Simultaneous Loss of Both Atypical Protein Kinase C Genes in the Intestinal Epithelium Drives Serrated Intestinal Cancer by Impairing Immunosurveillance.

Serrated adenocarcinoma, an alternative pathway for colorectal cancer (CRC) development, accounts for 15%-30% of all CRCs and is aggressive and treatment resistant. We show that the expression of atypical protein kinase C ζ (PKCζ) and PKCλ/ι was reduced in human serrated tumors. Simultaneous inactivation of the encoding genes in the mouse intestinal epithelium resulted in spontaneous serrated tumorigenesis that progressed to advanced cancer with a strongly reactive and immunosuppressive stroma. Whereas epithelial PKCλ/ι deficiency led to immunogenic cell death and the infiltration of CD8+ T cells, which repressed tumor initiation, PKCζ loss impaired interferon and CD8+ T cell responses, which resulted in tumorigenesis. Combined treatment with a TGF-ß receptor inhibitor plus anti-PD-L1 checkpoint blockade showed synergistic curative activity. Analysis of human samples supported the relevance of these kinases in the immunosurveillance defects of human serrated CRC. These findings provide insight into avenues for the detection and treatment of this poor-prognosis subtype of CRC.

Adipocyte p62/SQSTM1 Suppresses Tumorigenesis through Opposite Regulations of Metabolism in Adipose Tissue and Tumor.

Obesity is a leading risk factor for cancer. However, understanding the crosstalk between adipocytes and tumor cells in vivo, independently of dietary contributions, is a major gap in the field. Here we used a prostate cancer (PCa) mouse model in which the signaling adaptor p62/Sqstm1 is selectively inactivated in adipocytes. p62 loss in adipocytes results in increased osteopontin secretion, which mediates tumor fatty acid oxidation and invasion, leading to aggressive metastatic PCa in vivo. Furthermore, p62 deficiency triggers in adipocytes a general shutdown of energy-utilizing pathways through mTORC1 inhibition, which supports nutrient availability for cancer cells. This reveals a central role of adipocyte's p62 in the symbiotic adipose tissue-tumor collaboration that enables cancer metabolic fitness.

ATF4-Induced Metabolic Reprograming Is a Synthetic Vulnerability of the p62-Deficient Tumor Stroma.

Tumors undergo nutrient stress and need to reprogram their metabolism to survive. The stroma may play a critical role in this process by providing nutrients to support the epithelial compartment of the tumor. Here we show that p62 deficiency in stromal fibroblasts promotes resistance to glutamine deprivation by the direct control of ATF4 stability through its p62-mediated polyubiquitination. ATF4 upregulation by p62 deficiency in the stroma activates glucose carbon flux through a pyruvate carboxylase-asparagine synthase cascade that results in asparagine generation as a source of nitrogen for stroma and tumor epithelial proliferation. Thus, p62 directly targets nuclear transcription factors to control metabolic reprogramming in the microenvironment and repress tumorigenesis, and identifies ATF4 as a synthetic vulnerability in p62-deficient tumor stroma.

p62/SQSTM1 by Binding to Vitamin D Receptor Inhibits Hepatic Stellate Cell Activity, Fibrosis, and Liver Cancer.

Hepatic stellate cells (HSCs) play critical roles in liver fibrosis and hepatocellular carcinoma (HCC). Vitamin D receptor (VDR) activation in HSCs inhibits liver inflammation and fibrosis. We found that p62/SQSTM1, a protein upregulated in liver parenchymal cells but downregulated in HCC-associated HSCs, negatively controls HSC activation. Total body or HSC-specific p62 ablation potentiates HSCs and enhances inflammation, fibrosis, and HCC progression. p62 directly interacts with VDR and RXR promoting their heterodimerization, which is critical for VDR:RXR target gene recruitment. Loss of p62 in HSCs impairs the repression of fibrosis and inflammation by VDR agonists. This demonstrates that p62 is a negative regulator of liver inflammation and fibrosis through its ability to promote VDR signaling in HSCs, whose activation supports HCC.

Metabolic reprogramming of stromal fibroblasts through p62-mTORC1 signaling promotes inflammation and tumorigenesis.

