Invasive stratified mucin-producing carcinoma of the colorectum: expanding the morphologic spectrum of large bowel cancer.

BACKGROUND: Invasive stratified mucin-producing carcinoma is a recently recognized adenocarcinoma with distinctive features. It was first described in the cervix but similar tumors have since been reported in the penis, anus and prostate. In the gastrointestinal tract, the phenomenon of epithelial stratification has an interesting embryologic morphogenesis. Gastrointestinal mucosa starts off as nascent columnar epithelium that is subsequently patterned to confer regional specific functions. However, in disease states, normal architectural patterning can be disrupted by aberrant differentiation. Given this background and the phenotypic plasticity of neoplastic cells, we were interested in ascertaining whether invasive stratified mucin-producing carcinoma occurs in the colorectum. METHODS: This was a retrospective study of all 584 cases of colorectal carcinoma accessioned at our institution over a 2-year period (January 2021- December 2022). Cases were analyzed to determine which fulfilled the criteria for invasive stratified mucin-producing carcinoma. RESULTS: There were 9 cases of colorectal invasive stratified mucin-producing carcinoma-one pure form and 8 mixed. They showed the classic colorectal (CK20 + , CDX2 + , CK7-) immunostaining profile but, based on various morphologic criteria, they could be distinguished from conventional adenocarcinoma NOS, mucinous, signet ring cell, medullary, goblet cell and undifferentiated carcinomas. About half the cases were MLH1/PMS2 deficient and BRAF &/or PIK3CA mutated, which aligns with the hypermutated phenotype. CONCLUSIONS: Colorectal invasive stratified mucin-producing carcinoma appears to be a real entity, best recognized in its early stages. It appears to be a high-grade carcinoma. With tumor progression, it evolves into a mucinous adenocarcinoma with a proclivity towards signet ring cells. In summary, the study of this tumor, particularly in its early stages, provides useful clues to further understanding the biology and progression of large bowel cancer. Further studies are required to learn more about this tumor.

Gallbladder Cancer: Imaging Appearance and Pitfalls in Diagnosis [Formula: see text].

Gallbladder cancer is an uncommon malignancy with an overall poor prognosis. The clinical and imaging presentation of gallbladder cancer often overlaps with benign disease, making diagnosis difficult. Gallbladder cancer is most easily diagnosed on imaging when it presents as a mass replacing the gallbladder. At this stage, the prognosis is usually poor. Recognizing the features of gallbladder cancer early in the disease can enable complete resection and improve prognosis. Recognition of the patterns of wall enhancement on computed tomography can help differentiate gallbladder cancer from benign disease. Gallbladder wall thickening without pericholecystic fluid presenting in an older patient with raised alkaline phosphatase should raise concern regarding gallbladder cancer. Gallbladder polyps in high-risk individuals need close surveillance or surgery as per guidelines. Small gallbladder cancers in the neck can present as biliary dilatation or cholecystitis, and careful examination of this area is needed to assess for lesion. The imaging appearance of gallbladder cancer is reviewed and supported by local institutional data. Features that differentiate it from its common mimics enabling earlier diagnosis are described.

Pancreatic Neuroendocrine Tumors Complicated by Sinistral Portal Hypertension: Insights into Pathogenesis.

Purpose: To investigate the association between pancreatic neuroendocrine tumors (panNETs) and sinistral portal hypertension (SPH) and provide insights into the pathogenesis. Methods: A retrospective review of panNETs was conducted from our institution for 12 years. Medical imaging findings were analyzed to determine any association with splenic vein thrombosis (SVT) at diagnosis. The cases were further selected based on the criteria for SPH, namely, (1) presence of SVT, (2) gastric varices, (3) patent portal vein, and (4) normal liver function tests. Results: There were 61 patients with panNETs and 8 (8/61) had SVT and gastric varices at diagnosis. Four (4/8) met the strict criteria for SPH while the other four had more conventional portal hypertension. The four with SPH had large tumors located in the tail with splenic vein invasion and three of four presented with bleeding gastric varices. All four patients underwent surgical resection. Mean follow-up was 8.5 years and the hematemesis never recurred. The other four patients (four of eight) with gastric varices had unresectable disease and all died after a mean survival of 29 months. Conclusion: PanNETs appear to be more commonly associated with SVT and SPH compared with other tumors. This could be related to their relatively indolent nature and their intrinsic vascularity. From a surgical viewpoint, the decision to operate depends on many factors including but not limited to the size/stage, grade, and functionality of the tumor and comorbidities. These considerations notwithstanding, the association between panNETs and SPH suggests that there is benefit in timely resection of panNETs located in the tail.

