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BACKGROUND Mesothelin is a cell surface glycoprotein overexpressed in 28%-58% of colorectal cancer (CRC). We hypothesized that CRC mesothelin expression contributes to peritoneal spread and that it is selectively overexpressed in those with peritoneal metastasis versus distant metastasis. METHODS This case-controlled study involved mesothelin immunohistochemistry staining of tumor specimens from patients with metastatic CRC/appendiceal cancers between 2017 and 2019. Staining reactivity was graded from trace to 4+ (low ≤1+; high >1+). Staining patterns were characterized on global (focal/patchy/diffuse) and cellular (apical/cytoplasmic) levels. Immunostaining of normal mesothelial cells served as internal control. RESULTS Thirty-one patients were identified: 11 peritoneal (study) and 20 distant metastasis (control). The control group did not include appendiceal cancers. The study group had greater proportion of high staining reactivity (55% vs. 5%; odds ratio [OR] = 20.4, 95% confidence interval [CI] 1.96-211.8). The study group had more diffuse (36% vs. 0%; OR = 22.2, 95% CI 1.1-465.3) and cytoplasmic staining patterns (73% vs. 28%; OR = 6.9, 95% CI 1.3-37.2). CONCLUSION Mesothelin expression is higher in CRC/appendiceal cancers with peritoneal metastasis than those with distant metastasis. Immunohistochemistry staining patterns suggestive of propensity towards peritoneal metastasis include diffuse and cytoplasmic staining. Mesothelin may be a potential target for novel treatments of CRC/appendiceal carcinoma with peritoneal involvement.

Intra-tumoral heterogeneity (ITH) could represent clonal evolution where subclones with greater fitness confer more malignant phenotypes and invasion constitutes an evolutionary bottleneck. Alternatively, ITH could represent branching evolution with invasion of multiple subclones. The two models respectively predict a hierarchy of subclones arranged by phenotype, or multiple subclones with shared phenotypes. We delineate these modes of invasion by merging ancestral, topographic, and phenotypic information from 12 human colorectal tumors (11 carcinomas, 1 adenoma) obtained through saturation microdissection of 325 small tumor regions. The majority of subclones (29/46, 60%) share superficial and invasive phenotypes. Of 11 carcinomas, 9 show evidence of multiclonal invasion, and invasive and metastatic subclones arise early along the ancestral trees. Early multiclonal invasion in the majority of these tumors indicates the expansion of co-evolving subclones with similar malignant potential in absence of late bottlenecks and suggests that barriers to invasion are minimal during colorectal cancer growth., Invasion is a critical step in tumor development. Here, in colorectal cancer, the authors show that multiclonal invasion of the muscularis mucosae is pervasive, suggesting that invasive capacity is not a significant bottleneck in the evolution of the disease.

Checkpoint inhibitors have become a widely used and available immunotherapy option for treating a variety of malignancies, including hematological malignancies. Patients receiving these therapies may go on to receive a curative allogeneic hematopoietic stem cell transplant (allo-HSCT). This presents a clinical challenge as the safety and efficacy of HSCT is not well reported in this subset of patients and residual programmed death-ligand 1 inhibition could potentially enhance allogeneic T-cell responses, improving the graft-versus-tumor effect, but also increasing the incidence and severity of immune complications such as graft-versus-host disease (GVHD). Here, this report includes a detailed literature review summarizing all available data on HSCT outcomes in the setting of using checkpoint inhibitor therapy pre-transplant. Moreover, we report a case of acute GVHD after allo-HSCT in a patient with high-risk myelodysplastic syndrome who received prior atezolizumab therapy, highlighting the importance of further research into this specific population in order to improve transplant-related outcomes.

Inflammation of the appendix is one of the most common conditions requiring emergent surgical intervention. Computed tomography commonly demonstrates a dilated appendix with adjacent inflammation. Traditionally, luminal obstruction of the appendix has been thought to be the primary etiology of appendicitis. However, current evidence suggests that etiology of appendicitis is multifactorial and can involve a number of different pathogenic pathways. Here we present a case of acute eosinophilic appendicitis with radiologic-pathologic correlation from a hypersensitivity reaction pathway. Acute eosinophilic appendicitis may represent an early precursor to conventional acute suppurative (phlegmonous) appendicitis, or a variant form of acute appendicitis.

