201212892 六、發明說明： [相關申請之交叉引用] 本申請係將2008年8月19曰提出申請且發明名稱為 「將大分子物質導入活體標靶細胞之方法及系統」(METHOD AND SYSTEM FOR LEADING MACROMOLECULE SUBSTANCE INTO LIVING TARGET CELLS)之美國專利申請案第 12/194,497 號、2008年5月15日提出申請且發明名稱為「將大分子 物質導入活體標把細胞之方法及系統」之美國專利申請宰 第12/121，712號、2008年8月19日獲准專利且發明名稱 鲁 為「將大分子物質導入活體標把細胞之方法及系統」之美 國專利第7, 415, 302號、以及2003年1〇月15日提出申請 之台灣專利申請案第092128522號，上述全部專利申請案 及專利藉由引用併入本文。 【發明所屬之技術領域】 本發明通常係關於將大分子物質導入標靶細胞之方 法及系統’更特別地’係關於應用超音波以調節標乾細胞 細胞膜之穿透性，藉以有效地將低劑量之大分子物質導入 · 標靶細胞的方法及系統。 【先前技術】 人體之組織細胞有時受内部或外部有害因子刺激而 致使其染病，其結果為染病細胞之數目迅速增加，且染病 細胞轉移至健康組織’因而形成腫瘤。腫瘤包括良性腫瘤 及惡性腫瘤。與良性腫瘤相比，惡性腫瘤難以治愈，且對 人體危害更大。 111758 4 201212892 » 目前，每年有5, 000, 000人死於腫瘤，而惡性腫瘤為 主要殺手。隨著醫療科學之發展，已提供有多種先進之腫 瘤診斷方法及治療方法。腫瘤治療方法主要包括外科手 術、化學療法及放射療法。於化學療法之治療中，以低用 藥精確性向人體散佈藥物所產生之毒性，此仍待解決之局 限性及缺陷’往往吞噬著腫瘤患者的健康。因此，如何以 最小藥物劑量達成最大治療效果’以及如何提升用藥精確 性，為人們亟待克服之問題。 φ 近期之研究發現’體外震波碎石術（shock wave lithotripsy, SWL)可於細胞周圍產生微泡。此等微泡於細 胞膜中形成非永久性孔，因此，可提升細胞膜之穿透性， 並達成更好之藥物吸收性。美國專利第6 298,264號揭露 一種提升細胞膜穿透性的方法。該方法應用第一脈衝波 (pulsed wave，PW)及第二脈衝波以產生環繞細胞之微泡。 此等微泡於細胞膜中形成非永久性孔，以提升細胞膜之穿 φ透性。該方法可將細胞膜之穿透性增加至90%，因此，僅 需要低的藥物劑量。然而，該方法並未揭露如何精確定位 標靶細胞以及如何提升用藥精確度。因此，仍亟待需要精 確地定位標乾細胞並提升用藥精確度的方法。 【發明内容】 本發明之主要目的在於提供將大分子物質有效地導 入標靶細胞之方法及系統。 本發明之另-目的在於提供應用於基因輸送中之方 法及系統，以增加基因輸送之效率。 111758 5 201212892 本發明之再一目的在於提供應用於基因輸送中之方 法及糸統’以升基因療法之效率。 本發明之又一目的在於提供提升用藥精確度之方法 及系統。 本發明之再3 -目的在於提供降鋪_量並 地將藥物導入腫瘤細胞的方法及系統。 依照上述及其他目的，本發明提供將大分子物質導入 活體標乾細胞之方法及系統。該將大分子物質導入 起細胞之系統包含：影像絲單元，該影像擷取單元係^ 於榻取標無細胞所處之組織或n官的三維（3D)結構聲像 標乾細胞所處之組織或ϋ官的3Djk管攝影影像；影像 單元，該影像合成單元係用以將3D結構影像合併入汕血 管攝影影像，藉以選擇完全涵蓋傳輸大分子物質之標靶細 胞的金管通道；注射料’魅射單元係祕注射:體^ 傳輸大分子物質至該標把細胞；能量轉換模組，該能量轉 換模組係用於施加能量’以活化該液體並產生生物效鹿. 其中’該能量轉換模組係包含包括超音波轉換器或^擴 音器（tweeter)之超音波轉換模組’藉以於該絲細胞之: 胞膜中形祕永久性額；其中，大分子物質通過標說細 胞細胞膜中的非永久性孔進入標靶細胞中。 該將大分子物質導入活體標靶細胞之方法包含：首 先，擷取標靶細胞所處之組織或器官的三維（31))結構影像 及才示乾細胞所處之組織或器s的3D j6l管攝景;j产彡像.第 將3D結構影像合併入3D血管攝影影像，選^完全涵蓋傳 111758 6 201212892 r 輸大分子物質之標靶細胞的血管通道；第三，使用導管沿 著所選擇之血管通道注射微泡液體（超音波或人造血液）， 該微泡環繞著標靶細胞排列；第四，施加能量以活化該微 泡液體，以產生生物效應，藉以於該標靶細胞之細胞膜内 形成非永久性孔洞；以及最後，經由細胞膜内之非永久性 孔洞，沿著所選擇之血管通道，將大分子物質注射進入標 把細胞中。 與傳統醫療方法及系統相比，本發明之將大分子物質 # 導入活體標乾細胞之方法及系統擷取標乾細胞所處之組織 或器官的三維（3D)結構影像及標靶細胞所處之組織或器官 的3D血管攝影影像；將3D結構影像合併入3D血管攝影影 像，藉以精確定位該標靶細胞，以選擇完全涵蓋標靶細胞 的最有效血管通道；以及沿著所選擇之血管通道將大分子 物質注射進入標靶細胞中。隨後，該方法及系統施加能量 以活化環繞標靶細胞排列之微泡，以產生生物效應，藉以 於標靶細胞之細胞膜中形成非永久性孔洞。大分子物質係 通過標靶細胞細胞膜中之非永久性孔洞進入標乾細胞中。 因此，本發明之將大分子物質導入活體標靶細胞之方法及 系統具有諸多優點，如低藥物劑量、低成本、精確用藥及 有效治愈等效果。 為提供本發明之進一步理解，下述之詳細說明書例示 性說明本發明之具體實施態樣及實施例。應了解的是，此 詳細說明書僅提供本發明之例示性說明，而非作為本發明 範圍的限制。 7 111758 201212892 ' ^ 【實施方式】 本發明通常係關於將大分子物質引入標靶細胞之方 法及系統；更特別是關於應用超音波以調節標靶細胞之細 胞膜之穿透性，藉以有效地將低劑量之大分子物質導入標 靶細胞的方法及系統。下列敘述之呈現使具本技術領域中 具通常知識者可以完成及使用本發明，並提供於專利申請 案及其必要文件之内文中。熟悉本技藝之人士很容易對本 文揭示之較佳具體實施態樣、通常原理及特徵作出各種修 飾。因此，本發明並非欲限制為所顯示之該等具體實施態 樣，而係與本文所揭示之該等原理及特徵之最大範疇一致。 本發明之將大分子物質導入活體標靶細胞之方法及 系統可應用於多種不同領域，如基因輸送、基因療法、藥 物傳輸、部份用藥及腫瘤治療。本發明尤其適用於腫瘤治 療，更特別是實體瘤之治療。舉例而言，於實體瘤之治療 中，一般係將電腦斷層掃描（CT)或磁共振成像（MRI)作為預 備步驟。藉由該預備步驟取得腫瘤細胞所處之組織或器官 的三維（3D)結構影像，以作為後續治療（如外科手術、化學 療法及放射療法）之基礎。 請參閱第1A圖，依照本發明之較佳具體實施例之將 大分子物質導入活體標靶細胞之系統的基本結構係於第 1A圖中例示性說明，包括第2圖，僅係將大分子物質導入 活體標靶細胞之系統1之主要元件的簡明例示性說明。實 際使用之系統1可更為複雜。 將大分子物質導入活體標靶細胞之系統1包含影像擷 8 111758 201212892 =元刚、影像合成單元11G、注射單元m a能量轉換 模組13 0。於本具體音始你丨由 + 、貫關巾，影_取單元⑽、影像合 早凡 〆主射單A 120及能量轉換模’紐130受微處理 單元140所控制。 〜里 影像操取單元100用於掏取標無細胞所處之組織或器 官的二維（3D)結構影像及操取標乾細胞所處之血管的汕 攝影影像。於本具體實施例中，影㈣取單元100為下列 之/、中者CT裳置、觀裝置及血管攝影裝置。標乾細 •胞至少為一種腫瘤細胞。 通承CT |置利用扇$ X身十線從軸向掃描人體之斷 層’並利用一列檢測器接收穿透人體之訊號。當X射線發 射器固定在特定位置時，該等檢測器會從對應之特定層中201212892 VI. Description of the invention: [CROSS-REFERENCE TO RELATED APPLICATIONS] This application is filed on August 19, 2008, and the title of the invention is "Method and System for Introducing Macromolecular Substances into Living Target Cells" (METHOD AND SYSTEM FOR LEADING) U.S. Patent Application Serial No. 12/194,497, filed on May 15, 2008, which is hereby incorporated by reference in its entirety, the entire entire entire entire entire entire entire entire entire entire entire entire entire entire content US Patent No. 7, 415, 302, and 2003, which were granted patents on the basis of the patents and systems of No. 12/121,712, August 19, 2008, and the name of the invention, "Incorporating Macromolecular Substances into Living Standard Cells" Taiwan Patent Application No. 092128522, the entire disclosure of which is hereby incorporated by reference. FIELD OF THE INVENTION The present invention generally relates to a method and system for introducing macromolecular substances into target cells 'more particularly' with respect to the application of ultrasound to modulate the permeability of the stem cell membrane, thereby effectively lowering the dose. A method and system for introducing a macromolecular substance into a target cell. [Prior Art] Tissue cells of the human body are sometimes stimulated by internal or external harmful factors to cause disease, and as a result, the number of infected cells rapidly increases, and the infected cells are transferred to healthy tissues to form tumors. Tumors include benign tumors and malignant tumors. Compared with benign tumors, malignant tumors are difficult to cure and are more harmful to humans. 