Complete pathological response following neoadjuvant FOLFIRINOX in borderline resectable pancreatic cancer - a case report and review.

BACKGROUND: Pancreatic cancer is among the top 5 most common cancers worldwide, but is particularly devastating due to its insidious nature. Complete surgical resection remains the only potential curative treatment, although only 20 % of patients present with a resectable tumor. Patients may alternatively present with borderline resectable pancreatic cancer or locally advanced pancreatic cancer and can be offered treatment with neoadjuvant intent. The effectiveness of these treatments is unclear and there is a paucity of data to suggest one optimal treatment approach. CASE PRESENTATION: We describe a 61-year-old female who presented with a two-week history of obstructive jaundice in the context of vague abdominal pain that had been ongoing for years prior to her visit. CT scan of the abdomen confirmed a hypovascular mass in the uncinate process consistent with borderline resectable pancreatic cancer. Pancreatic adenocarcinoma was confirmed with endoscopic ultrasound guided fine-needle aspiration cytology. Following multidisciplinary discussion, it was recommended that she undergo treatment with FOLFIRINOX. After a total of 13 cycles, follow up CT revealed that the lesion had decreased in size and she was offered resection as a potentially curative treatment. She underwent pancreaticoduodenectomy. Final pathology report revealed no evidence of residual adenocarcinoma (ypT0 ypN0 (0/23)). The patient remains disease-free 15 months following surgery. CONCLUSION: To date, there have been very few reports of a complete pathological response following neoadjuvant therapy in borderline resectable or locally advanced pancreatic cancer. This report describes a unique case of a complete pathological remission in a patient with borderline resectable pancreatic cancer following FOLFIRINOX therapy alone and adds to the growing base of evidence meriting the initiation of clinical trials to assess the efficacy of FOLFIRINOX in these subsets of pancreatic cancer.

A case series of two glomus tumors of the gastrointestinal tract.

Two recent cases of glomus tumors (GTs) of the gastrointestinal tract presented with symptoms of GI bleeding. GTs, typically benign lesions of mesenchymal origin, are rarely seen in the GI tract, and most commonly involve the distal appendages. This case series discusses the tumor biology, presentation, imaging, endoscopic findings, pathology and management of GTs. While diagnosis of GTs is typically made on final surgical pathology, there are defining characteristics that can separate a GT from a gastrointestinal stromal tumor on endoscopic ultrasound (EUS) and CT imaging. The classic pathological findings are discussed, and surgical decision-making is reviewed. New developments in the form of EUS-guided biopsy and endoscopic submucosal dissection present new avenues for diagnosis and treatment of submucosal lesions of the GI tract, including GTs. While typically a benign tumor requiring no adjuvant therapy, this study discusses some very rare cases of metastatic GT in the literature.

Role of fine-needle aspiration in the surgical management of pancreatic neuroendocrine tumors: utility and limitations in light of the new World Health Organization classification.

CONTEXT: Pancreatic neuroendocrine tumors (Panc-NETs) are rare and tend to get overshadowed by their more prevalent and aggressive ductal adenocarcinoma counterparts. The biological behavior of PancNETs is unpredictable, and thus management is controversial. However, the new World Health Organization classification has significantly contributed to the prognostic stratification of these patients. Concurrently, there have been advances in surgical techniques for benign or low-grade pancreatic tumors. These procedures include minimally invasive and parenchyma-sparing operations such as laparoscopy and enucleation. OBJECTIVE: To report on the utility and limitations of fine-needle aspiration in the preoperative evaluation and management of PancNETs. DESIGN: This was a retrospective review of our institutional tumor database from 2002 to 2012. There were 25 cases of PancNETs that were localized and staged by medical imaging and diagnosed by fine-needle aspiration. RESULTS: Fourteen patients underwent laparotomy, with some requiring only limited surgery; 4 had laparoscopic resections; 4 were serially observed without surgical intervention; and another 3 were inoperable. After a mean follow-up of 37 months, more than half of the patients had no evidence of disease, including most of those who underwent minimally invasive surgery. CONCLUSIONS: Fine-needle aspiration is a useful diagnostic adjunct to medical imaging in the preoperative evaluation and management of PancNETs. However, there are limitations with regard to grading PancNETs using this technique.

