WWW.DISSERTATION.XLIBX.INFO
FREE ELECTRONIC LIBRARY - Dissertations, online materials
 
<< HOME
CONTACTS



Pages:     | 1 |   ...   | 7 | 8 || 10 | 11 |   ...   | 18 |

«COLLOIDAL GOLD NANOPARTICLES FOR CANCER THERAPY: EFFECTS OF PARTICLE SIZE ON TREATMENT EFFICACY, TOXICOLOGY, AND BIODISTRIBUTION A Dissertation ...»

-- [ Page 9 ] --

Maximum Drug Loading Efficiency using Fluorescence To find the optimum PEG density for colloidal gold stability, dynamic light scattering measurement was used for size change. After subtracting the gold surface area occupied by modified doxorubicin, available surface area was coated with various concentrations of PEG. As we increased the PEG concentration, saturation point was reached for gold-drug-PEG size measurement (Table 4.1).

Table 4.1.

PEG Coating and Gold Nanoparticle-Anticancer Agent-PEG (Au-DOX-PEG) System Size Change

–  –  –

Saturation of gold nanoparticle surface at low % PEG values indicate that most of the gold surface is coated with modified doxorubicin, and PEG is bound onto the gold surface in a “mushroom conformation”. Mushroom conformation is characterized by low surface grafting density and polymer tends to “lie” close to the surface that multiple points of a single polymer is covering the surface [115]. Despite the low PEG surface density, the resulting Au-DOX-PEG was colloidally stable in various mediums such as salt and serum solutions (APPX A Figure A.1). We suspect that in conjunction with DOX-PDPH coating, small-sized gold nanoparticle core (5nm) and long chain length (MW=5000) PEG resulted in a sufficient coverage of the gold surface for colloidal stability. It has been reported that nanoparticle stability increases with increasing PEG length and decreasing nanometer particle diameter [43]. In particular, PEG with a molecular weight of ≥ 2000 Daltons can significantly reduce protein adsorption and result in colloidal stability in highly ionic strength media due to increased steric repulsive forces [43, 170-175]. Supporting our results, Liu, Y. et al found that 1) for any given PEG length, decrease in nanoparticle diameter resulted in a decreased amount of PEG per nanoparticle input to the reaction mixture and 2) for a given nanoparticle diameter, increase in PEG length resulted in a decrease in the amount of PEG amount input to the reaction mixture for colloidal stability. Furthermore, in accordance with DLVO theory, small-sized gold nanoparticles are more stable in general than the larger-sized gold nanoparticles due to minimized van der Waals attraction energy.

Finally, we also wanted to ensure that bound doxorubicin is not affected by the addition of PEG. Similar to the method used for drug loading efficiency, the supernatant collected from centrifugation of various concentrations of PEG coated gold-drug-PEG systems indicated that addition of PEG, especially excess amount of PEG, did not affect bound doxorubicin and there was no detectable replacement of the bound drug.

Drug Release Profile Figure 4.3 shows pH-dependent release profile of doxorubicin linked to hydrazone bond of PDPH at 37°C. Initially, doxorubicin is slowly released for both acidic and basic conditions that no detectable release was observed up to 48 hours. More doxorubicin was released over time, especially for acidic condition, which led to approximately 4 times more doxorubicin release at the end of 96 hours for pH 5 condition compared to that of neutral condition. Previous studies also reported increased hydrolysis of hydrazone bond and rapid release of doxorubicin in acidic conditions compared to neutral conditions [130, 156].

Figure 4.3.

pH-Sensitive Drug Release The slow, delayed release of doxorubicin from Au-DOX-PEG system is due to (1) the diffusion barrier created by PEG coating and (2) interactions between the polymer and the drug. PEG chains interact with one another that complex is formed amongst PEG chains by hydrogen bonding [157]. As polymer chain length increases and more interpolymer complexes are formed, the release rate of the drug is decreased. Also, complexation affects PEG conformation that polymer coils provide additional diffusion barrier for more tortuous path for drug release [157, 158]. Because PEG is a hydrophilic polymer, as the hydrophobicity of the drug increases, the diffusion rate of the drug decreases. However, we believe that combination of the EPR effect and various in vivo conditions will affect the release profile of Au-DOX-PEG system to be more effective system.