The tumor microenvironment plays a critical role in cancer progression, but the precise mechanisms by which stromal cells influence the epithelium are poorly understood. Here we show that p62 levels were reduced in the stroma of several tumors and that its loss in the tumor microenvironment or stromal fibroblasts resulted in increased tumorigenesis of epithelial prostate cancer cells. The mechanism involves the regulation of cellular redox through an mTORC1/c-Myc pathway of stromal glucose and amino acid metabolism, resulting in increased stromal IL-6 production, which is required for tumor promotion in the epithelial compartment. Thus, p62 is an anti-inflammatory tumor suppressor that acts through the modulation of metabolism in the tumor stroma.

c-Myc phosphorylation by PKCζ represses prostate tumorigenesis.

Studies showing reduced PKCζ expression or enzymatic activity in different types of human cancers support the clinical relevance of PKCζ as a tumor suppressor. However, the in vivo role of PKCζ and its mechanisms of action in prostate cancer remain unclear. Here we demonstrate that the genetic inactivation of PKCζ in mice results in invasive prostate carcinoma in vivo in the context of phosphatase and tensin homolog deficiency. Bioinformatic analysis of human prostate cancer gene-expression sets revealed increased c-Myc transcriptional activity in PKCζ-inactive cells, which correlated with increased cell growth, invasion, and metastasis. Interestingly, PKCζ knockdown or the overexpression of a kinase-inactive mutant resulted in enhanced cell proliferation and invasion in vitro through increased c-Myc mRNA and protein levels and decreased Ser-373 phosphorylation of c-Myc. Analysis of prostate cancer samples demonstrated increased expression and decreased phosphorylation of c-Myc at Ser-373 in PKCζ knockout tumors. In vivo xenograft studies revealed that c-Myc phosphorylation by PKCζ is a critical event in the control of metastasis. Collectively, these results establish PKCζ as an important tumor suppressor and regulator of c-Myc function in prostate cancer.

Control of nutrient stress-induced metabolic reprogramming by PKCζ in tumorigenesis.

Tumor cells have high-energetic and anabolic needs and are known to adapt their metabolism to be able to survive and keep proliferating under conditions of nutrient stress. We show that PKCζ deficiency promotes the plasticity necessary for cancer cells to reprogram their metabolism to utilize glutamine through the serine biosynthetic pathway in the absence of glucose. PKCζ represses the expression of two key enzymes of the pathway, PHGDH and PSAT1, and phosphorylates PHGDH at key residues to inhibit its enzymatic activity. Interestingly, the loss of PKCζ in mice results in enhanced intestinal tumorigenesis and increased levels of these two metabolic enzymes, whereas patients with low levels of PKCζ have a poor prognosis. Furthermore, PKCζ and caspase-3 activities are correlated with PHGDH levels in human intestinal tumors. Taken together, this demonstrates that PKCζ is a critical metabolic tumor suppressor in mouse and human cancer.

Primary esophageal meningioma: first literature report.

We have described a primary esophageal meningioma (MG) clinical case diagnosed in a 62-year-old woman; also, we review the literature about extracranial MGs. To our knowledge, this is the first case report of an extracranial MG occurring primarily in the esophagus. These are benign neoplasms reported classically in the central nervous system (CNS). The extrancranial MGs have histopathologic and inmunohistochemical features identical to those observed in CNS MGs; thus, the main diagnostic hurdle is to keep it in the differential for lesions occurring outside the CNS.

Prediction of adenocarcinoma in esophagectomy specimens based upon analysis of preresection biopsies of Barrett esophagus with at least high-grade dysplasia: a comparison of 2 systems.

