Insulin-like Growth Factor II mRNA-binding Protein 3 is a Highly Sensitive Marker for Intravascular Large B-cell Lymphoma: Immunohistochemical Analysis of 152 Pathology Specimens From 88 Patients.

Intravascular large B-cell lymphoma (IVLBCL) is a rare type of aggressive extranodal large B-cell lymphoma characterized by the selective growth of lymphoma cells within the lumina of blood vessels, particularly capillaries. IVLBCL lacks mass formation, and its diagnosis can be challenging. We analyzed the utility of insulin-like growth factor II mRNA-binding protein 3 (IMP3) immunohistochemistry for IVLBCL diagnosis in various organs. Double staining with paired box 5 (PAX5) was performed for validation. Overall, 152 pathological specimens (111 positive and 41 negative for IVLBCL) obtained from 88 patients with a diagnosis of IVLBCL were stained for IMP3 and IMP3/PAX5. As negative controls, 40 pathology specimens from 38 patients with no history of IVLBCL or other B-cell lymphomas were stained for IMP3, which comprised 31 benign pathological specimens from 29 patients in whom malignancy was suspected, 7 cases of appendicitis with intravascular and/or intralymphatic lymphoid proliferations, and 2 cases of intravascular natural killer/T-cell lymphoma. All mononuclear cells with cytoplasmic staining were considered positive for IMP3 expression, but expression restricted to germinal center B cells was excluded from evaluation. All 111 IVLBCL pathological specimens were positive for IMP3 and IMP3/PAX5. In addition, 11 of the 41 specimens originally diagnosed as IVLBCL-negative showed IMP3/PAX5 double-positive cells, raising the suspicion of IVLBCL. However, of the 40 negative control samples, IMP3-positive non-germinal center B cells were detected in only 2 samples (P= 0.0131) and no intravascular IMP3-positive B cells suspicious for IVLBCL were identified. Altogether, IMP3 immunohistochemistry is a highly sensitive marker of IVLBCL and can be a helpful adjunct for IVLBCL diagnosis.

Evolutionary histories of breast cancer and related clones.

Recent studies have documented frequent evolution of clones carrying common cancer mutations in apparently normal tissues, which are implicated in cancer development1-3. However, our knowledge is still missing with regard to what additional driver events take place in what order, before one or more of these clones in normal tissues ultimately evolve to cancer. Here, using phylogenetic analyses of multiple microdissected samples from both cancer and non-cancer lesions, we show unique evolutionary histories of breast cancers harbouring der(1;16), a common driver alteration found in roughly 20% of breast cancers. The approximate timing of early evolutionary events was estimated from the mutation rate measured in normal epithelial cells. In der(1;16)(+) cancers, the derivative chromosome was acquired from early puberty to late adolescence, followed by the emergence of a common ancestor by the patient's early 30s, from which both cancer and non-cancer clones evolved. Replacing the pre-existing mammary epithelium in the following years, these clones occupied a large area within the premenopausal breast tissues by the time of cancer diagnosis. Evolution of multiple independent cancer founders from the non-cancer ancestors was common, contributing to intratumour heterogeneity. The number of driver events did not correlate with histology, suggesting the role of local microenvironments and/or epigenetic driver events. A similar evolutionary pattern was also observed in another case evolving from an AKT1-mutated founder. Taken together, our findings provide new insight into how breast cancer evolves.

[A Case of Malacoplakia of the Prostate which was Difficult to Distinguish from Prostate Cancer].

A 70-year-old-man was referred with urination pain and pyuria. Prostate specific antigen was 10.6 ng/ml, and urine culture was Escherichia coli. The abdominal ultrasonography showed irregular low echo at the right lobe of prostate. Prostate magnetic resonance imaging (MRI) showed an extracapsular infiltrated prostate tumor in the right lobe. Levofloxacin was administered and prostate biopsy was performed. The pathological examination revealed that the prostate tissue was filled with inflammatory cells and had lost its glandular duct structure. The patient was diagnosed with malacoplakia of the prostate. Four months after prostate biopsy, prostate MRI imaging showed disappearance of the extracapsular infiltration in right peripheral zone.

