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Objective Expression of Spam1/PH20 and its modulation of high/low molecular weight hyaluronan substrate have been proposed to play an important role in murine oligodendrocyte precursor cell (OPC) maturation in vitro and in normal and demyelinated central nervous system (CNS). We reexamined this using highly purified PH20. Methods Steady‐state expression of mRNA in OPCs was evaluated by quantitative polymerase chain reaction; the role of PH20 in bovine testicular hyaluronidase (BTH) inhibition of OPC differentiation was explored by comparing BTH to a purified recombinant human PH20 (rHuPH20). Contaminants in commercial BTH were identified and their impact on OPC differentiation characterized. Spam1/PH20 expression in normal and demyelinated mouse CNS tissue was investigated using deep RNA sequencing and immunohistological methods with two antibodies directed against recombinant murine PH20. Results BTH, but not rHuPH20, inhibited OPC differentiation in vitro. Basic fibroblast growth factor (bFGF) was identified as a significant contaminant in BTH, and bFGF immunodepletion reversed the inhibitory effects of BTH on OPC differentiation. Spam1 mRNA was undetected in OPCs in vitro and in vivo; PH20 immunolabeling was undetected in normal and demyelinated CNS. Interpretation We were unable to detect Spam1/PH20 expression in OPCs or in normal or demyelinated CNS using the most sensitive methods currently available. Further, “BTH” effects on OPC differentiation are not due to PH20, but may be attributable to contaminating bFGF. Our data suggest that caution be exercised when using some commercially available hyaluronidases, and reports of Spam1/PH20 morphogenic activity in the CNS may be due to contaminants in reagents.

Background We evaluated the antitumor activity and safety of avelumab, a human anti–PD-L1 IgG1 antibody, as first-line switch-maintenance (1 L-mn) or second-line (2 L) treatment in patients with advanced gastric/gastroesophageal cancer (GC/GEJC) previously treated with chemotherapy. Methods In a phase 1b expansion cohort, patients without (1 L-mn) or with (2 L) disease progression following first-line chemotherapy for advanced GC/GEJC received avelumab 10 mg/kg intravenously every 2 weeks. Endpoints included best overall response, progression-free survival (PFS), overall survival (OS), and safety. Results Overall, 150 patients were enrolled (1 L-mn, n = 90; 2 L, n = 60) and median follow-up in the 1 L-mn and 2 L subgroups was 36.0 and 33.7 months, respectively. The confirmed objective response rate was 6.7% in both subgroups (95% CI, 2.5–13.9% and 1.8–16.2%, respectively), including complete responses in 2.2% of the 1 L-mn subgroup (n = 2). In the 1 L-mn and 2 L subgroups, median duration of response was 21.4 months (95% CI, 4.0–not estimable) and 3.5 months (95% CI, 2.8–8.3) and disease control rates were 56.7 and 28.3%, respectively. Median PFS in the 1 L-mn and 2 L subgroups was 2.8 months (95% CI, 2.3–4.1) and 1.4 months (95% CI, 1.3–1.5), with 6-month PFS rates of 23.0% (95% CI, 14.7–32.4%) and 7.9% (95% CI, 2.6–17.2%), and median OS was 11.1 months (95% CI, 8.9–13.7) and 6.6 months (95% CI, 5.4–9.4), respectively. In the 1 L-mn subgroup, median OS measured from start of 1 L chemotherapy was 18.7 months (95% CI, 15.4–20.6). Across both subgroups, 20.7% had an infusion-related reaction of any grade. Other common treatment-related adverse events (TRAEs) of any grade included fatigue (10.0%) and nausea (6.7%). Treatment-related serious adverse events occurred in 4.0% of patients. Overall, 8.7% had a grade ≥3 TRAE, including 1 treatment-related death. Conclusion Avelumab showed clinical activity and an acceptable safety profile in patients with GC/GEJC. Trial registration ClinicalTrials.gov NCT01772004; registered 21 January 2013. Electronic supplementary material The online version of this article (10.1186/s40425-019-0508-1) contains supplementary material, which is available to authorized users.

