全國中小學科展

醫學與健康科學

治癌良「芝」—探討樟芝萃取物對口腔癌幹細胞的影響

研究旨在檢測牛樟芝菌絲萃取物4-Acetylantroquinonol B和Antrodin C對口腔癌幹細胞的影響。過去研究發現細胞膜蛋白CD44的表現與癌幹性有密切關係,因此本實驗著重於追蹤CD44的表現情況。透過3D懸浮培養獲得腫瘤球來擴增癌幹細胞群並用流式細胞儀分析。隨著兩種牛樟芝萃取物的濃度增加,CD44表現量下降,顯示此二化合物可能可以抑制其表現。實驗顯示牛樟芝萃取物不僅抑制癌幹細胞的存活率,且在低濃度下顯著抑制成球效率,還能促進癌幹細胞的凋亡。研究結果說明牛樟芝萃取物對癌幹細胞有影響,而這個發現可能可以提供潛在的治療靶點,有益未來口腔癌治療發展。

吲哚類化合物抑制神經膠質母細胞瘤及癌幹細胞之潛力

癌症幹細胞被認為是癌症會復發的主因,本研究著眼於探討新合成吲哚類化合物對癌細胞及癌幹細胞的效應與可能的作用機制。透過體外細胞實驗,我們使用不同神經膠質母細胞瘤細胞株,評估化合物對細胞增殖、存活和凋亡的作用。接著以類癌幹細胞球篩檢評估化合物對癌症幹細胞的影響。研究結果顯示,該化合物對神經膠質母細胞瘤及其癌幹細胞皆有一定程度的抑制效果。 同時,透過分子生物學技術,研究化合物的分子作用機制,結果顯示化合物能對細胞生長和凋亡的路徑產生影響。研究結果有望提供對候選抗癌藥物在細胞水平的效能、選擇性以及對癌幹細胞的特異性反應的深入理解。 期望本研究成果可為癌症治療藥物的開發提供重要參考,並促使對癌症治療新方法的探索。這將有助於確定更具潛力的藥物候選者,為癌症治療領域帶來更具前瞻性的解決方案。

Eradicating Cystic Fibrosis Biofilms by a Novel Non-Toxic, Multi-Pathway Salicylate Therapy

1.1. Cystic Fibrosis Biofilms Biofilms are bacterial aggregates in a matrix of polysaccharides, proteins and nucleic acids (Donlan, 2002). They account for 80% of all chronic infections and cause over 500,000 deaths annually. Cystic fibrosis (CF) is a genetic disorder characterized by mucus accumulation in the respiratory tracts (Morrison et al., 2020). This impairs mucociliary clearance, allowing chronic colonization by bacterial biofilms, leading to fatal respiratory failure, lung scarring, and necrosis of pulmonary epithelial tissues (Martin et al., 2021). 1.2. Obstacles in Current Treatments Three major therapies are used against CF biofilms: (1) aminoglycoside antibiotics like tobramycin, (2)non-aminoglycoside antibiotics such as ciprofloxacin and vancomycin, and (3) non-antibiotic therapies including flushing, chlorination, and ultraviolet disinfection. These have two major flaws. First, they are cytotoxic; 30% of patients experience acute kidney injury after three days of intravenous aminoglycoside therapy (Joyce et al., 2017). Furthermore, non-aminoglycoside therapies can cause phospholipid buildup in lysosomes of proximal tubule epithelial cells, accounting for 10-20% of acute renal failure cases. Second, antibiotic resistance due to horizontal gene transfer and mutations has significantly reduced treatment effectiveness. Therefore, cystic fibrosis biofilms remain a critical threat with few effective treatments. 1.3. Salicylate Derivatives This project tackled this issue using an innovative non-antibiotic approach with salicylate derivatives. Salicylates, a class of benzoic acids—benzene-based carboxylic acids (Figure 1)—used in painkillers and blood thinners, were investigated for their antibiofilm potential through a 3-step process: 1. Literature review: Identified three key biofilm therapeutic targets: quorum sensing, bacterial adhesion, and cell motility. Disrupting these pathways would result in biofilm eradication. 2. Molecule Identification: Recognized key molecules in each pathway: LasR, adhesins, and flagellin. Inhibiting these molecules would disrupt the pathways. 3. Screening: Found that salicylates could inhibit the identified molecules, though they had never been tested against cystic fibrosis biofilms.