111758 4 201212892 » Currently, 50,000 people die each year from cancer, and malignant tumors are the main killers. With the development of medical science, a variety of advanced tumor diagnosis methods and treatment methods have been provided. Tumor treatment methods mainly include surgical surgery, chemotherapy, and radiation therapy. In the treatment of chemotherapy, the toxicity caused by spreading the drug to the human body with low drug accuracy, and the limitations and defects still to be solved, often devour the health of the tumor patient. Therefore, how to achieve the maximum therapeutic effect with the minimum drug dose and how to improve the accuracy of medication is a problem that people need to overcome. φ Recent studies have found that shock wave lithotripsy (SWL) produces microvesicles around cells. These microbubbles form non-permanent pores in the cell membrane, thereby improving cell membrane penetration and achieving better drug absorption. A method for increasing the permeability of a cell membrane is disclosed in U.S. Patent No. 6,298,264. The method applies a first pulsed wave (PW) and a second pulsed wave to generate microbubbles surrounding the cell. These microbubbles form non-permanent pores in the cell membrane to enhance the permeability of the cell membrane. This method increases the permeability of the cell membrane to 90% and, therefore, requires only a low drug dose. However, this approach does not reveal how to accurately target target cells and how to improve drug accuracy. Therefore, there is still a need for a method of accurately positioning the stem cells and improving the accuracy of the medication. SUMMARY OF THE INVENTION A primary object of the present invention is to provide a method and system for efficiently introducing macromolecular substances into target cells. Another object of the present invention is to provide methods and systems for use in gene delivery to increase the efficiency of gene delivery. 111758 5 201212892 A further object of the present invention is to provide a method for applying gene delivery and a system for the efficiency of gene therapy. It is still another object of the present invention to provide a method and system for improving the accuracy of medication. Still another object of the present invention is to provide a method and system for down-regulating and introducing a drug into a tumor cell. In accordance with the above and other objects, the present invention provides methods and systems for introducing macromolecular substances into living stem cells. The system for introducing a macromolecular substance into a cell comprises: a video silk unit, wherein the image capturing unit is located in a tissue in which the cell is not located, or a tissue in which the three-dimensional (3D) structure of the standard imaged stem cell is located. Or the 3Djk tube photographic image of the eunuch; the image unit is used to merge the 3D structure image into the angiography image to select the gold tube channel that completely covers the target cell for transporting the macromolecular substance; Injection unit secret injection: body ^ transports macromolecular substances to the target cells; energy conversion module, which is used to apply energy 'to activate the liquid and produce biological effect deer. Where 'the energy conversion mode The group includes an ultrasonic transducer module including an ultrasonic transducer or a tweeter, for the purpose of the filament cell: a permanent shape in the cell membrane; wherein the macromolecular substance passes through the cell membrane of the cell Non-permanent pores enter the target cells. The method for introducing a macromolecular substance into a living target cell comprises: firstly, capturing a three-dimensional (31)) structural image of a tissue or an organ in which the target cell is located, and a 3D j6l tube showing the tissue or device in which the stem cell is located. Photographing; j production of 彡 image. The 3D structure image is merged into 3D angiography image, select ^ completely covers the vascular channel of the target cell of 111758 6 201212892 r transfusion; third, use the catheter along the selected The vascular channel is injected with a microbubble liquid (ultrasonic or artificial blood) which is arranged around the target cells. Fourth, energy is applied to activate the microbubble liquid to produce a biological effect, whereby the cell membrane of the target cell Non-permanent pores are formed therein; and finally, macromolecular substances are injected into the labeling cells along non-permanent pores in the cell membrane along the selected vascular channel. Compared with the conventional medical method and system, the method and system for introducing the macromolecular substance # into the living standard stem cell of the present invention captures the three-dimensional (3D) structure image