Promising Approach for Optimizing In Vivo Fluorescence Imaging in a Tumor Mouse Model: Precision in Cancer Research.

BACKGROUND/AIM: Cancer remains a major global health concern due to its high mortality rates. Advanced diagnostic imaging, such as in vivo near-infrared (NIR) fluorescence imaging, enhances early detection by reducing autofluorescence and enabling deeper tissue penetration, addressing some limitations of conventional methods. Understanding the underlying causes of autofluorescence, even in mouse model fluorescence imaging, is crucial for accurate interpretation. This study investigated the origins of autofluorescence observed in experimental animals under NIR wavelengths, achieving successful fluorescence imaging in a clinically relevant tumor mouse model. MATERIALS AND METHODS: Both fasting and non-fasting groups were evaluated to assess the dietary impact on autofluorescence, with various feeds tested. Subcutaneous and lung tumor models were established in C57BL/6 and BALB/c nude mice using LL/2-iRFP cells. Cryo-sectioning and lung tissue imaging were conducted to confirm tumor presence and assess fluorescence signals. RESULTS: It was found that autofluorescence, notably common in the abdomen, is attributed to dietary factors. By selecting feed that lacks autofluorescence, the impact of dietary fluorescence on imaging was evaluated, leading to the establishment of optimized imaging conditions suited to the presence or absence of autofluorescence. Subsequently, utilizing lung cancer cells expressing near-infrared proteins (LL/2-iRFP), intratracheal, and subcutaneous tumor mouse models were developed, and successful in vivo imaging was achieved using the optimized imaging protocols, effectively bypassing autofluorescence. CONCLUSION: This study emphasizes the importance of understanding and addressing autofluorescence in fluorescence imaging, presenting valuable insights for enhancing the reliability and accuracy of diagnostic imaging techniques in cancer research and clinical practice.

Anti-obesity and immunomodulatory effects of oil and fermented extract dried from Tenebrio molitor larvae on aged obese mice.

Preventing disease and maintaining the health of the elderly are crucial goals for an aging population, with obesity and immune function restoration being of paramount importance. Obesity, particularly visceral obesity characterized by excessive fat accumulation around the abdominal organs, is linked to chronic conditions such as diabetes, hypertension, cardiovascular diseases, and immune dysfunction. Globally, obesity is considered a disease, prompting significant research interest in its treatment. Therefore, it is essential to explore potential therapeutic and preventive strategies to address obesity and the decline in immune function brought about by aging. Tenebrio molitor larvae (TML), commonly known as 'mealworms,' are rich in unsaturated fatty acids, including oleic and linoleic acids, and essential amino acids, such as isoleucine and tyrosine. In this study, we aimed to investigate the effects of the consumption of TML oil and mealworm fermented extract (MWF-1) on obesity and immunological changes in aged obese mice. Our data showed reduced body fat in 23-week-old C57BL/6 mice fed processed TML products for 6 weeks. Additionally, the characteristically high levels of serum triglycerides decreased by treating with TML oil. The immune responsiveness results confirmed an increase in B cells by treating with MWF-1, while cytokine levels (interferon-gamma, tumor necrosis factor-alpha, interleukin-2, and -6) were restored to levels similar to young mice. These results suggest that TML oil and MWF-1 are promising dietary supplements for addressing obesity and restoring immune function.

Targeting Heme Oxygenase 2 (HO2) with TiNIR, a Theragnostic Approach for Managing Metastatic Non-Small Cell Lung Cancer.

Despite notable advancements in cancer therapeutics, metastasis remains a primary obstacle impeding a successful prognosis. Our prior study has identified heme oxygenase 2 (HO2) as a promising therapeutic biomarker for the aggressive subsets within tumor. This study aims to systematically evaluate HO2 as a therapeutic target of cancer, with a specific emphasis on its efficacy in addressing cancer metastasis. Through targeted inhibition of HO2 by TiNIR (tumor-initiating cell probe with near infrared), we observed a marked increase in reactive oxygen species. This, in turn, orchestrated the modulation of AKT and cJUN activation, culminating in a substantial attenuation of both proliferation and migration within a metastatic cancer cell model. Furthermore, in a mouse model, clear inhibition of cancer metastasis was unequivocally demonstrated with an HO2 inhibitor administration. These findings underscore the therapeutic promise of targeting HO2 as a strategic intervention to impede cancer metastasis, enhancing the effectiveness of cancer treatments.

