Detection of residual stones by peroral direct cholangioscopy in patients with intrahepatic stones after hepaticojejunostomy: A prospective study (with video).

BACKGROUND AND AIMS: The difficulty of radiographic confirmation of the presence of stones remains a challenge in the treatment of intrahepatic bile duct (IHBD) stones in patients after hepaticojejunostomy (HJ). Peroral direct cholangioscopy (PDCS) enables direct observation of the bile duct and is useful for detecting and removing residual stones; however, its effectiveness is not clearly established in this clinical context. METHODS: This single-center, single-arm, prospective study included 44 patients with IHBD who underwent bowel reconstruction with HJ during the study period. Stone removal was performed by short-type double-balloon enteroscopy (DBE). Following balloon-occluded cholangiography, the DBE was exchanged for an ultraslim endoscope through the balloon overtube for PDCS. The primary endpoint was the rate of residual stones detected by PDCS. Secondary endpoints were success rate of PDCS, residual stone removal with PDCS, procedure time for PDCS, procedure-related adverse events, and stone recurrence rate. RESULTS: PDCS was successful in 39/44 patients (89%), among whom residual stones were detected in 16 (41%) (95% CI: 28%-54%). Twelve patients (75%) had residual stones <5 mm. Stone removal was successful in 15 (94%) patients and median procedure time for PDCS was 16 (IQR: 10-26) min. The rate of procedure-related adverse events was 7% (3/44), all of which improved with conservative treatment. During median follow-up of 2.1 years (IQR: 1.4-3.3), the overall probability of recurrence-free status at 1, 2, and 3 years was 100%, 92%, and 86%, respectively. CONCLUSIONS: PDCS is a safe and effective procedure for complete stone removal in patients with IHBD stones after HJ.

A Case of Nonocclusive Mesenteric Ischemia During Bioradiotherapy With Cetuximab.

Nonocclusive mesenteric ischemia (NOMI) causes mesenteric ischemia and intestinal necrosis despite the absence of organic obstruction, such as thrombi and emboli in mesenteric blood vessels, and it has an extremely poor prognosis. We report a case of NOMI developed during bioradiotherapy (BRT) with cetuximab for cervical lymph node metastasis of tongue cancer. The patient was a 73-year-old man who underwent right radical neck dissection for neck lymph node metastasis after tongue cancer surgery. Postoperatively, the patient received BRT with cetuximab. On the 34th day after BRT, the patient had abdominal distension and a decreased level of consciousness. Contrast-enhanced computed tomography revealed mesenteric ischemia without thrombi and extensive intestinal emphysema. The patient was diagnosed with NOMI. Furthermore, he had septic shock and was treated with vasopressors and antibacterial agents; however, the condition of the patient did not improve, and he died on the same day.

Possible Role of miR-375-3p in Cervical Lymph Node Metastasis of Oral Squamous Cell Carcinoma.

No clinically useful predictors of latent cervical lymph node metastasis (LNM) in early oral squamous cell carcinoma (OSCC) are available. In this study, we focused on the microRNAs (miRNAs) involved in the expression of numerous genes and explored those associated with latent cervical LNM in early OSCC (eOSCC). First, microarray and RT-PCR analyses revealed a significant downregulation of miR-375-3p expression in primary eOSCC tissues with latent cervical LNM. Next, we examined the effects of miR-375-3p mimics on the growth and migration of four human OSCC cell lines that do not express miR-375-3p. The overexpression of miR-375-3p significantly suppressed the cell proliferation and migration of human OSCC cells in vitro. Furthermore, miR-375-3p mimics markedly inhibited the subcutaneously xenografted human OSCC tumors. Finally, we found the genes involved in the PI3K-AKT pathway and cell migration as target gene candidates of miR-375-3p in human OSCC cells. These findings suggest that miR-375-3p functions as a tumor suppressive-miRNA in OSCC and may serve as a potential biomarker for the prediction of latent cervical LNM in eOSCC and a useful therapeutic target to suppress OSCC progression.

MicroRNA-34a-5p: A pivotal therapeutic target in gallbladder cancer.