Summary Background The approval of anti-programmed death ligand 1 (PD-L1) and anti-programmed death 1 agents has expanded treatment options for patients with locally advanced or metastatic urothelial carcinoma. Avelumab, a human monoclonal anti-PD-L1 antibody, has shown promising antitumour activity and safety in this disease. We aimed to assess the safety profile in patients (both post-platinum therapy and cisplatin-naive) treated with avelumab and to assess antitumour activity of this drug in post-platinum patients. Methods In this pooled analysis of two cohorts from the phase 1 dose-expansion JAVELIN Solid Tumor study, patients aged 18 years and older with histologically or cytologically confirmed locally advanced or metastatic urothelial carcinoma that had progressed after at least one previous platinum-based chemotherapy were enrolled from 80 cancer treatment centres or hospitals in the USA, Europe, and Asia. Eligible patients had adequate end-organ function, an Eastern Cooperative Oncology Group performance status of 0 or 1, life expectancy of at least 3 months, and at least one measurable lesion. Cisplatin-ineligible patients who might have been previously treated in the perioperative setting, including platinum-naive patients, were also eligible. Patients unselected for PD-L1 expression received avelumab (10 mg/kg, 1 h intravenous infusion) every 2 weeks until confirmed disease progression, unacceptable toxicity, or other criterion for withdrawal. The primary endpoint for this efficacy expansion cohort was confirmed best overall response (according to RECIST version 1.1), adjudicated by independent review. Safety analysis was done in all patients who received at least one dose of avelumab. Antitumour activity was assessed in post-platinum patients who received at least one dose of avelumab. This trial is registered with ClinicalTrials.gov, number NCT01772004; enrolment in this cohort of patients with metastatic urothelial carcinoma is closed and the trial is ongoing. Findings Between Sept 3, 2014, and March 15, 2016, 329 patients with advanced metastatic urothelial carcinoma were screened for enrolment into this study; 249 patients were eligible and received treatment with avelumab for a median of 12 weeks (IQR 6·0–19·7) and followed up for a median of 9·9 months (4·3–12·1). Safety and antitumour activity were evaluated at data cutoff on June 9, 2016. In 161 post-platinum patients with at least 6 months of follow-up, a best overall response of complete or partial response was recorded in 27 patients (17%; 95% CI 11–24), including nine (6%) complete responses and 18 (11%) partial responses. The most frequent treatment-related adverse events (any grade in ≥10% patients) were infusion-related reaction (73 [29%]; all grade 1–2) and fatigue (40 [16%]). Grade 3 or worse treatment-related adverse events occurred in 21 (8%) of 249 patients, the most common of which were fatigue (four [2%]), and asthenia, elevated lipase, hypophosphataemia, and pneumonitis in two (1%) patients each. 19 (8%) of 249 patients had a serious adverse event related to treatment with avelumab, and one treatment-related death occurred (pneumonitis). Interpretation Avelumab showed antitumour activity in the treatment of patients with platinum-refractory metastatic urothelial carcinoma; a manageable safety profile was reported in all avelumab-treated patients. These data provide the rationale for therapeutic use of avelumab in metastatic urothelial carcinoma and it has received accelerated US FDA approval in this setting on this basis. Funding Merck KGaA, and Pfizer Inc.