of the tissue or organ in which the stem cell is located and the tissue of the target cell Or 3D angiographic images of organs; incorporating 3D structural images into 3D angiographic images to accurately locate the target cells to select the most effective vascular access that fully encompasses the target cells; and along the selected vascular access Molecular substances are injected into the target cells. Subsequently, the method and system apply energy to activate microbubbles arrayed around the target cells to produce a biological effect whereby non-permanent pores are formed in the cell membrane of the target cells. The macromolecular material enters the stem cells through non-permanent pores in the cell membrane of the target cell. Therefore, the method and system for introducing a macromolecular substance into a living target cell of the present invention have many advantages such as low drug dosage, low cost, precise administration, and effective healing. The detailed description set forth below is illustrative of specific embodiments and embodiments of the invention. It should be understood that the detailed description is only illustrative of the invention and is not intended to 7 111758 201212892 ' ^ Embodiments The present invention generally relates to methods and systems for introducing macromolecular species into target cells; more particularly with the application of ultrasound to modulate the permeability of cell membranes of target cells, thereby effectively A method and system for introducing a low dose of a macromolecular substance into a target cell. The following description is presented to enable a person of ordinary skill in the art to make and use the invention. A person skilled in the art will readily be able to make various modifications to the preferred embodiments, general principles and features disclosed herein. Therefore, the present invention is not intended to be limited to the specific embodiments shown, but the scope of the principles and features disclosed herein. The method and system for introducing a macromolecular substance into a living target cell of the present invention can be applied to various fields such as gene delivery, gene therapy, drug delivery, partial administration, and tumor treatment. The invention is particularly applicable to the treatment of tumors, and more particularly to the treatment of solid tumors. For example, in the treatment of solid tumors, computed tomography (CT) or magnetic resonance imaging (MRI) is generally used as a preparatory step. A three-dimensional (3D) structural image of the tissue or organ in which the tumor cells are located is obtained by this preliminary step as a basis for subsequent treatments such as surgery, chemotherapy, and radiation therapy. Referring to FIG. 1A, the basic structure of a system for introducing a macromolecular substance into a living subject cell according to a preferred embodiment of the present invention is exemplified in FIG. 1A, including FIG. 2, which is only a macromolecule. A concise illustrative illustration of the main components of the system 1 for introducing a substance into a living target cell. The system 1 actually used can be more complicated. The system 1 for introducing macromolecular substances into living target cells includes an image 撷 8 111758 201212892 = elementary, image synthesizing unit 11G, and injection unit m a energy conversion module 130. At the beginning of this specific sound, you will be controlled by the micro-processing unit 140 by +, the cross-cut towel, the shadow-taking unit (10), the image combination, the main shot A 120 and the energy conversion module 'New 130. The image manipulation unit 100 is configured to capture a two-dimensional (3D) structure image of a tissue or organ in which the cell is free, and a photographic image of a blood vessel in which the target stem cell is located. In the specific embodiment, the image (4) taking unit 100 is the following, the middle of the CT, the viewing device, and the angiography device. Standard dry cells • At least one tumor cell. Through the CT | use the fan $ X body ten lines to scan the human body's fault from the axial direction ' and use a column of detectors to receive signals that penetrate the human body. When the X-ray emitter is fixed at a specific location, the detectors will be from the corresponding specific layer