EGFR expression variance in paired colorectal cancer primary and metastatic tumors.

BACKGROUND: Previous studies indicate that drugs targeting the Epidermal Growth Factor Receptor (EGFR) signaling pathways can induce objective responses, prolong time to progression and improve survival of patients with metastatic colorectal cancer (mCRC). EGFR expression in the primary tumour may not predict response to these agents and data is conflicting regarding the correlation of EGFR expression in the primary tumour with the metastatic site. In other tumour sites, the presence of EGFR mutations was associated with efficacy in a subset of patients. OBJECTIVES: The goal of this study is to correlate tumour EGFR expression between primary and liver metastatic sites, and to assess the mutational status in the EGFR kinase domain. METHODS: This is a single center retrospective study of patients who underwent surgical resection of CRC, for whom paired paraffin-embedded tissue blocks of primary tumours and resected liver metastases were available. EGFR immunostaining and mutation analyses were preformed. RESULTS: Fifty six paired colorectal primaries and metastases were available for analysis. EGFR was detectable in 96.6% of the primary samples and in 89.7% of the metastatic samples. Perfect concordance in the intensity score between the primary and the metastases was found in 46.5% of the cases. While individual pairs were poorly concordant for intensity, the proportion of primaries with intense staining was similar to the proportion with intense staining in the metastatic samples. Overall survival did not correlate with either EGFR expression in the primary tumour, or with EGFR expression in the metastasis. There were 2 cases with mutations in the EGFR kinase domain. Both mutations were found in exon21 C>T. CONCLUSIONS: In this analysis, EGFR expression in the primary tumor site was not predictive of its level in the metastasis. EGFR expression levels in the primaries and in the metastases do not appear to be useful prognostic markers.

Defect of CD8+ memory T cells developed in absence of IL-12 priming for secondary expansion.

IL-12 priming plays an important role in stimulation of CD8+ effector T cells and development of CD8+ memory T (Tm) cells. However, the functional alteration of CD8+ Tm cells developed in the absence of IL-12 priming is elusive. In this study, we investigated the capacity of secondary expansion of CD8+ Tm cells developed from transgenic OT I CD8+ T cells. The latter cells were in vitro and in vivo stimulated by ovalbumin (OVA)-pulsed dendritic cells [DCOVA and (IL-12-/-)DCOVA] derived from wild-type C57BL/6 and IL-12 gene knockout mice, respectively. We demonstrated that IL-12 priming is important not only in CD8+ T cell clonal expansion, but also in generation of CD8+ Tm cells with the capacity of secondary expansion upon antigen re-encounter. However, IL-12 signaling is not involved in CD8+ Tm cell survival and recall responses. Therefore, this study provides useful information for vaccine design and development.

Review: cancer immunotherapy by exosome-based vaccines.

Exosomes (EXOs) are nanometer-sized membrane vesicles secreted from epithelial and hematopoietic cells. They display a spectrum of molecules involved in immune responses and signal transductions. Previous studies showed that tumor antigen-loaded dendritic cell (DC)- and tumor cell-derived EXOs (Dexo and Texo) induce tumor antigen-specific CD8(+) cytotoxic T-lymphocyte responses and antitumor immunity in experimental animal models and human clinical trials. This review will present the main biologic features of Dexo and Texo as cell-free cancer vaccines with emphasis on their immunostimulatory properties and their potential efficacy in cancer immunotherapy.

Type 1 CD8+ T cells are superior to type 2 CD8+ T cells in tumor immunotherapy due to their efficient cytotoxicity, prolonged survival and type 1 immune modulation.