Distinguishing Barrett esophagus with high-grade dysplasia (BE-HGD) from intramucosal and submucosal adenocarcinomas on biopsies is challenging, yet important, in the choice of therapy. The current study evaluates preresection biopsies from patients who underwent esophagectomy for at least BE-HGD, to compare the recently published histologic categories by the University of Michigan (UM) and Cleveland Clinic (CC), correlate preresection and final resection diagnosis, and identify histologic features in biopsies that might be predictive of adenocarcinoma on esophagectomy. A total of 112 cases with a consensus biopsy diagnosis (agreement by ≥4 of 7 gastrointestinal pathologists) were statistically analyzed to identify histologic features that predicted adenocarcinoma on resection. Applying the UM criteria to the biopsy series showed excellent agreement with the CC system (κ=0.86) and significant correlation between preoperative and esophagectomy diagnoses (P<0.001). The likelihood of finding carcinoma on resection was significantly higher with the category of HGD with marked glandular distortion cannot exclude intramucosal adenocarcinoma [CC; odd ratio (OR), 2.8; P=0.046] or HGD suspicious for adenocarcinoma (UM; OR, 4.3; P=0.008), compared to HGD alone. The presence of "never-ending" glands (OR, 3.7; P=0.008), sheet-like growth (P<0.001), angulated glands (OR, 8.5; P<0.001), ≥3 dilated glands with intraluminal debris (OR, 2.6; P=0.05), and >1 focus of single-cell infiltration into the lamina propria (OR, 8.9; P<0.001) increased the odds of finding carcinoma on resection. The latter 2 variables remained independent predictors of adenocarcinoma in multivariable analysis. In conclusion, the CC and UM systems show excellent agreement and define histologic categories that can improve prediction of adenocarcinoma on resection.

Simultaneous inactivation of Par-4 and PTEN in vivo leads to synergistic NF-kappaB activation and invasive prostate carcinoma.

Prostate cancer is one of the most common neoplasias in men. The tumor suppressor Par-4 is an important negative regulator of the canonical NF-kappaB pathway and is highly expressed in prostate. Here we show that Par-4 expression is lost in a high percentage of human prostate carcinomas, and this occurs in association with phosphatase and tensin homolog deleted from chromosome 10 (PTEN) loss. Par-4 null mice, similar to PTEN-heterozygous mice, only develop benign prostate lesions, but, importantly, concomitant Par-4 ablation and PTEN-heterozygosity lead to invasive prostate carcinoma in mice. This strong tumorigenic cooperation is anticipated in the preneoplastic prostate epithelium by an additive increase in Akt activation and a synergistic stimulation of NF-kappaB. These results establish the cooperation between Par-4 and PTEN as relevant for the development of prostate cancer and implicate the NF-kappaB pathway as a critical event in prostate tumorigenesis.

Protein kinase Czeta represses the interleukin-6 promoter and impairs tumorigenesis in vivo.

Gene alterations in tumor cells that confer the ability to grow under nutrient- and mitogen-deficient conditions constitute a competitive advantage that leads to more-aggressive forms of cancer. The atypical protein kinase C (PKC) isoform, PKCzeta, has been shown to interact with the signaling adapter p62, which is important for Ras-induced lung carcinogenesis. Here we show that PKCzeta-deficient mice display increased Ras-induced lung carcinogenesis, suggesting a new role for this kinase as a tumor suppressor in vivo. We also show that Ras-transformed PKCzeta-deficient lungs and embryo fibroblasts produced more interleukin-6 (IL-6), which we demonstrate here plays an essential role in the ability of Ras-transformed cells to grow under nutrient-deprived conditions in vitro and in a mouse xenograft system in vivo. We also show that PKCzeta represses histone acetylation at the C/EBPbeta element in the IL-6 promoter. Therefore, PKCzeta, by controlling the production of IL-6, is a critical signaling molecule in tumorigenesis.

The impact of genotype on outcome in oligodendroglioma: validation of the loss of chromosome arm 1p as an important factor in clinical decision making.