接收訊號。當X射線發射器環繞一斷層轉動時，定位於X 射線發射器反向之檢測器會接收來自同一層但不同方向之 訊號。電腦分析該等訊號並計算出組成該層組成點的密度 鲁为佈1¾後顯示具有不同灰階（gray level)之點圖形的景多 像’用以增強該層之解析度。就掃描大腦而言，約15張i 公分厚的層面即可以完全涵蓋整個大腦及小腦，且可顯示 大腦之微細結構。因此，可檢測大腦中是否有水腦或血塊。 目刖，快速全身型掃描儀可於患者屏住呼吸，以大幅降低 坪吸及腸移動干擾的境況下，於30秒内掃描肝臟。亦可使 用掃描儀快速檢測並清晰顯示其他疾病，如小肝癌、腎上 腺腫瘤或胰腺疾病。 MRI裝置係用於提供清晰之多層照片。MRI裝置利用 9 111758 1 ' 201212892 電磁波刺激患者，並利用檢測器接收自患者釋出之回波。 在多次複雜之刺激-回波過程之後，可根據龐大的回波資料 達成高解析度影像。不同組織雙刺激之後釋出不同的回 波，從而於所得之影像中產生截然不同之比較。相較於通 常從軸向（最多於大腦中加入一冠狀平面）掃描斷層之CT 裝置，MRI裝置可自不同角度掃描人體之部份，如類似腦 垂體或腦幹之特定部份，其結構可清晰顯示。於另一態樣 中’MRI裝置不利用X射線’可於15分鐘内完成掃描檢查， 因此，大幅降低對人體的輻射。再者’神經系統中很多疾 病，如腦幹之輕微中風、鄰近顧骨底部之小腫瘤或骨髓疾 病（如骨髓之急性創傷或椎間盤突出（lumbar* dise herniation，LDH)) ’ 一般為CT裝置所忽略，卻可藉由mri 裝置輕易地檢出。於骨骼及肌肉系統中，MRI |置尤其適 於檢查影響關節及薄臂組織之疾病，如運動傷害。MR I裝 置也可用以檢查膽管。於使用MRI裝置之膽管檢查中，可 於屏住呼吸之情況下於20秒内獲得該膽管之影像，從而可 免受内視鏡逆行胰膽管攝影（endoscopic retrograde cholangio pancreatography，ERCP)之苦。 儘管MRI裝置具有上述諸多優點，其用於檢查之成本 過高’使得MRI檢查無法廣泛使用。再者，若患者佩戴心 律調整器或其他生理監視器，將限制使用MR I裝置之檢查 效率。因此，擷取組織或器官之3D結構影像的適當方法應 根據腫瘤所處位置’以及患者個人情況而選擇。儘管CT 裝置及MRI裝置可有效地擷取組織或器官之3D結構影像， 10 111758 201212892 但使用注射方法之用藥中，一般無法控制藥物之輸送通 道，且使用導管注射之藥物是否有效地傳輸至全部腫瘤細 胞亦不確定，因此治愈的效果極差。爲了克服此等問題， 依照本發明之將大分子物質導入活體標靶細胞之系統1的 影像擷取單元100，進一步包含血管攝影裝置。 該血管攝影裝置將特定顯影劑注射入血管中，以生成 一系列血管影像。舉例而言，於心臟血管系統之檢查中， 先從腹股溝對股骨穿孔，隨後放入導管，再反向傳輸進入 • 特定血管中。隨後，通過導管將顯影劑快速注入，同時進 行連續圖像擷取。因此，可獲得血管流入之器官，如大腦、 心臟、肝臟或腎臟的血液流動情況。再者，可使用3D重建 血管照相術，如使用通用電氣公司（General Electric，GE) 所製造之診斷性及介入性血管攝影術系統（Advantx LCA+)、心血管及血管攝影術成像系統（Advantx LCV+)及雙 平面神經血管攝影術系統（Advantx LCN+)，以取得腫瘤細 胞所處之組織或器官的3D血管攝影影像。 * 影像合成單元110將藉由影像擷取單元100擷取之3D 結構影像合併於3D血管攝影影像中，以精確地定位腫瘤細 胞，以及選擇完全涵蓋腫瘤細胞之適當血管通道。如上所 述，於CT裝置及3D血管攝影裝置、及/或MRI裝置或3D 血管攝影裝置分別擷取腫瘤細胞之3D結構影像及3D血管 攝影影像之後，影像合成單元110執行影像合成操作（亦稱 為組織製圖）。經合成之影像用於精確地定位腫瘤細胞，以 及用於選擇最有效之血管通道。經由導管，沿著所選擇之 11 111758 201212892 ' 血管通道注射藥物，藉以確保將藥物有效地傳輸至腫瘤細 胞，並達成徹底治療及低的復發機會。 此外，影像合成之後，精確地顯示腫瘤及環繞腫瘤之 血管的相對位置。除了可精確地定位腫瘤細胞以外，還可 以選擇最有效之血管通道。因此，可通過導管將藥物沿著 最有效之血管通道傳輸至全部腫瘤細胞。 注射單元120利用導管將微泡液體及大分子物質注射 入標靶細胞中。大分子物質通過微泡於標乾細胞之細胞膜 中形成之非永久性孔洞並進入標乾細胞中。於本具體實施 例中，經由注射單元120之導管，沿著所選擇之血管通道， 注射微泡液體並環繞著腫瘤細胞分佈。爲了平順地通過血 管，氣泡之尺寸較佳為小於10微米。經由導管注射藥物之 步驟可於在細胞膜中形成非永久性孔洞之前或者之後進 行。由於藥物通過於細胞膜中所形成的孔洞進入腫瘤細 胞，因此藥物之劑量可減少至普通劑量之1 % 9且達成更加 有效之治癒效果，以避免由於藥物毒性對其他細胞的損 害，並節省大量成本。 能量轉換模組130用於施加能量以活化微泡液體，並 產生生物效應，藉以於標靶細胞之細胞膜中形成非永久性 孔洞。於本具體實施例中，能量轉換模組130可為超音波 轉換模組。具有超音波轉換器或擴音器之超音波轉換模組 施加20至50千赫（KHz)頻率之超音波，並於細胞膜中形成 非永久性孔洞，以幫助藥物進入腫瘤細胞中。 第1B圖至第1D圖分別為能量轉換之超音波能量轉換 12 111758 201212892 » 模組130的透視圖、正視圖及側浦 ㈣包括基底部請及成=機;:波能量轉換模 包括超音波傳播單元134,其係包含怠有^32。模組130 且用於輻射超音能量之圓盤。成像導^轉換器及擴音器 超音波妒旦值播i分渐田 1機器臂132控制低 曰波此里傳播早％ 134(用於超音波活化之分子輸送 於一具體實施例中，圓盤136之中心氣+ 斷轉換器（未顯示），以檢驗標乾位置。‘、，、超音波（Β模式）診 J1E圖係標示圓盤周圍具有數個低 器或擴音器150 (頻率範圍為20至5 心曰得轶 區域可調節之強度範圍為約〇 2 i Z ’且能量合併 (⑽2)(距圓盤約20cm)。· 。·3卿平方公分 第1F圖係標示在圓盤周圍對稱Μ 轉換器或擴音器（頻率範圍為2G至之低能量超音波 ^ on DU KHZ) ’處於如距圓 〇 cm遮的合併區域之内，合併區 〇2inqw/2 ^ 脅之超音波強度係約 々〇.3 W/cm之範圍内。藉由使 可對腫瘤等提供能量之有效輸送。 ㈣I且130， 第1G圖左側繪示了腫瘤實體及 成；右侧铷千睥人、土 A y入发山* 官之3D影像合 入蹄广、： 液全⑽奈米乳劑(微小白點)注射 溜金官中，以填充腫瘤細胞間隙。 ’ ' 第1H圖左側係顯示超音波機器 器臂之頭盤係具有8個對稱設置之』超音波機 (，圍為…一= 直徑。此等轉換器或擴音器之聚焦區域係自該 表面、·勺20 cm。頭盤之直徑約為15至2〇 ·盤中有 111758 13 I ' 201212892 一個設置於其中之B模式診斷性換能器（頻率3至8兆赫 (MHz)，直徑為3至5 cm，最大穿透深度為20至30 cm)。 第1H圖右側說明周圍轉換器之聚焦區域（合併區域） 係定位於距頭盤約20 cm處。應注意的是，聚焦區域之超 音波能量水準為約每平方公分0. 2至0. 3 W，其對於低頻 超音波空化（聲孔效應）效果係最佳，但於FDA超音波安全 性指南中為良好。該8個獨立之超音波波束之路徑具有非 常低之超音波能量，其既不能產生聲孔效應，亦無法獲得 任何非所欲之生理性效果。換句話說，僅聚焦區域可具有 治療性聲孔效應，且累積於該聚焦區域内之能量對於患者 是安全的。 第II圖係顯示經由電腦成像導引，藉由機器臂的辅 助，低能量超音波之聚焦區域被精確地定位在腫瘤實體内 之預定治療區域。 第1J圖係治療之前及之後之腫瘤的示意圖。腫瘤之 實體於治療之後大幅縮小。 第1K圖係顯示超音波機器臂可為獨立個體或其可連 接至或附裝於或安裝至成像裝置（如CT、MR、PET掃描儀） 上。 請參閱第2圖，繪示了使用上述之系統1將大分子物 質導入活體標靶細胞的步驟。 於步驟S201中，影像擷取單元100擷取腫瘤細胞所 處之組織或器官的3 D結構影像，以及腫瘤細胞所處之組織 或器官的3D血管攝影影像。隨後進行步驟S202。 14 111758 201212892 於步驟S202中，影像合成單元110將3D結構影像合 併入3D血管攝影影像，以精確地定位腫瘤細胞並選擇完全 涵蓋用於傳輸該大分子物質之標靶細胞的血管通道。隨後 進行步驟S203。 於步驟S203中，注射單元120經由所選擇之血管通 道，注射微泡液體以環繞腫瘤細胞。隨後進行步驟S204。 於步驟S204中，能量轉換模組130使用轉換器或擴 音器，施加用於活化微泡液體之超音波，以產生生物效應， • 藉以於腫瘤細胞之細胞膜中形成非永久性孔洞。隨後進行 步驟S205。 於步驟S205中，注射單元120經由腫瘤細胞之細胞 膜中的非永久性孔洞，將大分子物質注射入腫瘤細胞中。 於本發明之另一具體實例中，係將人造血液作為微泡 液體注射並環繞腫瘤細胞。人造血液所意指者係其滿足生 物學血液之某些功能，尤其是在人體中。因為人類血液除 了執行載氧功能之外，亦執行其他功能，所以稱為氧療法 更為精確。舉例而言，白血球防禦感染性疾病，以及血小 板參與血液凝結。人造血液之一實例為全氟碳（PFC)奈米乳 劑。該人造血液係具有約150奈米之非常小的體積，因此 不會堵塞毛細血管，且該人造血液不會進入血管間之裂 隙。因此，可改善使用導管時由於低血流所造成的缺氧。 亦可利用超音波顯影劑取得該3D血管攝影影像。該 超音波顯影劑係由包裹於特定保護殼體中的微泡組成。第 一代顯影劑由其内部包裹空氣之氣泡製成，例如具有4微 15 111758 201212892 米（um)之平均體積並由超音波振動之白蛋白， 我成的 albunex (mallinckrodt)。其他超音波顯影劑包 echovist、echogen、levovist、aerosomes 等。新一括 '-γχ ^37 音波顯影劑係由難溶於水的氣體如氟碳或四氟化硫製戍 將鱗脂類、白蛋白、聚合物、表面活性劑和其他物質力 該氣體中。新一代超音波顯影劑可延長其於血液 ^ 命，並強化超音波傳播效果。該超音波顯影劑之尺寸較％ 係不超過10微米，因此該超音波顯影劑可平滑地通過訪^ 血管’並可藉由靜脈注射或使用導管注射本發明之' 糸統中使用之超音波顯影劑。 當施加1兆帕（Mpa)強度之超音波時，顯影劑之氣、包 會產生非線性振動’並發射調諧訊號。因為氣泡之調^^ 號較組織之調諧訊號強很多’顯影劑之訊號截然不同於; 織之訊號’因此可清晰地顯示包括心肌及腎臟之灰流、、 的組織情況以及腫瘤之血管分佈。如上所述’將3D結構办 像合併入3D血管攝影影像之後，i擇最有效之血管通道 經由所選擇之通道，注射腫瘤治療之藥物以環繞腫瘤細胞。 將藥物注射為環繞腫瘤細胞之後，施加使用轉換器或 擴音器之至少1 Mpa強度之超音波或適當強度之震盪波， 以活化微泡或超音波顯影劑，並進行強烈之氣泡移動，藉 以於細胞膜中形成非永久性孔洞，藉以增加細胞膜之穿透 性’立即大幅地降低用藥劑量’並維持有效之治癒效果。 或者，可在腫瘤細胞之細胞膜中形成非永久性孔洞之前注 射藥物，藉以達成與上述相同之精確用藥效果。 16 111758 201212892 此外，本發明之將大分子物質導入活體標靶細胞之系 統1係進一步包含資料處理電子裝置或與資料處理電子裝 置一起運作，用於處理在系統1工作進程中產生之資料。 資料處理電子裝置可為個人電腦（PC)、筆記型電腦（NB)、 伺服器、工作站、個人數位助理（PDA)、液晶顯示器（LCD) 電腦或平板電腦等。資料處理電子裝置包含顯示單元及輸 入單元。顯示單元係用於顯示藉由影像合成單元110執行 之影像合成處理，藉由注射單元120執行之藥物注射過 φ 程，以及藉由能量轉換模組130執行之能量傳輸情況。輸 入單元係用於輸入本發明之將大分子物質導入活體標靶細 胞之系統1的指令及/或參數，至該資料處理電子裝置中。 顯然地，對於熟識該技藝之人士，上述說明書僅為本 發明之特定具體實施態樣及實施例之例示性說明。本發明 應因此涵蓋對本文揭示之本發明結構及操作所做的各種修 飾及變更，其落入於申請專利範圍所定義之本發明範圍内。 儘管業經根據所顯示之具體實施態樣揭示本發明，熟 * 識該技藝之人士將很容易認知，可對該等具體實施態樣進 行變更，且該變更將係處於本發明之精神及範圍内。藉此， 具有該技藝通常知識之人士可對本發明作出諸多修飾，而 不悖離後附申請專利範圍之精神及範圍。 【圖式簡單說明】 第1A圖為根據本發明之較佳具體實施例之將大分子 物質導入活體標靶細胞之系統之基本結構所繪示的方塊示 意圖。 17 111758 201212892 第1B圖至圖ID分別為用於能量轉換之超音波模組之 透視圖、正視圖及側視圖。 第1E圖繪示了圓盤周圍具有數個低能量超音波轉換 器。 第1F圖指出在圓盤周圍對稱設置之低能量超音波轉 換器處於合併區域之内。 