A detailed dosimetric comparison between Inverse Planning by Simulated Annealing (IPSA) and Dose Points (DP) optimized treatment plans has been performed for High Dose Rate (HDR) brachytherapy of skin lesions using Freiburg Flap applicator in order to find out whether or not IPSA offers better clinical dosimetric outcomes for lesions categorized into four different curvatures. Without compromising target coverage, IPSA reduced the volume of Planning Target Volume (lesion) that received at least 125% of the prescription dose on average by 41%. It also reduced the volume of the healthy skin surrounding the lesion that receives at least 100% of the prescription dose on average by 42%. IPSA did not show any advantage over DP in sparing normal structures underlying the lesions treated. Although DP optimization algorithm has been regularly used at Lynn Cancer Institute for HDR brachytherapy of skin lesions, recent upgrades in IPSA software have made IPSA more amenable to rapid treatment planning and therefore IPSA can be used either in place of DP or as its alternative. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2014. / FAU Electronic Theses and Dissertations Collection

The purpose of this study is to investigate the changes of the pericardial dose at different respiratory phases and statuses in accelerated partial breast irradiation (APBI) using Cyberknife M6™ multileaf collimators (CK-MLC). Anonymous 6 female patient files with respiration gated four-dimensional computed tomography (4DCT) sets, and 6 left breast cancer cases with CT images in free-breathing (FB) and deep inhalation breath-hold (BH) were selected. One CT image set from each patient was planned for APBI in Accuray Multiplan™ 5.2, and respectively compared its pericardial dose with those from CT sets of other respiratory phases. All the comparable CT images were fused in the planning system according to the left chest wall, among which the lung gap anterior to the pericardium varies by the lung expansion. For the purpose of this study, the tumor volume was outlined in the media-lower quadrant of the left breast where this lung gap is relatively small. All the plans in this study met the requirements set by the National Surgical Adjuvant Breast and Bowel Project/Radiation Therapy Oncology Group (NSABP/RTOG), specifically protocol B-39/RTOG 0413. From the comparisons in this investigation, the mean relative pericardial dose of the BH CT group showed significant or 45% (p < 0.01) lower value than that of FB CT group. However, in FB 4DCT group, 3 of 6 cases indicated a meaningful reduction (p < 0.05) in 100% inhalation phase when compared with the mean dose over other phases. The inconsistent pericardial doses were displayed in FB 4DCT group due to minimal changes in the anterior lung gap of the pericardium, when the diaphragmatic breathing was dominant in those patients. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2015. / FAU Electronic Theses and Dissertations Collection

Intensity Modulated Radiation Therapy (IMRT) is a well-known type of external beam radiation therapy. The advancement in technology has had an inevitable influence in radiation oncology as well that has led to a newer and faster dose delivery technique called Volumetric Modulated Arc Therapy (VMAT). Since the presence of the VMAT modality in clinics in the late 2000, there have been many studies in order to compare the results of the VMAT modality with the current popular modality IMRT for various tumor sites in the body such as brain, prostate, head and neck, cervix and anal carcinoma. This is the first study to compare VMAT with IMRT for breast cancer. The results show that the RapidArc technique in Eclipse version 11 does not improve all aspects of the treatment plans for the breast cases automatically and easily, but it needs to be manipulated by extra techniques to create acceptable plans thus further research is needed. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2015. / FAU Electronic Theses and Dissertations Collection

In addition, we investigated the cytotoxic effect of a widely used chemotherapeutic agent 5Fu on laryngeal squamous cell cancer cell lines and evaluated the role of p53 in 5Fu treatment. We found that the apoptosis and G1/S cell arrest mediated by 5Fu in laryngeal cancers is p53-independent but p21 WAF1/CIP1-dependent. We further demonstrated the effect of 5Fu on HPV16-associated laryngeal cancer cells. Using cytotoxicity assay and Annexin V staining, we proved that 5Fu induces apoptosis in all of the transfected cells in a dose- and time-dependent manner, suggesting that the process was not prevented by HPV16 E6 or E7. 5Fu induced the accumulation of active pRb and cyclin dependent kinase inhibitor p21WAF1/CIP1 together with an increase in Bak and Bax expression and a decrease in Bcl-2 levels in all the transfected cells. In addition, G1/S phase cell cycle arrest was associated with the antiproliferation activity of 5Fu in all cell lines. Through RT-PCR, 5Fu also presented some effects on the E6 and E7 oncoproteins of HPV16 in transfected UMSCC 12 cells. / Our results suggest that HPV16 E6 and E7 oncoproteins do not prevent 5Fu medicated apoptosis and G1/S cell arrest in laryngeal cancers. The anti-cancer effect of 5Fu is probably decided by the level of p21 WAF1/CIP1 while the sensitivity of laryngeal cancer cells responded to 5Fu treatment is associated with the increase of Bak or/and the decrease in Bcl-2, not with the HPV16 viral proteins and p53 status. 