[Indications and Outcomes of Laparoscopic Endoscopic Cooperative Surgery(LECS)for Gastric Gastrointestinal Stromal Tumor].

Laparoscopic and endoscopic cooperative surgery(LECS)for gastric gastrointestinal stromal tumor(GIST)has become a popular surgery with both curability and functional preservation. In this study, we examined the outcomes of 14 patients who underwent classical LECS or CLEAN-NET in our hospital. Until March 2022, classical LECS was performed in patients with intraluminal growth tumors or tumors close to the gastroesophageal junction. After April 2022, classical LECS was performed in patients with intraluminal growth tumors without ulceration, and CLEAN-NET was performed in patients with ulceration or intramural growth tumors. There were 10 males and 4 females with a median age of 80.5 years. Intraluminal growth tumor were 8 patients, close to the gastroesophageal junction tumor were 3, and intramural growth tumor were 4, respectively. Five of these patients had tumors with ulceration. Classical LECS was performed in 10 patients and CLEAN-NET in 4 patients, and the median operative time was 165.5 minutes. All patients underwent R0 resection, and no postoperative complications or recurrences were observed. LECS was performed safely, and it is important to select the surgical procedure according to the tumor site and growth type.

[A Case of Local Recurrence of Esophageal Cancer after Chemoradiation Therapy Successfully Treated with Nivolumab].

We report a case of a patient with locally recurrent esophageal cancer after chemoradiation therapy(CRT)who responded to nivolumab. The patient was an 86-year-old man with advanced esophageal cancer. Upper gastrointestinal endoscopy (EGD)revealed a type 2 lesion in the middle thoracic esophagus, and biopsy revealed squamous cell carcinoma(SCC). Contrast- enhanced CT showed invasion of the left main bronchi. The patient was diagnosed as Stage Ⅳa advanced esophageal cancer, and was treated with 5-FU plus cisplatin chemotherapy, and 60 Gy of radiation therapy. The tumor disappeared by CT and EGD, and the patient was followed up for observation. The patient experienced a feeling of tightness again, and EGD revealed an ulcerative lesion in the middle thoracic esophagus, and a biopsy detected SCC. Because of the early recurrence after CRT, the patient was judged to be resistant to 5-FU plus cisplatin chemotherapy, and 8 courses of nivolumab were administered as second-line treatment. Follow-up EGD confirmed disappearance of ulcerative lesions, and no tumors have been observed to date.

[A Case of Long-Term Survival after Multidisciplinary Treatment for Gastrointestinal Neuroendocrine Carcinoma].

A 55-year-old man was admitted to our hospital for examination and treatment of a transverse colon tumor detected at a nearby hospital. After CT, FDG-PET, and laparotomy biopsy, he was diagnosed with neuroendocrine cancer(Ki-67 index 40%)without distant metastasis. He underwent transverse colectomy. The pathological diagnosis was transverse colon neuroendocrine cancer(Ki-67 index 24.7%). Six courses of carboplatin and etoposide therapy as adjuvant chemotherapy were administered. Seven months after surgery, he developed lung metastasis that was surgically removed by partial lung resection. Eighteen months after the initial surgery, liver metastasis developed in S5 and S8. A right hepatic lobectomy was performed and there has been no recurrence after hepatectomy. The patient remains alive at 3 years and 4 months after initial treatment.

[Curative Resection of Pancreatic Head Cancer with Liver Metastasis after GEM plus Nab-PTX Therapy].

A 63-year-old man was admitted to our hospital for examination and treatment of a pancreatic head tumor detected at a nearby hospital. After CT, EUS-FNA, and PET-CT, he was diagnosed with unresectable pancreatic cancer with liver metastasis. After 9 courses of gemcitabine and nab-paclitaxel therapy, the primary tumor was dramatically reduced in size and the liver metastasis had disappeared. He underwent subtotal stomach-preserving pancreaticoduodenectomy. The postoperative diagnosis according to the General Rules of the Study of Pancreatic Cancer(7th edition)was Ph, TS1(15mm), adenosquamous carcinoma, ypT3, ypRP1, ypPL1, R0, ypN0(0/29), M0, CY0, ypStage ⅡA. The histological response was Grade 2. The patient remains alive without recurrence 5 months after surgical resection.

[A Case of Retroperitoneal Liposarcoma Resected 6 Times in 24 Years].

