Conversion surgery for initially unresectable locally advanced pancreatic ductal adenocarcinoma after chemotherapy followed by carbon-ion radiotherapy: a case report.

BACKGROUND: Recent advances in chemotherapy and chemoradiotherapy have enabled conversion surgery (CS) to be performed for selected patients with initially unresectable locally advanced (LA) pancreatic ductal adenocarcinoma (PDAC). Many studies indicate CS might extend the survival of patients with initially unresectable LA PDAC. However, several clinical questions concerning CS remain, such as the optimal preoperative treatment. Carbon-ion radiotherapy (CIRT) is a unique radiotherapy that offers higher biological effectiveness than conventional radiotherapy. Here, we report a long-term survival case with initially unresectable LA PDAC who underwent CS after chemotherapy followed by CIRT. CASE PRESENTATION: The patient was a 72-year-old Japanese woman with unresectable LA pancreatic head cancer with tumor contact to the superior mesenteric artery (SMA). She underwent four courses of chemotherapy (gemcitabine plus nanoparticle albumin-bound paclitaxel). However, the lesion did not shrink and tumor contact with the SMA did not improve after chemotherapy. Because the probability of achieving curative resection was judged to be low, she underwent radical dose CIRT, and chemotherapy was continued. She complained of vomiting 2 months after CIRT. Although imaging studies showed no tumor growth or metastasis, a duodenal obstruction which was speculated to be an adverse effect of CIRT was observed. She could not eat solid food and a trans-nasal feeding tube was inserted. Therapeutic intervention was required to enable enteral nutrition. We proposed several treatment options. She chose resection with the expectation of an anti-tumor effect of chemotherapy and CIRT rather than course observation with tube feeding or bypass surgery. Therefore, subtotal-stomach-preserving pancreatoduodenectomy with portal vein resection was performed as CS. Pathological examination of the resected specimen revealed an R0 resection with a histological response of Evans grade IIA. Postoperatively, she recovered uneventfully. Adjuvant chemotherapy with tegafur/gimeracil/oteracil (S1) was administrated. At the time of this report, 5 years have passed since the initial consultation and she has experienced no tumor recurrence. CONCLUSIONS: The present case suggests that multidisciplinary treatment consisting of a combination of recent chemotherapy and CIRT may be beneficial for unresectable LA PDAC. However, further studies are required to assess the true efficacy of this treatment strategy.

Neoadjuvant chemotherapy enhances anti-tumor immune response of tumor microenvironment in human esophageal squamous cell carcinoma.

Although chemotherapy has been an essential treatment for cancer, the development of immune checkpoint blockade therapy was revolutionary, and a comprehensive understanding of the immunological tumor microenvironment (TME) has become crucial. Here, we investigated the impact of neoadjuvant chemotherapy (NAC) on immune cells in the TME of human esophageal squamous cell carcinoma using single cell RNA-sequencing. Analysis of 30 fresh samples revealed that CD8+/CD4+ T cells, dendritic cells (DCs), and macrophages in the TME of human esophageal squamous cell carcinoma showed higher levels of an anti-tumor immune response in the NAC(+) group than in the NAC(-) group. Furthermore, the immune cells of the NAC(+) group interacted with each other resulting in enhanced anti-tumor immune response via various cytokines, including IFNG in CD8+/CD4+ T cells, EBI3 in DCs, and NAMPT in macrophages. Our results suggest that NAC potentially enhances the anti-tumor immune response of immune cells in the TME.

Involvement of angiogenesis in cancer-associated acinar-to-ductal metaplasia lesion of pancreatic cancer invasive front.