第1G圖左侧繪示了腫瘤實體及其血管之3D影像合 成；右侧繪示了注射人造血液全氟碳奈米乳劑（微小白點）Receive signals. When the X-ray emitter rotates around a fault, the detector positioned in the opposite direction of the X-ray emitter will receive signals from the same layer but in different directions. The computer analyzes the signals and calculates the density of the dots that make up the layer. After the cloth is printed, the scene image with different gray level dots is displayed to enhance the resolution of the layer. In terms of scanning the brain, about 15 i cm thick layers can completely cover the entire brain and cerebellum, and can display the fine structure of the brain. Therefore, it is possible to detect whether there is a water brain or a blood clot in the brain. It can be seen that the rapid whole body scanner can scan the liver within 30 seconds while the patient is holding his breath and greatly reducing the interference of the suction and intestinal movement. Scanners can also be used to quickly detect and clearly display other diseases such as small liver cancer, adrenal tumors or pancreatic diseases. The MRI device is used to provide clear multi-layer photos. The MRI device utilizes 9 111758 1 '201212892 electromagnetic waves to stimulate the patient and uses the detector to receive echoes released from the patient. High resolution images can be achieved based on large echo data after multiple complex stimulus-echo processes. Different tissue double stimuli release different echoes, resulting in distinct contrasts in the resulting images. Compared to CT devices that scan the fault from the axial direction (up to a coronal plane in the brain), the MRI device can scan parts of the human body from different angles, such as a specific part of the pituitary gland or brainstem. Clearly displayed. In another aspect, the 'MRI apparatus does not utilize X-rays' can complete the scanning inspection within 15 minutes, thereby greatly reducing the radiation to the human body. Furthermore, many diseases in the nervous system, such as mild stroke of the brain stem, small tumors near the bottom of the bone, or bone marrow diseases (such as acute trauma of the bone marrow or lumbar* dise herniation (LDH)) are generally CT devices. Ignore it, but it can be easily detected by the mri device. In bone and muscle systems, MRI | is especially suitable for examining diseases that affect joints and thin arm tissue, such as sports injuries. The MR I device can also be used to inspect the bile duct. In the bile duct examination using the MRI device, the image of the bile duct can be obtained within 20 seconds while holding the breath, thereby avoiding the endoscopic retrograde cholangio pancreatography (ERCP). Although MRI devices have many of the above advantages, their cost for inspection is too high, making MRI inspections unusable. Furthermore, if the patient wears a heart rate adjuster or other physiological monitor, the efficiency of the MR I device will be limited. Therefore, an appropriate method for capturing 3D structural images of tissues or organs should be selected based on the location of the tumor and the individual patient's condition. Although the CT device and the MRI device can effectively capture the 3D structural image of the tissue or organ, 10 111758 201212892, in the use of the injection method, it is generally impossible to control the drug delivery channel, and whether the drug injected using the catheter is effectively transmitted to all. Tumor cells are also uncertain, so the cure is extremely poor. In order to overcome such problems, the image capturing unit 100 of the system 1 for introducing a macromolecular substance into a living subject cell according to the present invention further includes a blood vessel imaging device. The angiography device injects a specific developer into a blood vessel to generate a series of blood vessel images. For example, in the examination of the cardiovascular system, the femur is perforated from the groin, then placed into the catheter, and then reversed into a specific blood vessel. Subsequently, the developer was quickly injected through the catheter while continuous image capture was performed. Therefore, blood flow to the organs in which blood vessels flow in, such as the brain, heart, liver or kidneys, can be obtained. Furthermore, 3D reconstruction angiography can be used, such as the diagnostic and interventional angiography system (Advantx LCA+) manufactured by General Electric (GE), and the cardiovascular and angiographic imaging system (Advantx LCV+). And a biplane neurovascular photography system (Advantx LCN+) to obtain 3D angiographic images of tissues or organs in which the tumor cells are located. * The image synthesizing unit 110 combines the 3D structured images captured by the image capturing unit 100 into the 3D angiographic images to accurately locate the tumor cells and select appropriate vascular channels