We report a long-surviving case of retroperitoneal liposarcoma requiring 6 operations in 24 years. A 56-year-old female was diagnosed with liposarcoma and it was resected for the first time in 1991. The pathological diagnosis was well-differentiated liposarcoma. Thereafter, we resected recurrences in the retroperitoneum in July 2008, with the left half of the colon in June 2011. Then, we resected the pancreatic tails, spleen, accessory spleen, left adrenal gland, left kidney, and part of the diaphragm in October 2012, and part of the diaphragm, descending colon, and jejunum in October 2014. At this time, there were growing recurrences removed from the pancreatic tails. There is no evidence of recurrence after 24 years after the first resection. In this case, it was thought that the factor correlated with long survival was actively resecting recurrences and recognizing high-grade dedifferentiated type liposarcoma at an early stage.

[Three Cases of Long-Term Surviving Stage IV Gastric Cancer after Conversion Surgery].

The standard therapy for Stage IV gastric cancer is chemotherapy. It is not certain, but conversion surgery is expected to be effective. We report the cases of 3 patients who achieved long-term survival after conversion surgery. Case 1 was of a 59- year-old woman. The tumor was classified as L-Less Post, Type 2, tub1, cT3N2M1(#16a2lat), Stage IV. Then, we initiated S-1 plus cisplatin and the LN achieved PRafter 4 courses. We performed distal gastrectomy with D2 lymph node dissection in February 2011. It was classified as ypT2N2 and the primary lesion was histologically classified as Grade 1a. Case 2 was of a 74- year-old man. The tumor was classified as UM-Less Ant, Type 3, por1, cT3N2H0P1CY1, Stage IV. Then, we initiated docetaxel plus cisplatin plus S-1 and the primary tumor achieved PRafter 6 courses. There were no new tumors and we conducted a laparoscopic examination. After the decision of P0CY0, we performed total gastrectomy with D2 lymph node dissection in April 2012. It was classified as ypT3N1 and the primary lesion was histologically classified as Grade 2. Case 3 was of a 64-yearold woman. The tumor was classified as UM-Less, Type 3, por1, cT3N2H1M0(liver), Stage IV. Then, we initiated capecitabin plus cisplatin and liver metastasis achieved PRafter 6 courses. We performed total gastrectomy with D2 lymph node dissection in July 2012. It was classified as ypT3N1 and the primary lesion was histologically classified as Grade 1b. All postoperative chemotherapy courses were of only S-1. In case 1, the para aortic LN exhibited recurrence 6 months postoperatively. We initiated weekly paclitaxel as second-line therapy. It achieved CRafter 6 courses, and the same trend was maintained. In cases 2 and 3, no therapy was administered after 8 S-1 courses, but no recurrences occurred. All patients survived after 62-77 months postoperatively. A new clinical trial is needed to prove the improvement in prognosis for Stage IV gastric cancer after conversion surgery.

[A Case of Long-Term Survival with Multidisciplinary Treatment in Unresectable Advanced Gastric Cancer Found with Ovarian Tumor].

Although chemotherapy is the standard treatment for unresectable advanced gastric cancer, its prognosis is poor and the median survival time is only around 10 months. With some literature consideration, we report that ovarian metastasis triggered the diagnosis of unresectable advanced gastric cancer with long-term survival through multidisciplinary treatment. This is the case of a 69-year-old woman, who was diagnosed with ovarian tumor and underwent right extracorporectomy and omentum resection in 2011. Pathological diagnosis suspected adenocarcinoma. In February 2012, EGD found a type 4 tumor in the upper portion of the pyloric area of the stomach. She was diagnosed with gastric cancer with Stage IV(T4aN0M1 [ovary])ovarian metastasis. Because the primary tumor was HER2 positive, XP plus HER therapy(capecitabine 1,000mg/m2 twice a day for 14 days, CDDP 80mg/m2 every 3 weeks on the first day, and trastuzumab 8 mg/kg every 3 weeks on the first day)was administered since March 2012. No metastasis was found in the imaging examination after 8 courses of chemotherapy, and we also confirmed the reduction of the primary tumor in the EGD. Based on the images, primary resection was already possible. In December 2012, after a diagnostic laparoscopy, total gastrectomy with D2 lymph node dissection was performed. Postoperative pathology was diagnosed as Stage IV(pT4aN0M1). To control postoperative micrometastasis, capecitabine therapy(1,000mg/m2 twice a day for 14 days)was administered for 12 months starting from February 2013. Then, recurrence and metastasis were not observed during follow-up. However, in January 2017, a circumstellar stenotic tumor was found in the rectum and was diagnosed as recurrence of peritoneal dissemination through images. In February 2017, artificial ostomy(in the sigmoid colon, double-mouth type)was made. Then, the patient underwent an outpatient chemotherapy, with hospital visits, and she survives.