Visualization of Metastatic Lung Cancer with TiNIR.

The development of efficient biomarkers and probes for monitoring and treating cancer, specifically metastatic cancer, is a critical research area that can have a significant impact on both patient outcomes and drug discovery. In this context, TiNIR has been developed to detect tumor-initiating cells (TICs), with heme oxygenase 2 (HO2) as a promising therapeutic biomarker for tumor-initiating cells. In this study, TiNIR has demonstrated its effectiveness as an in vivo metastatic lung cancer tracker, highlighting its potential as a valuable tool in cancer research and therapy. The development of innovative approaches that selectively target metastatic cancers represents a promising avenue for improving survival rates and enhancing the quality of life of cancer patients.

A preliminary study for the development of cleavable linkers using activatable fluorescent probes targeting leucine aminopeptidase.

Ligand-targeted drugs (LTDs) such as antibody-drug conjugates (ADCs) are currently attracting great attention as an alternative class of therapeutics to conventional chemotherapy for the clinical treatment of cancer. The linker is one of important factors determining the efficacy and toxicity of LTDs. The linker for LTDs should have enough stability during blood circulation, effectively release the payload, and leave no polar moieties in the released payload. However, the drug release activity and plasma stability of cleavable linkers are generally evaluated by complex and sophisticated in vivo techniques containing LC-MS, and the designing of new clinically applicable linkers remains a challenge. In this work, leucine aminopeptidase (LAP)-responsive fluorescent probes were designed as a simple preliminary model to verify whether a peptidase-responsive fluorescent probe can be used as a facile tool for the development of cleavable linkers although LAP is an exopeptidase and can't be a real target for cleavable linkers. LAP-responsive fluorescent probes were prepared by conjugation of a leucine to several xanthene fluorophores through a few linkages with a p-aminobenzyl spacer. The stability tests, kinetic study and live cell imaging of LAP-responsive activatable fluorescent probes demonstrated that the chemical stability and intrinsic activity of the linker for the release of drug can be easily evaluated by a fluorogenic assay. The ex vivo plasma stability test using mice suggested that an enzyme-responsive activatable fluorescent probe can be used as a feasible platform to evaluate the plasma stability of cleavable linkers during blood circulation.

Visualizing inflammation with an M1 macrophage selective probe via GLUT1 as the gating target.

Macrophages play crucial roles in protecting our bodies from infection and cancers. As macrophages are multi-functional immune cells, they have diverse plastic subsets, such as M1 and M2, derived from naïve M0 cells. Subset-specific macrophage probes are essential for deciphering and monitoring the various activation of macrophages, but developing such probes has been challenging. Here we report a fluorescent probe, CDr17, which is selective for M1 macrophages over M2 or M0. The selective staining mechanism of CDr17 is explicated as Gating-Oriented Live-cell Distinction (GOLD) through overexpressed GLUT1 in M1 macrophages. Finally, we demonstrate the suitability of CDr17 to track M1 macrophages in vivo in a rheumatoid arthritis animal model.

Anti-Inflammatory Effects of Phlebia sp. Extract in Lipopolysaccharide-Stimulated RAW 264.7 Macrophages.

Lichens are a life form in which algae and fungi have a symbiotic relationship and have various biological activities, including anti-inflammatory and antiproliferative activities. This is the first study to investigate the anti-inflammatory activity of a Phlebia sp. fungal extract (PSE) isolated from Peltigera neopolydactyla in lipopolysaccharide- (LPS-) stimulated RAW 264.7 macrophage. PSE reduced the production of the proinflammatory cytokine (tumor necrosis factor-α, interleukin-6, and interleukin-1ß), chemokine (granulocyte-macrophage colony-stimulating factor), nitric oxide, and prostaglandin E2 in the LPS-stimulated RAW264.7 macrophages. Especially, PSE inhibits the phosphorylation of activator protein-1 (AP-1) signaling (c-Fos and c-Jun) and their upstream mitogen-activated protein kinase kinases/mitogen-activated protein kinases (MKK/MAPKs: MKK4, MKK7, and JNK) and finally reduced the production of the inflammatory cytokines. The inhibitory effects mainly act via suppressing JNK-mediated AP-1 rather than the NF-κB pathway. Furthermore, PSE inhibited the production of final inflammatory effector molecules involved in AP-1 signaling, including nitric oxide (NO) and prostaglandin E2 (PGE2). Here, we report that PSE has the potential to be developed as an anti-inflammatory agent.