On the origin of gastric tumours: analysis of a case with intramucosal gastric carcinoma and oxyntic gland adenoma.

Most gastric cancers develop in inflamed gastric mucosa due to Helicobacter pylori infection, typically with metaplastic changes. However, the origins of gastric cancer remain unknown. Here, we present a case of intramucosal gastric carcinoma (IGC) and oxyntic gland adenoma (OGA) derived from spasmolytic polypeptide-expressing metaplasia (SPEM). Early gastric cancer adjacent to a polyp was found in the upper corpus of a 71-year-old woman without H. pylori infection and was endoscopically resected. Histological examination showed IGC and OGA, both of which had predominant MUC6 expression. Interestingly, gastric glands with enriched MUC6-positive mucous cells, referred to as SPEM, expanded between them. Whole-exome sequencing analysis revealed a truncating KRAS(G12D) mutation in IGC, OGA, and SPEM. In addition, TP53 and CDKN2A mutations and a loss of chromosome 17p were found in the IGC, whereas a GNAS mutation was observed in the OGA. These results indicated that IGC and OGA originated from the KRAS-mutated SPEM. © 2023 The Pathological Society of Great Britain and Ireland.

Squamous cell and urothelial carcinomas in an ileal neobladder.

Introduction: In cases of ileal neobladder following radical cystectomy for the treatment of bladder cancer, tumor development in the isolated gut segment is extremely rare. Herein, we report a case of squamous cell and urothelial carcinomas in the ileal neobladder 23 years after radical cystectomy. Case presentation: A 71-year-old man was referred to our hospital for further examination of a solitary tumor in an ileal neobladder. At the age of 48 years, he underwent radical cystectomy with ileal neobladder reconstruction. Transurethral resection of the bladder tumor was performed, and histopathological findings showed squamous cell carcinoma with high-grade urothelial carcinoma. Conclusion: To our knowledge, this is the first report of squamous cell and urothelial carcinomas in an ileal neobladder. While secondary tumor development in an ileal neobladder is rare, it is a cause for concern as a late postoperative adverse event. Therefore, long-term follow-ups are recommended.

A rare intracranial fusiform thrombosed aneurysm of the distal middle cerebral artery: A case report.

BACKGROUND: Intracranial aneurysms of the distal middle cerebral artery are rare, and most etiologies are infection or dissection. We present an extremely rare intracranial fusiform thrombosed aneurysm of the distal middle cerebral artery with histopathological confirmation of a pseudoaneurysm. CASE DESCRIPTION: Our patient, a 68-year-old female, was previously healthy and had no history of infection or trauma. A fusiform thrombosed aneurysm of the distal middle cerebral artery was detected incidentally. The patient was treated successfully with trapping and resection of the aneurysm followed by superficial temporal artery to middle cerebral artery anastomosis. Xanthochromic and hypertrophic arachnoid membranes around the aneurysm were noticed, and a thrombus was detected inside the lesion. The aneurysmal wall had hyalinized connective tissue incompletely surrounded with intima, with no media or adventitia. Pathologically, it was a pseudoaneurysm. CONCLUSION: We report an extremely rare case of a pseudoaneurysm of the distal middle cerebral artery. We discuss the etiology of the lesion, with a literature review, and propose that the appearance and increase of the pseudoaneurysm was followed by microbleed of an aneurysm unrelated to the branching zone.

Acute Tubulointerstitial Nephritis in Rosai-Dorfman Disease Mimicking IgG4-related Disease.