OBJECT: Oligodendrogliomas are rare primary brain tumors. They comprise approximately 5 to 33% of all glial tumors but differ from astrocytomas by being associated with a more favorable prognosis, making their correct identification important. Allelic loss of chromosome arms 1p and 19q is found in a substantial subpopulation of tumors with an oligodendroglioma phenotype. Anaplastic oligodendrogliomas with allelic loss of 1p have been associated with chemosensitivity and a longer patient survival period. METHODS: Oligodendroglial neoplasms were studied using fluorescence in situ hybridization of formalin-fixed, paraffin-embedded tissue specimens; reference and target probe sets were used to map the telomeric regions of 1p and 19q. The results were correlated with the clinical characteristics of patients treated at our institution between 1993 and 2003. Data obtained in 96 patients were analyzed. This included 63 patients (65.6%) with World Health Organization (WHO) Grade II oligodendroglioma, 22 (23%) with Grade III oligodendroglioma, and 11 (11.4%) with mixed oligoastrocytoma. Analysis of 1p in patients with pure oligodendroglioma revealed a loss of 1p in 42 patients (49.4%). In 46 of these patients 19q was lost and in 70 (82.3%) there was concordance for combined loss or retention of both 1p and 19q (p < 0.0001). Patients with oligodendroglioma in whom a loss of 1p was present fared significantly better, and this outcome was unrelated to the treatment modality or WHO grade, compared with patients in whom 1p was intact (p < 0.05). CONCLUSIONS: To the authors' knowledge, this study includes the largest published series of WHO Grade II oligodendroglioma and 1p analysis. The results suggest that the association between long-term survival and 1p loss in oligodendroglioma is unrelated to treatment. The authors of further prospective studies may better determine the true value of the allelic loss of 1p and its implication for clinical decision making.

Correlation between viral loads of cytomegalovirus in blood and bronchoalveolar lavage specimens from lung transplant recipients determined by histology and immunohistochemistry.

Cytomegalovirus (CMV) is an important pathogen in lung transplant recipients. Early detection of CMV end-organ disease should help with treatment management. We determined the CMV viral load by hybrid capture in bronchoalveolar lavage (BAL) fluid samples from patients who had undergone lung transplantation. For 39 of these samples (from 25 patients), corresponding transbronchial biopsy samples were available for CMV immunohistochemistry (IHC). The CMV IHC results were interpreted and categorized as positive or negative, and the positive results were subcategorized as typical if cells with both significant nuclear enlargement or Cowdry A-type inclusions and positive staining were present or as atypical if definitive nuclear staining was seen but significant nuclear enlargement was not. Diagnostic CMV viral inclusions were reported in the anatomic diagnosis, based on hematoxylin-eosin staining alone, for three (8%) of the biopsy samples. CMV was detected by IHC in 13 (33%) samples (5 typical, 8 atypical). The median CMV viral load in BAL samples was 0 copies/ml for BAL samples from patients with IHC-negative biopsy samples; 47,678 copies/ml for BAL samples from patients with biopsy samples with positive, atypical staining; and 1,548,827 copies/ml for BAL samples from patients with biopsy samples with positive, typical staining (P < 0.001). Compared to routine pathology of biopsy samples, the use of IHC increased the diagnostic yield of CMV. Also, the CMV viral load in BAL fluid samples increased along with immunoreactivity from negative to positive, atypical staining to positive, typical staining. The CMV viral load determined with the end-organ sample, the BAL fluid sample, was higher than the corresponding viral load determined with blood. Both IHC and determination of the CMV viral load in BAL samples may be useful for the detection of individuals at risk for the development of fulminant invasive CMV disease.

Cyclooxygenase-2 in oligodendroglial neoplasms.

BACKGROUND: Although increased expression of cyclooxygenase-2 (COX-2) has been described in association with a variety of neoplasms, including tumors of astrocytic derivation, limited data are available on COX-2 expression in oligodendrogliomas. METHODS: The current study retrospectively reviewed 53 oligodendrogliomas and 7 oligodendroglioma-predominant oligoastrocytomas (mixed gliomas) for COX-2 expression and MIB-1 proliferative index (by immunohistochemistry) and for chromosome 1p status (by fluorescence in situ hybridization). RESULTS: Patients included 35 males and 25 females, with a mean age of 41 years (range, 12-73 years) at the time of surgery. Forty-four tumor specimens were classified as World Health Organization (WHO) Grade II neoplasms and 16 as WHO Grade III tumors. MIB-1 labeling indices (marker of cell proliferation) ranged from 0 to 22.3 (mean 4.5). Twenty-eight tumor specimens demonstrated allelic loss on chromosome 1p. Positive staining was observed in 17 tumor specimens with COX-2 antibody. COX-2-positive tumor specimens were also evaluated with CD68 (macrophage/microglial cell marker) by coimmunolabeling to confirm that the observed COX-2 immunostaining was not due to immunoreactive macrophages or microglial cells. COX-2 expression, lack of allelic loss at chromosome 1p, and high proliferation indices were associated with decreased survival (P = 0.002, P = 0.009, and P = 0.015, respectively). No correlation with outcome was found with patient gender, age at diagnosis, or histologic grade. CONCLUSIONS: Chromosome 1p, COX-2 immunoreactivity, and MIB-1 labeling indices correlated with outcome and were associated with decreased survival. There was not a one-to-one correspondence between COX-2 immunoreactivity and lack of allelic loss at chromosome 1p. Tumors with expression of COX-2 by immunohistochemistry may, in theory, benefit from treatment with therapeutic agents that inhibit COX-2.