Psychological Impact of Type of Breast Cancer Surgery: A National Cohort Study.

BACKGROUND: The present study assessed the impact of different types of breast surgery on rates of psychological disorders in breast cancer patients. METHODS: This nationwide cohort study, based on Korean Health Insurance Review and Assessment Service claims data, included 26,259 breast patients who underwent surgery from June 1, 2017, to December 31, 2018. Associations between the incidence of psychological disorders and variables were evaluated by time dependent Cox regression analyses. RESULTS: Of the 26,259 patients, 9394 (35.8%) underwent total mastectomy (TM) and 16,865 (64.2%) underwent partial mastectomy (PM); of the former, 4056 (43.2%) underwent breast reconstruction surgery (RS). A total of 4685 patients (17.84%) were newly diagnosed with psychological disorders after surgery. Multivariable analysis showed that axillary lymph node dissection was significantly associated with increased rates of overall psychological disorders (p < 0.0001), depression (p = 0.0462), anxiety (p < 0.0001) and insomnia (p < 0.0001). The rates of overall psychological disorders (p = 0.0002) and insomnia (p = 0.01) were significantly lower in patients who underwent TM than PM. RS tended to associated with reduced rates of overall psychological disorders in patients who underwent TM. Subgroup analysis showed that, compared with PM, RS after TM significantly associated with a reduced incidence of overall psychological disorders and insomnia in younger patients (< 50 years) and those who underwent sentinel lymph node biopsy. CONCLUSION: In contrast to general belief, rates of overall psychological disorders and insomnia were lower in patients who underwent TM than PM. Moreover, RS after TM confers psychological benefit in younger patients with early stage breast cancer compared with PM.

Racial differences in predictive value of the 21-gene recurrence score assay: a population-based study using the SEER database.

PURPOSE: The 21-gene recurrence score (RS) assay is currently used for predicting chemotherapeutic benefits for hormone receptor-positive (HR +) early-stage breast cancer patients without consideration regarding racial differences in that predictive value. This study aimed at demonstrating racial differences in the predictive values of the 21-gene RS assay. METHODS: The study cohort was selected from the Surveillance, Epidemiology, and End Results (SEER) database. Breast cancer-specific mortality (BCSM) was compared between patients who received chemotherapy (the "CTx group") and those who did not (the "no CTx group") to estimate the predictive value of the assay. This comparison was repeated for each racial group. RESULTS: Among 88,498 T1 - 2N0 HR + breast cancer patients who had results of 21-gene RS, 13,123 patients had RS > 25, which included 10,697 Whites, 1282 Blacks, and 1,144 Asian Americans/Pacific Islanders (AAPIs). Chemotherapy was administered to 8364 patients (63.4%). The adjusted hazard ratio for BCSM in the CTx group (vs. no CTx group) was 0.734 (95% confidence interval [CI] 0.588-0.917) in Whites, 0.748 (95% CI 0.428-1.307) in Blacks, and 1.343 (95% CI 0.558-3.233) in AAPIs. No subgroup within patients with RS > 25 among non-White women showed a significant predictive value of the 21-gene RS assay, except for Black women with grade 3 tumors. CONCLUSION: The predictive value of the 21-gene RS assay for assessing chemotherapy benefit was validated in White women based on the SEER database, although the predictive value was not warranted in non-White women.

Microbiota of Breast Tissue and Its Potential Association with Regional Recurrence of Breast Cancer in Korean Women.