CD8+ cytotoxic T (Tc) cells play a crucial role in host immune responses to cancer, and in this context, adoptive CD8+ Tc cell therapy has been studied in numerous animal tumor models. Its antitumor efficacy is, to a large extent, determined by the ability of Tc cells to survive and infiltrate tumors. In clinical trials, such in vitro-activated T cells often die within hours to days, and this greatly limits their therapeutic efficacy. CD8+ Tc cells fall into two subpopulations based upon their differential cytokine secretion. In this study, we in vitro generated that ovalbumin (OVA)-pulsed dendritic cell (DCOVA)-activated CD8+ type 1 Tc (Tc1) cells secreting IFN-gamma, and CD8+ type 2 Tc (Tc2) cells secreting IL-4, IL-5 and IL-10, which were derived from OVA-specific T cell receptor (TCR) transgenic OT I mice. We then systemically investigated the in vitro and in vivo effector function and survival of Tc1 and Tc2 cells, and then assessed their survival kinetics after adoptively transferred into C57BL/6 mice, respectively. We demonstrated that, when compared to CD8+ Tc2, Tc1 cells were significantly more effective in perforin-mediated cytotoxicity to tumor cells, had a significantly higher capacity for in vivo survival after the adoptive T cell transfer, and had a significantly stronger therapeutic effect on eradication of well-established tumors expressing OVA in animal models. In addition, CD8+ Tc1 and Tc2 cells skewed the phenotype of CD4+ T cells toward Th1 and Th2 type, respectively. Therefore, the information regarding the differential effector function, survival and immune modulation of CD8+ Tc1 and Tc2 cells may provide useful information when preparing in vitro DC-activated CD8+ T cells for adoptive T cell therapy of cancer.

Alpha tumor necrosis factor contributes to CD8(+) T cell survival in the transition phase.

Cytokine and costimulation signals determine CD8(+) T cell responses in proliferation phase. In this study, we assessed the potential effect of cytokines and costimulations to CD8(+) T cell survival in transition phase by transferring in vitro ovalbumin (OVA)-pulsed dendritic cell-activated CD8(+) T cells derived from OVA-specific T cell receptor transgenic OT I mice into wild-type C57BL/6 mice or mice with designated gene knockout. We found that deficiency of IL-10, IL-12, IFN-gamma, CD28, CD40, CD80, CD40L, and 41BBL in recipients did not affect CD8(+) T cell survival after adoptive transfer. In contrast, TNF-alpha deficiency in both recipients and donor CD8(+) effector T cells significantly reduced CD8(+) T cell survival. Therefore, our data demonstrate that the host- and T cell-derived TNF-alpha signaling contributes to CD8(+) effector T cell survival and their transition to memory T cells in the transition phase, and may be useful information when designing vaccination.

Bidirectional membrane molecule transfer between dendritic and T cells.

The acquisition of dendritic cell (DC) molecules by T cells has been previously reported. However, it remains unclear whether the transfer is only mono- or bidirectional. In this study, we incubated CMFDA-labeled ovalbumin (OVA)-pulsed DC2.4 (DC2.4(OVA)) cells with Dil-labeled OT II CD4(+) T cells and analyzed the potential bidirectional molecule transfer. We also assessed the distribution of internalized membrane using two engineered DC2.4/Ia(b)GFP and MF4/TCRCFP DC lines. Our findings showed that membrane molecule transfer is bidirectional. CD4(+) T cells acquired Ia(b), CD11c, CD40, and CD80 from DC2.4(OVA) cells, and conversely DC2.4(OVA) cells took up CD4, CD25, CD69, and T cell receptor from T cells. The internalized molecules acquired by T cells and DCs mostly localized in endosomes and lysosomes, respectively. Taken together, this study demonstrated a novel phenomenon of bidirectional membrane molecule transfer between DCs and T cells.

Interferon gamma stimulates cellular maturation of dendritic cell line DC2.4 leading to induction of efficient cytotoxic T cell responses and antitumor immunity.