Trojan Horses in the Fight against Skin Cancer

In photodynamic therapy (PDT), reactive oxygen species are generated within the cytoplasm to destroy cancer cells selectively. Using porphyrinic structures (PS) as photosensitizers holds promise for targeting cancer cells. However, direct incorporation of the porphyrins into cancer cells remains elusive. Hence, Dr. Martina Vermathen’s research introduced specific membranous phospholipid nanocarriers for topical porphyrin applications. However, since a sufficiently high enough concentration of PS in cancer cells has not yet been achieved, this study aimed to improve skin uptake of the nanocarriers. Two approaches were examined: (1) comparing polar and nonpolar porphyrins and (2) assessing the effect of a penetration enhancer, DMSO, through a neat and diluted application. The polarity of the porphyrins was first quantified with a log P test. The nanocarriers were assembled by incorporating two different PS compounds, either the mono- or tetra-4-carboxy substituted phenyl porphyrin. They were then characterized by 1D and 2D-NMR analysis. The porphyrin permeation was tested by Franz diffusion tests on pig ear skin. For the second approach, DMSO was added in the Franz diffusion test, either directly applied on the skin (“neat“) or diluted in the nanocarriers (“diluted”). The log P test for the mono- and the tetra-carboxyphenyl porphyrin resulted in values of 4.5 and -1.1, respectively. The more polar tetra-carboxyphenyl porphyrin exhibited 2.8 times better skin uptake compared to the mono-carboxyphenyl porphyrin. The neat DMSO application increased uptake by a factor of 5.5. The diluted DMSO application worsened skin uptake slightly. Analytical techniques revealed differences in porphyrin encapsulation: The mono-carboxyphenyl porphyrins were encapsulated in the centre, whereas tetra-carboxyphenyl porphyrins were localised around the nanocarriers. Results indicated potential instability of the nanocarriers. The more polar tetra-substituted porphyrins showed superior skin diffusion than the mono-substituted derivative. The neat DMSO application facilitated enhanced skin uptake by inducing membrane destabilization and pore formation but may have limited applicability. Further research is suggested to explore porphyrinic PS with alternative polar substitution patterns and tailored penetration enhancers for lipid-based delivery systems. Overall, the study underscores the importance of molecular properties of the PS system and demonstrates the potential of penetration enhancers in optimizing PDT for skin cancer treatment.

降脂轉肌–將脂肪轉變成肌肉的可能性探討

先前研究發現一種多元不飽和脂肪酸 15-keto-PGE2 能減少肌肉慢性發炎以及增加肌肉生長因子胰島素的敏感性,具有增加肌肉的潛力。我們探討此脂肪酸在小鼠中將脂肪組織轉成肌肉組織的可能性。 我們先以 15-keto-PGE2 治療肥胖型肌少症小鼠後,再以全基因組mRNA定序,發現在小鼠皮下脂肪中,肌肉特異基因表現量高度上升。透過基因表現路徑分析軟體 GSEA證實此脂肪酸會高度引發皮下脂肪中,與肌肉分化、肌肉收縮與肌肉結構相關的生物路徑。另外, 15-keto-PGE2 也可以誘導脂肪前驅細胞3T3-L1中與肌肉生成相關的基因表現。 最後,我們發現以 15-keto-PGE2 治療的老年肥胖型肌少症小鼠,體重、血糖、脂肪比例下降,肌肉質量及力量上升,證實了它在體內同時減少脂肪並增加肌肉量的效果。