PURPOSE: This study aimed to demonstrate the involvement of angiogenesis in cancer-associated acinar-to-ductal metaplasia (CA-ADM) lesion of invasive front pancreatic ductal adenocarcinoma (PDAC) and investigate the possible mechanism. METHODS: Tissue samples from 128 patients with PDAC and 36 LSL-KrasG12D/+; LSL-Trp53R172H/+; Pdx-1-Cre mice were analyzed. Immunohistochemical assay was performed using HE, anti-CK19 and anti-amylase to confirm the presence of CA-ADM lesions, using anti-CD34 and anti-CD31 to measure microvessel density (MVD), and using anti-CD68, anti-CD163, anti-iNOS, or anti-MMP9 to evaluate the immune microenvironment. We performed multiplex immunohistochemical assay to detect the co-expression of MMP9 and CD68 on macrophage. We examined clinical outcomes and other clinicopathological factors to determine the significance of high-level MVD of CA-ADM on survival and liver metastasis. We performed tube formation assay to evaluate the effect of macrophage on angiogenic capacity in vitro. RESULTS: Angiogenesis was significantly abundant in CA-ADM lesions compared with that in PDAC lesions in human and mouse tissues. High-level MVD in CA-ADM lesions was an independent predictor of poor prognosis (P = 0.0047) and the recurrence of liver metastasis (P = 0.0027). More CD68-positive and CD163-positive macrophages were detected in CA-ADM lesions than in PDAC. The percentage of CD68-positive macrophages was positively correlated with MVD in CA-ADM lesions. Multiplex-immunostaining revealed that MMP9 was expressed in CD68-positive macrophages of CA-ADM lesions. In CA-ADM lesions, the percentage of macrophages was positively correlated with MMP9 expression, which positively correlated with microvessel density. CONCLUSION: CA-ADM related angiogenesis is a promising predictive marker for poor prognosis of PDAC and may provide an attractive therapeutic target for PDAC.

Single-cell transcriptome analysis reveals functional changes in tumour-infiltrating B lymphocytes after chemotherapy in oesophageal squamous cell carcinoma.

BACKGROUND: Tumour immune microenvironment is related with carcinogenesis and efficacy of immunotherapy. B cells play major roles in humoral immunity, but detailed functions of tumour-infiltrating B lymphocytes (TIL-Bs) are unknown. Therefore, our aim was to investigate the functional heterogeneity of TIL-Bs in oesophageal squamous cell carcinoma (ESCC) and lymph nodes (LNs) during chemotherapy. METHODS: Single-cell transcriptome analysis was performed on 23 specimens. We also performed immunohistochemical analysis of immunoglobulin κ C (IGKC), an antibody-secreting cell (ASC) marker, in 166 ESCC samples and evaluated the implication of IGKC in 2-year recurrence free survival (RFS) and 3-year overall survival (OS). RESULTS: A total of 81,246 cells were grouped into 24 clusters. We extracted B cell clusters based on canonical markers and identified 12 TIL-B subtypes in ESCC. We found that several functions, such as co-stimulation and CD40 signalling, were enhanced in TIL-Bs after chemotherapy. The proportion of naive B cells (NBCs) decreased and B cell activation genes were up-regulated in NBCs after chemotherapy. The proportion of ASCs in tumours increased with the loss of migratory abilities and antibody production in ASCs was promoted after chemotherapy. Differentially expressed genes up-regulated with chemotherapy in ASCs correlated with prolonged survival with oesophageal cancer (p = .028). In a metastatic LN, the ASC proportion increased and B cell differentiation was enhanced. In immunohistochemical analysis, RFS and OS of high IGKC expression cases were significantly better than those of low IGKC expression cases (RFS: p < .0001, OS: p < .0001). And in multivariable analysis, the expression of IGKC was an independent favourable prognostic factor for RFS (hazard ratio (HR): 0.23, 95% confidence interval (CI): 0.12-0.45, p < .0001) and OS (HR: 0.20, 95% CI: 0.086-0.47, p = .0002) in ESCC. CONCLUSIONS: Our findings provide novel insights for the heterogeneity of TIL-Bs during chemotherapy and will be useful to understand the clinical importance of TIL-Bs.