5Fu also presented some effects on the E6 and E7 oncoproteins of HPV16 in laryngeal cancer. However, the anti-viral effect of 5Fu still needs further investigation. / Our study indicated that (1) the evasion of apoptosis mediated by HPV16 E6 and E7 plays a critical role in laryngeal carcinogenesis; (2) HPV16 E6 or E7 plays an important role in regulating the expression of Bak, Bax and Bcl-2; (3) The degradation of Bak by HPV16 E6 is not caused by interacting with the promoter of Bak; (4) The induction of Bcl-2 is mediated through HPV16 E7; (5) HPV16 transfection does not interfere with the apoptosis and cell cycle arrest mediated by 5Fu in human laryngeal squamous cancer cells. / There is a growing body of evidence that human papillomavirus type 16 (HPV16) is involved in the development of human laryngeal cancer, especially in Chinese population. The two oncoproteins, HPV16 E6 and E7 that target host cell tumor suppressor proteins p53 and Rb respectively, may generate antiapoptotic effects and induce cell immortalization. However, the effect of both oncoproteins on apoptosis in laryngeal cancers is not completely clear. In this study, we demonstrated the possible mechanism of high risk HPV16 in laryngeal carcinogenesis and evaluated the effect of 5Fu on HPV16-positive laryngeal cancer cells. / We employed two human laryngeal cancer cell lines---UMSCC12 (with truncated p53) and UMSCC11A (with mutant but functional p53) in this study. These two cell lines were stably transfected with HPV16 E6, E7 or empty vector, pcDNA3.1, which provided a good foundation for further study on the carcinogenic mechanism of HPV16 E6 or E7 in human laryngeal cancers. Through Annexin V staining and protein stability assay, we found that the transfection of HPV16 E6 and E7 induced fewer spontaneous apoptosis in both UMSCC11A and UMSCC12 cells accompanied with enhanced protein stability of Bcl-2 and increased protein degradation of Bak. Similar results were obtained when E6- and E7-transfected cells exposed to apoptosis stimuli---TNF-alpha/CHX. These results indicate that stable transfection of E6 and E7 in human laryngeal cancer cells on one hand shortened the half-life of Bak protein, and on the other hand, enhanced the steady-state levels of Bcl-2 protein. In order to gain insight into the role of Bak and Bcl-2 in regulating apoptosis in HPV-associated laryngeal cancer cells, we performed transient transfection of Bcl-2 into E6- and E7-transfected cells. It is found that HPV16 E7 statistically enhanced the expression of Bcl-2 in laryngeal cancer, indicating that the induction of Bcl-2 require the transfection of HPV16 E7. Furthermore, Luciferase assay was performed to investigate whether the viral proteins E6 and E7 altered the stability of Bak through interaction with the promoter of Bak. Negative results were obtained, suggesting that E6 or E7 do not alter the transcription activity of Bak, indicating the degradation of Bak by E6 or E7 may be mediated through other mechanisms. / Liu Han-ching. / "August 2006." / Advisers: C. A. van Hasselt; George G. Chen. / Source: Dissertation Abstracts International, Volume: 68-03, Section: B, page: 1569. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (p. 245-274). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

Hepatocellular carcinoma (HCC) is the fifth most common cancer worldwide and the third most common cause of cancer-related deaths. Systemic therapies are the main treatment options for HCC patients with advanced disease (&sim; 80% of all cases). However, only very moderate clinical responses are achieved with most of the conventional therapies. Thus, more effective therapeutic strategies are much needed. The PI3K/Akt/mTOR signaling pathway, which plays a critical role in controlling cell proliferation and survival, is aberrantly activated in &sim; 45% HCC, suggesting it to be a potential target for HCC treatment. Moreover, emerging evidences indicate that activation of the PI3K/Akt/mTOR pathway may be associated with resistance to many cytotoxic chemotherapies, including microtubule targeting agents. In this study, by gene expression profiling and gene ontology analysis, "microtubule-related cellular assembly" was identified to be the major biological/functional process involved in HCC development, suggesting that microtubule is also an important therapeutic target for HCC. With these understandings, it is hypothesize in this thesis that combined targeting of a key component ofthe PI3K/Akt/mTOR pathway, namely the mammalian target of rapamycin (mTOR) and the microtubule would be an effective therapeutic strategy for HCC. The objectives of the thesis are to examine the therapeutic potential of microtubule targeting, mTOR targeting, and combined targeting of the microtubule and mTOR in both in vitro and in vivo models of HCC. / In summary, the PI3K/Akt/mTOR pathway and the microtubule represent promising therapeutic targets for HCC treatment. The findings from this thesis offer a rationale for combining mTOR inhibitors with microtubule targeting agents for effective HCC treatment. / In the second part, the effect of mTOR inhibition, either alone or in combination with an additional microtubule targeting agent (vinblastine) was investigated in HCC. Temsirolimus, an mTOR inhibitor, suppressed HCC cell proliferation in as early as 24 hrs with an IC50 of 1.27+/-0.06muM (Huh7), 8.77+/-0.76muM (HepG2), and 52.95+/-17.14muM (Hep3B). Vinblastine (1nM) alone caused 30--50% growth inhibition in 3 HCC cell lines. In these HCC cell lines, it was found that temsirolimus/vinblastine combination resulted in an additive to synergistic effect (when compared to single agents alone) with maximum growth inhibition of 80--90% as early as 24 hrs upon treatment. This marked growth inhibition was accompanied with cell cycle arrest at both G1 and G2/M phases, and PARP cleavage (a hallmark for apoptosis). Moreover, the combination specifically caused concerted down-regulation of several important anti-apoptotic and