Gallbladder cancer incidence has been increasing globally, and it remains challenging to expect long prognosis with the current systemic chemotherapy. We identified a novel nucleic acid-mediated therapeutic target against gallbladder cancer by using innovative organoid-based gallbladder cancer models generated from KrasLSL-G12D/+; Trp53f/f mice. Using comprehensive microRNA expression analyses and a bioinformatics approach, we identified significant microRNA-34a-5p downregulation in both murine gallbladder cancer organoids and resected human gallbladder cancer specimens. In three different human gallbladder cancer cell lines, forced microRNA-34a-5p expression inhibited cell proliferation and induced cell-cycle arrest at the G1 phase by suppressing direct target (CDK6) expression. Furthermore, comprehensive RNA sequencing revealed the significant enrichment of gene sets related to the cell-cycle regulators after microRNA-34a-5p expression in gallbladder cancer cells. In a murine xenograft model, locally injected microRNA-34a-5p mimics significantly inhibited gallbladder cancer progression and downregulated CDK6 expression. These results provide a rationale for promising therapeutics against gallbladder cancer by microRNA-34a-5p injection, as well as a strategy to explore therapeutic targets against cancers using organoid-based models, especially for those lacking useful genetically engineered murine models, such as gallbladder cancer.

Prognostic Significance of Serum Interleukin-6 Levels in Oral Squamous Cell Carcinoma.

Introduction The prognosis of oral squamous cell carcinoma (OSCC) is often poor despite standard treatments. Additionally, no useful prognostic markers are available. Therefore, we aimed to investigate the relationship between serum Interleukin-6 (IL-6) levels and prognosis and explore its local and systemic effects in patients with OSCC. Methods Ninety-five new cases of OSCC were included, and the prognosis was compared between high and low serum IL-6 groups. The localization of IL-6 in OSCC tissues was examined. Furthermore, a comprehensive gene expression analysis was performed in OSCC tissues and compared between the two groups. Results A significant difference in overall survival and disease-free survival was observed. Furthermore, a substantial expression of IL-6 was localized in the stroma. Comprehensive gene expression analysis of tumor localization showed increased expression of genes related to oxidoreductase and lipid metabolism in the primary tissues of the group with high serum IL-6 levels. Regarding the correlation between blood tests and serum IL-6 levels, a strong positive correlation was observed between inflammatory responses and nutritional factors. Conclusion These results suggest that serum IL-6 may be a prognostic factor for metabolic abnormalities in patients with OSCC and that aggressive nutritional interventions may contribute to prognosis.

The association between high-dose allopurinol and erythropoietin hyporesponsiveness in advanced chronic kidney disease. JOINT-KD study.

INTRODUCTION: To explore the association between urate-lowering agents and reduced response to erythropoietin-stimulating agents in patients suffering from chronic kidney disease G5. METHODS: We conducted a cross-sectional, multicenter study in Japan between April and June 2013, enrolling patients aged 20 years or older with an estimated glomerular filtration rate of ≤15 mL/min/1.73 m2. Exclusion criteria encompassed patients with a history of hemodialysis, peritoneal dialysis, or organ transplantation. The patients were categorized into four groups based on the use of urate-lowering drugs: high-dose allopurinol (>50 mg/day), low-dose allopurinol (≤50 mg/day), febuxostat, and no-treatment groups. We used a multivariable logistic regression model, adjusted for covariates, to determine the odds ratio (OR) for erythropoietin hyporesponsiveness, defined by an erythropoietin resistance index (ERI) of ≥10, associated with urate-lowering drugs. RESULTS: A total of 542 patients were included in the analysis, with 105, 36, 165, and 236 patients in the high-dose allopurinol, low-dose allopurinol, febuxostat, and no-treatment groups, respectively. The median and quartiles of ERIs were 6.3 (0, 12.2), 3.8 (0, 11.2), 3.4 (0, 9.8), and 4.8 (0, 11.2) in the high-dose allopurinol, low-dose allopurinol, febuxostat, and no-treatment groups, respectively. The multivariate regression model showed a statistically significant association between the high-dose allopurinol group and erythropoietin hyporesponsiveness, compared to the no-treatment group (OR=1.98, 95% confidence interval: 1.10-3.57). CONCLUSIONS: Our study suggests that the use of high-dose allopurinol exceeding the optimal dose may lead to hyporesponsiveness to erythropoiesis-stimulating agents.

KRT13 and UPK1B for differential diagnosis between metastatic lung carcinoma from oral squamous cell carcinoma and lung squamous cell carcinoma.