In Vitro Therapeutic Efficacy of Au-DOX- PEG System MTT assay with 4T1 cell line was used to study the anticancer efficacy of AuDOX-PEG system. Cell viability was inversely related to doxorubicin activity that absence or minimal efficacy of doxorubicin resulted in increased cell survival. Cell viability for each group (pure doxorubicin, Au-PEG, and Au-DOX-PEG) was compared to the control group which was free of doxorubicin (Figure 4.4 (b)).

Compared to the pure doxorubicin 96 hour toxicity, the slightly lower toxicity of Au-DOX-PEG could have resulted from slow release of doxorubicin within PEG shell (also seen in Figure 4.3) and different cellular localization of Au-DOX-PEG system compared to the pure drug.

The slow release of doxorubicin from Au-DOX-PEG system was also evident in the fluorescence images of the 4T1 murine breast cancer cells incubated with the gold conjugate system (Figure 4.4 (a)). When equal concentration of doxorubicin was used, cells that contained Au-DOX-PEG displayed less fluorescence intensity compared to the pure doxorubicin incubated cells. Also, when equal concentration of gold nanoparticle was used, only Au-DOX-PEG displayed fluorescence for doxorubicin and Au-PEG fluorescence images resembled that of CONTROL where no fluorescence was detected for doxorubicin. The actual uptake of gold nanoparticle by 4T1 cells is evident in TEM images in next section.





(a) Figure 4.4. In vitro Therapeutic Efficacy of Au-DOX-PEG and Cellular Uptake of Gold Nanoparticle (a) 4T1 cells were incubated with doxorubicin, Au-DOX-PEG, and Au-PEG at 37°C for 2.5 hours and fluorescence images were taken for DAPI (nucleus staining) and DOX (Doxorubicin) (equal amount of gold was used for Au-DOX-PEG and AUPEG groups; 5μg/mL of doxorubicin and equivalent was used for doxorubicin and AuDOX-PEG, respectively) (b) MTT assay results with 4T1 murine breast cancer cells (equal amount of gold was used for Au-DOX-PEG and AU-PEG groups; 7 μg/mL of doxorubicin and equivalent was used for doxorubicin and Au-DOX-PEG, respectively) In vivo Therapeutic Efficacy of Au-DOX-PEG Drug Delivery System for solid tumor Body Weight, Tumor Volume, and Tumor Raw Weight Changes To investigate the therapeutic efficacy of Au-DOX-PEG drug delivery system in vivo, we conducted a comparison study between pure doxorubicin versus Au-DOX-PEG system on 4T1 murine breast cancer mice model. 4T1 tumor cells were injected subcutaneously at the mammary fat pad of female BALB/c mice. Treatments were carried out by injecting saline (control), pure doxorubicin, Au-DOX-PEG (at an equal doxorubicin dose of 2.4 mg/ kg), and Au-PEG into tail-vein for every 3 days over ~2 weeks (n=5 for each group). As shown in Figure 4.5, tumor volume was measured every other day by a caliper and it was observed that Control group (untreated) and gold nanoparticle only group (Au-PEG) displayed average fractional tumor volumes (i.e. final tumor volume divided by initial tumor volume) of 6.3 ± 1.8 and 6.6 ± 2.4, respectively, on day 16. Statistical analysis results indicated that there was no significant difference between the Control and Au-PEG groups (P0.05 for Control versus Au-PEG). Pure doxorubicin treated group resulted in fractional tumor volume of 3.1 ± 1.2 on day 16, which represents tumor group inhibition or TGI of ~50 % (i.e. TGI= 100 * {[Control fractional tumor volume – Group of Interest fractional tumor volume]/ Control fractional tumor volumes}). The statistical analysis result indicated that there were no significant differences between the Control versus pure Doxorubicin groups and Au-PEG versus pure Doxorubicin groups (P0.05 for doxorubicin versus control; P 0.05 for doxorubicin versus Au-PEG). In contrast, Au-DOX-PEG treatment resulted in a fractional tumor volume of 1.8 ± 0.3 on day 16 (P0.01 Au-DOX-PEG versus Control;

P0.05 Au-DOX-PEG versus Au-PEG; P0.05 Au-DOX-PEG versus pure doxorubicin) with tumor group inhibition of ~71%, which was more effective than the pure Doxorubicin group.