survival proteins (survivin, Bcl-2 and Mcl-1), which was not observed in single agent treatments. It was hypothesized that inhibition of these key anti-apoptotic/survival proteins may represent a novel mechanistic action of this highly effective combination approach of dual targeting of mTOR and microtubule by temsirolimus/vinblastine in HCC cells. Indeed, transient over-expression of each of these genes (survivin, Bcl-2 or Mcl-1) in HCC cells did partially rescue the growth inhibitory effect of the temsirolimus/vinblastine combination. More importantly, this novel combination significantly suppressed the growth of HCC xenografts in nude mice (when compared with single agents alone). / In the third part, the anti-tumor effect of another mTOR inhibitor everolimus in combination with microtubule targeting agents, vinblastine and patupilone (a microtubule-stabilizing agent), was investigated in HCC cells. Everolimus/vinblastine combination resulted in an additive to synergistic effect accompanied with cell cycle arrest at both G1 and G2/M phases, and PARP cleavage. The combination also caused concerted down-regulation of anti-apoptotic and survival proteins (survivin, Bel-2 and Mel-1) as observed with the temsirolimus/vinblastine combination. However, everolimus only moderately enhanced the sensitivity of patupilone for reasons unknown. / Taxanes are the major chemotherapeutic agents that target the microtubule. In the first part of the thesis, the anti-tumor activity of two taxanes, paclitaxel and docetaxel (which are known to stabilize microtubules) was examined and compared with doxorubicin (a DNA intercalating agent). Across all three HCC cell lines tested, it was found that the microtubule targeting agents, taxanes, were more efficacious than doxorubicin. This supports the initial finding that microtubule assembly process is functionally important in HCC. Recent studies demonstrated that using nanoparticles for drug delivery can greatly enhance therapeutic efficacy and reduce side-effects. Therefore, the nanoparticle albumin-bound (nab)-paclitaxel was employed to further evaluate the therapeutic efficacy of such a delivery strategy in HCC models. In all three HCC cell lines tested, nab-paclitaxel was found to be the most effective agent, with an average IC50 value of 0.16--10.42nM, when compared to non-conjugated taxanes (paclitaxel, docetaxel) and doxorubicin. In vitro analysis showed that nab-paclitaxel was able to induce cell cycle arrest at G2/M phase and apoptosis in HCC cells. In vivo study demonstrated that nab-paclitaxel readily inhibited the growth of HCC xenografts with lower toxicity when compared to paclitaxel, docetaxel and doxorubicin. Moreover, specific silencing of a key regulatory protein for microtubule dynamics, Stathmin 1, by siRNA significantly enhanced the effect of nab-paclitaxel in HCC cells, resulting in synergistic growth inhibition in vitro. / Zhou, Qian. / Advisers: Winnie Yeo; Vivian Lui; Nathalie Wong. / Source: Dissertation Abstracts International, Volume: 73-06, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 148-164). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Nasopharyngeal cancer (NPC) is endemic in Southern China and Hong Kong. It has traditionally been treated by local radiotherapy with great success especially for early stage disease. However the treatment outcome in advanced stage disease is unsatisfactory. / Results from this series of combined clinical, translational and laboratory studies have redefined the role of hypoxia, angiogenesis and metastasis as new therapeutic targets in NPC. Novel biomarkers and new therapeutic approaches were developed based on these targets. / To develop new therapies in NPC, we demonstrated in a randomized controlled phase 2 clinical trial that sequential therapy of neoadjuvant chemotherapy followed by chemoradiotherapy was well tolerated with a manageable toxicity profile that allowed subsequent delivery of full dose chemoradiotherapy. This strategy reduced distant metastasis which translated into improved patient survival. In preclinical studies, the antiangiogenesis agent sunitinib demonstrated potent in vitro and in vivo growth inhibition in NPC. In a phase 2 clinical trial, sunitinib demonstrated modest clinical activity in heavily pretreated NPC patients. However, the unexpected high incidence of severe hemorrhage from upper aero-digestive tract in NPC patients who received prior high dose RT to the region is of concern. We propose to exclude NPC patients with disease recurrence within previous radiation field and/or with vascular invasion from future antiangiogenesis therapy. / To investigate potential new therapeutic targets and biomarkers in NPC, we first confirmed from the Hong Kong NPC study group of 2915 patients' database that distant metastasis was the leading cause of NPC failure after primary radiotherapy. We further showed that hypoxia induced broad changes of both up- and down-regulated gene expressions involved in diverse biological processes in NPC cells. Over-expression of biomarkers of hypoxia and angiogenesis (including HIF-1alpha, CA IX and VEGF) is common in NPC and is associated with poor prognosis. Elevated plasma osteopontin is a biomarker of distant metastasis, and pre-treatment plasma osteopontin level may be a useful biomarker of response to radiotherapy in NPC. / Hui, Pun. / "September 2010." / Adviser: Anthony Chan. / Source: Dissertation Abstracts International, Volume: 73-04, Section: B, page: . / Thesis (M.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 269-293). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web.