Intramucosal esophageal adenocarcinoma can be reliably treated endoscopically. Controversy exists about the use of endotherapy versus esophagectomy for submucosal tumors. Increasingly endotherapy is considered for submucosal tumors in part because of the presumed high mortality with esophagectomy and the perceived poor prognosis in patients with nodal disease. This study was designed to assess survival following primary en bloc esophagectomy (EBE) in patients with submucosal esophageal adenocarcinoma (EAC). This is a retrospective review of all patients who underwent EBE for submucosal EAC between 1998 and 2015. No patient had neoadjuvant therapy. There were 32 patients (28M/4F; median age 64 years). The median tumor size was 1.5 cm (0.4–8.0), and the median number of resected nodes was 48 (23–85). There was one perioperative death. Lymph node metastases were present in 7 patients (22%). There was one involved node in four patients and 2, 3, and 31 nodes in one patient each. The one N3 patient received adjuvant therapy. The median follow-up was 87 months. Overall survival at 5 and 10 years was 84 and 70% respectively. Disease-specific survival at 10 years was 90%. Eight patients died, but only three deaths (9%) were related to EAC. Disease-specific survival at 10 years in node-positive patients was 71%. Survival after primary en bloc esophagectomy for submucosal adenocarcinoma was excellent even in node-positive patients. Mortality with esophagectomy was low and far less than the 22% risk of node metastases in patients with submucosal tumor invasion. Esophagectomy should remain the preferred treatment for T1b esophageal adenocarcinoma.

Summary Cancer is one of the major non-communicable diseases posing a threat to world health. Unfortunately, improvements in socioeconomic conditions are usually associated with increased cancer incidence. In this Commission, we focus on China, India, and Russia, which share rapidly rising cancer incidence and have cancer mortality rates that are nearly twice as high as in the UK or the USA, vast geographies, growing economies, ageing populations, increasingly westernised lifestyles, relatively disenfranchised subpopulations, serious contamination of the environment, and uncontrolled cancer-causing communicable infections. We describe the overall state of health and cancer control in each country and additional specific issues for consideration: for China, access to care, contamination of the environment, and cancer fatalism and traditional medicine; for India, affordability of care, provision of adequate health personnel, and sociocultural barriers to cancer control; and for Russia, monitoring of the burden of cancer, societal attitudes towards cancer prevention, effects of inequitable treatment and access to medicine, and a need for improved international engagement.

Background Gastric adenocarcinoma is a rare diagnosis in childhood. A 14-year old male patient presented with metastatic gastric adenocarcinoma, and a strong family history of colon cancer. Clinical sequencing of CDH1 and APC were negative. Whole exome sequencing was therefore applied to capture the majority of protein-coding regions for the identification of single-nucleotide variants, small insertion/deletions, and copy number abnormalities in the patient’s germline as well as primary tumor.

This is a case report of the anesthetic management for the hepatic resection of a metastatic paraganglioma in a patient with a history of prior orthotopic liver transplantation. Of interest, the metastatic paraganglioma originated from the donor organ. The patient is an 80-year-old woman with multiple medical problems including a history of cryptogenic cirrhosis who underwent successful orthotopic liver transplantation 9 years prior. She later presented with signs and symptoms of catecholamine excess suggestive of a catecholamine-producing tumor (paraganglioma or pheochromocytoma). Elevated urine catecholamine levels and radiographic evidence of a paraganglioma in the transplanted liver metastatic from the donor organ confirmed the diagnosis. Radiofrequency ablation of the tumor and surgical resection was previously attempted without success. We describe the anesthetic management for the successful resection of the metastatic hepatic paraganglioma, which was complicated by profound intraoperative hypertension and hypotension that necessitated the use of multiple vasoactive infusions, extensive surgical blood loss requiring blood transfusion, and difficult glycemic control in an insulin-dependent diabetic patient. The postoperative course is also described. This unique case presented the anesthesia team with challenges specific to both surgery for hepatic resection as well as for catecholamine-secreting tumors. We are not aware of any reports of paragangliomas of either donor or recipient origin involving a transplanted liver, making this the first such report to the best of our knowledge.

