity 20/25 or better. Of those patients (80%) implanted with a lens available in 0.25 D increments (manufactured at a tolerance of 0.11 D) 40.9%, 69.8% and 93.8% of patients were within ±0.25 D, ±0.50 D and ±1.0 D of predicted target refraction respectively. Overall incidence of cumulative and persistent IOL Grid AEs was 2.2% with no AE meeting or exceeding the FDA Grid of Historical Controls. CONCLUSIONS: The Softec HD IOL is a safe and effective lens. The high manufacturing tolerance of the lens appears to enhance refractive outcomes.

Effect of deguelin on UVB-induced skin carcinogenesis.

Nonmelanoma skin cancer afflicts more than one million people in the U.S. annually, highlighting the need for more effective preventive regimens. We have investigated the ability of deguelin, a plant-derived rotenoid with cancer chemopreventive activity, to inhibit UVB-induced skin carcinogenesis with the SKh-1 mouse model. Topically-applied deguelin significantly inhibited the multiplicity of UVB-induced skin tumors, indicating potential as a human skin cancer chemopreventive agent. Mechanistic studies to determine the potential of deguelin to block a number of established UVB-induced molecular events yielded negative results [including UVB-induced AP-1 DNA binding, c-fos and TNFalpha mRNA induction, arachidonic acid release and UVB-induced phosphorylation of mTOR (Ser2448), akt (Ser473) and erk (Thr202/Tyr204)]. These results are of interest as they contradict a major hypothesis for the mode of action of deguelin, i.e., a general down regulation of signal transduction based on inhibition of NADH dehydrogenase and depletion of ATP levels. In the current work, however, deguelin was found to activate 5' AMP-activated kinase (AMPK), a protein that acts as a cellular energy sensor. This is the first report of a chemopreventive agent having this effect and suggests a possible role for AMPK in cancer chemoprevention.

Activity-guided isolation of constituents of Cerbera manghas with antiproliferative and antiestrogenic activities.

Two new cardenolides, (-)-14-hydroxy-3beta-(3-O-methyl-6-deoxy-alpha-L-rhamnosyl)-11a lpha, 12alpha-epoxy-(5beta,14beta,17betaH)-card-20 (22)-enolide (1), (-)-14-hydroxy-3beta-(3-O-methyl-6-deoxy-alpha-L-glucopyranosyl)-11al pha,12alpha-epoxy-(5beta,14beta,17betaH)-card -20(22)-enolide (2), and a known cardenolide, (-)-17beta-neriifolin (3), were isolated from the roots of Cerbera manghas as antiproliferative and antiestrogenic principles when evaluated against a human colon cancer cell line (Col2) and the Ishikawa cell line, respectively. Two known lignans, (-)-olivil (4) and (-)-cycloolivil (5), were also isolated but were inactive in the assay systems used.

Piggyback intraocular lens implantation.

The piggyback method of implanting two intraocular lenses in one eye has been successfully expanded to address pseudophakic refractive error in normal eyes and eyes that have undergone post-penetrating keratoplasty. Piggyback implantation has been combined with the use of newly available minus-power lenses to provide appropriate power for a cataract patient with keratoconus, as well as to correct pseudophakic myopia. The phenomenon of increased depth of focus in piggybacks may be explained by a contact zone between the lenses. The late complication of inter-lenticular cellular growth with resultant hyperopic shift, opacification, and loss of vision has recently become a concern.

Evaluation of selected lichens from iceland for cancer chemopreventive and cytotoxic activity.

Cancer chemopreventive effects of organic extracts from 29 species of lichens collected in Iceland were evaluated using a panel of in vitro bioassays whereby extracts were tested for potential to induce quinone reductase (QR) and differentiation of human promyelocytic leukemia (HL-60) cells, inhibit cyclooxygenase-1 (COX-1), phorbol ester-induced ornithine decarboxylase (ODC), aromatase and sulfatase, as well as for antioxidant, estrogenic/anti-estrogenic and antiproliferative activity. In addition, the extracts were tested for cytotoxicity against 12 cancer cell lines. The most significant results were exhibited by extracts from Xanthoria elegans and Alectoria nigricans , which respectively, induced QR activity (concentration to double activity = 4.8 µg/ml) and inhibited phorbol ester-induced ODC activity with mouse 308 cells in culture (IC 50 = 2.6 µg/ml). Moderate inhibition of [ 3 H]thymidine incorporation with HL-60 cells was exhibited by the Peltigera leucophlebia extract. Several extracts prevented estrogen formation from estrogen precursors by inhibiting the enzymatic activities of aromatase ( Sphaerophorus globosus , Cetrariella delisei , Melanelia hepatizon ) and sulfatase ( Cladonia gracilis , Sphaerophorus fragilis , S. globosus ). None of the extracts demonstrated significant cytotoxic effects with selected cell lines.