Recent studies have reported dysbiosis of the microbiome in breast tissue collected from patients with breast cancer and the association between the microbiota and disease progression. However, the role of the microbiota in breast tissue remains unclear, possibly due to the complexity of breast cancer and various factors, including racial and geographical differences, influencing microbiota in breast tissue. Here, to determine the potential role of microbiota in breast tumor tissue, we analyzed 141 tissue samples based on three different tissue types (tumor, adjacent normal, and lymph node tissues) from the same patients with breast cancer in Korea. The microbiota was not simply distinguishable based on tissue types. However, the microbiota could be divided into two cluster types, even within the same tissue type, and the clinicopathologic factors were differently correlated in the two cluster types. Risk of regional recurrence was also significantly different between the microbiota cluster types (p = 0.014). In predicted function analysis, the pentose and glucuronate interconversions were significantly different between the cluster types (q < 0.001), and Enterococcus was the main genus contributing to these differences (q < 0.01). Results showed that the microbiota of breast tissue could interact with the host and influence the risk of regional recurrence. Although further studies would be recommended to validate our results, this study could expand our understanding on the breast tissue microbiota, and the results might be applied to develop novel prediction methods and treatments for patients with breast cancer.

Application of patch stimulator for intraoperative neuromonitoring during thyroid surgery: maximizing surgeon's convenience.

BACKGROUND: Intraoperative neuromonitoring (IONM) is frequently used in thyroid surgery to reduce recurrent laryngeal nerve (RLN) injury by providing the surgeon with real-time feedback on nerve stimulation during dissection. We applied a disposable adhesive patch electrode to a dissecting instrument to transfer electrical stimulation to the dissecting instrument for IONM during thyroid surgery. This study aimed to evaluate the feasibility of using the patch stimulator approach for IONM during thyroid surgery. METHODS: We reviewed the medical records of patients who underwent thyroidectomy using both conventional stimulator and adhesive patch stimulator for IONM. The electromyography (EMG) amplitudes of the vagal and the RLNs before (V1, R1) and after thyroid resection (V2, R2) were alternatively checked with each type of stimulator at the same location of each nerve. RESULTS: Fifteen consecutive patients (4 males, 11 females) were included in this analysis, and a total of 38 nerves (19 vagus nerves and 19 RLNs) were evaluated. No statistically significant differences were seen in the mean amplitudes evoked by the patch stimulator and the conventional probe stimulator for the V1 signal (825.5±394.6 vs. 821.8±360.9 µV, P=0.954), R1 signal (1,044.8±471.2 vs. 1,039.2±507.4 µV, P=0.898), R2 signal (1,037.8±495.0 vs. 938.2±415.8 µV, P=0.948), or V2 signal (812.5±391.9 vs. 787.3±355.7 µV, P=0.975). CONCLUSIONS: The patch stimulator was safely and effectively used for IONM during thyroid surgery and provided similar nerve monitoring responses as the conventional stimulator. This approach may be used to enhance the surgeon's convenience during thyroid surgery.

Prognostic influences of BCL1 and BCL2 expression on disease-free survival in breast cancer.

We investigated the prognostic influences of BCL1 and BCL2 expression on disease-free survival in breast cancer patients. BCL1 and BCL2 expression statuses were assessed by immunohistochemistry using tissue microarrays from 393 breast cancer patients. The Kaplan-Meier estimator and log-rank test were used for survival analyses. The Cox proportional hazards model was used to calculate hazard ratio (HR) and the 95% confidence interval (CI) of survival analyses. BCL1 expression revealed no impact on survival. The high BCL2 group showed superior disease-free survival compared with the low BCL2 group (p = 0.002), especially regarding local recurrence-free survival (p = 0.045) and systemic recurrence-free survival (p = 0.002). BCL2 expression was a significant prognostic factor by univariable analysis (HR, 0.528; 95% CI, 0.353-0.790; p = 0.002) and by multivariable analysis (HR, 0.547; 95% CI, 0.362-0.826; p = 0.004). High BCL2 expression was associated with higher disease-free survival in the hormone receptor (HRc)-positive and human epidermal growth factor receptor 2 (HER2)-negative (HRc(+)/HER2(-)) subtype only (p = 0.002). The high BCL2 group was associated with positive estrogen receptor (ER), positive progesterone receptor (PR), low histologic grade, and age ≤ 50 years. BCL1 expression had no prognostic impact, but BCL2 expression was a significant independent prognostic factor. High BCL2 expression was associated with higher disease-free survival, especially regarding local recurrence and systemic recurrence. The prognostic effect of BCL2 expression was effective only in the HRc(+)/HER2(-) subtype. Favorable clinicopathologic features and a strong association with the ER/PR status could partly explain the superior prognosis of the high BCL2 group. BCL2 expression could be utilized to assess the prognosis of breast cancer patients in clinical settings.