Rosai-Dorfman-Destombes disease (RDD) is a non-Langerhans cell histiocytosis characterized by the accumulation of histiocytes inside the lymph nodes or extranodally. The association between RDD and IgG4-related disease (IgG4-RD) is discussed. We herein report a case of RDD manifesting as acute tubulointerstitial nephritis mimicking IgG4-RD. The first renal biopsy showed severe tubulointerstitial nephritis with infiltration of S100-positive histiocytes and IgG4-positive plasma cells; storiform fibrosis and obliterative phlebitis were not confirmed. After prednisolone therapy, IgG4-positive cells and S100-positive histiocytes were decreased, but the IgG4/IgG ratio increased despite clinical improvement. These findings indicated extranodal RDD in the kidney presenting as tubulointerstitial nephritis.

Indolent feature of Helicobacter pylori-uninfected intramucosal signet ring cell carcinomas with CDH1 mutations.

BACKGROUND: In Helicobacter pylori (Hp)-uninfected individuals, diffuse-type gastric cancer (DGC) was reported as the most common type of cancer. However, the carcinogenic mechanism of Hp-uninfected sporadic DGC is largely unknown. METHODS: We performed whole-exome sequencing of Hp-uninfected DGCs and Hp-uninfected normal gastric mucosa. For advanced DGCs, external datasets were also analyzed. RESULTS: Eighteen patients (aged 29-78 years) with DGCs and nine normal subjects (28-77 years) were examined. The mutation burden in intramucosal DGCs (10-66 mutations per exome) from individuals aged 29-73 years was not very different from that in the normal gastric glands, which showed a constant mutation accumulation rate (0.33 mutations/exome/year). Unbiased dN/dS analysis showed that CDH1 somatic mutation was a driver mutation for intramucosal DGC. CDH1 mutation was more frequent in intramucosal DGCs (67%) than in advanced DGCs (27%). In contrast, TP53 mutation was more frequent in advanced DGCs (52%) than in intramucosal DGCs (0%). This discrepancy in mutations suggests that CDH1-mutated intramucosal DGCs make a relatively small contribution to advanced DGC formation. Among the 16 intramucosal DGCs (median size, 6.5 mm), 15 DGCs were pure signet ring cell carcinoma (SRCC) with reduced E-cadherin expression and a low proliferative capacity (median Ki-67 index, 2.4%). Five SRCCs reviewed endoscopically over 2-5 years showed no progression. CONCLUSIONS: Impaired E-cadherin function due to CDH1 mutation was considered as an early carcinogenic event of Hp-uninfected intramucosal SRCC. Genetic and clinical analyses suggest that Hp-uninfected intramucosal SRCCs may be less likely to develop into advanced DGCs.

[Complete Transurethral Resection of Large Non-Muscle-Invasive Bladder Cancer Preoperatively Suspected to Be Muscle-Invasive Disease : A Case Report and Literature Review].

A 69-year-old man presented with gross hematuria. Cystoscopy revealed a large papillary tumor occupying the bladder. Magnetic resonance imaging showed a large bladder tumor more than 8cm in maximum diameter,suspected to be muscle-invasive disease. We performed the 1st transurethral resection of bladder tumor (TURBT) for the main purpose of pathological confirmation. Histopathological findings of the resected specimens showed urothelial carcinoma,low grade pTa. We performed subsequent treatments with TURBT twice,resulting in complete resection. The histopathological findings showed the same results as those of the 1st TURBT conclusively,which was consistent with non-muscle-invasive bladder cancer. He received intravesical instillation of pirarubicin eight times in total and has remained free from recurrence for more than 26 months after the final TURBT.

Evaluation of Malignant Breast Lesions Using High-resolution Readout-segmented Diffusion-weighted Echo-planar Imaging: Comparison with Pathology.