丹參為雙子葉植物唇形科鼠尾草族植物的乾燥根及根莖。在中國，丹參為廣泛用於治療心血管疾病的藥用植物，而在西方，丹參也常作為一種輔助性藥物。《中國藥典2010版》收錄了35個以上含有丹參的複方或者方劑。在這些複方中，採用了富含丹酚酸和丹參酮的丹參水提物、乙醇提取物或兩者的混合物。丹參提取物具有較強的抗氧化作用，被認為在化學預防和化療的輔助治療中有一定用途。作為主要的丹參水溶性成分，熱敏感的丹酚酸在提取與加熱過程中可能會降解為其他丹酚酸。丹參水提取物的化學組成可能會在不同熱水提取溫度下有所不同，進而影響其藥理活性。在本研究中，通過加熱回流提取和在不同溫度下的微波提取（MAE-W）獲得了6種丹參水提取物，並對這些提取物進行化學成分和藥理分析，考察它們的抗氧化、抗凋亡和血管舒張作用。在這些提取物中，第三輪的微波提取物（100 oC）含有最多的丹酚酸和丹參酮，在1，1-二苯基-2-三硝基苯肼（DPPH）法和鐵還原/抗氧化能力（FRAP）法中具有最強的抗氧化活性，在2,2'-偶氮二(2-脒基丙烷)二鹽酸鹽（AAPH）誘導人血紅細胞的溶血實驗和過氧化氫誘導大鼠心肌細胞H9c2凋亡實驗中還顯示了最強的抑制作用，對大鼠腦基底動脈有最強的鬆弛效應。這些丹參水提取物的抗氧化作用與它們的血管舒張效應呈一定的線性關係（回歸係數r = 0.895 - 0.977）。通過多元線性回歸分析發現，丹參素可以作為丹參水提物的抗氧化和血管舒張功能的顯著性標記物，而丹參酮IIA則是抑制過氧化氫誘導大鼠心肌H9c2細胞凋亡的標記物。 / 作為丹參中主要的脂溶性成分，丹參酮在不同的腫瘤細胞系和荷瘤小鼠模型中展示了抗癌潛力。這些丹參酮的抗癌機制包括細胞週期阻滯，觸發半胱天冬酶（Caspase）依賴的內源性和外源性的凋亡途徑和絲裂原激活的蛋白激酶（MAPK）信號通路等。丹參新酮（miltirone）是從丹參中分離得到的松香烷型二萜醌類化合物，具有多種的藥理活性，如抗氧化，抗焦慮和抗腫瘤等。本研究評估了丹參新酮在人肝癌HepG2細胞系和P-糖蛋白（P-gp）過表達的阿霉素耐藥HepG2細胞系（R-HepG2）中的凋亡作用及其機制。丹參新酮在HepG2細胞中顯示了細胞毒性（EC₅₀值為7.06 微摩），而丹參新酮在抑制HepG2和R-HepG2細胞增殖中的濃度依賴性沒有顯著性差異。丹參新酮（1.56 - 6.25微摩）與阿霉素（DOX）對R-HepG2細胞的增殖具有協同效應，在達到50的生長抑制時，它們的聯合用藥指數為0.3至0.5。流式細胞術分析表明，丹參新酮降低了R-HepG2細胞中P-gp介導的阿霉素外排，分子對接研究表明該效果是通過抑制P-gp的藥物結合位點。在非壞死濃度（25微摩或以下），丹參新酮在HepG2和R-HepG2細胞中活化了Caspase依賴的凋亡途徑，誘導產生活性氧（ROS）和氧化應激，且觸發ROS介導的包括p38 MAPK，應激活化蛋白激酶/c-Jun氨基末端激酶（SAPK / JNK）以及細胞外調節激酶1和2在內的MAPK信號通路。綜上所述，在R-HepG2中丹參新酮是P-gp和細胞增殖的雙重抑制劑，顯示了其在治療肝癌（HCC）的潛力。 / 為了增加藥物開發的成功率，在藥物發現的早期階段應考察新化學實體（NCEs）的蛋白結合率，清除率，藥動學參數，以及藥物代謝相互作用等體內代謝參數。以往的研究已經顯示了從丹參中分離得到的四種主要丹參酮對人和大鼠的細胞色素P450酶介導的探針底物的代謝具有不同程度的抑制作用，需要注意丹參和其他藥物間的相互作用。本研究的另一目的是在人類肝微粒體中探討丹參新酮與探針底物間的細胞色素P450酶介導的代謝相關的相互作用。人肝微粒體孵育實驗結果表明丹參新酮對CYP1A2（IC₅₀值為 1.73微摩）和CYP2C9（IC₅₀值為8.61微摩）有中等強度的抑制，對CYP2D6（IC₅₀值為30.20微摩）和CYP3A4（IC₅₀值為33.88微摩）有弱的抑制。酶動力學和分子對接研究的結果進一步表明，丹參新酮為CYP1A2（Ki值為3.17微摩）的中等強度混合型抑制劑，是CYP2C9（Ki值為1.48微摩）的中等強度競爭型抑制劑，也是CYP2D6（Ki值為24.25微摩）和CYP3A4（Ki值為35.09微摩）的弱的混合型抑制劑。這些結果表明，應考慮丹參新酮與CYP1A2和CYP2C9代謝的藥物間的相互作用，但是可認為其與CYP2D6及CYP3A4代謝的藥物間幾乎不存在相互作用。 / 總之，本研究考察了不同提取方法對丹參提取物成份及其藥效的影響，確定了不同用途的丹參提取物的質控標記物。本研究還考察了丹參新酮體外抗肝癌的能力及其藥物代謝相互作用為基礎的類藥性,為其進一步的體內試驗提供了依據。 / Danshen, the dried root and rhizome of Salvia miltiorrhiza Bg. (Fam. Labiatae), is a widely used medicinal plant for the treatment of cardiovascular diseases in China and also a complementary medicine in the West. Danshen is indexed in the Pharmacopoeia of People’s Republic of China (2010 Edition), with more than 35 formulations and concoctions containing Danshen water-extracts, ethanolic extracts or their combination which are rich in phenolic acids and tanshinones with various contents. Danshen extracts have been considered for the use as an adjunct in chemoprevention and chemotherapy due to their strong antioxidant effects. Phenolic acids, the major water-soluble components in Danshen, are thermosensitive and may degrade to other phenolic acids during extractions upon heating. The chemical profiles of Danshen water-extracts may vary with different heat water extraction at different temperatures, affecting the composition and bioactivity of the extracts obtained. In this study, six water-extracts of Danshen obtained from heat reflux water extraction and microwave-assisted extraction with water (MAE-W) at different temperatures were prepared for evaluation of their composition and pharmacological effects such as antioxidant, anti-apoptosis and vascular relaxation. Among these extracts obtained, the third-round MAE-W (100 °C) product, which was the last round product obtained by extracting the same crude material three times, had the highest contents of phenolic acids and tanshinones, with the strongest antioxidant activity estimated by 2, 2-diphenyl-1-(2, 4, 6-trinitrophenyl) hydrazyl (DPPH) assay and ferric reducing / antioxidant potential (FRAP) assay. This extract also possessed the strongest inhibitory effects on 2, 2'-azobis-2-amidino-propane (AAPH)-induced haemolysis in human red blood cells, hydrogen peroxide-induced apoptosis in rat heart H9c2 cells and the highest relaxation effects on rat basilar artery. The antioxidant effects of Danshen water-extracts linearly correlated to their relaxation effects (r = 0.895 to 0.977). Through multiple linear regression analysis, danshensu was found to be the most significant marker in the antioxidant and vasodilation effects of Danshen water-extract, while tanshinone IIA as the marker on hydrogen peroxide-induced apoptosis in rat heart H9c2 cells. Danshensu is, therefore, a useful marker for the quality control of Danshen water-extracts in antioxidant and vasodilation, while tanshinone IIA for anti-apoptotic potential of water-extracts. / Tanshinones, the major lipid-soluble components isolated from Danshen, have been reported for their anti-cancer potential in various cell lines and tumor-bearing mice models. Their anti-cancer mechanisms are also well-studied, mainly through cell cycle arrest, caspase-dependent apoptotic pathways and mitogen activated protein kinase (MAPK) signaling pathways. Miltirone, another abietane type-diterpene quinone isolated from Danshen, has been reported for its anti-oxidative, anxiolytic and anti-cancer effects. This study evaluated the apoptotic effect of miltirone and the underlying mechanisms in a human hepatoma HepG2 cell line and its p-glycoprotein (P-gp)-overexpressed doxorubicin-resistant counterpart (R-HepG2). Miltirone showed similar cytotoxicity in HepG2 (EC₅₀ = 7.06 μM) and R-HepG2 (EC₅₀ = 12.0 μM), demonstrated synergistic effects (1.56 - 6.25 μM) with doxorubicin (DOX) on the growth inhibition of R-HepG2 (synergism: 0.3 < CI < 0.5 at 50 % inhibition). Flow cytometric analysis showed that miltirone decreased P-gp-mediated DOX efflux in R-HepG2, and molecular docking studies illustrated that this effect was through inhibition on the active site of P-gp. At non-necrotic concentrations (25 μM or below), miltirone activated caspase-dependent apoptotic pathways, and induced the generation of reactive oxygen species (ROS) and oxidative stress which triggered ROS-mediated MAPK signaling