Heterogeneity of Cancer-Associated Fibroblasts and the Tumor Immune Microenvironment in Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers, with a 5-year survival rate of 9%. Cancer-associated fibroblasts (CAFs) have historically been considered tumor-promoting. However, multiple studies reporting that suppression of CAFs in PDAC mouse models resulted in more aggressive tumors and worse prognosis have suggested the existence of a tumor-suppressive population within CAFs, leading to further research on heterogeneity within CAFs. In recent years, the benefits of cancer immunotherapy have been reported in various carcinomas. Unfortunately, the efficacy of immunotherapies in PDAC has been limited, and the CAF-driven cancer immunosuppressive microenvironment has been suggested as the cause. Thus, clarification of heterogeneity within the tumor microenvironment, including CAFs and tumor immunity, is urgently needed to establish effective therapeutic strategies for PDAC. In this review, we report the latest findings on the heterogeneity of CAFs and the functions of each major CAF subtype, which have been revealed by single-cell RNA sequencing in recent years. We also describe reports of tumor-suppressive CAF subtypes and the existence of CAFs that maintain a differentiated PDAC phenotype and review the potential for targeted therapy.

Subtypes in pancreatic ductal adenocarcinoma based on niche factor dependency show distinct drug treatment responses.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is characterized by abundant stroma in which microenvironmental (niche) factors promote PDAC progression. In mouse models, reduction of the stroma increased the proportion of poorly differentiated PDAC with a worse prognosis. Here, we aimed to clarify the effects of stroma on PDAC that may define the PDAC phenotype and induce distinct therapeutic responses. METHODS: The molecular features of PDAC based on differentiation grade were clarified by genome and transcriptome analysis using PDAC organoids (PDOs). We identified the dependency on niche factors that might regulate the differentiation grade. A three-dimensional co-culture model with cancer-associated fibroblasts (CAFs) was generated to determine whether CAFs provide niche factors essential for differentiated PDAC. PDOs were subtyped based on niche factor dependency, and the therapeutic responses for each subtype were compared. RESULTS: The expression profiles of PDOs differed depending on the differentiation grade. Consistent with the distinct profiles, well differentiated types showed high niche dependency, while poorly differentiated types showed low niche dependency. The three-dimensional co-culture model revealed that well differentiated PDOs were strongly dependent on CAFs for growth, and moderately differentiated PDOs showed plasticity to change morphology depending on CAFs. Differentiated PDOs upregulated the expression of mevalonate pathway-related genes correlated with the niche dependency and were more sensitive to simvastatin than poorly differentiated PDOs. CONCLUSIONS: Our findings suggest that CAFs maintain the differentiated PDAC phenotype through secreting niche factors and induce distinct drug responses. These results may lead to the development of novel subtype-based therapeutic strategies.

Repositioning of duloxetine to target pancreatic stellate cells.

Pancreatic cancer cells (PCCs) are surrounded by an abundant stroma, which is produced by pancreatic stellate cells (PSCs). PSCs promote tumor cell proliferation and invasion. The objective of the current study was to identify compounds that suppress PSC activation. Gene expression profiles of cancer-derived fibroblasts and normal fibroblasts were used, and the pathway analysis suggested altered pathways that were chosen for validation. It was found that the 'neuroactive ligand-receptor interaction' pathway from the Kyoto Encyclopedia of Genes and Genomes pathway analysis was one of the altered pathways. Several compounds related with this pathway were chosen, and changes in PSC activity were investigated using fluorescence staining of lipid droplets, reverse transcription-quantitative PCR, western blotting, and invasion and migration assays. Among these candidates, duloxetine, a serotonin-noradrenaline reuptake inhibitor, was found to suppress PSC activation and disrupt tumor-stromal interaction. Thus, duloxetine may be a potential drug for suppressing PSC activation and pancreatic cancer growth.