Alternatively, therapeutic efficacy of Au-DOX-PEG system was studied by extracting tumor from mice and raw tumor weight was measured from each group on day

16. Similar to the tumor volume change results, there was no significant difference between the Control and Au-PEG average tumor raw weights (P0.05 for Control versus

–  –  –

Figure 4.5.

In Vivo Results (a) Weight Change: there was no visible weight changes amongst the group over the course of treatment (b) Tumor Volume Change: Tumor

Volume Change for Control, Doxorubicin, Au-DOX-PEG, and Au-PEG (*: p 0.05; # :

p 0. 01; §: p 0.05; pink arrow indicates treatment); Tumor Weight (**: p 0.05 from control only) (c) Extracted Tumor Raw Weight [n=5 and standard error of mean were used for all figures] Au-PEG). Compared to the average Control tumor raw weight, Doxorubicin group resulted in an average tumor weight of 0.73 ± 0.22 g, which was ~25% lighter than the Control group (P0.05 for Doxorubicin versus Control). Au-DOX-PEG group had an average tumor weight of 0.38 ± 0.11 g, which was significantly lighter than the Control group (~61% lighter than the Control group with P0.05 for Au-DOX-PEG versus Control). Compared to the pure Doxorubicin group, Au-DOX-PEG group was ~47% lighter but it was not statistically significant enough (P 0.05 for Au-DOX-PEG versus pure Doxorubicin). Thus, consistent with the tumor volume change results, Au-DOXPEG group resulted in a tumor raw weight that was significantly differently (all P0.05) from the Control and Au-PEG groups at the end of the therapy. In contrast, Doxorubicin group did not display statistically significant difference between Control group or between the Au-PEG group tumor raw weights.

Finally, body weights amongst the Control, pure Doxorubicin, Au-DOX-PEG, and Au-PEG groups were measured throughout the therapy. The statistical analysis results indicates that there was no significant differences amongst the body weights for all four groups (P0.05).

In summary, there was no significant difference between Control and Doxorubicin, whereas Au-DOX-PEG group showed statistically significant difference from the Control group. Moreover, Au-DOX-PEG group showed statistically significant difference from the Au-PEG group, whereas pure Doxorubicin group did not. Although Au-DOX-PEG group exhibited somewhat higher tumor group inhibition (TGI) index than the Doxorubicin group, there was no statistically significant difference (P0.05) seen between Doxorubicin and Au-DOX-PEG groups. However, it should be noted that AuDOX-PEG displayed no apparent toxicity to the vital organs in contrast to pure Doxorubicin group as shown in the upcoming section.

Spatial Distribution of Au-DOX-PEG in Tumor For the drug delivery nanoparticle system to exert its full therapeutic efficacy, it must reach the cancer cells in the solid tumor by (1) crossing (i.e. extravasate) the tumor blood vessel wall into the tumor interstitium (i.e vascular permeability), (2) migrating through the tumor interstitial space or penetration away from the blood vessel through the extracellular matrix, and then (3) penetrating or entering the cancer cell (i.e. cellular uptake) [176, 177]. Our system resulted in a successful extravasation by diffusion through the tumor blood vessel wall due to small size scale of Au-DOX-PEG (Figure 4.6). Tumor accumulation is a function of both the rate of extravasation from the blood to the tumor space and also the rate of clearance from the tumor. Hobbs et al. showed that the rate of extravasation ofs”small-sized” bovine serum albumin (BSA) of 7nm was independent of pore size over a variety of tumor models [28]. This demonstrates that for a ~7 nm molecule, which is much smaller than the transvascular pore, extravasation is not dependent on pore size but is instead a diffusive process that will depend on the concentration gradient between blood and tumor. Chilkoti et al. demonstrated that dextran drug carrier of molecular weights of 40 and 70 kDa (diameters of 11.2-14.6nm) resulted in a successful extravasation and penetration into the tumor [23]. Similarly, Tang et al. demonstrated that doxorubicin containing PEG-PE micelles with size of 10-20nm resulted in a successful extravasation, accumulation, and penetration of the nanoparticle at the tumor site [178].