pathways, including p38 MAPK, stress-activated protein kinase / c-Jun N-terminal kinase (SAPK/JNK) and extracellular regulated kinase 1/2, in both HepG2 and R-HepG2 cells. It is therefore concluded that miltirone is a dual inhibitor on P-gp and cell proliferation in R-HepG2 cells, with potential for the treatment of human hepatocellular carcinoma (HCC). / In order to improve the successful rates in drug development, the in vivo metabolic parameters of new chemical entities (NCEs), such as protein bindings, clearance rate, pharmacokinetic parameters and metabolism-based drug-drug interactions, should be considered at the early stage of drug discovery. Previous studies have shown that major tanshinones isolated from Danshen inhibited the metabolism of model probe substrates of human and rat CYP450 enzymes, with potential in causing herb-drug interactions. The aim of this study was to study the effect of miltirone on the metabolism of model probe substrates of CYP1A2, 2C9, 2D6 and 3A4 in pooled human liver microsomes. Miltirone showed moderate inhibition on CYP1A2 (IC₅₀ = 1.73 μM) and CYP2C9 (IC₅₀ = 8.61 μM), and weak inhibition on CYP2D6 (IC₅₀ = 30.20 μM) and CYP3A4 (IC₅₀ = 33.88 μM). Enzyme kinetic studies showed that miltirone competitively inhibited CYP2C9 (Ki = 1.48 μM), and displayed mixed type inhibitions on CYP1A2, CYP2D6 and CYP3A4 with Ki values of 3.17 μM, 24.25 μM and 35.09 μM, respectively. Molecular docking study further confirmed the ligand-binding conformations of miltirone in the active sites of human CYP450 isoforms. These findings suggested that miltirone may have potential drug-drug interactions with CYP1A2- and CYP2C9-metabolized drugs, and to a lesser extent with CYP2D6- and CYP3A4-metabolized drugs. / In conclusion, this study investigated the effects of Danshen water-extracts produced by different extraction methods on the chemical compositions and pharmacological activities, and consequently confirmed the biomarkers for the quality control of Danshen water-extracts for different medicinal uses. This study also demonstrated the anti-cancer potential of miltirone for HCC in vitro and the metabolism-based drug-drug interactions for its drug-likeness, which may provide useful and promising data for in vivo anti-cancer study of miltirone and further pre-clinical studies. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Zhou, Xuelin / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 195-224). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Abstract --- p.i / 論文摘要 --- p.v / Publications based on the work in this thesis --- p.viii / Acknowledgements --- p.x / Abbreviations --- p.xii / Table of Contents --- p.xv / Chapter Chapter 1 --- General introduction --- p.1 / Chapter 1.1 --- Reactive oxygen species and carcinogenesis --- p.1 / Chapter 1.2 --- Reactive oxygen species and tumor progression & metastasis --- p.2 / Chapter 1.3 --- Antioxidant enzymes in chemoprevention and chemotherapy --- p.3 / Chapter 1.3.1 --- Glutathione and Glutathione reductase --- p.5 / Chapter 1.3.2 --- Glutathione Peroxidase --- p.5 / Chapter 1.3.3 --- Glutathione S-transferases --- p.6 / Chapter 1.3.4 --- NAD(P)H: quinone reductase 1 --- p.7 / Chapter 1.3.5 --- Heme oxygenase-1 --- p.8 / Chapter 1.3.6 --- Thioredoxin reductase --- p.9 / Chapter 1.3.7 --- Superoxide Dismutase --- p.10 / Chapter 1.3.8 --- Catalase --- p.11 / Chapter 1.4 --- Medicinal uses of Danshen --- p.12 / Chapter 1.5 --- Analysis of Danshen and its components --- p.14 / Chapter 1.6 --- Antioxidant effects of Danshen extract and its bioactive compounds in chemoprevention and chemotherapy-related disease --- p.19 / Chapter 1.7 --- Anti-cancer effects of tanshinones isolated from Danshen --- p.21 / Chapter 1.7.1 --- Tanshinone IIA --- p.22 / Chapter 1.7.2 --- Tanshinone I --- p.26 / Chapter 1.7.3 --- Cryptotanshinone --- p.27 / Chapter 1.7.4 --- Dihydrotanshinone --- p.27 / Chapter 1.8 --- Metabolism / disposition of Danshen and its major active ingredients --- p.28 / Chapter 1.9 --- Herb-drug interactions with Danshen --- p.31 / Chapter 1.10 --- Effects of Danshen (and its major active ingredients) on model probe substrates of CYP isoforms --- p.33 / Chapter 1.11 --- CYPs induction by Danshen and its active components --- p.38 / Chapter 1.12 --- Effects of Danshen / active ingredients on drug transporter proteins --- p.40 / Chapter 1.13 --- CYP450 inhibition screening for new chemical entity --- p.42 / Chapter 1.14 --- Molecular docking analysis --- p.44 / Chapter 1.15 --- The Aim of this study --- p.45 / Chapter Chapter 2 --- Quantitative and qualitative studies to evaluate the efficiency of different heat water-extractions --- p.48 / Chapter 2.1 --- Introduction --- p.48 / Chapter 2.2 --- Materials and methods --- p.51 / Chapter 2.2.1 --- Materials and apparatus --- p.51 / Chapter 2.2.2 --- Extraction procedures --- p.51 / Chapter 2.2.3 --- HPLC analysis --- p.54 / Chapter 2.2.4 --- DPPH assay and FRAP assay --- p.54 / Chapter 2.2.5 --- Inhibition of 2,2'-azobis-2-amidinopropane (AAPH)-induced haemolysis in human red blood cells --- p.55 / Chapter 2.2.6 --- Protective effects on hydrogen peroxide-induced apoptosis in rat heart H9c2 cells --- p.56 / Chapter 2.2.7 --- Vasodilation effects on rat basilar artery --- p.57 / Chapter 2.2.8 --- Statistical analysis --- p.58 / Chapter 2.3 --- Results and Discussion --- p.59 / Chapter 2.3.1 --- Chemical profiles analyzed by HPLC analysis --- p.59 / Chapter 2.3.2 --- DPPH assay and FRAP assay --- p.63 / Chapter 2.3.3 --- Inhibition of AAPH-induced haemolysis --- p.65 / Chapter 2.3.4 --- Protective effects on hydrogen peroxide-induced apoptosis --- p.69 / Chapter 2.3.5 --- Vasodilation effects on rat basilar artery --- p.71 / Chapter 2.3.6 --- Multiple linear regression