We have previously reported a topological model of the electrogenic Na+–HCO3− cotransporter (NBC1) in which the cotransporter spans the plasma membrane 10 times with N- and C-termini localized intracellularly. An analysis of conserved amino acid residues among members of the SLC4 superfamily in both the transmembrane segments (TMs) and intracellular/extracellular loops (ILs/ELs) provided the basis for the mutagenesis approach taken in the present study to determine amino acids involved in NBC1-mediated ion transport. Using large-scale mutagenesis, acidic and basic amino acids putatively involved in ion transport mediated by the predominant variant of NBC1 expressed in the kidney (kNBC1) were mutated to neutral and/or oppositely charged amino acids. All mutant kNBC1 cotransporters were expressed in HEK-293T cells and the Na+-dependent base flux of the mutants was determined using intracellular pH measurements with 2′,7′-bis-(carboxyethyl)-5(6)-carboxyfluorescein (BCECF). Critical glutamate, aspartate, lysine, arginine and histidine residues in ILs/ELs and TMs were detected that were essential for kNBC1-mediated Na+-dependent base transport. In addition, critical phenylalanine, serine, tyrosine, threonine and alanine residues in TMs and ILs/ELs were detected. Furthermore, several amino acid residues in ILs/ELs and TMs were shown to be essential for membrane targeting. The data demonstrate asymmetry of distribution of kNBC1 charged amino acids involved in ion recognition in putative outward-facing and inward-facing conformations. A model summarizing key amino acid residues involved in kNBC1-mediated ion transport is presented.

We have recently shown that carbonic anhydrase II (CAII) binds in vitro to the C-terminus of the electrogenic sodium bicarbonate cotransporter kNBC1 (kNBC1-ct). In the present study we determined the molecular mechanisms for the interaction between the two proteins and whether kNBC1 and CAII form a transport metabolon in vivo wherein bicarbonate is transferred from CAII directly to the cotransporter. Various residues in the C-terminus of kNBC1 were mutated and the effect of these mutations on both the magnitude of CAII binding and the function of kNBC1 expressed in mPCT cells was determined. Two clusters of acidic amino acids, L958DDV and D986NDD in the wild-type kNBC1-ct involved in CAII binding were identified. In both acidic clusters, the first aspartate residue played a more important role in CAII binding than others. A significant correlation between the magnitude of CAII binding and kNBC1-mediated flux was shown. The results indicated that CAII activity enhances flux through the cotransporter when the enzyme is bound to kNBC1. These data are the first direct evidence that a complex of an electrogenic sodium bicarbonate cotransporter with CAII functions as a transport metabolon.

The transmembrane topography of the human pancreatic electrogenic sodium bicarbonate cotransporter pNBC1 was investigated using in vitro transcription/translation of HK-M0 and HK-M1 fusion vectors designed to test membrane insertion properties of pNBC1 hydrophobic sequences (H). These vectors encode N-terminal 101 (HK-M0) or 139 (HK-M1) amino acids of the H,K-ATPase alpha-subunit, a linker region and the C-terminal 177 amino acids of the H,K-ATPase beta-subunit that contain five N-linked glycosylation consensus sites (Bamberg, K., and Sachs, G. (1994) J. Biol. Chem. 269, 16909-16919). The glycosylation status of the beta-region was used as a reporter to determine whether a given hydrophobic sequence possesses signal anchor and/or stop transfer properties in the HK-M0 and HK-M1 vectors. The linker region of each vector was replaced either with individual hydrophobic sequences or combinations thereof. The transcription/translation products of these fusion vectors in reticulocyte lysate system +/- microsomal membranes were identified by [(35)S]-autoradiography following separation using SDS-PAGE. The results of the in vitro transcription/translation analysis indicated that 10 (H1, H2N, H3, H5, H6, H7, H8, H9, H11, and H12) out of 12 hydrophobic sequences were able to insert into the plasma membrane. Two hydrophobic sequences, H4 and H10, had no membrane insertion activity even when upstream and downstream sequences were present. These data and immunocytochemical studies indicate that pNBC1 contains 10 transmembrane domains with N- and C-termini oriented intracellularly. This is the first characterization of the membrane topography of a sodium bicarbonate cotransporter.