Oral squamous cell carcinomas unusually show distant metastasis to the lung after primary treatment, which can be difficult to differentiate from primary squamous cell carcinoma of the lung. While the location and number of tumor nodules is helpful in diagnosing cases, differential diagnosis may be difficult even with histopathological examination. Therefore, we attempted to identify molecules that can facilitate accurate differential diagnosis. First, we performed a comprehensive gene expression analysis using microarray data for OSCC-LM and LSCC, and searched for genes showing significantly different expression levels. We then identified KRT13, UPK1B, and nuclear receptor subfamily 0, group B, member 1 (NR0B1) as genes that were significantly upregulated in LSCC and quantified the expression levels of these genes by real-time quantitative RT-PCR. The expression of KRT13 and UPK1B proteins were then examined by immunohistochemical staining. While OSCC-LM showed no KRT13 and UPK1B expression, some tumor cells of LSCC showed KRT13 and UPK1B expression in 10 of 12 cases (83.3%). All LSCC cases were positive for at least one of these markers. Thus, KRT13 and UPK1B might contribute in differentiating OSCC-LM from LSCC.

MicroRNA-451a inhibits gemcitabine-refractory biliary tract cancer progression by suppressing the MIF-mediated PI3K/AKT pathway.

Gemcitabine is an effective chemotherapeutic agent for biliary tract cancers (BTCs), including gallbladder cancer (GBC) and cholangiocarcinoma (CCA). However, few other effective agents are currently available, particularly for GEM-refractory BTCs. We previously identified microRNA-451a (miR-451a) as a potential therapeutic target in GBC. To elucidate the antineoplastic effects of miR-451a and its underlying mechanisms, we transfected miR-451a into GBC, gemcitabine-resistant GBC (GR-GBC), and gemcitabine-resistant CCA (GR-CCA) cell lines. Furthermore, mimicking in vivo conditions, tumorigenic GBC organoids and three-dimensional (3D) cell culture systems were employed to investigate the anti-proliferative effects of miR-451a on BTCs, and its effect on stem cell properties. We found that miR-451a significantly inhibited cell proliferation, induced apoptosis, and reduced chemoresistant phenotypes, such as epithelial-mesenchymal transition, in both GBC and GR-GBC. The principal mechanism is probably the negative regulation of the phosphatidylinositol 3-kinase/AKT pathway, partially accomplished by directly downregulating macrophage migration inhibitory factor. The Gene Expression Omnibus database revealed that miR-451a was the most significantly downregulated microRNA in CCA tissues. The introduction of miR-451a resulted in similar antineoplastic effects in GR-CCA. Furthermore, miR-451a reduced cell viability in 3D spheroid models and tumorigenic GBC organoids. These findings suggest that the supplementation of miR-451a is a potential treatment strategy for GEM-refractory BTCs.

Cysts or necrotic components in pancreatic ductal adenocarcinoma is associated with the risk of EUS-FNA/B complications including needle tract seeding.

BACKGROUND: EUS-FNA/B for pancreatic ductal adenocarcinoma (PDAC) is generally considered to be safe; however, while the incidence is low, there are occurrences of complications. Among these complications, there are serious ones like needle tract seeding (NTS), and it is not known than which types of tumors have the risks of EUS-FNA/B complications. This study aimed to evaluate the risk of EUS-FNA/B complications in patients with PDAC, focusing on morphological features. METHODS: Overall, 442 patients who underwent EUS-FNA/B for solid pancreatic masses between January 2018 and May 2022 in four institutions were retrospectively surveyed. Finally, 361 patients histopathologically diagnosed with PDAC were analyzed. Among these patients, 79 tumors with cysts or necrotic components were compared with 282 tumors without cysts or necrotic components. The incidence and risk of EUS-FNA/B complications including NTS were evaluated. RESULTS: There were 9 (2.4 %) of total EUS-FNA/B complications and 3 (0.8 %) of NTS. The incidence of total complication rate and NTS in tumors with cysts or necrotic components were significantly higher than in those without cysts or necrotic components (total complication 6.3 % vs. 1.4 %, p = 0.026, NTS 3.7 % vs. 0 %, p = 0.01). The transgastric route of puncture (OR: 93.3, 95 % CI: 3.81-2284.23) and the existence of cysts or necrotic components (OR: 7.3, 95 % CI: 1.47-36.19) were risk factors for EUS-FNA/B complications identified by the multivariate analysis. CONCLUSIONS: We should pay attention to the risks of EUS-FNA/B complications, including NTS, when the tumor has cysts or necrotic components.