analysis --- p.76 / Chapter Chapter 3 --- Effects of miltirone on cell proliferation in a hepatoma HepG2 cell line and its doxorubicin-resistant counterpart --- p.83 / Chapter 3.1 --- Introduction --- p.83 / Chapter 3.2 --- Materials and Methods --- p.87 / Chapter 3.2.1 --- Chemicals --- p.87 / Chapter 3.2.2 --- Cell culture --- p.87 / Chapter 3.2.3 --- Cell viability test --- p.88 / Chapter 3.2.4 --- Drug-efflux study by flow cytometry --- p.89 / Chapter 3.2.5 --- Molecular docking study and Ligand-based prediction --- p.90 / Chapter 3.2.6 --- Measurement of ROS generation by confocal microscopy and flow cytometry --- p.91 / Chapter 3.2.7 --- GSH and GSSG determination for oxidative stress --- p.93 / Chapter 3.2.8 --- Apoptosis-related proteins expression detected by Western blotting analysis --- p.94 / Chapter 3.2.9 --- Data analysis --- p.96 / Chapter 3.3 --- Results --- p.97 / Chapter 3.3.1 --- Cytotoxicity in hepatoma cells --- p.97 / Chapter 3.3.2 --- Drug-efflux study by flow cytometry --- p.104 / Chapter 3.3.3 --- Molecular docking study and Ligand-based prediction --- p.108 / Chapter 3.3.4 --- ROS generation --- p.113 / Chapter 3.3.5 --- Determination of GSH/GSSG ratio --- p.117 / Chapter 3.3.6 --- Caspase-dependent apoptosis. --- p.121 / Chapter 3.3.7 --- Phosphorylation of MAPKs --- p.126 / Chapter 3.4 --- Discussion --- p.134 / Chapter Chapter 4 --- Enzyme kinetic and molecular docking studies of miltirone on major human cytochrome P450 isozymes inhibitions --- p.139 / Chapter 4.1 --- Introduction --- p.139 / Chapter 4.2 --- Material and Methods --- p.141 / Chapter 4.2.1 --- Materials and Reagents --- p.141 / Chapter 4.2.2 --- Incubation conditions --- p.142 / Chapter 4.2.3 --- Samples preparation --- p.143 / Chapter 4.2.4 --- HPLC analysis --- p.143 / Chapter 4.2.5 --- CYP inhibition and enzymatic kinetic study --- p.144 / Chapter 4.2.6 --- Molecular docking analysis --- p.145 / Chapter 4.2.7 --- Data analysis --- p.146 / Chapter 4.3 --- Results --- p.148 / Chapter 4.3.1 --- CYP inhibition and enzymatic kinetic study --- p.148 / Chapter 4.3.2 --- Molecular docking study of miltirone --- p.167 / Chapter 4.4 --- Discussions --- p.184 / Chapter Chapter 5 --- General discussion --- p.188 / References --- p.195

惡性腫瘤嚴重威脅著人類的身體健康，其治療也成為人類關注的焦點。傳統的化學療法和放射療法由於缺乏特異性，取得療效的同時往往也帶來較大的毒副作用。隨著對腫瘤發生發展分子機制認識的不斷深入，腫瘤的基因治療已成為攻克和治愈腫瘤最具希望和挑戰的研究領域。近年來研究發現骨髓間充斥幹細胞(MSCs)可被募集至腫瘤或損傷部位并參與腫瘤生長或組織修復，研究證明間充斥幹細胞通過靜脈注入帶瘤鼠（比如乳腺癌、膠質瘤、結腸癌及黑色素瘤）體內后，特異性的分佈于生長中的腫瘤中。這種特異性向腫瘤組織趨化轉移的特性使得骨髓間充斥幹細胞成為腫瘤基因靶向治療的載體的理想細胞。酶蛋白基因如單純皰疹病毒胸苷激酶(HSV-TK)可以使一些無毒或低毒的前藥轉化為強細胞毒性物質，殺死腫瘤細胞。我們前期實驗結果表明，通過遺傳改造后的表達TK基因的MSCs在GCV的存在下，具有殺傷腫瘤細胞抑制腫瘤生長的能力。但沒有改造的MSCs遷移至腫瘤之後可能會分化成成纖維細胞或者腫瘤基質細胞等支持腫瘤生長，但其命運和影響到底如何，我們怎麼樣進一步促進其向腫瘤的遷移以提高殺傷腫瘤的效率是本研究需要解決的問題。 / 本研究擬採用免疫螢光組織化學技術和分子生物學等技術研究和觀察MSCs對腫瘤（以乳腺癌，前列腺癌為例）的趨化過程及其在腫瘤生長中的作用，在在此基礎上研究促進攜帶HSV-TK自殺基因的MSCs的腫瘤靶向性細胞治療策略，採用分子和細胞生物學等方法評估其對荷瘤鼠體內腫瘤殺傷的原理，為利用TK-MSCs腫瘤的靶向治療奠定基礎。 / 研究結果顯示體外共培養的條件下，小鼠骨髓間充斥幹細胞可促進小鼠乳腺癌細胞增長，且增長速度同培養體系中間充斥幹細胞數目呈正相關。將兩種細胞混合注射于裸鼠體內，相比共注射小鼠皮膚成纖維細胞，間充斥幹細胞可促進體內腫瘤生長。使用人胚胎骨髓間充斥幹細胞和前列腺癌細胞可得出類似的效果。將腫瘤組織切片分析發現間充斥幹細胞促進體內腫瘤細胞增殖的同時，提高了腫瘤組織內血管密度。體外實驗發現共培養前列腺癌細胞和間充斥幹細胞可促進血管生成且在間充斥細胞內同血管增生相關的蛋白表達量都有相應提高，進一步證實間充斥幹細胞可能通過促進血管增生從而促進腫瘤生長。另外，我們利用人胚胎來源的骨髓間充斥幹細胞建立了穩定表達TK自殺基因的細胞系，且在GCV的存在下具有抑制腫瘤生長的能力。為了促進它們向腫瘤遷移的能力，我們用多柔比星預處理腫瘤細胞，和沒處理過的對照組相比，能增強對表達TK的間充斥幹細胞的招募能力。且在聯合利用多柔比星和TK的條件下，腫瘤生長能得到較大程度的抑制，這種抑制作用强於單獨使用多柔比星和表達自殺基因的間充斥幹細胞系統。初步認為是多柔比星的處理能增強腫瘤組織內炎性介質的分泌從而增強間充斥幹細胞的遷移達到增強自殺基因系統殺死腫瘤細胞的目的。 / 總的來說，雖然間充質幹細胞對腫瘤的生長存在一定的促進作用，但我們仍能對其進行遺傳改造，且在其它抗腫瘤藥的配合下達到最大的抗腫瘤效果。 / Eradication of cancer, especially when it has metastasized is extremely difficult and conventional cancer therapies are simply unable to specifically target tumors/cancers, thus causing unwanted side effects and complications. Recently, it has been shown that bone marrow mesenchymal stem cells (MSCs) are able to migrate specifically to tumors and contribute to the formation of tumor-associated stroma. These properties make MSCs good candidates as anti-tumor agent delivery vehicles and lead to a great deal of interest in the possibility of genetically modifying MSCs to express anticancer molecules and using them as specific targeted anticancer agents. We and others have showed that MSCs have the ability to migrate towards various cancer cells including breast, colon, fibrosarcoma and prostate cancer cells. Suicide gene therapy is widely used in cancer gene therapy. When stably infected with herpes simplex virus thymidine kinase gene by lentivirus, TK-MSCs maintained their MSCs characters and tumor tropism potential and significantly inhibited tumor growth, in the presence of the pro-drug ganciclovir (GCV). Improve MSCs homing to tumor tissue as anti-tumor gene therapy vehicles and maximizing their tumor killing effects is highly warranted. Furthermore, MSCs interact with tumor cells in numerous ways, which have the potential to support or suppress tumor growth. Therefore the fate and role of MSCs engrafted in tumor sites need to be clarified in order to making better use of these cells as anti-cancer agent delivery vehicles. / The aims of the current study are: (1) to study the role and fate of MSCs homed into the tumors; (2) to establish human bone marrow MSCs that stably express the TK genes; (3) to investigate the methods that enhance the anti-tumor efficiency of TK-MSCs. / In this study, bone marrow-derived mesenchymal stem cells from mice or human fetus were isolated and characterized. Effects of BM-MSCs on tumor cell proliferation in vitro were analyzed in a co-culture system with mouse breast cancer cell 4T1 cells. Both co-culture with BM-MSCs and treatment with MSC-conditioned medium led to enhanced growth of 4T1 cells. Co-injection of 4T1 cells and MSCs into nude mice led to increased tumor size compared with injection of 4T1 cells