The development of a functional human small intestinal epithelium model for drug absorption.

Advanced technologies are required for generating human intestinal epithelial cells (hIECs) harboring cellular diversity and functionalities to predict oral drug absorption in humans and study normal intestinal epithelial physiology. We developed a reproducible two-step protocol to induce human pluripotent stem cells to differentiate into highly expandable hIEC progenitors and a functional hIEC monolayer exhibiting intestinal molecular features, cell type diversity, and high activities of intestinal transporters and metabolic enzymes such as cytochrome P450 3A4 (CYP3A4). Functional hIECs are more suitable for predicting compounds metabolized by CYP3A4 and absorbed in the intestine than Caco-2 cells. This system is a step toward the transition from three-dimensional (3D) intestinal organoids to 2D hIEC monolayers without compromising cellular diversity and function. A physiologically relevant hIEC model offers a novel platform for creating patient-specific assays and support translational applications, thereby bridging the gap between 3D and 2D culture models of the intestine.

Multigene Panel Testing for Hereditary Cancer and Genetic Counseling.

As sequencing technology and information of the genomic causes for cancer development expand, multi-gene panel testing for hereditary cancer is increasing in clinical practice. In this chapter, we reviewed the application of multi-gene panel with pre-/post- testing considerations and summarized genetic counseling based on panel testing results in clinical field. In addition, we introduce multi-gene panel for hereditary cancer developed in Seoul National University Hospital.

Metachronous Sporadic Sextuple Primary Malignancies Including Bilateral Breast Cancers.

Multiple primary malignancies are defined as the presence of more than one malignant neoplasm with a distinct histology occurring at different sites in the same individual. They are classified as synchronous or metachronous according to the diagnostic time interval of different malignancies. Diagnosis of multiple primary malignancies should avoid misclassification from multifocal/multicentric tumors or recurrent/metastatic lesions. In multiple primary malignancies, with increase in the number of primary tumors, the frequency rapidly decreases. Here, we report an exceptionally rare case of a woman who was diagnosed with metachronous sporadic sextuple primary malignancies including bilateral breast cancers (gastric cancer, ovarian Sertoli-Leydig cell tumor, left breast cancer, thyroid cancer, right breast cancer, and rectal neuroendocrine tumor). The sextuple primary malignancies in this case involved 5 different organs: the stomach, ovary, thyroid, rectum, and bilateral breasts. Further studies are needed to elucidate the current epidemiologic status of patients with multiple primary malignancies.

Influence of Metabolic Syndrome on Risk of Breast Cancer: A Study Analyzing Nationwide Data from Korean National Health Insurance Service.

BACKGROUND: To investigate the influence of metabolic syndrome and its components on the risk of breast cancer. METHODS: Retrospective nationwide cohort study analyzing data of 13,377,349 women older than 19 years from Korean National Health Insurance Service was performed. Cox proportional hazards model was used to calculate HR and 95% confidence interval (CI) of breast cancer risk. RESULTS: The presence of metabolic syndrome decreased the risk of all breast cancer types in all subjects (HR, 0.954; 95% CI, 0.939-0.970). In women with age ≤50 years, metabolic syndrome decreased the risk of all breast cancer types, with similar findings for all subject groups (HR, 0.915; 95% CI, 0.892-0.939). In women with age >50 years, metabolic syndrome increased the risk of all breast cancer types (HR, 1.146; 95% CI, 1.123-1.170), especially in age groups of more than 55 years. In women with age >50 years, HRs increased as the number of metabolic syndrome components increased, while HRs decreased as the number of metabolic syndrome components increased in women with age ≤50 years. CONCLUSIONS: The presence of metabolic syndrome increased the risk of breast cancers in postmenopausal women, but decreased the risk in premenopausal women. Every metabolic syndrome component played similar roles on the risk of breast cancer as metabolic syndrome, and their effects became stronger when the number of components increased. IMPACT: Metabolic syndrome is associated with the risk of breast cancer having different effect according to age groups.

The 21-Gene Recurrence Score Assay and Prediction of Chemotherapy Benefit: A Propensity Score-Matched Analysis of the SEER Database.