Minus-power intraocular lenses to correct refractive errors in myopic pseudophakia.

PURPOSE: To evaluate the effectiveness of a secondary, piggyback, minus-power intraocular lens (IOL) to correct the refractive error in patients with myopic pseudophakia. METHODS: In this prospective noncomparative cohort study, 51 myopic pseudophakic patients received implantation of a minus-power IOL as a secondary procedure to correct residual pseudophakic myopia. RESULTS: The mean residual myopia of -3.05 diopters (D) was reduced to -0.38 D. All eyes were within +/- 1.00 D of the desired refraction. Uncorrected visual acuity was 20/40 or better in 72% of eyes, and best corrected visual acuity was 20/40 or better in 96%. Uncorrected visual acuity improved by 2 or more lines in 85% of eyes and by 5 or more lines in 65%. CONCLUSION: Clinical outcomes can now be improved in patients with myopic pseudophakia whose previous options (i.e., lens exchange or refractive surgery) were more traumatic or less predictable.

Refractive and visual outcome of hyperopic cataract cases operated on before and after implementation of the Holladay II formula.

OBJECTIVE: The primary objective was to evaluate the refractive and visual outcomes in a series of hyperopic cataract cases in which the Holladay II intraocular lens (IOL) power formula was used in conjunction with added eye measurements (measured anterior chamber depth [ACD], lens thickness, and corneal diameter) to improve predictability of refractive outcome. In addition, the impact of use of a double ("piggyback") IOL on refractive outcome was evaluated. DESIGN: Prospective, nonrandomized comparative clinical trial. PARTICIPANTS: A total of 136 consecutive hyperopic primary cataract-IOL cases operated on at in an outpatient eye surgery center were evaluated. The main inclusion criterion was the requirement of at least 30 D of emmetropia power. INTERVENTION: Implantation of a total implanted power calculated using a newly developed (Holladay II) formula, which uses additional eye measurements (measured ACD, lens thickness, corneal diameter) in addition to the axial length and keratometry normally used, was performed. In the first series, IOL powers were chosen using the Lloyd-Gills formula with modifiers; in the second series, powers were chosen using the Holladay II formula option in the Holladay IOL Consultant software. Selection criteria for both series were the same (requiring at least 30 diopters [D] of power for emmetropia). Keratometry and axial length measurements (by immersion) were taken using the same instrumentation and methodology in both series. Predicted postoperative refraction based on the IOL implanted and the method of power calculation used were computed for each case in both groups and compared to the actual achieved refraction. MAIN OUTCOMES MEASUREMENTS: Main clinical outcome parameters evaluated were the postoperative spherical equivalent (compared with the predicted spherical equivalent) and the best-corrected vision. These outcome parameters were evaluated within each surgical series, in the total group of cases (regardless of power calculation method). Further stratification according to the use of single or double implants also was done. RESULTS: In the group using an older formula system, mean preoperative spherical equivalent of 4.79 D was reduced to -0.67 D. Similarly, in the Holladay II group, the preoperative mean of 5.60 D was reduced to -0.58 D. However, there were fewer large deviations between predicted and achieved spherical equivalent in the Holladay II group as indicated by a smaller standard deviation of the absolute deviation (0.47 vs. 0.59), and the range of postoperative refractions was smaller with fewer large overcorrections or undercorrections. However, almost 90% of both groups were within a diopter of the predicted refraction. Visual results were comparable in the two groups. CONCLUSION: Both IOL calculation systems showed good predictability in these extremely short eyes. The Holladay II formula was simpler because it is incorporated into a user-friendly software package (Holladay IOL Consultant) and required only the input of IOL constants and preoperative measurements with no "fudge factor" modifiers. Results within the series using this formula had a tendency toward a smaller standard deviation with fewer outliers.

Piggyback minus-power lens implantation in keratoconus.

A 53-year-old man with keratoconus and an axial length of 32.59 mm had cataract extraction by phacoemulsification. The Holladay II formula called for -14.00 diopters (D) of power. Two negative-power intraocular lenses (IOLs) were implanted to optimize visual results. A 1 day postoperative refraction of +1.50 D sphere necessitated an exchange of the anterior IOL. Six days after the exchange, the patient had a refraction of -1.25 D sphere and best corrected visual acuity of 20/50.

Microtubule disruption leads to cellular contraction in human trabecular meshwork cells.