The increased accumulation and retention of nanoparticle at the tumor interstitium increases the success rate of the drug delivery system. For normal vasculature, macromolecules and cell debris are cleared from the interstitium through lymphatic drainage. However, lymphatics in tumors are either poorly developed or nonfunctional that leads to decreased rate of clearance of particles [179, 180]. As seen in Figure 4.7, statistical analysis (P0.05, Kruskal-Wallis Test) shows that there was no significant difference amongst the total gold amount accumulated in tumor over time after a single tail-vein injection of gold nanoparticle system. This indicates that the retention of our AuDOX-PEG system within the tumor mass for prolonged period time for successful tumor therapy. Accumulation of Au-DOX-PEG in tumor was also evident by the color change of tumor mass itself as seen in Figure 4.6. Au-DOX-PEG displayed high accumulation in the tumors that they penetrated relatively long distance (radially) into the tumor (dark thick halo seen in the sliced open tumor mass picture of Figure 4.6 (a)), rather than being concentrated only near the vascular surface of short distance. Also, TEM Images from Figure 4.6 shows that gold nanoparticle are present in the extravascular compartment or the tumor interstitium, where concentration in this compartment represents the cumulative exposures of cancer cells to drug. It has been reported that when the particles are too small (molecular weight 40kDa), nanoparticles are rapidly cleared from the tumor extravascular compartment [23]. Thus, our Au-DOX-PEG system has the ideal size (5nm core with ~13nm shell) that resulted in successful vascular permeability and retention in the tumor interstitium.

(a) (b)

–  –  –

Figure 4.6.



Pages:     | 1 |   ...   | 7 | 8 || 10 | 11 |   ...   | 18 |


Similar works:

«Sensationalism, Cinema and the Popular Press in Mexico and Brazil, 1905-1930 By Rielle Edmonds Navitski A dissertation submitted in partial satisfaction of the requirements for the degree of Doctor of Philosophy in Film and Media in the Graduate Division of the University of California, Berkeley Committee in charge: Professor Kristen Whissel (chair) Professor Natalia Brizuela Professor Mary Ann Doane Professor Mark Sandberg Spring 2013 Sensationalism, Cinema and the Popular Press in Mexico and...»

«POLARIZATION-SENSITIVE OPTICAL COHERENCE TOMOGRAPHY USING POLARIZATION-MAINTAINING FIBERS A dissertation submitted to the Faculty of the Graduate School of the UNIVERSITY OF MINNESOTA by MUHAMMAD K. AL-QAISI in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Adviser: Taner Akkin December 2010 © Muhammad K. Al-Qaisi December 2010 ACKNOWLEDGEMENTS I want to start by thanking my adviser Prof. Taner Akkin for his valuable guidance. Working in his lab was an...»

«No God, No Laws Nancy Cartwright Philosophy LSE and UCSD Introduction. My thesis is summarized in my title, ‘No God, No Laws’: the concept of a law of Nature cannot be made sense of without God. It is not as dramatic a thesis as it might look, however. I do not mean to argue that the enterprise of modern science cannot be made sense of without God. Rather, if you want to make sense of it you had better not think of science as discovering laws of Nature, for there cannot be any of these...»

«University of Alberta A Magnetotelluric Investigation of the Arabia – Eurasia Collision in Eastern Anatolia by Er an Türko lu A thesis submitted to the Faculty of Graduate Studies and Research in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Geophysics Department of Physics Edmonton, Alberta Spring 2009 ABSTRACT Eastern Anatolia is the location of a young continent – continent collision between the Arabian and Eurasian plates in Eastern Turkey. The first...»