PURPOSE: We aimed to investigate the performance of high resolution-diffusion-weighted imaging (HR-DWI) using readout-segmented echo-planar imaging in visualizing malignant breast lesions and evaluating their extent, using pathology as a reference. METHODS: This retrospective study included patients who underwent HR-DWI with surgically confirmed malignant breast lesions. Two radiologists blinded to the final diagnosis evaluated HR-DWI independently and identified the lesions, measuring their maximum diameters. Another radiologist confirmed if those lesions were identical to the pathology. The maximum diameters of the lesions between HR-DWI and pathology were compared, and their correlations were calculated using Spearman's correlation coefficient. Apparent diffusion coefficient (ADC) values of the lesions were measured. RESULTS: Ninety-five mass/64 non-mass lesions were pathologically confirmed in 104 females. Both radiologists detected the same 93 mass lesions (97.9%). Spearman's correlation coefficient for mass lesions were 0.89 and 0.90 (P < 0.0001 and 0001) for the two radiologists, respectively. The size differences within 10 mm were 90.3% (84/93) and 94.6% (88/93) respectively. One radiologist detected 35 non-mass lesions (54.7%) and another radiologist detected 32 non-mass lesions (50.0%), of which 28 lesions were confirmed as identical. Spearman's correlation coefficient for non-mass lesions were 0.59 and 0.22 (P = 0.0002 and 0.22), respectively. The mean ADC value of mass lesions and non-mass lesions were 0.80 and 0.89 × 10-3 mm2/s, respectively. CONCLUSION: Using HR-DWI, malignant mass lesions were depicted with excellent agreement with the pathological evaluation. Approximately half of the non-mass lesions could not be identified, suggesting a current limitation of HR-DWI.

Perihilar cholangiocarcinoma in an explanted liver after Kasai operation for biliary atresia: A case report and literature review.

Kasai operation is widely performed for biliary atresia (BA), as it improves the prognosis. Biliary tract cancer has rarely been reported as a complication after Kasai operation. A 17-year-old man underwent liver transplantation for progressive jaundice and liver dysfunction after Kasai operation for BA. A macroscopic examination of the explanted liver revealed a 3.6-cm-diameter mass in the hilus of the explanted liver. The tumor consisted of an atypical proliferation of glandular cells in the collagenous stroma. The differential diagnosis included a florid ductular reaction and primary adenocarcinoma. Immunohistochemistry revealed that the glandular cells were diffusely positive for IGF2BP3 and S100P, but negative for CDX2. A diagnosis of extrahepatic cholangiocarcinoma arising from the liver hilus was made. To our knowledge, this is the third case of perihilar cholangiocarcinoma after Kasai operation for BA. The previously reported cases had a poor prognosis, but the current case did not recur during a 15-year follow-up. Cholangiocarcinoma and hepatocellular carcinoma can occur as a late complication of BA after Kasai operation, and early detection and liver transplantation may improve the prognosis.

The comparison of high-resolution diffusion weighted imaging (DWI) with high-resolution contrast-enhanced MRI in the evaluation of breast cancers.

PURPOSE: We sought to investigate the performance of high resolution (HR) diffusion-weighted imaging (DWI) using readout-segmented echo-planar imaging (rs-EPI), compared with high-resolution contrast-enhanced MRI (HR CE-MRI) in terms of morphological accuracy, on the basis of the Breast Imaging and Reporting and Data System (BI-RADS) MRI descriptors and lesion size. METHODS: This retrospective study included the image data of 94 patients with surgically confirmed malignant breast lesions who had undergone high resolution diffusion-weighted imaging (HR-DWI) and HR CE-MRI. Two radiologists blinded to the final diagnosis independently identified the lesions on HR-DWI, described the morphology of the lesions according to BI-RADS descriptors, and measured lesion size. HR CE-MRI was subsequently evaluated using the same procedure. The inter-method agreement of the morphology was assessed using kappa statistics. Correlation on size was also assessed. RESULTS: Reader A detected 79 mass lesions and 37 non-mass lesions on HR-DWI and HR CE-MRI. Reader B detected 81 mass lesions and 33 non-mass lesions on HR-DWI and HR CE-MRI. Very high agreement (kappa = 0.81-0.89, p < .05) was observed in the shape and margin assessment of mass lesions, where agreement on internal enhancement/signals was moderate to substantial (kappa = 0.43-0.61, p < .05). Disagreement was mostly seen in the evaluation of rim enhancement. High agreement was observed for non-mass lesion distribution (kappa = 0.76-0.84, p < .05), and agreement on internal enhancement/signals was moderate to fair (kappa = 0.34-0.49, p < .05). Agreement among heterogeneous, clumped, and clustered-ring patterns was variable. Size assessment showed very strong correlation both in mass (Spearman's rho = 0.90-0.96, p < .0001) and non-mass lesions (Spearman's rho = 0.86, p < .0001). CONCLUSIONS: The findings in morphology and lesion extent showed high agreement between HR-DWI and HR CE-MRI for malignant breast lesions. These results imply the potential of applying HR-DWI for evaluation of malignant breast lesions using BI-RADS MRI.