Purpose We assessed the safety and antitumor activity of avelumab, a fully human anti–programmed death-ligand 1 (PD-L1) IgG1 antibody, in patients with refractory metastatic urothelial carcinoma. Methods In this phase Ib, multicenter, expansion cohort, patients with urothelial carcinoma progressing after platinum-based chemotherapy and unselected for PD-L1 expression received avelumab 10 mg/kg intravenously every 2 weeks. The primary objectives were safety and tolerability. Secondary objectives included confirmed objective response rate (Response Evaluation Criteria in Solid Tumors [RECIST] version 1.1), progression-free survival, overall survival (OS), and PD-L1–associated clinical activity. PD-L1 positivity was defined as expression by immunohistochemistry on ≥ 5% of tumor cells. Results Forty-four patients were treated with avelumab and followed for a median of 16.5 months (interquartile range, 15.8 to 16.7 months). The data cutoff was March 19, 2016. The most frequent treatment-related adverse events of any grade were fatigue/asthenia (31.8%), infusion-related reaction (20.5%), and nausea (11.4%). Grades 3 to 4 treatment-related adverse events occurred in three patients (6.8%) and included asthenia, AST elevation, creatine phosphokinase elevation, and decreased appetite. The confirmed objective response rate by independent central review was 18.2% (95% CI, 8.2% to 32.7%; five complete responses and three partial responses). The median duration of response was not reached (95% CI, 12.1 weeks to not estimable), and responses were ongoing in six patients (75.0%), including four of five complete responses. Seven of eight responding patients had PD-L1–positive tumors. The median progression-free survival was 11.6 weeks (95% CI, 6.1 to 17.4 weeks); the median OS was 13.7 months (95% CI, 8.5 months to not estimable), with a 12-month OS rate of 54.3% (95% CI, 37.9% to 68.1%). Conclusion Avelumab was well tolerated and associated with durable responses and prolonged survival in patients with refractory metastatic UC.

Tumor necrosis factor-Stimulated Gene 6 protein (TSG-6) is a hyaluronan (HA)-binding glycoprotein containing an HA-binding Link module. Because of its well-defined structure, HA binding properties and small size, TSG-6 is an excellent candidate as an alternative to animal-derived HA-binding protein (HABP) for the detection of HA. The present work describes the generation and characterization of a novel recombinant HA-binding probe obtained by fusion of a modified TSG-6 Link module with mutationally inactivated heparin-binding sequence and the Fc portion of human IgG1 (TSG-6-ΔHep-Fc) for tissue HA detection in histological samples. Direct binding assays indicated strong binding of TSG-6-ΔHep-Fc to HA, with little residual binding to heparin. Histolocalization of HA in formalin-fixed, paraffin-embedded tissue sections using biotin-TSG-6-ΔHep-Fc resulted in hyaluronidase-sensitive staining patterns similar to those obtained with biotin-HABP, but with improved sensitivity. HA was detected in many human tissues, and was most abundant in soft connective tissues such as the skin dermis and the stroma of various glands. Digital image analysis revealed a linear correlation between biotin-HABP and biotin-TSG-6-ΔHep-Fc staining intensity in a subset of normal and malignant human tissues. These results demonstrate that TSG-6-ΔHep-Fc is a sensitive and specific probe for the detection of HA by histological methods.

Reduced phosphatase and tensin homolog (PTEN) expression in breast cancer confers susceptibility to phosphatidylinositol 3 kinase/Akt pathway inhibitors. As inhibitors of this pathway are in development for treatment of breast and other cancers, it is of interest to minimize the frequency at which PTEN signal is reduced because of poor antigen preservation, rather than to decreased protein expression. Methods for preserving antigenicity include coating sections in paraffin, refrigerated storage, desiccation, and storage in nitrogen. This study examined various combinations of the first 2 methods with respect to PTEN immunoreactivity. This was assessed semiquantitatively in freshly cut sections of paraffin blocks from 26 breast cancer specimens. The histoscore was compared with that observed in cut sections stored for 2 to 6 months with or without paraffin coating at room temperature (RT; 20 to 25°C) or at 2 to 8°C. At 2 months, the histoscore was strongest in freshly cut sections. Among the other treatments, refrigerated paraffin-coated sections had the highest histoscore followed by refrigerated uncoated sections. It was lower in RT paraffin-coated sections. Uncoated sections stored at RT showed the greatest signal loss. The greatest magnitude of changes between the various storage conditions and freshly cut slides ranged from 15% for refrigerated paraffin-coated slides to 75% for RT uncoated slides at 4 months. Although freshly cut paraffin-embedded sections were optimal, sections can be stored with adequate antigen preservation for 2 months using refrigeration with or without paraffin coating, and for 4 months using refrigeration with paraffin coating.