«DESIGN OF FAULT TOLERANT CONTROL SYSTEMS by Benjamin Erik Walker A dissertation submitted to the faculty of The University of North Carolina at Charlotte in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Electrical Engineering Charlotte 2012 Approved by: Dr. Bharat Joshi Dr. Yogendra Kakad Dr. James Conrad Dr. Mehdi Miri Dr. Edgar Munday ii ©2012 Benjamin Erik Walker ALL RIGHTS RESERVED iii ABSTRACT BENJAMIN ERIK WALKER. Design of fault tolerant control...»

«The Techneducator Effect: Colliding Technology and Education in the Conceptualization of Virtual Learning Environments Daniel Jason Nolan A thesis subrnitted in conforrnity with the requirements for the degree of Doctor of Philosophy Department of Curriculum, Teaching and Learning Ontario Institute for Studies in Education of the University of Toronto O Copyright by Daniel Jason Nolan (2001) Bibliothèque nationale 1*1 National Library du Canada o Canada f Acquisitions and Acquisitions et...»

«The New Zealand Dressmaker: Experiences, Practices and Contribution to Fashionability, 1940 to 1980. A thesis submitted in (partial) fulfilment of the requirements for the degree of Doctor of Philosophy Jan Hamon MA (Fashion & Textile Studies) School of Design and Architecture RMIT University July 2007 i Declaration I certify that except where due acknowledgement has been made, the work is that of the author alone; the work has not been submitted previously, in whole or in part, to qualify for...»

«HIGH QUALITY LIQUID CRYSTAL TUNABLE LENSES AND OPTIMIZATION WITH FLOATING ELECTRODES A dissertation submitted to Kent State University in partial fulfillment of the requirements for the degree of Doctor of Philosophy by Liwei Li December, 2013 UMI Number: 3618874 All rights reserved INFORMATION TO ALL USERS The quality of this reproduction is dependent upon the quality of the copy submitted. In the unlikely event that the author did not send a complete manuscript and there are missing pages,...»

«BWANA ASIFIWE IN THE USA: LANGUAGE AND IDENTITY IN EAST AFRICAN CHRISTIAN FELLOWSHIPS IN THE UNITED STATES By Candis Driver Smith A DISSERTATION Submitted to Michigan State University in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY English 2011 ABSTRACT BWANA ASIFIWE IN THE USA: LANGUAGE AND IDENTITY IN EAST AFRICAN CHRISTIAN FELLOWSHIPS IN THE UNITED STATES By Candis Driver Smith In an effort to expand the sociolinguistic knowledge regarding the East African...»

«EXPLORATORY STUDY OF RFID APPLICATIONS FOR AIR CARGO OPERATIONS By MAGALIE LANIEL A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2010 1 © 2010 Magalie Laniel 2 To all who guided me throughout my lifetime, making this milestone possible 3 ACKNOWLEDGMENTS This work has been carried out under the guidance of my advisor Dr. Jean-Pierre Émond. His brilliant ideas...»

«Department of Philosophy Undergraduate Careers Guide Contents: Introduction 3 Part 1: Medium to long-term plans 4 Jobs Philosophers Do 4 Part 2: Going On the Job Market 6 A Cautionary Tale 6 Preparing yourself 8 Preparing your CV 9 The Interview 13 And If I Don’t Get the Job? 14 Part 3: Further Study 15 Professional Training 15 Post-Graduate Study in Philosophy 15 Introduction A university education has at least three aspects: academic work; social life; and preparation for a career. Nothing...»

«Transit for National Parks and Gateway Communities: Impacts and Guidance A Dissertation Presented to The Academic Faculty By Anne E. Dunning In Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in Civil and Environmental Engineering Georgia Institute of Technology January 2005 Copyright © Anne E. Dunning, 2005.Transit for National Parks and Gateway Communities: Impacts and Guidance Approved by: Dr. Michael D. Meyer, Advisor Dr. Michael O. Rodgers School of Civil and...»





 
<<  HOME   |    CONTACTS
2016 www.dissertation.xlibx.info - Dissertations, online materials

Materials of this site are available for review, all rights belong to their respective owners.
If you do not agree with the fact that your material is placed on this site, please, email us, we will within 1-2 business days delete him.