Evaluation of short-term outcomes of laparoscopic-assisted surgery for colorectal cancer in elderly patients aged over 75 years old: a multi-institutional study (YSURG1401).

BACKGROUND: The short-term outcomes of laparoscopic-assisted surgery for colorectal cancer (LAC) have not been fully evaluated in elderly patients. The aim of this study was to compare the short term surgical outcomes of LAC between the patients older than 75 years and those with non-elderly patients. METHODS: This retrospective multi-institutional study selected patients who underwent LAC between April 2013 and March 2014 at Yokohama City University Hospital and its related general hospitals. The patients were categorized into two groups: elderly patients (>75 years of age: group A) and non-elderly patients (<75 years of age: group B). Surgical outcomes and post operative complications were compared between the two groups. RESULTS: A total of 237 patients were evaluated in the present study. Eighty-four patients were classified into group A, and 153 into group B. Preoperative clinicopathological outcomes demonstrated no significant differences except for the ASA score. When comparing the surgical outcomes between group A and group B, the rate of conversion to open procedure (3.6% vs 5.2%, P = 0.750), median operation time (232 min vs 232 min, P = 0.320), median blood loss (20 ml vs 12 ml, P = 0.350). The differences were not significantly different in the surgical outcomes. The incidences of > grade 2 post operative surgical complications were similar between two groups ((19.0% vs 15.7%, p = 0.587). No mortality was observed in this study. The length of postoperative hospital stay was also similar (10 days vs 10 days, p = 0.350). CONCLUSIONS: The present study suggested that LAC is safe and feasible, regardless of the age of the patient, especially for elderly patients who may be candidates for colon cancer surgery.

[A Case of Surgical Resection of Hepatic Leiomyosarcoma].

A 72-year-old woman was referred to our hospital due to an abdominal mass. Ultrasound sonography and computed tomography revealed liver tumors in liver segments 2 and 3. The preoperative diagnosis was hepatocellular carcinoma. The patient underwent an extended left lobectomy. Histological examination of resected specimens revealed that the tumors were hepatic leiomyosarcoma. We report our experience of this hepatic leiomyosarcoma, along with the literature on this tumor type.

[A Case of Rectal Cancer with Unresectable Liver Metastasis Responding to Rechallenge with FOLFIRI].

A 63-year-old man underwent low anterior resection for rectal cancer.A synchronous liver metastasis located in segment 8 was 12 cm in diameter and unresectable due to its proximity to the inferior vena cava(IVC).The postoperative pathological findings revealed a T3(SS), N0, M1(liver)Stage IV tumor, and wild type K-RAS was expressed.We chose FOLFIRI plus cetuximab(Cmab)for first-line chemotherapy.After 6 courses, we changed the molecular target drug from Cmab to bevacizumab( Bmab)because the liver metastasis remained unresectable.The patient had long-term stable disease(SD)for approximately 30 months with the FOLFIRI-based regimen.We then changed the regimen to mFOLFOX6 plus Bmab for second-line, Cmab for third-line, and trifluridine/tipiracil hydrochloride for fourth-line chemotherapy to treat progressive disease(PD).After treatment with these chemotherapies, the patient wished to continue treatment.We restarted FOLFIRI plus Bmab for fifth-line chemotherapy as his general condition was still good.Consequently, his tumor markers levels decreased with stabilization of the disease on CT scans, and he continued therapy for 6 months while maintaining a good quality of life.This case suggested that rechallenge with anti-cancer agents could be effective and improve the prognosis of colorectal cancer patients after using all key drugs.

[A case of intractable fistula after low anterior resection repaired by transsacral direct suture].

We report a case of an intractable fistula repaired by transsacral direct suture. A 65-year-old man underwent low anterior resection for rectal cancer. He subsequently underwent ileostomy due to anastomosis leakage. The fistula of the anastomosis persisted 3 months after surgery. He underwent surgery to repair the fistula using a transsacral approach. After removing the coccyx, the fistula in the postrectal space was exposed directly. The presence of the fistula was confirmed by an air leak test and was closed by direct suture. After 33 days, the patient underwent ileostomy closure.

Small extracellular vesicles carrying reovirus, tumor antigens, interferon-ß, and damage-associated molecular patterns for efficient tumor treatment.