that completely encompass the tumor cells. As described above, after the 3D structural image and the 3D angiographic image of the tumor cell are respectively captured by the CT device, the 3D angiography device, and/or the MRI device or the 3D angiography device, the image synthesizing unit 110 performs an image synthesizing operation (also referred to as an image synthesizing operation). Drawing for the organization). The synthesized images are used to accurately locate tumor cells and to select the most effective vascular access. The drug is injected via the catheter along the selected 11 111758 201212892 ' vascular channel to ensure efficient delivery of the drug to the tumor cells and to achieve complete treatment and low chance of recurrence. In addition, after image synthesis, the relative positions of the tumor and the blood vessels surrounding the tumor are accurately displayed. In addition to accurately positioning tumor cells, the most effective vascular access can be selected. Thus, the drug can be delivered through the catheter to the entire tumor cell along the most efficient vascular access. The injection unit 120 uses a catheter to inject microbubble liquid and macromolecular substances into the target cells. The macromolecular material passes through the non-permanent pores formed by the microbubbles in the cell membrane of the stem cells and enters the stem cells. In this embodiment, the microbubble liquid is injected and distributed around the tumor cells along the selected vascular access via a catheter of the injection unit 120. In order to smoothly pass through the blood vessel, the size of the bubble is preferably less than 10 μm. The step of injecting the drug via the catheter can be performed before or after the formation of non-permanent pores in the cell membrane. Since the drug enters the tumor cells through the pores formed in the cell membrane, the dose of the drug can be reduced to 1% of the common dose and a more effective healing effect is achieved to avoid damage to other cells due to drug toxicity, and a large cost is saved. . The energy conversion module 130 is for applying energy to activate the microbubble liquid and to produce a biological effect whereby non-permanent pores are formed in the cell membrane of the target cell. In this embodiment, the energy conversion module 130 can be an ultrasonic conversion module. Ultrasonic transducer modules with ultrasonic transducers or amplifiers apply ultrasonic waves at frequencies of 20 to 50 kHz and form non-permanent holes in the cell membrane to help the drug enter the tumor cells. 1B to 1D are respectively the ultrasonic energy conversion of energy conversion 12 111758 201212892 » The perspective view, front view and side of the module 130 (4) include the base part and the machine =; the wave energy conversion mode includes the ultrasonic wave Propagation unit 134, which includes 怠32. The module 130 is also used for a disk that radiates ultrasonic energy. Imaging guide converter and loudspeaker ultrasonic wave value broadcast i point gradual field 1 robot arm 132 control low 此 wave propagation here as early as 134 (molecular transport for ultrasonic activation in a specific embodiment, circle The center gas + interrupt converter (not shown) of the disc 136 is used to verify the dry position. ',,, ultrasonic (Β mode) diagnosis J1E diagram indicates that there are several low-level or loudspeakers around the disc 150 (frequency The range is 20 to 5. The adjustable range of the 曰 曰 area is about i 2 i Z ' and the energy is combined ((10) 2) (about 20 cm from the disc). ·························· Symmetrical Μ converter or loudspeaker around the disk (frequency range 2G to low energy ultrasound ^ on DU KHZ) 'In the combined area of the circle 〇cm cover, the merged area 〇2inqw/2 ^ The intensity of the sound wave is in the range of about W3 W/cm. It can effectively deliver energy to tumors and the like. (4) I and 130, the left side of the 1G figure shows the tumor entity and the formation; , soil A y into the mountain * official 3D image combined into the hoof,: liquid full (10) nano emulsion (small white spots) injection slip gold In order to fill the gap of the tumor cells. ' ' The left side of the 1H figure shows that the head of the ultrasonic machine arm has 8 symmetrical settings of the ultrasonic machine (, surrounded by ... a = diameter. These converters or expansion The focus area of the sounder is 20 cm from the surface, the spoon. The diameter of the head disk is about 15 to 2 〇. The disk has 111758 13 I ' 201212892 A B mode diagnostic transducer set in it (frequency 3 to 8 megahertz (MHz) with a diameter of 3 to 5 cm and a maximum penetration depth of 20 to 30 cm. The right side of Figure 1H illustrates the focal area of the surrounding converter (combined area) positioned approximately 20 cm from the head. It should be noted that the ultrasonic energy level of the focus area is about 0.2 to 0.3 W per square centimeter, which is the best for low frequency ultrasonic cavitation (sound hole effect), but the FDA ultrasonic safety. The guide is good. The path of the eight independent ultrasonic beams has very low ultrasonic energy, which neither produces a sonic effect nor any undesired physiological effects. In other words, only the focus area Can have a therapeutic sonic effect and accumulate in the poly The energy in the focal region is safe for the patient. Figure II shows the imaging guidance via computer, with the aid of the robotic arm, the focal region of the low-energy ultrasound is precisely positioned within the predetermined treatment area within the tumor entity. 