Cavitary mass lesion and recurrent pneumothoraces due to Paragonimus kellicotti infection: North American paragonimiasis.

North American paragonimiasis is well described in omnivorous and carnivorous animals on this continent. Humans are rarely infected, largely because of dietary customs, but are at risk for infection if raw or undercooked crayfish are consumed. We describe a patient with a pleuropulmonary infection due to Paragonimus kellicotti that presented as recurrent pneumothoraces and a cavitary lesion. This is the first case of North American paragonimiasis in which the diagnosis was based on the morphology of the eggs present in histologic sections.

Initial evaluation of Cyberknife technology for extracorporeal renal tissue ablation.

OBJECTIVES: To determine whether Cyberknife technology can be applied to renal tissue safely and effectively. The goal was to achieve the high efficacy of a surgical treatment, with the low morbidity of a noninvasive intervention. METHODS: The Cyberknife is a frameless, image-guided radiosurgical device. This innovative extracorporeal treatment combines a linear accelerator mounted on a highly maneuverable robotic arm. The Cyberknife is unique in that it divides the high-dose radiation necessary to ablate the lesion completely into up to 1200 beams. Each one of these beams of radiation has a significantly reduced dose. Therefore, the individual dose of each beam is essentially benign to the pathway and surrounding tissue. However, at the focal point of these beams, the dose is additive, and the desired ablative dose is attained. Predetermined "lesions" in 16 kidneys were treated in vivo in the porcine model. Complete treatment was accomplished in one session per animal, with no complications. Gross and histologic evaluations were completed at 4, 6, or 8 weeks. RESULTS: The degree of radiation changes correlated with longer treatment intervals. After 8 weeks, the lesions showed complete fibrosis. The zones of complete fibrosis were characterized by dense, paucicellular connective tissue completely devoid of all normal kidney elements, including tubules and glomeruli. CONCLUSIONS: This initial preclinical evaluation of the Cyberknife for extracorporeal renal tissue ablation appears to be very promising and demonstrated its ability to ablate a targeted area precisely and completely with relative sparing of the surrounding tissue. This innovative technology introduces an exciting approach as a potential treatment option of renal masses in the future.

Interobserver variability in determining MIB-1 labeling indices in oligodendrogliomas.

Several studies have shown that MIB-1 labeling indices correlate well with tumor grade and prognosis in a variety of tumor types. Several factors are responsible for some degree of variability in the determination of labeling indices. Interobserver variability is one of the factors often cited as responsible for this variability. A slide from each of 30 oligodendrogliomas, stained with MIB-1 antibody, was distributed to six pathologists. The same set of slides was reviewed by each individual. Each pathologist was instructed to determine a MIB-1 labeling index by evaluating 1,000 tumor cell nuclei from the area of the slide with the most staining. The labeling index record reflected a percentage of positive-staining tumor cells. Interobserver agreement was compared. MIB-1 labeling indices ranged from 0 to 45.7. Overall agreement was good (> or =0.75) with a concordance coefficient of 0.832 (confidence interval, 0.700 to 0.909). Variability was greater among tumors with higher labeling indices as compared with tumors with labeling indices closer to 0. The overall agreement of MIB-1 labeling indices, while not perfect, was good. The generally minor variability among observers may be related to differences in the area of the slide evaluated and in differing lower thresholds for interpreting positivity. Further improvement of concordance may theoretically be attainable by further training and discussion among observers.