Bone marrow-derived macrophages converted into cancer-associated fibroblast-like cells promote pancreatic cancer progression.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by a desmoplastic reaction caused by cancer-associated fibroblasts (CAFs), which provokes treatment resistance. CAFs are newly proposed to be heterogeneous populations with different functions within the PDAC microenvironment. The most direct sources of CAFs are resident tissue fibroblasts and mesenchymal stem cells, however, the origins and functions of CAF subtypes remain unclear. Here, we established allogeneic bone marrow (BM) transplantation models using spontaneous PDAC mice, and then investigated what subtype cells derived from BM modulate the tumor microenvironment and affect the behavior of pancreatic cancer cells (PCCs). BM-derived multilineage hematopoietic cells were engrafted in recipient pancreas, and accumulated at the invasive front and central lesion of PDAC. We identified BM macrophages-derived CAFs in tumors. BM-derived macrophages treated with PCC-conditioned media expressed CAF markers. BM-derived macrophages led the local invasion of PCCs in vitro and enhanced the tumor invasive growth in vivo. Our data suggest that BM-derived cells are recruited to the pancreas during carcinogenesis and that the specific subpopulation of BM-derived macrophages partially converted into CAF-like cells, acted as leading cells, and facilitated pancreatic cancer progression. The control of the conversion of BM-derived macrophages into CAF-like cells may be a novel therapeutic strategy to suppress tumor growth.

Correction to: Long-term survival after hepatectomy for metachronous liver metastasis of pancreatic ductal adenocarcinoma: a case report.

An amendment to this paper has been published and can be accessed via the original article.

Development of Wernicke's encephalopathy long after subtotal stomach-preserving pancreatoduodenectomy: a case report.

BACKGROUND: Wernicke's encephalopathy (WE) is an acute neuropsychiatric disorder resulting from thiamine (vitamin B1) deficiency, frequently associated with chronic alcoholism and total parenteral nutrition without thiamine. However, only a few reports have focused on the relationship between WE and subtotal stomach-preserving pancreatoduodenectomy (SSPPD). CASE PRESENTATION: A 71-year-old woman underwent SSPPD for an adenocarcinoma of the ampulla of Vater. Although there had been no evidence of recurrence, the patient was treated with antibiotics for cholangitis at 12 and 31 months, respectively, post-surgery. Thereafter, the patient presented with vomiting and disorientation 33 months after surgery. Although she was admitted and underwent closer inspection by a neurologist and a psychiatrist, the exact cause of these syndromes remained unknown. The psychiatrist measured thiamine concentration to examine the cause of disorientation. After 6 days, her level of consciousness worsened. Magnetic resonance imaging of the head showed symmetrically multiple abnormal hyperintense signals on fluid-attenuated inversion-recovery and diffusion weighted image, compatible with WE. An administration of intravenous thiamine was immediately initiated. After 8 days of the measurement of the thiamine level, the patient's serum thiamine level was found to be 6 µg/mL (reference range, 24-66 µg/mL). Accordingly, the patient was diagnosed with WE. Shortly after starting the treatment, blood thiamine value reached above normal range with significant improvement of her confusional state. However, short-term memory and ataxia remained. CONCLUSIONS: Development of WE after SSPPD is uncommon. However, to prevent an after-effect, the possibility of development of WE after SSPPD should be recognized.