以果蠅建立單純型表皮水皰症(EBS)模型、建立藥物篩選流程並以雙醋瑞因(Diacerein)進行測試

遺傳性表皮分解性水泡症(EB)是種罕見疾病,因突變使角蛋白異常,造成表皮組織脆弱易形成水泡,單純型水泡症(EBS)是最常見的類型。此計畫旨在:(一)建立果蠅EBS疾病模型;(二)探討溫度對病徵的影響;(三)以此果蠅EBS疾病模型發展藥物篩選平台。初步使用Diacerein測試,評估對EBS症狀的改善效果。 目前顯示突變角蛋白K5/K14R125C會形成積聚體,與正常K5/K14形成的角蛋白網絡不同;全程25℃培養,約32%果蠅翅膀有水泡,亦符合EBS病徵。篩藥平台建置已完成色素和溶劑DMSO劑量測試,初步顯示Diacerein有助病徵緩解。目前將擴大統計不同溫度對 EBS果蠅死亡率、水泡發生率和角蛋白積聚形成比例。希望以本研究建立的果蠅EBS疾病模型與篩藥平台,能為罕見遺傳疾病療程開發奠定基礎。

Natural resources utilization for the in-house production of fluorescence lipid nanoparticles

Nanotechnology, a transformative force, has steadily gained traction across multiple scientific disciplines, including physics, chemistry, engineering, and biology. It offers unprecedented capabilities, especially in the realm of nanoscale particles, ushering in new paradigms in various applications. One of the most revolutionary applications of nanotechnology is in the pharmaceutical sector. Here, nanoparticles have transformed drug and vaccine delivery systems, offering both efficacy and precision. Among these nanoparticles, lipid nanoparticles (LNPs) have stood out, especially for their role in delivering nucleic acid-based drugs and vaccines. These LNPs are intricate assemblies composed of lipids and nucleic acid complexes, offering an amalgamation of stability and deliverability. Such properties have rendered LNPs as invaluable tools in enhancing therapeutic efficacy while minimizing off-target side effects. The myriad of nanoparticles available includes the likes of silver, gold, and lipid nanoparticles. However, the emphasis of this research lies with lipid nanoparticles, given their widespread success in the pharmaceutical arena. LNPs have showcased their potential in delivering drugs with low therapeutic indices, emphasizing their capability to act as versatile platforms for novel drug development. Recent advances have further expanded the horizons of LNPs, paving the way for novel antisense oligonucleotides, innovative vaccines, and complex lipid nanoparticle formations. Characterizing these nanoparticles is paramount, not only for the development of novel drugs but also to comprehend their in vivo behavior. Their multifaceted nature, stemming from their unique excipients, core-bilayer design, and varying sizes, makes their characterization a critical step in the research and development pipeline.

YKT6與癌纖維母細胞的「泌」密關係

本研究以人類肺癌A549細胞株和纖維母細胞模擬體內腫瘤微環境,挖掘纖維母細胞如何促進癌細胞的生長。從病人的正常和癌組織提取癌相關纖維母細胞(cancer-associated fibroblasts, CAFs) 和 正常纖維母細胞(Normal Fibroblasts, NFs),經過基因序列一對對作分析,開發新的治療策略和潛在的靶點。利用核糖核酸定序(RNA-Seq)分析發現CAFs會比NFs分泌更多SNARE 蛋白 YKT6,而更深入地探究獲悉YKT6會透過活化YKT6+CAFs途徑促進肺癌A549細胞惡化,此惡化過程包括誘導及提升癌細胞的生殖(proliferation),轉移(migration)和入侵(invasion)能力。 此外,在 CAFs 中敲除 YKT6基因,減弱CAFs 的外泌體(exosome)釋放,從而調節了其對肺癌細胞A549的腫瘤促進作用。本研究發現靶向YKT6並抑制外泌體分泌,從而降低CAFs對肺腺癌細胞的腫瘤支援功能可以為肺癌治療提供一種新的策略。