The efficacy of non-anesthesiologist-administered propofol sedation with a target-controlled infusion system during double-balloon endoscopic retrograde cholangiopancreatography.

BACKGROUND: The sedation method used during double-balloon endoscopic retrograde cholangiopancreatography (DB-ERCP) differs among countries and/or facilities, and there is no established method. This study aimed to evaluate the efficacy of non-anesthesiologist-administered propofol (NAAP) sedation using a target-controlled infusion (TCI) system during DB-ERCP. METHODS: This retrospective study was conducted between May 2017 and December 2020 at an academic center. One hundred and fifty-six consecutive patients who underwent DB-ERCP were sedated by gastroenterologists using diazepam (n = 77) or propofol with a TCI system (n = 79), depending on the period. The primary endpoint was a comparison of poor sedation rates between the two groups. Poor sedation was defined as a condition requiring the use of other sedative agents or discontinuation of the procedure. Secondary endpoints were sedation-related adverse events and risk factors for poor sedation. RESULTS: Poor sedation occurred significantly more often in the diazepam sedation group (diazepam sedation, n = 12 [16%] vs. propofol sedation, n = 1 [1%]; P = 0.001). Vigorous body movements (3 or 4) (diazepam sedation, n = 40 [52%] vs. propofol sedation, n = 28 [35%]; P = 0.038) and hypoxemia (< 85%) (diazepam sedation, n = 7 [9%] vs. propofol sedation, n = 1 [1%]; P = 0.027) occurred significantly more often in the diazepam sedation group. In the multivariate analysis, age < 70 years old (OR, 10.26; 95% CI, 1.57-66.98; P = 0.015), BMI ≥ 25 kg/m2 (OR, 11.96; 95% CI, 1.67-85.69; P = 0.014), and propofol sedation (OR, 0.06; 95% CI, 0.01-0.58; P = 0.015) were associated factors for poor sedation. CONCLUSIONS: NAAP sedation with the TCI system during DB-ERCP was safer and more effective than diazepam sedation.

MicroRNA-1289 Functions as a Novel Tumor Suppressor in Oral Squamous Cell Carcinoma.

Recently, numerous tumor-suppressive microRNAs (TS-miRs) have been identified in human malignancies. Here, we attempted to identify novel TS-miRs in oral squamous cell carcinoma (OSCC). First, we transfected human OSCC cells individually with 968 synthetic miRs mimicking human mature miRs individually, and the growth of these cells was evaluated using the WST-8 assay. Five miR mimics significantly reduced the cell growth rate by less than 30%, and the miR-1289 mimic had the most potent growth inhibitory effect among these miRs. Subsequently, we assessed the in vivo growth-inhibitory effects of miR-1289 using a mouse model. The administration of the miR-1289 mimic-atelocollagen complex significantly reduced the size of subcutaneously xenografted human OSCC tumors. Next, we investigated the expression of miR-1289 in OSCC tissues using reverse transcription-quantitative PCR. The expression level of miR-1289 was significantly lower in OSCC tissues than in the adjacent normal oral mucosa. Furthermore, 15 genes were identified as target genes of miR-1289 via microarray and Ingenuity Pathway Analysis (IPA) microRNA target filtering. Among these genes, the knockdown of magnesium transporter 1 (MAGT1) resulted in the most remarkable cell growth inhibition in human OSCC cells. These results suggested that miR-1289 functions as a novel TS-miR in OSCC and may be a useful therapeutic tool for patients with OSCC.

A Comparison of the Efficacy of Plastic Stent Placement Above and Across the Sphincter of Oddi for Benign Biliary Hilar Stricture.