Background: Patients (pts) with advanced GC/GEJC have a poor prognosis and standard chemotherapy options provide limited efficacy with significant toxicity. Avelumab, a human anti–PD-L1 IgG1 monoclonal antibody, has shown durable efficacy and acceptable safety in various tumor types. Here, we report updated data from JAVELIN Solid Tumor (NCT01772004) for avelumab treatment in pts with previously treated GC/GEJC. Methods: Pts with advanced GC/GEJC without or with disease progression after first-line combination chemotherapy in the metastatic setting (1L mn and 2L subgroups, respectively) received avelumab 10 mg/kg IV Q2W until confirmed progression, unacceptable toxicity, or withdrawal. Best overall response and progression-free survival (PFS) per RECIST v1.1 and overall survival (OS) were evaluated from the start of avelumab treatment. Time-to-event endpoints were estimated using the Kaplan–Meier method. Results: As of Sep 30, 2017, pts in the 1L mn subgroup (n=90) and 2L subgroup (n=60) had a median (range) follow-up of 36.0 mo (27–42) and 33.7 mo (25–42), respectively. In the 1L mn subgroup, best response on prior chemotherapy was partial response in 27.8%, stable disease in 65.6%, and not evaluable in 6.7%. Confirmed objective response rate (ORR) during avelumab 1L mn therapy (additional effect after end of chemotherapy) was 6.7% (95% CI 2.5–13.9), including complete response in 2.2%. Median duration of response was 21.4 months (95% CI 4.0–not estimable) and responses to avelumab lasted ≥12 mo in an estimated 50.0% of responders (95% CI 11.1–80.4). In evaluable pts with PD-L1+ (n=26) and PD-L1− (n=51) tumors (≥1% tumor cell cutoff; Dako PD-L1 IHC 73-10 pharmDx assay), ORR was 7.7% (95% CI 0.9–25.1) vs 3.9% (95% CI 0.5–13.5). The disease control rate (DCR) was 56.7% and median PFS was 2.8 mo (95% CI 2.3–4.1); median OS was 11.1 mo (95% CI 8.9–13.7) when measured from the start of avelumab therapy (12-mo rate 46.2%; 95% CI 35.6–56.1), and 18.7 mo (95% CI 15.4–20.6) when measured from start of 1L chemotherapy. In the 2L subgroup, confirmed ORR was 6.7% (95% CI 1.8–16.2), DCR was 28.3%, median PFS was 1.4 mo (95% CI 1.3–1.5), and median OS was 6.8 mo (95% CI 5.4–9.5). Across both subgroups, 56.7% had a treatment-related adverse event (TRAE) of any grade, most commonly (>10%) infusion-related reaction (12.7%) and fatigue (10.0%). 8.7% had a grade ≥3 TRAE, with 1 treatment-related death (hepatic failure/autoimmune hepatitis). 15.3% had an immune-related AE (grade ≥3 in 2.0%). Conclusion: Avelumab showed promising clinical activity and an acceptable safety profile in pts with advanced GC/GEJC with prior chemotherapy. A phase 3 trial of avelumab as 1L mn therapy vs continuation of 1L chemotherapy in pts with advanced GC/GEJC is ongoing (NCT02625610). Citation Format: Hyun Cheol Chung, Hendrik-Tobias Arkenau, Jeeyun Lee, Sun Young Rha, Do-Youn Oh, Lucjan Wyrwicz, Yoon-Koo Kang, Keun-Wook Lee, Todd M. Bauer, Sung Sook Lee, Margaret Kemeny, Ulrich Keilholz, Bohuslav Melichar, Alain Mita, Ruth Plummer, Denis Smith, Arnold B. Gelb, Huiling Xiong, Janet Hong, Vikram Chand, Howard Safran. Avelumab (anti-PD-L1) as first-line maintenance (1L mn) or second-line (2L) therapy in patients with advanced gastric or gastroesophageal junction cancer (GC/GEJC): updated phase Ib results from the JAVELIN Solid Tumor trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr CT111.