A randomized, 3-arm, neoadjuvant, phase 2 study comparing docetaxel + carboplatin + trastuzumab + pertuzumab (TCbHP), TCbHP followed by trastuzumab emtansine and pertuzumab (T-DM1+P), and T-DM1+P in HER2-positive primary breast cancer.

PURPOSE: The standard of care in the neoadjuvant setting for human epidermal growth factor receptor 2 (HER2)-positive breast cancer is dual HER2-targeted therapy. However, a need to minimize treatment-related toxicity and improve pathological complete response (pCR) rates, particularly in luminal HER2-positive disease, exists. METHODS: Neopeaks, a randomized, phase 2 study, compared docetaxel + carboplatin + trastuzumab + pertuzumab (TCbHP; 6 cycles; group A), TCbHP (4 cycles) followed by trastuzumab emtansine + pertuzumab (T-DM1+P; 4 cycles; group B), and T-DM1+P (4 cycles; group C) regimens in HER2-positive primary breast cancer patients; concurrent hormone therapy with T-DM1+P was administered in case of estrogen receptor positivity (ER+). Based on tumor shrinkage, nonresponders in group C were switched to 5-fluorouracil + epirubicin + cyclophosphamide (FEC; 4 cycles). Primary endpoint was pCR (comprehensive pCR ypN0 [ypT0-TisypN0]). RESULTS: Of 236 patients enrolled, 204 were randomized to groups A (n = 51), B (n = 52), and C (n = 101). In group C, 80 (79%) patients continued T-DM1+P following favorable response, whereas 21 (21%) nonresponders switched to FEC. pCR rate was numerically higher with the TCbHP → T-DM1+P regimen (71%) versus the standard TCbHP (57%) and T-DM1+P (57%) regimens. The rate in group C was higher among responders continuing T-DM1+P (63%) versus nonresponders who switched to FEC (38%). pCR rates after initial 4 cycles of T-DM1+P (group C; 57%) and standard TCbHP regimen (57%) were equivalent. pCR rate in patients with ER+ was significantly higher in group B (69%) than groups A (43%) and C (51%), but was comparable in patients with ER- (67-76%). Compared with the T-DM1-based arm, the incidence of adverse events was higher in the taxane-based arms. CONCLUSION: In the neoadjuvant setting, the pCR rate with the standard TCbHP → T-DM1+P regimen was numerically better than the TCbHP regimen alone and significantly better in patients with ER+. Personalization of the T-DM1+P regimen could serve as a reasonable approach to minimize toxicity while maintaining efficacy. Trial registration ID: UMIN-CTR: UMIN000014649.

A novel endoscopic imaging system for quantitative evaluation of colonic mucosal inflammation in patients with quiescent ulcerative colitis.