alone. Identical experiments using human prostate cancer cell DU145 cells and hBM-MSCs instead of 4T1 cells and mBM-MSCs yielded similar results. Compared with tumors induced by injection of cancer cells alone, tumor vessel area was greater in tumors from co-injection of 4T1 or DU145 with BM-MSCs, which correlated with decreased central tumor necrosis and increased tumor cell proliferation. Furthermore, both conditioned medium from co-cultures of hBM-MSCs and DU145 cells or hBM-MSCs alone was able to induce angiogenesis in human umbilical vein endothelial cells (HUVEC). When hBM-MSCs exposed to DU145 cells environment, the expression of markers associated with neovascularization (α-SMA, VEGF, TGF-β and IL6) were increased. Together, these results indicate that MSCs promote tumor growth both in vitro and in vivo and suggest that tumor promotion in vivo may be attributable in part to enhanced angiogenesis. / Immortalized human fetal bone marrow-derived MSCs (hfBMSCs) expressing herpes simplex virus thymidie kinase was established by conventional lentiviral transduction method. Functional expression of TK was evaluated by cytotoxicity in the presence of its prodrug GCV. SV40-TK-hfBMSCs exhibited comparable proliferation, surface phenotype expression, multi-differentiation potential and tumor-tropic migration ability as hfBMSCs. By measurement of tumor volume, repeated injection of the SV40-TK-hfBMSCs and subsequent consecutive GCV administration could suppress tumor growth in DU145 or PC3 human prostate tumor xenograft nude mice model without causing weight loss. However, its clinical applications are still limited. Alternative strategies have been pursued in this study by the use of combination therapy with cytotoxic chemotherapy to improve the overall efficacy of the TK-hfBMSCs/GCV system. / TK-hfBMSCs/GCV was evaluated alone or combined with low-dose doxorubicin in human prostate carcinoma DU145 xenografts in nude mice, testing for effects on local growth and overall survival. Tissues were evaluated through immunofluorescence and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining (TUNEL) for treatment effects on tumor cell proliferation and apoptosis. Transwell migration assay was used to access the migration ability of TK-hfBMSCs to tumor cells upon doxorubicin treatment and caspase-3 activity was conducted for test the tumor cells apoptosis under TK-hfBMSCs/GCV, doxorubicine, or combination of the two compound treatments respectively. Only minimal growth inhibition was observed in DU145 after treatment with TK-hfBMSCs/GCV or doxorubicin alone at doses and time points as indicated. In contrast, the combination of both agents resulted in a significant growth inhibition. Caspase-3, plays a central role in the execution-phase of cell apoptosis, was increased by TK-hfBMSCs/GCV or doxorubicine and also to a much greater extent by the combination treatment. Treatment by TK-hfBMSCs/GCV resulted in only a slight decrease in tumor growth compared with controls. Treatment with low-dose doxorubicin alone resulted in a small, nonstatistically significant decrease in tumor growth; In contrast, combined low-dose doxorubicin and TK-hfBMSCs/GCV was markedly inhibitory compared with control, doxorubicin alone, or TK-hfBMSCs/GCV alone. During the whole treatment process, no significant weight loss was observed. Furthermore, combined therapy induced increased area of necrosis, significant apoptosis and decreased tumor cell proliferation in treated tumors. Taken together, low dosage of doxorubicin could be used in combination with TK-hfBMSCs based suicide gene therapy. / In conclusion, we have demonstrated that BM-MSCs could increase the growth of human prostate cancer and mouse breast cancer. The promotion effect may partly attribute to the increased expression of pro-angiogenic factors in BM-MSCs in tumor microenvironment and subsequent enhancement in angiogenesis and tumor growth. The current study also suggests combination of TK-hfBMSCs/GCV and doxorubicin was more effective in inhibiting prostate cancer cells growth than TK-hfBMSCs/GCV or doxorubicin alone. Although many problems need to be resolved for further application, our study provided the possibility of a new strategy of suicide gene-based therapy accompanied by low dosage of chemotherapy in treating prostate cancer. Therefore MSCs were described as a “double-edged sword in their interaction with tumors. However, if MSCs are suitably engineered with anticancer genes they could be employed as a valuable “single-edged sword“ against cancers. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Zhang, Ting. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 120-158). / Abstracts also in Chinese. / ACKNOWLEDGEMENT --- p.ii / PUBLICATIONS --- p.vii / ABSTRACT --- p.xiii / Chapter CHAPTER 1 --- Introduction --- p.1 / Chapter 1.1 --- Mesenchymal stem cells (MSCs) --- p.2 / Chapter 1.2 --- Tumor microenvironment and involvement of MSCs in tumor establishment --- p.5 / Chapter 1.3 --- Tumors-tropic characteristics of MSCs --- p.15 / Chapter 1.4 --- Impact of MSCs on in vivo tumors --- p.21 / Chapter 1.5 --- In vivo imaging demonstrating MSCs tumor-homing potentials --- p.25 / Chapter 1.6 --- Evidence for use of MSCs as anti-tumor agents delivery vehicles --- p.26 / Chapter 1.7 --- Homing strategies to enhance efficacy and safety of MSCs therapy --- p.32 / Chapter 1.8 --- Summary --- p.35 / Chapter CHAPTER 2 --- Hypotheses, Objectives and Study Design --- p.35 / Chapter 2.1 --- Hypothesis --- p.35 / Chapter 2.2 --- Objective --- p.36 / Chapter 2.3 --- Study design --- p.37 / Chapter CHAPTER 3 --- Bone Marrow-derived Mesenchymal Stem Cells Promote Growth and Angiogenesis of Breast and Prostate Tumors (Study I) --- p.40 / Chapter 3.1 --- Materials