Purpose: To evaluate the therapeutic and biological effects of CHIR-258, an orally bioavailable, potent inhibitor of class III-V receptor tyrosine kinases, in colon cancer models. Experimental Design: The pharmacologic activity of CHIR-258 was characterized by monitoring target modulation as well as by evaluating the antitumor and antiangiogenic effects in human colon xenograft models. Results: CHIR-258 inhibits vascular endothelial growth factor receptor 1/2, fibroblast growth factor receptor 1/3, and platelet-derived growth factor receptor β (PDGFRβ) and shows both antitumor and antiangiogenic activities in vivo. Treatment of KM12L4a human colon cancer cells with CHIR-258 resulted in a dose-dependent inhibition of vascular endothelial growth factor receptor 1 and PDGFRβ phosphorylation and reduction of phosphorylated extracellular signal-regulated kinase (ERK) levels, indicating modulation of target receptors and downstream signaling. In vivo administration of CHIR-258 resulted in significant tumor growth inhibition and tumor regressions, including large, established tumors (500-1,000 mm3). Immunohistochemical analysis showed a reduction of phosphorylated PDGFRβ and phosphorylated ERK in tumor cells after oral dosing with CHIR-258 compared with control tumors. These changes were accompanied by decreased tumor cell proliferation rate and reduced intratumoral microvessel density. CHIR-258 inhibited the phosphorylation of PDGFRβ and ERK phosphorylation in tumors within 2 hours following dosing and the inhibitory activity was sustained for >24 hours. Significant antitumor activity was observed with intermittent dosing schedules, indicating a sustained biological activity. Conclusion: These studies provide evidence that biological activity of CHIR-258 in tumors correlates with efficacy and aids in the identification of potential biomarkers of this multitargeted receptor tyrosine kinase inhibitor. CHIR-258 exhibits properties that make it a promising candidate for clinical development in a variety of solid and hematologic malignancies.

e20581 Background: Response to anti–PD-L1/PD-1 therapy in patients with NSCLC has been associated with tumor PD-L1 expression. Avelumab is a fully human anti‒PD-L1 IgG1 antibody that has shown antitumor activity in advanced NSCLC, with a trend for greater efficacy in patients with PD-L1+ vs PD-L1– NSCLC tumors, as assessed using a proprietary PD-L1 assay in development (Dako, Carpinteria, CA; based on antibody clone 73-10, Merck KGaA, Darmstadt, Germany). This study compares the analytical performance of the 73-10 assay with the FDA-approved PD-L1 IHC 22C3 pharmDx diagnostic assay (Dako) in NSCLC tumors. Methods: Formalin-fixed paraffin-embedded NSCLC tumor samples were obtained from a commercial source and from a study of first-line (1L) avelumab in patients with advanced NSCLC (NCT01772004). Tumor membrane PD-L1 expression was assessed with the 73-10 and 22C3 assays by immunohistochemistry. Correlation between assays was determined at different PD-L1 cut-off levels. Results: In initial staining of commercial NSCLC samples, the 73-10 assay showed a broad dynamic range. Of 148 commercial samples, the 73-10 assay with ≥1%, ≥50% and ≥80% cut-offs classified 64.2%, 36.5% and 23.6% of samples as PD-L1+, respectively, whereas 20.3% were PD-L1+ with the 22C3 assay at a pre-specified ≥50% cut-off. The overall percentage agreement between assays was 83.8% using the ≥50% cut-off for both assays, and 93.9% using the ≥80% cut-off for 73-10 and ≥50% for 22C3. In follow-up studies, 83 study samples from the 1L NSCLC cohort were evaluable with both assays. With the 73-10 assay at ≥1%, ≥50%, and ≥80% cut-offs, 79.5%, 45.8%, and 31.3% of study tumors, respectively, were PD-L1+; with the 22C3 assay at ≥1% and ≥50% cut-offs, 59.0% and 21.7% were PD-L1+. Conclusions: Using a high tumor cell PD-L1 staining cut-off, the 73-10 and 22C3 assays showed highly concordant staining of NSCLC samples, with similar sensitivity observed with an ≥80% cut-off for 73-10 and ≥50% for 22C3. Using a low frequency of tumor PD-L1 expression, data suggested that the 73-10 assay had greater sensitivity than the 22C3 assay. These results provide a rationale for additional studies using the 73-10 assay at various cut-offs within the avelumab trial program. Clinical trial information: NCT01772004.