Background and study aims Mucosal healing (MH) is associated with clinical outcome in ulcerative colitis (UC) patients. In most clinical trials, a Mayo endoscopic subscore (MES) of 0 or 1 is defined as MH. However, several recent studies have reported that clinical outcome is different between UC patients with MES 0 and those with MES 1. In addition, the MES is subjective and may differ among endoscopists. Therefore, a repeatable and objective scoring system is required to distinguish MES 0 from MES 1, even in clinically quiescent UC. Here, we assessed the usefulness of new image-enhancing endoscopic technology, the i-scan TE-c, to quantitatively evaluate colonic inflammation in patients with quiescent UC. Methods We retrospectively reviewed the data from 52 UC patients in clinical remission who had undergone routine colonoscopy with standard white light. The white-light images were reassessed using the new system, and the degree of colonic mucosal inflammation was quantified according to the MAGIC (Mucosal Analysis of Inflammatory Gravity by i-scan TE-c Image) score. We used the i-scan TE-c system to investigate the association among the MAGIC score, MES, and histologic activity (Geboes score). Results The MAGIC score was significantly higher in the MES 1 group than in the MES 0 group ( P  = 0.0034). The MAGIC score significantly correlated with the Geboes score ( P  = 0.015). Conclusions Our novel image-enhancing endoscopic system was useful for objective and quantitative evaluation of MH in patients with quiescent UC. Further clinical studies using this imaging system are required to confirm its clinical benefit for the management of UC patients.

Frequent mutations that converge on the NFKBIZ pathway in ulcerative colitis.

Chronic inflammation is accompanied by recurring cycles of tissue destruction and repair and is associated with an increased risk of cancer1-3. However, how such cycles affect the clonal composition of tissues, particularly in terms of cancer development, remains unknown. Here we show that in patients with ulcerative colitis, the inflamed intestine undergoes widespread remodelling by pervasive clones, many of which are positively selected by acquiring mutations that commonly involve the NFKBIZ, TRAF3IP2, ZC3H12A, PIGR and HNRNPF genes and are implicated in the downregulation of IL-17 and other pro-inflammatory signals. Mutational profiles vary substantially between colitis-associated cancer and non-dysplastic tissues in ulcerative colitis, which indicates that there are distinct mechanisms of positive selection in both tissues. In particular, mutations in NFKBIZ are highly prevalent in the epithelium of patients with ulcerative colitis but rarely found in both sporadic and colitis-associated cancer, indicating that NFKBIZ-mutant cells are selected against during colorectal carcinogenesis. In further support of this negative selection, we found that tumour formation was significantly attenuated in Nfkbiz-mutant mice and cell competition was compromised by disruption of NFKBIZ in human colorectal cancer cells. Our results highlight common and discrete mechanisms of clonal selection in inflammatory tissues, which reveal unexpected cancer vulnerabilities that could potentially be exploited for therapeutics in colorectal cancer.

High-resolution imaging mass spectrometry combined with transcriptomic analysis identified a link between fatty acid composition of phosphatidylinositols and the immune checkpoint pathway at the primary tumour site of breast cancer.

BACKGROUND: The fatty acid (FA) composition of phosphatidylinositols (PIs) is tightly regulated in mammalian tissue since its disruption impairs normal cellular functions. We previously found its significant alteration in breast cancer by using matrix-assisted laser desorption and ionisation imaging mass spectrometry (MALDI-IMS). METHODS: We visualised the histological distribution of PIs containing different FAs in 65 primary breast cancer tissues using MALDI-IMS and investigated its association with clinicopathological features and gene expression profiles. RESULTS: Normal ductal cells (n = 7) predominantly accumulated a PI containing polyunsaturated FA (PI-PUFA), PI(18:0/20:4). PI(18:0/20:4) was replaced by PIs containing monounsaturated FA (PIs-MUFA) in all non-invasive cancer cells (n = 12). While 54% of invasive cancer cells (n = 27) also accumulated PIs-MUFA, 46% of invasive cancer cells (n = 23) accumulated the PIs-PUFA, PI(18:0/20:3) and PI(18:0/20:4). The accumulation of PI(18:0/20:3) was associated with higher incidence of lymph node metastasis and activation of the PD-1-related immune checkpoint pathway. Fatty acid-binding protein 7 was identified as a putative molecule controlling PI composition. CONCLUSIONS: MALDI-IMS identified PI composition associated with invasion and nodal metastasis of breast cancer. The accumulation of PI(18:0/20:3) could affect the PD-1-related immune checkpoint pathway, although its precise mechanism should be further validated.

Hepatitis-associated Aplastic Anemia with Rapid Progression of Liver Fibrosis Due to Repeated Hepatitis.