We investigated the efficacy and safety of endoscopic plastic stent (PS) placement for hilar benign biliary strictures (BBSs) and compared cases with PS placement above (inside stent, IS) and across (usual stent, US) the sphincter of Oddi. Patients who underwent initial endoscopic PS placement for hilar BBSs between August 2012 and December 2021 were retrospectively analyzed. Hilar BBSs in 88 patients were investigated. Clinical success was achieved in 81 of these cases (92.0%), including 38 patients in the IS group and 43 patients in the US group. Unexpected stent exchange (uSE) before the first scheduled PS exchange occurred in 18 cases (22.2%). The median time from first stent placement to uSE was 35 days. There was no significant difference in the rate and median time to uSE between the two groups. The rates of adverse events such as pancreatitis or cholangitis in the two groups did not significantly differ. However, the rate of difficult stent removal in the IS group (15.8%) was significantly higher than that in the US group (0%) (p=0.0019). US placement is preferable to IS placement for scheduled stent exchange, as it offers the same effectiveness and risk of adverse events with easier stent removal.

Risk Factors for Ceftriaxone-Associated Pseudolithiasis in Adults.

INTRODUCTION: Ceftriaxone (CTRX) is known to occasionally cause pseudolithiasis. This condition is often observed in children; however, few studies have reported the incidence and risk factors for CTRX-associated pseudolithiasis. METHODS: In this single-center retrospective study, we investigated the incidence of and risk factors for CTRX-associated pseudolithiasis in adults. All patients underwent computed tomography to confirm pseudolithiasis before and after CTRX administration. RESULTS: The study included 523 patients. Pseudolithiasis was detected in 89 patients (17%). Data analysis showed that abdominal area-related biliary diseases at the site of infection (odds ratio [OR] 0.19, 95% confidence interval [CI] 0.064-0.53, p = 0.0017), CTRX administration for >3 days (OR 5.0, 95% CI: 2.5-9.9, p < 0.0001), CTRX dose of 2 mg (OR 5.2, 95% CI: 2.8-9.6, p < 0.0001), fasting period >2 days (OR 3.2, 95% CI: 1.6-6.4, p = 0.0010), and estimated glomerular filtration rate <30 mL/min/1.73 m2 (OR 3.4, 95% CI: 1.6-7.5, p = 0.0022) were independent factors for pseudolithiasis. CONCLUSIONS: CTRX-associated pseudolithiasis may occur in adults and should be considered in the differential diagnosis in patients who develop abdominal pain or liver enzyme elevation after CTRX administration, particularly in patients with chronic kidney disease, in those who are fasting, in and those who receive high-dose CTRX therapy.

The mutational spectrum in whole exon of p53 in oral squamous cell carcinoma and its clinical implications.

Mutations in p53 are common in human oral squamous cell carcinoma (OSCC). However, in previous analyses, only detection of mutant p53 protein using immunohistochemistry or mutations in some exons have been examined. Full length mutant p53 protein in many cases shows a loss of tumor suppressor function, but in some cases possibly shows a gain of oncogenic function. In this study, we investigate relationships of outcomes with the mutational spectrum of p53 (missense and truncation mutations) in whole exon in OSCC. Specimens from biopsy or surgery (67 cases) were evaluated using next-generation sequencing for p53, and other oncogenic driver genes. The data were compared with overall survival (OS) and disease-free survival (DFS) using univariate and multivariate analyses. p53 mutations were detected in 54 patients (80.6%), 33 missense mutations and 24 truncation mutations. p53 mutations were common in the DNA-binding domain (43/52) and many were missense mutations (31/43). Mutations in other regions were mostly p53 truncation mutations. We detected some mutations in 6 oncogenic driver genes on 67 OSCC, 25 in NOTCH1, 14 in CDKN2A, 5 in PIK3CA, 3 in FBXW7, 3 in HRAS, and 1 in BRAF. However, there was no associations of the p53 mutational spectrum with mutations of oncogenic driver genes in OSCC. A comparison of cases with p53 mutations (missense or truncation) with wild-type p53 cases showed a significant difference in lymph node metastasis. DFS was significantly poorer in cases with p53 truncation mutations. Cases with p53 truncation mutations increased malignancy. In contrast, significant differences were not found between cases with p53 missense mutations and other mutations. The p53 missense mutation cases might include cases with mostly similar function to that of the wild-type, cases with loss of function, and cases with various degrees of gain of oncogenic function.

Gastric linitis plastica with autoimmune pancreatitis diagnosed by an endoscopic ultrasonography-guided fine-needle biopsy: A case report.