BACKGROUND: To evaluate the performance of the 21-gene recurrence score (RS) assay in predicting chemotherapy benefit in the Surveillance, Epidemiology, and End Results population, we aimed to assess breast cancer-specific mortality (BCSM) by chemotherapy use within each of the RS categories. METHODS: Data on breast cancer (BC) cases diagnosed between 2004 and 2015 with available RS results were released. Our analysis included patients with hormone receptor-positive, node-negative early-stage BC (n = 89,402), and three RS groups were defined; RS < 11, low; RS 11-25, intermediate; RS > 25, high. A propensity score matched-analysis was performed to assess and compare BCSM. RESULTS: Chemotherapy was significantly associated with a reduced risk of BC death among patients in the high RS group (hazard ratio = 0.782; 95% CI, 0.618-0.990; p = 0.041). However, in the low and intermediate RS groups, there were no significant differences in BCSM between patients who received chemotherapy and those who did not. Among those with RS 11-25, chemotherapy benefit varied with tumor size (p = 0.001). CONCLUSIONS: Our findings provide real-world evidence that the 21-gene RS assay is predictive of chemotherapy benefit among patients in clinical practice. More refined risk estimates would be needed for patients with an intermediate RS.

Identifying Long-Term Survival Candidates among Patients with Isolated Locoregionally Recurrent Breast Cancer: Implications of the Use of Systemic Chemotherapy.

PURPOSE: We aimed to investigate the clinicopathologic factors associated with distant metastasis (DM) and post-recurrence overall survival (OS) after salvage treatments for isolated locoregional recurrence (ILRR) of breast cancer and identify long-term surviving patients for providing a more personalized therapy. METHODS: We analyzed 125 patients who underwent salvage local treatments for ILRR after initial curative breast surgery. RESULTS: Fifty-two (41.6%) patients experienced secondary recurrence or disease progression, of which 20 (38.5%) experienced a secondary locoregional recurrence and 40 (76.9%) experienced DM as the first site of failure. In multivariate analysis of distant metastasis free survival (DMFS) and post-recurrence OS, the initial pN2-3 stage, a disease-free interval of < 36 months, and non-curative resection for recurrent disease were independently poor prognosticators. The score for patients stratified according to the number of risk factors increased from 0 to 3; the corresponding 5-year DMFS rates were 91.4%, 53.0%, 35.9%, and 0% and the 5-year OS rates were 97.3%, 70.4%, 32.7%, and 25.0%, respectively (p < 0.001). Systemic chemotherapy reduced DM in patients with a score of 2-3, but it did not in those with a score of 0-1. CONCLUSION: Our collective stratification can help with prognosis prediction for ILRR of breast cancer. Depending on the DM risk of patients, the potential combination of systemic therapy should be discussed further.

Feasibility of Attachable Ring Stimulator for Intraoperative Neuromonitoring during Thyroid Surgery.

OBJECTIVE: Stimulator-attached dissecting instruments are useful for intraoperative nerve monitoring during thyroidectomy. The aim of this study was to evaluate the feasibility of an attachable ring stimulator (ARS) by comparing the electromyography (EMG) amplitudes evoked by an ARS and a conventional stimulator. METHODS: Medical records of fourteen patients who underwent thyroidectomy using intraoperative neuromonitoring between June and August 2019 were retrospectively reviewed. The amplitudes of V1, R1, R2, and V2 signals were checked using both the ARS and a conventional stimulator, at the same point. RESULTS: Both stimulators were tested on 20 recurrent laryngeal nerves (RLNs) and 20 vagus nerves (VNs). In all the nerves, the amplitudes of V1, R1, R2, and V2 were greater than 500 µV. The mean amplitudes of V1, R1, R2, and V2 checked with the ARS were 1175, 1432, 1598, and 1279 µV, respectively. The mean amplitudes of V1, R1, R2, and V2 checked with the conventional stimulator were 1140, 1425, 1557, and 1217 µV, respectively. Difference between amplitudes evoked by the two stimulators for V1, R1, R2, and V2 was 77, 110, 102, and 99 µV, respectively. There was no statistical difference in the amplitudes between the two groups for V1, R1, R2, and V2. CONCLUSION: The ARS transferred electric stimulation as effectively as the conventional stimulator. It is an effective tool for repeated stimulation and facilitates continuous feedback regarding the functional integrity of nerves during thyroid surgery.