Small extracellular vesicles (SEV) have attracted much attention both as mediators of intercellular communication and as drug delivery systems. In addition, recent studies have shown that SEV containing virus components and virus particles are released from virus-infected cells. Oncolytic viruses, which efficiently kill tumor cells by tumor cell-specific replication, have been actively studied as novel anticancer agents in clinical and preclinical studies. However, it remains to be fully elucidated whether SEV released from oncolytic virus-infected cells are involved in the antitumor effects of oncolytic viruses. In this study, we examined the tumor cell killing efficiencies and innate immune responses following treatment with SEV released from oncolytic reovirus-infected tumor cells in vitro and in vivo. Reovirus-infected B16 cells secreted SEV associated with or containing reovirus particles (Reo-SEV) with a diameter of approximately 130 nm and a zeta potential of -17 mV, although death of reovirus-infected B16 cells was not observed. The secreted Reo-SEV also contained interferon (IFN)-ß, tumor antigens, and damage-associated molecular patterns (DAMPs), including heat shock proteins (HSPs). Reo-SEV were secreted from the tumor tissues of reovirus-injected mice. Inhibition of the SEV secretion pathway using GW4869, which is a neutral sphingomyelinase inhibitor, resulted in significant reduction in the infectious titers of reovirus in the culture supernatants, suggesting that the cells released progeny virus via the SEV secretion pathway. Reo-SEV more efficiently killed mouse tumor cells and induced innate immune responses in mouse bone marrow-derived dendritic cells than reovirus. Reovirus and Reo-SEV mediated efficient and comparable levels of growth suppression of B16 subcutaneous tumors and induction of tumor infiltration of CD8+ T cells following intravenous administration. These results indicate that Reo-SEV are a promising oncolytic agent and that SEV are an effective delivery vehicle for oncolytic virus.

Synthesis of a novel adapter lipid using Fc-region mediated antibody modification for post-insert preparation of transferrin receptor targeted messenger RNA-loaded lipid nanoparticles.

Lipid nanoparticles (LNPs) are promising delivery systems with the ability to deliver small interfering RNA (siRNA) and messenger RNA (mRNA) in diseased tissues and intracellular sites of action. However, delivery to non-hepatic tissues via systemic administration remains challenging. Antibody modification of LNPs is a hopeful approach for improving their selectivity to target tissues. The conventional method of antibody modification via thiol-maleimide linkage is concerned with reduced recognition efficiency of the disease-related target molecules owing to variations in antibody orientation on the surface of the LNPs. In this study, we developed a novel adapter lipopeptide for antibody modification of LNPs via the Fc-region. Here, we selected RI7-217, an anti-transferrin receptor antibody, as the ligand. Through optimization of spacer peptides, we found a FcBP-EKGG-lipid exhibits high water-dispersibility for post-insertion method to LNPs. We prepared RI7-217-modified LNPs by modifying LNPs with FcBP-EKGG-lipids and mixing the antibodies. We found that the luciferase protein expression of RI7-217-modified LNPs was significantly enhanced in an antibody-specific manner against transferrin receptor-expressing U-87 MG cells. This information would be valuable in the development of antibody-modified LNPs for cell-selective targeting.

Non-invasive intraductal oncocytic papillary neoplasm forming a protruding lesion toward the duodenum from the accessory papilla: a case report.

BACKGROUND: Intraductal oncocytic papillary neoplasm (IOPN), previously classified as a subtype of intraductal papillary mucinous neoplasm (IPMN), has been described as an independent disease by the WHO since 2019. IOPN is a rare tumor, with few reported cases. Herein, we report a case of resected non-invasive IOPN that formed a lesion protruding toward the duodenum from the accessory papilla. CASE PRESENTATION: An 80-year-old woman was referred to our hospital because of a giant mass in the pancreatic head detected on abdominal contrast-enhanced computed tomography (CT) performed for a close examination of a mass in the right breast. CT revealed a 90-mm-sized tumor with a mixture of solid and cystic components, with contrast enhancement in the pancreatic head, and a dilated main pancreatic duct. Esophagogastroduodenoscopy revealed a semi-circumferential papillary tumor protruding toward the duodenal lumen, which did not protrude from the papilla of Vater. Transpapillary biopsy led to a preoperative diagnosis of IPMN with an associated invasive carcinoma. As there were no distant metastasis, open subtotal stomach-preserving pancreaticoduodenectomy was performed. Analysis of the surgical specimen and histopathological examination revealed that the tumor was an IOPN that protruded toward the duodenal mucosa from the accessory papilla while replacing the duodenal mucosa with no obvious stromal invasion. CONCLUSION: IOPN is a rare and poorly recognized tumor with few reported cases. There have been no reports describing IOPN forming a protruding lesion toward the duodenum from the accessory papilla. Therefore, further accumulation of cases such as this one is important to advance the study of IOPN.