Dendritic cells (DCs) are the most potent antigen-presenting cells (APCs) for the initiation of antigen (Ag)-specific immune responses. In most studies, mature DCs are generated from bone marrow cells or peripheral monocytes; in either case, the harvested cells are then cultured in medium containing recombinant GM-CSF, IL-4 and TNF-alpha for 7-10 days and stimulated with lipopolysaccharide (LPS). However, this approach is time-consuming and expensive. There is another less cost approach of using immobilized DC cell lines, which can easily grow in the medium. A disadvantage with the immobilized DC cell lines, however, is that they are immature DCs and lack expression of MHC class II and costimulatory CD40 and CD80 molecules. This, therefore, limits their capacity for inducing efficient antitumor immunity. In the current study, we investigated the possible efficacy of various stimuli (IL-1beta,IFN-gamma, TNF-alpha, CpG and LPS) in converting the immature dendritic cell line DC2.4 to mature DCs. Our findings were quite interesting since we demonstrated for the first time that IFN-gamma was able to stimulate the maturation of DC2.4 cells. The IFN-gamma-activated ovalbumin (OVA)-pulsed DC2.4 cells have capacity to upregulate MHC class II, CD40, CD80 and CCR7, and to more efficiently stimulate in vitro and in vivo OVA-specific CD8+ T cell responses and antitumor immunity. Therefore, IFN-gamma-activated immortal DC2.4 cells may prove to be useful in the study of DC biology and antitumor immunity.

Combinational adenovirus-mediated gene therapy and dendritic cell vaccine in combating well-established tumors.

Recent developments in tumor immunology and biotechnology have made cancer gene therapy and immunotherapy feasible. The current efforts for cancer gene therapy mainly focus on using immunogenes, chemogenes and tumor suppressor genes. Central to all these therapies is the development of efficient vectors for gene therapy. By far, adenovirus (AdV)-mediated gene therapy is one of the most promising approaches, as has confirmed by studies relating to animal tumor models and clinical trials. Dendritic cells (DCs) are highly efficient, specialized antigen-presenting cells, and DC-based tumor vaccines are regarded as having much potential in cancer immunotherapy. Vaccination with DCs pulsed with tumor peptides, lysates, or RNA, or loaded with apoptotic/necrotic tumor cells, or engineered to express certain cytokines or chemokines could induce significant antitumor cytotoxic T lymphocyte (CTL) responses and antitumor immunity. Although both AdV-mediated gene therapy and DC vaccine can both stimulate antitumor immune responses, their therapeutic efficiency has been limited to generation of prophylactic antitumor immunity against re-challenge with the parental tumor cells or to growth inhibition of small tumors. However, this approach has been unsuccessful in combating well-established tumors in animal models. Therefore, a major strategic goal of current cancer immunotherapy has become the development of novel therapeutic strategies that can combat well-established tumors, thus resembling real clinical practice since a good proportion of cancer patients generally present with significant disease. In this paper, we review the recent progress in AdV-mediated cancer gene therapy and DC-based cancer vaccines, and discuss combined immunotherapy including gene therapy and DC vaccines. We underscore the fact that combined therapy may have some advantages in combating well-established tumors vis-a-vis either modality administered as a monotherapy.

Conversion of tolerogenic CD4-8- dendritic cells to immunogenic ones inducing efficient antitumor immunity.

Culturing conditions may affect dendritic cell (DC) maturation status and functional effects. We have previously demonstrated that different DC subsets play distinct roles in immune responses. The splenic CD4-8- DC subset that secretes transforming growth factor (TGF)-beta stimulates CD4+ regulatory T type 1 (Tr1) cell responses, and this leads to antitumor immune tolerance. In this study, we investigated the potential effect of culturing conditions, namely: (1) duration of culturing and (2) the dose of antigen ovalbumin (OVA) for DC pulsing, respectively, in the conversion of tolerogenic CD4-8- DC into immunogenic DCs. Our data showed that isolated CD4-8- DCs cultured for an additional 18 hours in medium containing 15-20 ng/mL granulocyte macrophage colony-stimulating factor (GM-CSF) became more mature compared to the freshly isolated CD4-8- DCs. When pulsed with OVA at the relatively high concentration of 1 mg/mL, but not at 0.1 mg/mL, the CD4-8- DCs could be converted into immunogenic CD4-8- DCs, which stimulated CD4+ T-cell differentiation into type 1 helper T (Th1) cells. Vaccination of mice with converted CD4-8- DCs induced strong OVA-specific cytotoxic T-lymphocyte (CTL) responses and protective immunity against OVA-expressing BL6-10OVA B16 melanoma. Taken together, our findings indicate that the conversion of DCs from a tolerogenic to an immunogenic state can be achieved by the elongation of DC culturing time in combination with a high-dose antigen for DC pulsing. Therefore, our results may have a significant impact in designing DC-based antitumor vaccines.