1J is a schematic diagram of a tumor before and after treatment. The body of the tumor is greatly reduced after treatment. Figure 1K shows that the ultrasonic robotic arm can be an individual or it can be attached to or attached to or mounted to an imaging device (eg CT, MR, PET scanner). Referring to Figure 2, the procedure for introducing macromolecular substances into living target cells using the system 1 described above is illustrated. In step S201, the image capturing unit 100 captures a 3D structure image of the tissue or organ in which the tumor cell is located, and a 3D angiogram of the tissue or organ in which the tumor cell is located. Then step S202 is performed. 14 111758 201212892 In step S202, the image synthesizing unit 110 incorporates the 3D structural image into the 3D angiographic image to accurately locate the tumor cells and select a vascular channel that completely covers the target cells for transporting the macromolecular substance. Then, step S203 is performed. In step S203, the injection unit 120 injects microbubble liquid to surround the tumor cells via the selected blood vessel passage. Then step S204 is performed. In step S204, the energy conversion module 130 applies a transducer or a microphone to apply an ultrasonic wave for activating the microbubble liquid to generate a biological effect, and to form a non-permanent hole in the cell membrane of the tumor cell. Subsequently, step S205 is performed. In step S205, the injection unit 120 injects the macromolecular substance into the tumor cells via non-permanent holes in the cell membrane of the tumor cells. In another embodiment of the invention, the artificial blood is injected as a microbubble liquid and surrounds the tumor cells. Artificial blood means that it meets certain functions of biological blood, especially in the human body. Because human blood performs other functions in addition to performing oxygen carrying functions, it is called oxygen therapy more precise. For example, white blood cells protect against infectious diseases and blood platelets participate in blood clotting. An example of artificial blood is a perfluorocarbon (PFC) nanoemulsion. The artificial blood system has a very small volume of about 150 nm, so that the capillaries are not blocked, and the artificial blood does not enter the gap between the blood vessels. Therefore, the lack of oxygen due to low blood flow when using the catheter can be improved. The 3D angiography image can also be obtained using an ultrasonic developer. The ultrasonic developer consists of microbubbles encased in a specific protective casing. The first generation of the developer is made of air bubbles that are internally encased in air, such as albunex (mallinckrodt) which has an average volume of 4 micro 15 111758 201212892 m (um) and is vibrated by ultrasonic waves. Other ultrasonic developer packages are echovist, echogen, levovist, aerosomes, etc. A new type of '-γχ ^37 sonic developer is made from a water-insoluble gas such as fluorocarbon or sulfur tetrafluoride. Selenium, albumin, polymers, surfactants and other substances are used in this gas. A new generation of ultrasonic developers extends their blood and enhances ultrasonic transmission. The size of the ultrasonic developer is not more than 10 micrometers, so the ultrasonic developer can smoothly pass through the blood vessel' and can inject the ultrasonic wave used in the present invention by intravenous injection or using a catheter. Developer. When a supersonic wave of 1 MPa (Mpa) intensity is applied, the gas and the package of the developer generate a nonlinear vibration' and emit a tuning signal. Because the tone of the bubble is much stronger than the tuning signal of the tissue, the signal of the developer is completely different from the signal of the weave, so that the ash flow including the heart muscle and the kidney, the tissue condition, and the blood vessel distribution of the tumor can be clearly displayed. After incorporating the 3D structural image into the 3D angiographic image as described above, the most effective vascular channel is selected to inject the tumor-treating drug to surround the tumor cell via the selected channel. After injecting the drug into the surrounding tumor cells, apply a supersonic wave of at least 1 Mpa intensity or a shock wave of appropriate intensity using a transducer or a microphone to activate the microbubble or ultrasonic developer and perform strong bubble movement. Non-permanent pores are formed in the cell membrane to increase the permeability of the cell membrane 'immediately reduce the dose" and maintain an effective healing effect. Alternatively, the drug can be injected before a non-permanent hole is formed in the cell membrane of the tumor cell, thereby achieving the same precise drug effect as described above. 