Clinical Impact of Preoperative Neutrophil-to-Lymphocyte Ratio in Surgical Patients With Pancreatic Cancer.

BACKGROUND/AIM: The prognosis of patients with pancreatic cancer remains poor, despite recent advances in surgical techniques, perioperative care, neoadjuvant and adjuvant chemotherapy. This study aimed to investigate the preoperative neutrophil-to-lymphocyte ratio (NLR) as a prognostic factor and determine the optimal cutoff value in surgical patients with pancreatic cancer. PATIENTS AND METHODS: We retrospectively enrolled 461 patients with pancreatic cancer who underwent resection between January 2013 and December 2022 in the Department of Gastrointestinal Surgery at Kanagawa Cancer Center. The association between continuous or categorical variables and NLR was analyzed using the Mann-Whitney U-test and Fisher's exact test. Overall survival (OS) and relapse-free survival (RFS) rates were calculated using the Kaplan-Meier method. Univariate and multivariate analyses were performed using Cox proportional-hazard regression models. RESULTS: The optimal cutoff value for the preoperative NLR was 3.2. The NLR≥3.2 was associated with a large tumor size (p=0.005), poor histological differentiation (p=0.002), and less adjuvant chemotherapy (p=0.048). The NLR≥3.2 had an important influence on the decreased OS (21.6 vs. 25.8 months), and RFS (10.3 vs. 14.3 months). In univariate and multivariate analyses, the preoperative NLR was an independent prognostic factor for OS (p=0.022) and RFS (p=0.002). CONCLUSION: Preoperative NLR (cutoff value: 3.2) within two weeks before surgery is a prognostic factor for OS and RFS in surgical patients with pancreatic cancer. This study could help establish evidence on the immune system's impact and a unified treatment strategy pre-surgery, potentially improving the prognosis for patients with pancreatic cancer.

Effect of Cholesterol Content of Lipid Composition in mRNA-LNPs on the Protein Expression in the Injected Site and Liver After Local Administration in Mice.

Delivery of messenger RNA (mRNA) using lipid nanoparticles (LNPs) is expected to be applied to various diseases following the successful clinical use of the mRNA COVID-19 vaccines. This study aimed to evaluate the effect of the cholesterol molar percentage of mRNA-LNPs on protein expression in hepatocellular carcinoma-derived cells and in the liver after intramuscular or subcutaneous administration of mRNA-LNPs in mice. For mRNA-LNPs with cholesterol molar percentages reduced to 10 mol% and 20 mol%, we formulated neutral charge particles with a diameter of approximately 100 nm and polydispersity index (PDI) <0.25. After the intramuscular or subcutaneous administration of mRNA-LNPs with different cholesterol molar percentages in mice, protein expression in the liver decreased as the cholesterol molar percentage in mRNA-LNPs decreased from 40 mol% to 20 mol% and 10 mol%, suggesting that reducing the cholesterol molar percentage in mRNA-LNPs decreases protein expression in the liver. Furthermore, in HepG2 cells, protein expression decreased as cholesterol in mRNA-LNPs was reduced by 40 mol%, 20 mol%, and 10 mol%. These results suggest that the downregulated expression of mRNA-LNPs with low cholesterol content in the liver involves degradation in systemic circulating blood and decreased protein expression after hepatocyte distribution.

Induction of liver-resident memory T cells and protection at liver-stage malaria by mRNA-containing lipid nanoparticles.

Recent studies have suggested that CD8+ liver-resident memory T (TRM) cells are crucial in the protection against liver-stage malaria. We used liver-directed mRNA-containing lipid nanoparticles (mRNA-LNPs) to induce liver TRM cells in a murine model. Single-dose intravenous injections of ovalbumin mRNA-LNPs effectively induced antigen-specific cytotoxic T lymphocytes in a dose-dependent manner in the liver on day 7. TRM cells (CD8+ CD44hi CD62Llo CD69+ KLRG1-) were induced 5 weeks after immunization. To examine the protective efficacy, mice were intramuscularly immunized with two doses of circumsporozoite protein mRNA-LNPs at 3-week intervals and challenged with sporozoites of Plasmodium berghei ANKA. Sterile immunity was observed in some of the mice, and the other mice showed a delay in blood-stage development when compared with the control mice. mRNA-LNPs therefore induce memory CD8+ T cells that can protect against sporozoites during liver-stage malaria and may provide a basis for vaccines against the disease.