Long-term survival after hepatectomy for metachronous liver metastasis of pancreatic ductal adenocarcinoma: a case report.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is among the most aggressive malignancies. The prognosis for recurrence after surgery is extremely unfavorable, and liver metastasis of PDAC confers poor prognosis despite resection. CASE PRESENTATION: A 69-year-old man was admitted to our hospital for further examination and treatment, including surgery for a pancreatic tumor. On close inspection, he was suspected to have pancreatic head cancer without enlarged lymph nodes or distant metastasis, and pancreatoduodenectomy with D2 lymph node dissection was performed. A postoperative pathological examination revealed well-differentiated invasive ductal adenocarcinoma with lymph node metastasis (stage IIB; pT2N1M0). Postoperatively, he received adjuvant chemotherapy containing gemcitabine for 1 year. Eight years after the radical surgery, his serum carbohydrate antigen 19-9 level was elevated, and computed tomography (CT) and magnetic resonance imaging revealed a well-circumscribed 10-mm mass in liver segment 5. Positron emission tomography/CT also revealed high fluorine-18-fluorodeoxyglucose uptake only in this hepatic tumor. Accordingly, the patient was diagnosed with a solitary liver metastasis of PDAC. As the liver metastasis was isolated and identified long after the initial surgery, we decided to resect it using laparoscopic partial hepatectomy of segment 5. Histopathological examination confirmed liver metastasis of PDAC and the patient received adjuvant chemotherapy containing S-1. No evidence of recurrence has been seen for 11 years since the pancreatoduodenectomy and 3 years since the hepatic resection. CONCLUSIONS: Cases of metachronous liver metastasis of PDAC after radical surgery, in which patients exhibit long-term survival without recurrence after hepatectomy, are extremely rare. Hepatectomy may confer long-term survival, and the time to postoperative recurrence and the number of liver metastases may be useful criteria for deciding whether to perform hepatic resection.

Neutrophil extracellular traps promote liver micrometastasis in pancreatic ductal adenocarcinoma via the activation of cancer­associated fibroblasts.

Cancer­associated fibroblasts (CAFs) promote the progression of pancreatic ductal adenocarcinoma (PDAC) via tumor­stromal interactions. Neutrophil extracellular traps (NETs) are extracellular DNA meshworks released from neutrophils together with proteolytic enzymes against foreign pathogens. Emerging studies suggest their contribution to liver metastasis in several types of cancer. Herein, in order to investigate the role of NETs in liver metastasis in PDAC, the effects of NET inhibitors on spontaneous PDAC mouse models were evaluated. It was demonstrated that DNase I, a NET inhibitor, suppressed liver metastasis. For further investigation, further attention was paid to liver micrometastasis and an experimental liver metastasis mouse model was used that was generated by intrasplenic tumor injection. Furthermore, DNase I also suppressed liver micrometastasis and notably, CAFs accumulated in metastatic foci were significantly decreased in number. In vitro experiments revealed that pancreatic cancer cells induced NET formation and consequently NETs enhanced the migration of hepatic stellate cells, which was the possible origin of CAFs in liver metastasis. On the whole, these results suggest that NETs promote liver micrometastasis in PDAC via the activation of CAFs.

Hepatic inflammatory pseudotumor associated with primary biliary cholangitis and elevated alpha-fetoprotein lectin 3 fraction mimicking hepatocellular carcinoma.

BACKGROUND: Hepatic inflammatory pseudotumor (IPT) is a rare benign lesion. Because there is no specific laboratory marker or radiographic appearance, the majority of reported cases of hepatic IPT have been diagnosed after surgery or at autopsy. The etiology of hepatic IPT remains unclear but several mechanisms have been postulated such as infection or immune reaction. CASE PRESENTATION: A 79-year-old woman had been seeing her family doctor for hypertension, and she had been diagnosed with liver dysfunction for about 10 years. She continued attending follow-ups because of her drinking habit. Two months before her visiting our institution, further elevation of hepatobiliary enzymes was noted, and abdominal ultrasonography showed a hepatic tumor 4 cm in diameter in the lateral segment, so she was referred to our hospital. Hepatocellular carcinoma (HCC) was suspected because alpha-fetoprotein (102 ng/ml) (AFP) and lectin 3 (L3) fraction (85.4%) were elevated and the appearance on enhanced computed tomography was not inconsistent with HCC. Thus, we performed laparoscopic hepatectomy. She recovered uneventfully and was discharged on postoperative day 7. Pathological diagnosis revealed that the tumor was hepatic IPT and that the background liver condition was primary biliary cholangitis (PBC). AFP and L3 fraction decreased to normal ranges after surgery. CONCLUSIONS: In 7 of 29 patients (24.1%) with reported cases of tumor markers in liver IPT, carbohydrate antigen 19-9 was elevated and AFP was elevated in 2 of 58 patients (3.4%). AFP is also frequently elevated in benign liver diseases such as hepatitis and liver cirrhosis, and L3 fraction has been used as a tumor marker for HCC with high specificity. To our knowledge, this is the first report of a case diagnosed with liver IPT in which AFP and L3 fraction increased before surgery and decreased to the normal range after resection. This confirms the rarity of hepatic IPT associated with PBC and elevated AFP and L3 fraction.