MicroRNA-21 (miR-21) is upregulated and inversely associated with survival in many cancer types, including pancreatic ductal adenocarcinoma (PDAC). We studied the predictive value of miR-21 levels for gemcitabine or 5-fluorouracil (5-FU) response in tumor cells (TCs) or cancer associated fibroblasts (CAFs) in a cohort of PDAC patients from the RTOG 9704 trial.MiR-21 expression in CAFs and TCs, determined by in situ hybridization, of the 229 PDAC subset from RTOG 9704 was correlated with (i) histopathology characteristics using a chi-square test; and (ii) patient overall survival (OS) using the Cox proportional hazards model.MiR-21 was strongly expressed in TCs and CAFs in 137/182 (75%) and 152/181 (84%) PDACs, respectively. MiR-21 expression in CAFs for the group given 5-FU for OS: (i) approached significance in a univariate analysis (hazard ratio [HR], 1.57; 95% confidence interval [CI], 0.95-2.57; P = 0.07); and (ii) was significant in the multivariate model (HR, 1.70; 95% CI, 1.03-2.82; P = 0.038).MiR-21 expression in CAFs was associated with decreased OS in PDAC patients who received 5-FU, but not gemcitabine. These findings begin to identify stromal miR-21 as a marker to guide chemotherapy choice in PDAC patients.

The optimal surgical management of small nonfunctional pancreatic neuroendocrine tumors (NF-PNETs) remains controversial. We sought to identify (1) clinicopathologic factors associated with survival in NF-PNETs and (2) preoperative tumor characteristics that can be used to determine which lesions require resection and lymph node (LN) harvest. The records of all 116 patients who underwent resection for NF-PNETs between 1989 and 2012 were reviewed retrospectively. Preoperative factors, operative data, pathology, surgical morbidity, and survival were analyzed. The overall 5- and 10-year survival rates were 83.9 and 72.8 %, respectively. Negative LNs (p = 0.005), G1 or G2 histology (p = 0.033), and age

Previous studies provided functional evidence for electrogenic Na+-HCO3−cotransport in hepatocytes and in intrahepatic bile duct cholangiocytes. The molecular identity of the transporters mediating electrogenic sodium-bicarbonate cotransport in the liver is currently unknown. Of the known electrogenic Na+-HCO3−cotransporters (NBC1 and NBC4), we previously showed that NBC4 mRNA is highly expressed in the liver. In the present study, we performed RT-PCR, immunoblotting, and immunohistochemistry to characterize the expression pattern of NBC4 in rat liver and kidney. For immunodetection, a polyclonal antibody against rat NBC4 was generated and affinity purified. Of the known human NBC4 variants, only the rat NBC4c ortholog was detected by RT-PCR in rat liver, and the molecular mass of the NBC4c protein was ∼145 kDa. NBC4c protein was expressed in hepatocytes and in the cholangiocytes lining the intrahepatic bile ducts. In hepatocytes, NBC4c was localized to the basolateral plasma membrane, whereas intrahepatic cholangiocytes stained apically. The NBC1 electrogenic sodium cotransporter variants kNBC1 and pNBC1 were not detected by immunoblotting and immunohistochemistry in rat liver. The pattern of localization of NBC4c in the liver suggests that the cotransporter plays a role in mediating Na+-HCO3−cotransport in hepatocytes and intrahepatic cholangiocytes. Unlike the liver, the rat kidney expressed electrogenic sodium-bicarbonate cotransporter proteins kNBC1 and NBC4c. In kidney, NBC4c also had a molecular mass of ∼145 kDa and was immunolocalized to uroepithelial cells lining the renal pelvis, where the cotransporter may play an important role in protecting the renal parenchyma from alterations in urine pH.

The electrogenic Na+-HCO3 − cotransporters play an essential role in regulating intracellular pH and extracellular acid-base homeostasis. Of the known members of the bicarbonate transporter superfamily (BTS), NBC1 and NBC4 proteins have been shown to be electrogenic. The electrogenic nature of these transporters results from the unequal coupling of anionic and cationic fluxes during each transport cycle. This unique property distinguishes NBC1 and NBC4 proteins from other sodium bicarbonate cotransporters and members of the bicarbonate transporter superfamily that are known to be electroneutral. Structure-function studies have played an essential role in revealing the basis for the modulation of the coupling ratio of NBC1 proteins. In addition, the recent transmembrane topographic analysis of pNBC1 has shed light on the potential structural determinants that are responsible for ion permeation through the cotransporter. The experimentally difficult problem of determining the nature of anionic species being transported by these proteins (HCO3 − versus CO3 2−) is analyzed using a theoretical equilibrium thermodynamics approach. Finally, our current understanding of the molecular mechanisms responsible for the regulation of ion coupling and flux through electrogenic sodium bicarbonate cotransporters is reviewed in detail.