Hepatitis-associated aplastic anemia (HAAA) is a variant of acquired aplastic anemia and characterized by bone marrow failure that follows the development of acute hepatitis. We herein report a rare case of HAAA with rapid progression of liver fibrosis due to repeated hepatitis. A pathological examination of liver specimens revealed liver fibrosis progression over a short period. Immunosuppressive therapy with cyclosporine effectively cured both the pancytopenia and hepatitis. Our case suggests that the pathological examination of the liver tissue is useful for determining a treatment plan and that immunosuppressive therapy is a promising treatment for both aplastic anemia and persistent hepatitis.

A ganglion-rich gastrointestinal stromal tumor: A case report.

We report a case of an extremely rare type of duodenal gastrointestinal stromal tumor (GIST) that included neuronal components. Although gastrointestinal autonomic nerve tumors (GANTs), a subtype of GISTs, exhibit ultrastructural features of the nerve plexus, neuronal cells have not been observed within GANTs or GISTs. GISTs originate from interstitial cells of Cajal (ICCs), which are markedly different from the progenitor cells of neural elements and neural-crest-derived stem cells. This may explain why GISTs typically lack neuronal elements. It remains unclear that the neuronal components of this tumor are neoplastic or hyperplastic, but proliferation and survival of ICCs have recently been reported to be closely related to neurons. Although we could not find the KIT, PDGFR, and BRAF mutation as far as we examined, it may have had a rare mutation in NF1, a fusion of EVT6-NTRK3, or an as-yet-unknown KIT mutation that affected neurogenesis. Further investigation of related genetic mutations and accumulation of data from other similar cases is needed.

Elevated levels of circulating ITIH4 are associated with hepatocellular carcinoma with nonalcoholic fatty liver disease: from pig model to human study.

BACKGROUND: Noninvasive biomarkers are urgently needed for optimal management of nonalcoholic fatty liver disease (NAFLD) for the prevention of disease progression into nonalcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC). In order to identify the biomarkers, we generated the swine hepatocellular carcinoma (HCC) model associated with NAFLD and performed serum proteomics on the model. METHODS: Microminipigs were fed a high-fat diet to induce NAFLD and a normal diet as the control. To induce HCC, diethylnitrosamine was intraperitoneally administered. Biopsied liver samples were histopathologically analyzed every 12 weeks. Serum proteins were separated by blue native two-dimensional gel electrophoresis and proteins of interest were subsequently identified by MALDI-TOF MS/MS. Human serum samples were analyzed to validate the candidate protein using antibody-mediated characterization. RESULTS: In the NAFLD pigs, hepatic histology of nonalcoholic steatohepatitis (NASH) was observed at 36 weeks, and HCC developed at 60 weeks. Among serum proteins identified with MALDI-TOF MS/MS, serum inter-alpha-trypsin inhibitor heavy chain 4 (ITIH4), an acute response protein which is secreted primarily by liver, was identified as the most characteristic protein corresponding with NAFLD progression and HCC development in the NAFLD pigs. With immunoassay, serum ITIH4 levels in the NAFLD pigs were chronologically increased in comparison with those in control animal. Furthermore, immunohistochemistry showed ITIH4 expression in hepatocytes also increased in both the cancer lesions and parenchyma as NAFLD progressed. Human study is also consistent with this observation because serum ITIH4 levels were significantly higher in HCC-NAFLD patients than in the simple steatosis, NASH, and virus-related HCC patients. Of note, HCC-NAFLD patients who had higher serum ITIH4 levels exhibited poorer prognosis after hepatectomy. CONCLUSIONS: We established an HCC pig model associated with NAFLD. Serum proteomics on the swine HCC with NAFLD model implicated ITIH4 as a non-invasive biomarker reflecting NAFLD progression as well as subsequent HCC development. Most importantly, the results in the swine study have been validated in human cohort studies. Dissecting speciation of serum ITIH4 promises to have clinical utility in monitoring the disease.