16 111758 201212892 Furthermore, the system 1 of the present invention for introducing macromolecular substances into living target cells further comprises or operates in conjunction with data processing electronics for processing data generated during the course of operation of system 1. The data processing electronic device can be a personal computer (PC), a notebook computer (NB), a server, a workstation, a personal digital assistant (PDA), a liquid crystal display (LCD) computer, or a tablet computer. The data processing electronic device includes a display unit and an input unit. The display unit is for displaying the image synthesizing process performed by the image synthesizing unit 110, the drug injection process performed by the injection unit 120, and the energy transfer performed by the energy conversion module 130. The input unit is for inputting instructions and/or parameters of the system 1 for introducing a macromolecular substance into a living subject cell of the present invention into the data processing electronic device. It is apparent that the above description is only illustrative of specific embodiments and embodiments of the invention. The present invention is therefore intended to cover various modifications and variations of the structure and operation of the invention disclosed herein. Although the present invention has been disclosed in terms of the specific embodiments shown, it will be obvious to those skilled in the art that the present invention may be modified and the changes are within the spirit and scope of the present invention. . Accordingly, a person skilled in the art can make various modifications to the invention without departing from the spirit and scope of the appended claims. BRIEF DESCRIPTION OF THE DRAWINGS Fig. 1A is a block diagram showing the basic structure of a system for introducing a macromolecular substance into a living subject cell according to a preferred embodiment of the present invention. 17 111758 201212892 Figure 1B to Figure ID are perspective, front and side views, respectively, of the ultrasonic module for energy conversion. Figure 1E depicts several low energy ultrasonic transducers around the disk. Figure 1F indicates that the low energy ultrasonic transducer symmetrically placed around the disk is within the merged region. The left side of Figure 1G shows the 3D image synthesis of the tumor entity and its blood vessels; the right side shows the injection of artificial blood perfluorocarbon nanoemulsion (small white spots)
進入腫瘤jk管中，以填充腫瘤細胞間隙空間。 I 第1H圖左側係顯示超音波機器臂之設計；右側係說 明周圍轉換器之聚焦區域（合併區域）定位在離頭盤約 cm遠之處。 第II圖係顯示經由電腦成像導引，藉由機器臂的輔 助，低旎量超音波之聚焦區域被精確地定位在腫瘤實體内 之預疋治療區域。 第1J圖係治療之前及之後之腫瘤的示意圖。 第1K圖係顯示超音波機器臂可為獨立個體或其可連 φ 接至或附裝或安裝至成像裝置上。 第2圖為使用第1圖之系統將大分子物質導入活體標 乾細胞之步驟所繪示的流程圖。 【主要元件符號說明】 1 將大分子物質導入活體標靶細胞之系統 100 影像擷取單元 U0 影像合成單元 111758 18 201212892 120 注射單元 130 能量轉換模組 131 基底部份 132 成像導引機器臂 134 超音波傳播單元 136 圓盤 140 微處理單元 150 低能量超音波轉換器或擴音器 A 聲孔效應轉換器或擴音器 B 成像轉換器Enter the tumor jk tube to fill the interstitial space of the tumor cells. I The left side of Figure 1H shows the design of the ultrasonic robot arm; the right side shows that the focus area (combined area) of the surrounding converter is located about cm away from the head. Figure II shows the imaging guidance via computer, with the aid of the robotic arm, the focus area of the low volume ultrasound is accurately positioned in the pre-treatment area of the tumor entity. Figure 1J is a schematic representation of tumors before and after treatment. Figure 1K shows that the ultrasonic robotic arm can be an individual or its φ can be attached to or attached to or mounted to the imaging device. Fig. 2 is a flow chart showing the steps of introducing macromolecular substances into living stem cells using the system of Fig. 1. [Main component symbol description] 1 System for introducing macromolecular substances into living target cells 100 Image capturing unit U0 Image synthesizing unit 111758 18 201212892 120 Injection unit 130 Energy conversion module 131 Base portion 132 Imaging guiding robot arm 134 Ultra Sound wave propagation unit 136 disk 140 micro processing unit 150 low energy ultrasonic transducer or loudspeaker A sound hole effect converter or loudspeaker B imaging converter
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