PURPOSE: To determine whether microtubule- and actin-altering drugs, which have been shown to increase aqueous humor outflow, cause cellular contraction in human trabecular meshwork (HTM) cells. METHODS: HTM cells were plated in culture dishes containing a polymerized deformable silicone substrate. After 48 hours, the dishes were placed on an inverted microscope and treated with ethacrynic acid, colchicine, vinblastine, cytochalasin B, or 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7) and then recorded on videotape for 15 minutes. An increase in silicone substrate wrinkle size and/or number indicated a contraction. Sham controls were used. RESULTS: Cellular contraction was observed with ethacrynic acid, colchicine, and vinblastine in the 10(-5) to 10(-4) M dosage range. Pretreatment with H-7 blocked these effects. Cytochalasin B did not produce cellular contraction. CONCLUSIONS: Microtubule disruption causes cellular contraction in HTM cells, and this effect depends on an intact actin cytoskeleton network. Contraction of trabecular meshwork cells in response to various stimuli is an attractive hypothesis for possible homeostatic mechanisms in the outflow pathway, and this may serve as a focus for novel glaucoma drug development.

Unpreserved lidocaine to control discomfort during cataract surgery using topical anesthesia.

PURPOSE: To determine whether intraoperative unpreserved lidocaine further decrease discomfort or pain during sutureless small incision cataract surgery and intraocular lens (IOL) implantation under topical anesthesia. SETTING: Outpatient ambulatory surgical center. METHODS: In this prospective controlled study, comparable eligible patients were randomized to receive 0.1 cc unpreserved lidocaine 1% or 0.1 cc balanced salt solution (BSS) (control group) in double-masked fashion. Study drugs were injected intracamerally 1 minute before phacoemulsification. A predefined uniform pain/discomfort scale was used for assessment during phacoemulsification and IOL insertion. A secondary study using a 0.5 cc dose was also performed. RESULTS: Twenty-six percent in the control group and 9% in the lidocaine group had discomfort pain scores of 2 or more; 10% in the BSS group felt increased pressure or pain during phacoemulsification. In the lidocaine group, discomfort was felt mainly during IOL insertion, possibly as a result of wound manipulation. During phacoemulsification, no patient in the lidocaine group reported pain; 2% felt increased pressure during phacoemulsification. A dose increase to 0.5 cc reduced any intraocular sensation to 3% in the lidocaine group. No patient in either group had significant cell loss or adverse events. CONCLUSION: Intraoperative lidocaine is safe and effective in controlling intraoperative discomfort.

Achieving emmetropia in extremely short eyes with two piggyback posterior chamber intraocular lenses.

PURPOSE: To examine the refractive results and limitations of current intraocular lens power formulas when implanting two posterior chamber lenses in-the-bag to achieve emmetropia in extremely short eyes. METHODS: Preoperative measurements (corneal diameter, axial length, keratometry, anterior chamber depth, and lens thickness) and postoperative measurements (refraction, corneal vertex to iris depth, and iris to front anterior lens surface) were taken in six eyes from three patients, with axial lengths ranging from 15.09 to 19.95 mm. These data were used to calculate the prediction error for three current third-generation formulas (Holladay, Hoffer Q, SRK/T) and two older formulas (SRK2 and SRK1). RESULTS: None of the formulas accurately predicted the refractions using the optimized lens constants for normal eyes. The third-generation formulas were not different (P > or = 0.602) and averaged 5 diopters (D) of absolute error (Hoffer Q = 4.64 +/- 1.57 D; Holladay = 5.07 +/- 1.28 D; SRK/T = 5.12 +/- 1.43 D). The older formulas were significantly worse (P = 0.0006), with average mean absolute errors of 10.93 +/- 5.09 D for the SRK2 and 13.33 +/- 5.09 D for the SRK1. When the formulas were optimized for these six eyes, the mean absolute errors were Holladay = 1.33 +/- 1.25 D; SRK/T = 2.10 +/- 1.31 D; Hoffer Q = 4.54 +/- 2.00 D; SRK2 = 4.71 +/- 1.94 D; and SRK1 = 4.71 +/- 1.94 D. The Holladay and SRK/T formulas were statistically better (P = 0.0068) than the Hoffer Q and the two older formulas. CONCLUSION: Current third-generation formulas are better than older formulas for extremely short eyes, but still are not acceptable for the desired clinical accuracy. Newer formulas that will use additional anterior segment measurements (corneal diameter, anterior chamber depth, and lens thickness) will be required for improved accuracy, because the anterior segment often is not proportional to the axial length.