Early gastric cancer with diffuse heterotopic gastric glands and granular cell tumors mimicking advanced gastric cancer.

INTRODUCTION: Heterotopic gastric grands (HGGs) are gastric grands that are observed in the submucosa and are considered to be paracancerous lesions or precursors of gastric cancer (GC). Granular cell tumors (GCTs) are benign neural origin tumors. Gastrointestinal GCTs are rare and gastric GCTs are seldom seen. We report the case of a patient who was diagnosed with early GC with diffuse HGGs affecting the whole stomach and two GCTs mimicking advanced GC. PRESENTATION OF CASE: The patient is a 71-year-old male with epigastric discomfort. Gastrointestinal endoscopy revealed an ulcerated lesion at the mid-gastric body. A biopsy specimen indicated adenocarcinoma. Moreover, gastrointestinal endoscopy revealed a submucosal tumor at the posterior wall and multiple transparent protuberances across the entire stomach. Computed tomography demonstrated diffuse gastric wall thickening with lymphadenopathies. Total gastrectomy was performed under the preoperative diagnosis of advanced GC with lymph node metastases. The pathological diagnosis was adenocarcinoma invading submucosal stroma without lymph node metastasis, two GCTs, and diffuse HGGs affecting whole stomach. DISCUSSION: Preoperative diagnosis of GC depth or range associated with HGGs is often difficult. Although diffuse HGGs are sometimes observed, there is no previous report of a case of HGGs with whole gastric wall thickening observed by computed tomography. As a result, this case was overdiagnosed as advanced GC. Although the relationship between GCTs and HGGs or GC is unclear, there is no case report of GCTs accompanied by HGGs or GC. CONCLUSION: This case report suggested that cautious preoperative assessment for GC co-occurring with HGGs is required.

Long-Term Outcomes After Pancreatectomy for Pancreatic Ductal Adenocarcinoma in Elderly Patients: Special Reference to Postoperative Adjuvant Chemotherapy.

BACKGROUND: The benefit of pancreatectomy for elderly patients with pancreatic ductal adenocarcinoma (PDAC) remains controversial. Moreover, adjuvant chemotherapy (AC) for elderly patients has not been fully evaluated. We investigated the long-term outcomes after pancreatectomy for PDAC in elderly patients with special reference to AC. METHODS: The medical records of 123 patients who underwent pancreatectomy for PDAC from 2007 to 2016 were retrospectively reviewed. The patients were divided into two groups: young (<75 years) and elderly patients (≥75 years). RESULTS: The study population comprised 91 young and 32 elderly patients. The postoperative clinical courses were not different between the two groups. AC was more frequently administered to young (85%) than elderly patients (66%; P = 0.04). The weekly dose of tegafur/gimeracil/oteracil potassium (S1) for AC was significantly lower in elderly (median 423 mg/m2) than young patients (median 491 mg/m2; P = 0.02). The prevalence of adverse events and the completion rate of AC were not significantly different between the two groups. There were no significant differences in recurrence-free survival (P = 0.73) or overall survival (P = 0.68) between the two groups in univariate analysis. Receipt of AC was not a significant independent factor for survival, and completion of planned AC was a significant independent factor for recurrence-free survival and overall survival in multivariate analysis. CONCLUSIONS: The benefit of pancreatectomy for PDAC was the same between young and elderly patients. Completion of planned AC was important, and lowered-dose AC using S1 for elderly patients might be safe and therapeutically useful.