BACKGROUND: Gastric linitis plastica (GLP) is a subset of gastric cancer with a poor prognosis. It is difficult to obtain a definitive diagnosis by endoscopic mucosal biopsies, and the usefulness of an endoscopic ultrasonography-guided fine-needle biopsy (EUS-FNB) for GLP has been recently reported. Meanwhile, autoimmune diseases are occasionally known to coexist with malignant tumors as paraneoplastic syndrome. We herein report the usefulness of an EUS-FNB for detecting GLP and the possibility of paraneoplastic syndrome coexisting with GLP. CASE SUMMARY: An 81-year-old man was admitted to our hospital for a 1-mo history of epigastric pain that increased after eating. His laboratory data revealed high levels of serum carbohydrate antigen 19-9 and immunoglobulin-G4. Endoscopic examinations showed giant gastric folds and reddish mucosa; however, no epithelial changes were observed. The gastric lumen was not distensible by air inflation, suggesting GLP. Computed tomography showed the thickened gastric wall, the diffuse enlargement of the pancreas, and the peripancreatic rim, which suggested autoimmune pancreatitis (AIP) coexisting with GLP. Because the pathological findings of the endoscopic biopsy showed no malignancy, he underwent an EUS-FNB and was diagnosed with GLP. He received chemotherapy for unresectable gastric cancer due to peritoneal metastasis, after which both the gastric wall thickening and diffuse enlargement of the pancreas were improved. CONCLUSION: An EUS-FNB for GLP with a negative endoscopic biopsy is useful, and AIP may develop as a paraneoplastic syndrome.

Diversity of the Origin of Cancer Stem Cells in Oral Squamous Cell Carcinoma and Its Clinical Implications.

Oral squamous cell carcinoma (OSCC) histopathologically accounts for ≥90% of oral cancer. Many clinicopathological risk factors for OSCC have also been proposed, and postoperative therapy is recommended in guidelines based on cancer stage and other risk factors. However, even if the standard treatment is provided according to the guidelines, a few cases rapidly recur or show cervical and distant metastasis. In this review article, we focus on the diversity of the origin of OSCC. We also discuss cancer stem cells (CSCs) as a key player to explain the malignancy of OSCC. CSCs are a subset of cancer cells that occupy a very small portion of the cancer mass and have characteristics of stem cells. When gene abnormalities accumulate in somatic stem cells, those cells transform into CSCs. CSCs as the origin of cancer then autonomously grow and develop into cancer. The histopathological phenotype of cancer cells is determined by the original characteristics of the somatic stem cells and/or surrounding environment. OSCC may be divided into the following three categories with different malignancy based on the origin of CSCs: cancer from oral epithelial stem cell-derived CSCs, cancer from stem cells in salivary gland-derived CSCs, and cancer from bone marrow-derived stem cell-derived CSCs.

Plasma KRAS mutations predict the early recurrence after surgical resection of pancreatic cancer.

BACKGROUND: The technique to analyze circulating tumor DNA (ctDNA) in body fluid (so-called "liquid biopsy") is recently developed. AIMS: Our aim was to assess the utility of liquid biopsy for predicting progression of pancreatic ductal adenocarcinoma (PDAC) after surgical resection or chemotherapy. METHODS: A total of 72 patients with PDAC were retrospectively enrolled for this study, 33 treated surgically and 39 given chemotherapy, either FOLFIRINOX (oxaliplatin/irinotecan/fluorouracil/leucovorin) or gemcitabine plus nab-paclitaxel. Prior to treatment, patients were screened for the presence of KRAS mutations (G12D and G12V) in plasma using droplet digital polymerase chain reaction, and outcomes were compared. RESULTS: KRAS mutations were identified in plasma samples of 12 patients (36%) underwent surgical resection. Patients with plasma KRAS mutations had significantly shorter disease-free survival (DFS) and overall survival (p < .01 and p = .01, respectively). Of 10 clinical variables analyzed, plasma KRAS mutation was the factor predictive of DFS in multivariate analysis (RR = 3.58, 95% CI: 1.36-9.60; p = .01). Although 12 patients (31%) given chemotherapy tested positive for plasma KRAS mutations, there was no demonstrable relation between plasma KRAS mutations and progression-free survival (PFS) or overall survival (OS) (p = .35 and p = .68, respectively). CONCLUSIONS: In patients with PDAC, detection of KRAS mutations in plasma proved independently predictive of early recurrence after surgical resection but did not correlate with PFS following chemotherapy.