Incidence and Risk of Venous Thromboembolism in Patients With Resectable Pancreatic Cancer Receiving Neoadjuvant Chemotherapy.

BACKGROUND/AIM: Pancreatic cancer has the highest risk of venous thromboembolism (VTE). Additionally, chemotherapy for cancer patients increases the risk of developing VTE. Due to recent advances in neoadjuvant chemotherapy (NAC) regimens, more patients with resectable pancreatic cancer will receive NAC. However, the incidence, risk, and predictors of developing VTE in these patients have not been fully evaluated. PATIENTS AND METHODS: We retrospectively evaluated the incidence, risk, and predictors of VTE among 67 consecutive patients with resectable pancreatic cancer who received neoadjuvant combination therapy with gemcitabine+S-1 (NAC-GS) followed by surgery and 45 patients with resectable pancreatic cancer who underwent upfront surgery (Up-S). RESULTS: The incidence of VTE in the NAC-GS and Up-S groups was 10.4% and 6.6%, respectively. Preoperative D-dimer levels were significantly higher in the NAC-GS group, and D-dimer levels were significantly increased during NAC-GS. Preoperative D-dimer level was the only predictor for VTE in patients with resectable pancreatic cancer who received NAC-GS. CONCLUSION: There is an increased risk of developing VTE during NAC. Screening with D-dimer and taking appropriate measures to suppress critical VTE is essential to provide NAC to patients with resectable pancreatic cancer.

Outcomes of feeding jejunostomy after pancreaticoduodenectomy: A single-center experience.

Aims: Feeding jejunostomy tube (FJT) is one option for enteral nutrition after pancreaticoduodenectomy (PD); however, controversy regarding its clinical outcome(s) persists. The aim of the present study was to determine the safety and efficacy of FJT management. Materials and Methods: Data from 156 consecutive patients, who underwent PD between January 2015 and December 2017, were retrospectively reviewed. Safety was assessed according to postoperative and tube-related complications. Nutritional efficacy was evaluated based on improvement in serum albumin levels. Results: Thirty-day morbidity and mortality rates were 61.0% (n = 95) and 1.9% (n = 3), respectively. The rates of clinically relevant postoperative pancreatic fistula and delayed gastric emptying were 30.8% and 9.0%, respectively. In total, nine (5.8%) patients experienced complications directly related to FJT. Eight patients experienced surgical site infection adjacent to the catheter/skin interface. Although all required catheter removal at the bedside or in the office, none required reoperation. The improvement in serum albumin level 1 month after PD was 40.7% compared with 1 week after PD. Conclusion: FJT was useful in improving nutritional intake and status. Although FJT was associated with minor self-limiting complications, they could be managed by simple bedside or office treatment. As such, results of this study support the safety and efficacy of the FJT protocol used in the authors' department for nutritional management.

Stability Study of mRNA-Lipid Nanoparticles Exposed to Various Conditions Based on the Evaluation between Physicochemical Properties and Their Relation with Protein Expression Ability.

Lipid nanoparticles (LNPs) are currently in the spotlight as delivery systems for mRNA therapeutics and have been used in the Pfizer/BioNTech and Moderna COVID-19 vaccines. mRNA-LNP formulations have been indicated to require strict control, including maintenance at fairly low temperatures during their transport and storage. Since it is a new pharmaceutical modality, there is a lack of information on the systematic investigation of how storage and handling conditions affect the physicochemical properties of mRNA-LNPs and their protein expression ability. In this study, using the mRNA-LNPs with standard composition, we evaluated the effects of temperature, cryoprotectants, vibration, light exposure, and syringe aspiration from the vials on the physicochemical properties of nanoparticles in relation to their in vitro/in vivo protein expression ability. Among these factors, storage at -80 °C without a cryoprotectant caused a decrease in protein expression, which may be attributed to particle aggregation. Exposure to vibration and light also caused similar changes under certain conditions. Exposure to these factors can occur during laboratory and hospital handling. It is essential to have sufficient knowledge of the stability of mRNA-LNPs in terms of their physical properties and protein expression ability at an early stage to ensure reproducible research and development and medical care.