Immunohistochemical expression of cathepsin D in meningiomas.

We retrospectively studied the expression of cathepsin D by immunohistochemical analysis in 86 meningiomas (World Health Organization [WHO] grade I, n = 44; WHO grade II, n = 21; WHO grade III, n = 21) and correlated the results with tumor grade and outcome. Staining was scored semiquantitatively based on distribution among neoplastic cells as follows: 0, no staining; 1+, 5% or less of the cells; 2+, 6% to 20%; 3+, 21% to 50%; and 4+, more than 50% of the cells. Cathepsin D expression was observed as follows: 0, 10 cases (12%); 1+, 25 cases (29%); 2+, 15 cases (17%); 3+, 12 cases (14%); and 4+, 24 cases (28%). A higher degree of cathepsin D immunostaining was associated with low tumor grade (P = .0014), low mitotic count (P < .0001), low apoptotic count (P < .0001), and the development of recurrence (P = .035). There was no correlation with outcome or MIB-1 proliferation index. Cathepsin D expression by immunohistochemical analysis was identified in the majority (88% [76/86]) of meningiomas studied. A greater degree of immunoreactivity was observed in the WHO grade I group.

Prognostic importance of resection margin width after nephron-sparing surgery for renal cell carcinoma.

OBJECTIVES: To examine the relationship between the width of the resection margin and disease progression in renal cell carcinoma (RCC) after nephron-sparing surgery (NSS). During NSS for RCC, it is standard practice to excise the tumor along with a surrounding margin of normal parenchyma (margin of resection) to ensure complete resection of the neoplasm. However, no agreement has been reached on how wide the margin of resection should be. METHODS: We retrospectively reviewed the histopathologic sections and medical records of 69 patients with localized RCC who had undergone NSS between 1976 and 1988 to determine whether the resection margin, tumor size, TNM stage, and Fuhrman nuclear grade were associated with disease progression (defined as local tumor recurrence or metastasis). The mean postoperative follow-up interval was 8.5 years. RESULTS: No association was found between the width of the resection margin and disease progression (P = 0.98, log-rank test). Both TNM stage and Fuhrman nuclear grade correlated with disease progression. Patients with T1-T2 tumors had lower progression (P <0.001, log-rank test), and increased Fuhrman nuclear grade correlated with more disease progression (P <0.001, log-rank test). CONCLUSIONS: The width of the resection margin after NSS for RCC does not correlate with long-term disease progression. A histologic tumor-free margin of resection, irrespective of the width of the margin is sufficient to achieve complete local excision of RCC.

Chromosome 1p allelic loss by fluorescence in situ hybridization is not observed in dysembryoplastic neuroepithelial tumors.

Differentiation of dysembryoplastic neuroepithelial tumor (DNT) from cystic low-grade oligodendroglioma, particularly in a limited biopsy orfragmented specimen, may be impossible. Research has shown that allelic loss of chromosome 1p is a relatively common finding in oligodendrogliomas. Little is known about chromosome 1p status in DNT. We retrospectively evaluated 14 DNTs for loss of heterozygosity (LOH) on chromosome 1p by fluorescence in situ hybridization (FISH) and compared the results with 1p FISH analysis in 57 low-grade oligodendrogliomas (World Health Organization grade II). The 14 DNTs arose in 8 females and 6 males (mean age, 20.9 years at the time of surgery). All 14 DNTs were 1p intact by FISH analysis. The 57 low-grade oligodendrogliomas arose in 31 males and 26 females (mean age, 43.2 years). LOH on chromosome 1p was present in 31 (54%) of 57 tumors; the remaining 26 tumors were 1p intact. LOH on chromosome 1p is not a feature of DNTs. LOH on chromosome 1p may be a useful differential diagnostic feature (favoring oligodendroglioma) in a subset of cases in which specimen fragmentation or size raises the differential diagnosis of DNT vs oligodendroglioma.