The Gastric Cancer Novel Drug Screening And Targeting Detection Based On Synthesis Of Optical Moleuclar Imaging Probes

Recently, with the completion of the Human Genome Project and breakthroughs inrelated fields of biological sciences and technologies, including cell biology, molecularbiology and proteomics etc., the new generation of medical imaging, i.e. molecularimaging(MI), was born due to the combination of traditional imaging methods and theadvances. With the ability of monitoring the physiological process at cellular andmolecular levels in vivo, MI provides new techniques of acquiring and analyzinginformation for various applications, such as qualitative and quantitative gene functionalresearch, metabolism, occurance and development mechanisms of diseases, monitoringreport, targeted treatment, therapeutic effects evaluation and so on, and holds thepromise of realizing early and accurate diagnosis and effective treatment. As one of themodalities of MI, optical imaging has been exploited widely due to its low energy, noionizing radiation, high sensity, high time resolution, low-cost and ease of use etc.Molecular probe is the preliminary requirement of MI. Constructing gastric cancer(GC) targeted optical molecular probes and utilizing optical MI on early diagnosis,monitoring, targeted treatment, effect evaluation and novel drug development and so on,has significant impact on helping making breakthrough on GC research. The work ofthis dissertation based on constructing targeted optical molecular probes for GC relatedresearch: constructing a GC cell line stablely expressing firefly luciferase (FLuc) forbioluminescence imaging (BLI), therapeutic effect evalutation of novel targetinganti-GC drug on the cell line based on BLI, assessing the liability of real-time tracingGC cells in vivo and the effectiveness of research on GC progrssion, metastasis andnovel drug screening with BLI using the constructed cell line as the biological model.On the other hand, a series of new promising cyanine dyes were constructed andconjugated with a GC specific molecule, consisting a GC specific fluorescent probe. GCvisualized detection based on fluorescent molecule imaging (FMI) in vivo was exploited.The modifiability of the series of probes, specificity of the GC specific molecule andliability of specific diagnosis of GC using the series of specific-target probes wasdiscussed. The main work of the dissertation includes:1. Constructing a GC cell subline SGC-7901-luc which express FLuc reporter genestably. GC cell subline SGC-7901-luc expressing exogenous Fluc reporter gene wasconstructed by lentiviral transfection. And it provides a reliable biological model fortracing the growth of GC cells in vivo.2. In vitro inhibition effect of ApoG2which is a novel Bcl-2/Bcl-xL small-molecule inhibitor for gastric carcinoma cells was explored. Defective apoptosis is an importantprocess for tumorigenesis. Chemotherapy induced apoptosis is an effective treatmentstrategy for GC. However, failure of the treatment usually occur. One of the mainreasons is the resistance of traditional chemotherapeutic drugs. Thus, discovering newchemotherapeutic drugs is important to the management of GC. And several Bcl-2small-molecule inhibitors were used for anti-cancer treatment studies in recent years.Therefore, ApoG2，a non-peptide small-molecule inhibitors of Bcl-2family proteins,was used to evaluate the anti-cancer effect in vitro on GC. The results showed ApoG2could effectively inhibit the growth and proliferation of GC cells through themitochondrial pathway of apoptosis.3. Furthermore, in vivo anti-tumor activity of ApoG2was explored. Based onSGC-7901-luc cell subline mentioned above, real-time tracing ApoG2-treated GC cellsin vivo was performed by BLI. The results revealed that ApoG2induced apoptosis inGC cells was associated with the endoplasmic reticulum stress-induced apoptosispathway, and then had anti-cancer effects with little toxicity in vivo. In general, ApoG2,being used as a potent specific-target chemotherapeutic drug, has clinical transitionalpotent.4. A series of novel penta-methine cyanine dyes with low-toxicity, stronglight-stability, membrane-permeability and high biological-activity were synthesizedand a new effective method of synthesis of cyanine dyes was promoted. For the sake ofsmall molecular weight, less toxicity, broad spectra tunability and large molar extinctioncoefficient, cyanine dyes were widely used as fluorescent marker. But, for the limitedchoices, poor light-stability, low fluorescence quantum yield, complicated synthesisroute, large percentage of byproduct, burden of purification and therefore high cost,there’re only a few kinds available (usually IRDye800and Cy5.5), hence application intumor research based on FMI was limited. Cyanine dye synthesis and enhancementmethods were surveyed. With symmetrical synthesis method, the novel penta-methinecyanine dyes were synthesized via the subsitution of central chloro-atoms in thepolymethine chain by mercapto group and introduction bioactivity group (carboxylgroup:COOH) in its structure. Variation of cyanine dyes were enhanced and the basis ofFMI-related research was formed. The promoted novel effective synthesis method ofcyanine dyes solves the problems of complex route, high-yield of byproduct, low yieldof product, burden of purification and high cost.5. A GC specific-target fluorescent probe targeting GC vasculature was constructed and targeting tracing of GC in vivo with FMI was studied. In the dissertation, the GCvasculature specific-target peptide GX1, which has good bioaffinity, was chosen as GCspecific-target marker. GX1was conjugated with our novel penta-methine cyanine dyeCyIC, consisting GC specific-target CyIC-GX1. With in vitro and in vivo FMI andtumor cell versus nude mice competitive inhibition experiments, the synthesized probewas found be able to bind with GC vascular endothelial cell specifically. Therefore,newly constructed penta-methine cyanine dye is suitable for FMI and supplements thechoice of fluorescent marker. In addition, GX1, which was screened out by CancerBiology State Key Laboratory, Digest Department, Xijing Hospital, has great cancervasculature target-specificity, especially GC, and can be used as GC specific marker forrelated GC diagnosis research, and has clinical transitional potent.