The electroneutral sodium bicarbonate cotransporter 3 (NBC3) coimmunoprecipitates from renal lysates with the vacuolar H+-ATPase. In renal type A and B intercalated cells, NBC3 colocalizes with the vacuolar H+-ATPase. The involvement of the COOH termini of NBC3 and the 56-kDa subunit of the proton pump in the interaction of these proteins was investigated. The intact and modified COOH termini of NBC3 and the 56-kDa subunit of the proton pump were synthesized, coupled to Sepharose beads, and used to pull down kidney membrane proteins. Both the 56- and the 70-kDa subunits of the proton pump, as well as a PDZ domain containing protein Na+/H+exchanger regulatory factor 1 (NHERF-1), were bound to the intact 18 amino acid NBC3 COOH terminus. A peptide truncated by five COOH-terminal amino acids did not bind these proteins. Replacement of the COOH-terminal leucine with glycine blocked binding of both the proton pump subunits but did not affect binding of NHERF-1. The 18 amino acid COOH terminus of the 56-kDa subunit of the proton pump bound NHERF-1 and NBC3, but the truncated and modified peptide did not. A complex of NBC3, the 56-kDa subunit of the proton pump, and NHERF-1 was identified in rat kidney. The data indicate that the COOH termini of NBC3 and the 56-kDa subunit of the vacuolar proton pump are PDZ-interacting motifs that are necessary for the interaction of these proteins. NHERF-1 is involved in the interaction of NBC3 and the vacuolar proton pump.

Incremental Value of EUS-FNA Compared With CT/MRI in the Management of Pancreatic Neuroendocrine Tumors (pNETs): Defining the Pivotal Role of EUS-FNA Justin A. Reynolds*, Alireza Sedarat, Irma Oliva, Sergei Tatishchev, Joseph R. Pisegna, James J. Farrell Division of Digestive Diseases, Department of Medicine, David Geffen School of Medicine at UCLA, Los Angeles, CA; Department of Pathology, David Geffen School of Medicine at UCLA, Los Angeles, CA; Division of Gastroenterology and Hepatology, VA Greater Los Angeles Healthcare System, Los Angeles, CA Background: Pancreatic neuroendocrine tumors (PNETs) are rare but are increasingly being identified in patients due to greater awareness and improved imaging modalities. Endoscopic ultrasound (EUS) with fine needle aspiration (FNA) allows for highly sensitive and specific localization and diagnosis of PNETs. However, the role of EUS-FNA in the pre-operative staging of PNETs is not clearly defined. Accordingly, the current study was undertaken to investigate the impact of EUS-FNA on the surgical management of patients with newly diagnosed PNETs. Methods: We reviewed a clinical-pathology database of all pancreatic specimens at the UCLA Medical Center from January 1999 through October 2011 that included neuroendocrine tumor, neoplasm, or carcinoma, including all patients who underwent EUS-guided FNA. Patient demographics, clinical presentation, indication for EUS-FNA, and possible impact on surgical management were assessed. Results: 80 patients (mean age 57 16 years, 55% male) were identified with PNETs by EUS-FNA. Mean tumor size by EUS was 20.8mm 16.3mm (range 4-70mm). Tumor locations in the pancreas included the head/neck (43), body/tail (35), or multifocal (2). EUS-FNA diagnosing PNET impacted on the surgical management of 40 patients by: (a) finding resectable PNET incidentally despite negative CT or MRI imaging (19); (b) confirming the diagnosis of PNET in patients previously suspected as having lymphoma or metastases (8); (c) confirming PNET in the setting of a pancreatic cyst (2); (d) precluding surgery for PNET by finding vascular invasion or metastases not seen on MRI or CT (3); and (e) helping medical decision-making favoring observation rather than surgery because of patient comorbidities and the likelihood of slower tumor progression (8). Conclusions: EUS-FNA affects the surgical management of PNETs by identifying resectable tumors not seen by CT or MRI, identifying PNETs in patients previously suspected as having lymphoma or metastases, and by identifying metastatic disease not seen by CT or MRI. The finding of PNET on EUS-FNA also leads some patients to observation rather than surgery because of comorbidities and the expected slow progression of disease, while offering the potential for use of newer chemotherapeutic agents (e.g. sunitinib, everolimus) in patients with unresectable disease. Further efforts to expand on the role of EUS in the management and surveillance of PNETs should be pursued.