and Methods --- p.40 / Chapter 3.2 --- Results --- p.49 / Chapter 3.3 --- Discussion --- p.64 / Chapter 3.4 --- Conclusions --- p.67 / Chapter CHAPTER 4 --- Immortalized human fetal bone marrow-derived mesenchymal stem cell expressing anti-tumor suicide gene for anti-tumor therapy in vitro and in vivo (Study II) --- p.68 / Chapter 4.1 --- Materials and Methods --- p.68 / Chapter 4.2 --- Results --- p.73 / Chapter 4.3 --- Discussion --- p.85 / Chapter CHAPTER 5 --- Enhanced antitumor effects by combination therapy using mesenchymal stem cell expressing anti-tumor suicide gene and Doxorubicin in a xenograft mouse model (Study III) --- p.89 / Chapter 5.1 --- Materials and Methods --- p.89 / Chapter 5.2 --- Results --- p.97 / Chapter 5.3 --- Discussion --- p.111 / Chapter CHAPTER 6 --- General discussion and conclusions --- p.116 / Chapter 6.1 --- General discussion --- p.116 / Chapter 6.2 --- General conclusions --- p.119 / FUNDING --- p.120 / REFERENCE --- p.120

Personalized cancer medicine is a cancer care paradigm in which diagnostic and therapeutic strategies are customized for individual patients. Microsystems that are created by Micro-Electro-Mechanical Systems (MEMS) technology and integrate various diagnostic and therapeutic methods on a single chip hold great potential to enable personalized cancer medicine. Toward ultimate realization of such microsystems, this thesis focuses on developing critical functional building blocks that perform genetic variation identification (single-nucleotide polymorphism (SNP) genotyping) and specific, efficient and flexible cell manipulation on microfluidic surfaces. For the identification of genetic variations, we first present a bead-based approach to detect single-base mutations by performing single-base extension (SBE) of SNP specific primers on solid surfaces. Successful genotyping of the SNP on exon 1 of HBB gene demonstrates the potential of the device for simple, rapid, and accurate detection of SNPs. In addition, a multi-step solution-based approach, which integrates SBE with mass-tagged dideoxynucleotides and solid-phase purification of extension products, is also presented. Rapid, accurate and simultaneous detection of 4 loci on a synthetic template demonstrates the capability of multiplex genotyping with reduced consumption of samples and reagents. For cell manipulation, we first present a microfluidic device for cell purification with surface-immobilized aptamers, exploiting the strong temperature dependence of the affinity binding between aptamers and cells. Further, we demonstrate the feasibility of using aptamers to specifically separate target cells from a heterogeneous solution and employing environmental changes to retrieve purified cells. Moreover, spatially specific capture and selective temperature-mediated release of cells on design-specified areas is presented, which demonstrates the ability to establish cell arrays on pre-defined regions and to collect only specifically selected cell groups for downstream analysis. We also investigate tunable microfluidic trapping of cells by exploiting the large compliance of elastomers to create an array of cell-trapping microstructures, whose dimensions can be mechanically modulated by inducing uniform strain via the application of external force. Cell trapping under different strain modulations has been studied, and capture of a predetermined number of cells, from single cells to multiple cells, has been achieved. In addition, to address the lack of aptamers for targets of interest, which is a major hindrance to aptamer-based cell manipulation, we present a microfluidic device for synthetically isolating cell-targeting aptamers from a randomized single-strand DNA (ssDNA) library, integrating cell culturing with affinity selection and amplification of cell-binding ssDNA. Multi-round aptamer isolation on a single chip has also been realized by using pressure-driven flow. Finally, some perspectives on future work are presented, and strategies and notable issues are discussed for further development of MEMS/microfluidics-based devices for personalized cancer medicine.

The overall goal of this thesis was to determine whether optical tomographic imaging can be employed to predict treatment outcome in women with breast cancer (BC) who undergo neoadjuvant chemotherapy (NACT). NACT is widely applied as a standard treatment for patients with newly diagnosed operable invasive BC. Only about 13-30% of women have a response to this treatment. Furthermore, NACT is an expensive and toxic treatment that takes several months to completely administer. In order to know the response of the patient, physicians usually need to wait until the months of NACT has finished and the patient has undergo surgery in which they receive the pathology. If the long-term treatment response could be predicted early into the treatment regimen, the patient would be relieved of any unnecessary side effects and alternative treatments could be initiated. We have used a novel dynamic DOT system to study the effects of targeted NACT. Unlike X-ray imaging, which requires potentially harmful ionizing radiation, DOT can be applied without side effects, which is particularly important in the case of multiple imaging sessions to be performed over the course of treatment. We have tracked 40 subjects and imaged them at 6 different time points during their NACT. For this study, two different types of data were collected: static (single 3D image) and dynamic (3D movies). The combination of the data may be used to accurately determine the response of the patient. With non-invasive, non-ionizing DOT imaging we have been able to determine within two weeks if the patient will respond to treatment with an accuracy as high as 94.1%.