Nuclear expression of thymidylate synthase in colorectal cancer cell lines and clinical samples.

Thymidylate synthase (TS) [TYMS; OMIM reference number (188,350)] is normally considered to be a cytoplasmic enzyme. However, a few reports have suggested it may also be present in the nucleus. To explore this in more detail, we used a highly specific polyclonal antibody to TS and a combination of techniques, including immunocytochemistry, confocal microscopy, cell fractionation, and Western blotting. We developed cell line HeLa-55, a HeLa derivative that grossly overexpresses TS. Although the vast majority of TS was in the cytoplasm, some TS also was seen in the nucleus. TS in parental HeLa cells and in normal human fibroblasts was seen exclusively in the cytoplasm. HeLa-55 cells exposed to 5-fluorodeoxyuridine were fractionated and examined by Western blotting. Interestingly, both free TS and the ternary complex of TS were seen in the cytoplasmic fraction but only free TS was detected in the nuclear fraction. Amongst different cell lines examined, HCT-15 and normal fibroblasts showed no nuclear TS, HCC-2998 and SW-620 showed a small amount of nuclear TS, and HT-29, RKO, and HCT-116 showed a strong nuclear TS signal. Nuclear staining was clearly evident in some clinical colorectal specimens, both normal and malignant. This staining was definitively shown to be TS by competition with recombinant TS protein. A putative leucine-rich nuclear export sequence was identified but its function could not be confirmed. We conclude that small amounts of TS protein is present in the nucleus of some cell types but further work is needed to determine the significance of this observation.

Molecular and immunophenotypical characterization of progressive and regressive leukemia cell lines.

The P815 and P198 cell lines are clonally related mouse mastocytoma cell lines. They differ in their biologic behavior in that P815 is a progressive tumor cell line, whereas P198 is a regressive one. These cell lines have been extensively used as models for the study of tumor-host relationships and tumor immunology. Although some of their biological properties have been well documented, the molecular mechanisms underlying tumor progression or regression have not been completely elucidated. In this study, we characterized the growth behavior and immunophenotype of these two cell lines, and analyzed their gene profiles using a complementary deoxynucleic acid (cDNA) microarray composed of 514 immunologically relevant genes. Our data showed that the two cell lines exhibited quite dissimilar and contrasting growth characteristics when inoculated into syngeneic mice. P815 tumors grew unremittingly, while P198 tumors gradually regressed. From a molecular viewpoint, P815 cells showed a higher expression of genes promoting tumor growth, such as IGF-1, IL-8R, FGFR1, VEGF-A, and VEGF-B. On the other hand, P198 tumor cells expressed CD11b and CD80, which favor the recruitment of lymphocytes and antigen-presenting cells (APCs), as well as the elicitation of antitumor immunity. P198 tumor cells also depicted a higher expression of genes inhibiting tumor growth, such as TNF-alpha, SOCS-1, CIS1, 4-1BB, and GDF-10. In conclusion, our results contribute further information in the understanding of the molecular mechanisms associated with the regression and progression of P815 and P198 tumor cells.

A novel approach to tumor suppression using microencapsulated engineered J558/TNF-alpha cells.

AIM: Immunoisolation technology using microencapsulated nonautologous cells is a novel alternative approach to the treatment of cancer. The work was aimed on investigation of the effect of implantation of microencapsulates on tumor growth in vivo. METHODS: In this study, we constructed an engineered tumor cell line J558/TNF-alpha that secreted functional tumor necrosis factor-alpha (TNF-alpha) (2 ng/ml), and went on to encapsulate the engineered cells into microencapsules. RESULTS: Our data showed that the microencapsulates thus produced could release functional TNF-alpha (1.2 ng/ml), which then diffused through the microencapsule membrane into the supernatant, and produced a cytotoxic effect on L929 cells in vitro. Microencapsulated cells were intratumorally (i.t.) implanted into athymic nude mice bearing the human breast cancer MCF-7. The results showed that the i.t. implantation induced extensive tumor cell apoptosis and necrosis leading to significant tumor regression and slower tumor growth than in the control groups that were i.t. injected with microencapsulated J558 or PBS respectively (p < 0.05). CONCLUSION: This study provides further evidence that the microencapsulation of recombinant tumor cells secreting cytokines may be an alternative approach in treatment of cancer.