運用細胞水膠化技術製作微流道晶片進行抗原專一性T細胞之篩選 Flow-induced Mechanical Screening of Antigen Specific T cells with Biomimetic Microfluidic Chip

積極發展癌症相關治療策略極為重要,其中T細胞免疫療法(adaptive T cell therapy)是一深具臨床價值的選項。即是將T細胞自體內取出後並增殖到一定數量,而後將其回輸病人體內使得T細胞攻擊癌細胞。此方式關鍵的步驟在於必須要能夠篩選出足量的抗原專一性T細胞。現行主要的篩選方法雖然方便,但會誤捕不相關抗原專一性T細胞,降低治療效率。為解決此問題,本研究將利用水膠細胞技術,運用其完整保存生物膜的特性,模仿細胞膜之免疫突觸現象,搭配微流控晶片可控制流速改變沖刷力的特性,成功開發一可篩選親和力較高之T細胞微流道晶片。已在晶片內建立可置換任意抗原之水膠細胞單層,並以SIINFEKEL抗原作為模擬,達到極高之置換率。預期能在未來的研究中提升篩選專一性與數量,進而提升其臨床價值。

Utilizing Flavonoids From the Invasive Species Pilea Melastomoides and Daucus Carota as Well as the Protein PTK-2 to Create a Skin Gel Aimed for Burn Wound Healing.

Burns are a major global health concern especially in developing countries like 印尼, where southeast asian women experience the highest burn incidents globally. Burns can cause severe physical and psychological impacts, with treatments that are critical to reduce complications. This study focuses on the development of organic, cost-effective burn gels using flavonoid compounds which are Quercetin and Myrecetin which are taken from pilea melastomoides leaves, a wild 印尼n plant and carrot (Daucus Carota). These skin extracts aim to accelerate wound healing, minimize pain and prevent infection. The gel formation involves extracting active compounds using 96% ethanol as it has been effectively used for extracting a wide range of bioactive compounds to preserve their quality by preventing microbial contamination, and ensures a high yield of active ingredients suitable for topical applications. Then it goes through a process of Phytochemical screening to confirm the presence of flavonoids by using the Shinoda test. The formulation process included dissolving the HPC-m (Hydroxypropyl Cellulose) as a gelling agent, then adding plant extracts (pilea melastomoides leaves and carrot), as well as combining other ingredients such as propylene glycol, sodium benzoate, sodium metabisulfite, and disodium EDTA. The gel was stirred thoroughly to ensure uniformity and left at room temperature for 48 hours to attain the required consistency. The gel that was formatted went under various quality assessments, first being organoleptic testing. This test is used to evaluate its physical characteristics which includes color aroma, and consistency which confirms a stable dark green appearance and a natural strong scent from the plant extracts. The homogeneity test is used to verify the uniformity distribution of active compounds across the gel, to ensure a consistent efficacy. The pH test showed the gel’s acidity level which remained the safe range for skin application. Additionally, the spreading ability test demonstrated the gel’s excellent application properties, with consistent results across trials. Subsequently, the in silico analysis was conducted to predict the behaviour of specific flavonoid compounds used which is the myricetin and quercetin, highlighting their potential anti-inflammatory and antimicrobial activities. Further bacterial contamination tests confirmed the gel’s antimicrobial efficacy, reducing the risk of infection in wounds. This study demonstrates that the gel, formulated with pilea melastomoides leaves and carrot skin extracts, effectively utilizes flavonoids and other phytochemicals to reduce inflammation, promote tissue regeneration and retain moisture, which fosters an optimal condition for wound healing. This organic and sustainable burn treatment utilizes locally sourced ingredients, providing a natural solution that speeds up recovery, reduces pain and prevents infections. The results highlight its significant potential for wider healthcare use, especially in resource-limited environments.