4528 Background: Avelumab, a fully human anti‒PD-L1 IgG1 antibody, has shown promising efficacy and safety in 2 cohorts of patients (pts) with mUC. We now report updated data from a pooled analysis of these pts with mUC from JAVELIN Solid Tumor (NCT01772004) and further characterize the clinical activity of avelumab in this disease. Methods: Pts with mUC progressed after platinum-based therapy or cisplatin ineligible received avelumab 10 mg/kg 1-hour IV Q2W. Tumors were assessed every 6 weeks by independent review (RECIST v1.1). Endpoints included objective response rate (ORR), duration of response (DOR), progression-free survival (PFS), overall survival (OS), safety (NCI CTCAE v4.0), and tumor PD-L1 expression. Results: As of Jun 9, 2016, 249 pts had received avelumab for a median of 12 weeks (range 2-92) and were followed up for a minimum of 6 weeks. Primary tumor site was upper tract (renal pelvis/ureter) in 23.3% and lower tract (bladder/urethra) in 76.7%. 242 pts (97.2%) had progressed on prior platinum therapy and 7 pts (2.8%) were platinum naive. In 161 post-platinum pts with ≥6 months of follow-up, confirmed ORR was 17.4% (95% CI 11.9-24.1; complete response in 6.2%) with a disease control rate of 39.8%. Response was ongoing in 23/28 responders at data cut (82.1%; median DOR not reached), and the 24-week durable response rate was 92.3% (95% CI 72.6-98.0). Responses occurred across PD-L1 expression levels tested (≥5% and < 5% tumor cell‒staining [25.4% and 13.2%]). In all post-platinum pts (n = 242), median PFS was 6.6 weeks (95% CI 6.1-11.6), median OS was 7.4 months (95% CI 5.7-10.3) and 6-month OS rate was 54.9 (95% CI 47.7-61.7). Treatment-related adverse events (TRAE) of any grade occurred in 166/249 pts (66.7%); most common (≥10%) were infusion-related reaction (22.9%, all grade ≤2) and fatigue (16.1%). 21 pts (8.4%) had a grade ≥3 TRAE (fatigue [1.6%] and asthenia [0.8%] in > 1 pt). 34 pts (13.7%) had an immune-related AE (grade ≥3 in 2.4%). There was 1 treatment-related death (pneumonitis). Conclusions: Avelumab was well tolerated and showed durable responses in heavily pretreated pts with mUC, irrespective of tumor PD-L1 expression status. Clinical trial information: NCT01772004.

330 Background: Avelumab is a fully human anti‒PD-L1 IgG1 antibody with promising efficacy and safety in patients (pts) with metastatic urothelial carcinoma (mUC). To further characterize the clinical activity of avelumab in mUC, we report a planned interim pooled analysis of 2 cohorts from a large phase 1b trial (NCT01772004). Methods: Pts with mUC progressed after platinum-based therapy or cisplatin-ineligible, and unselected for tumor PD-L1 expression, received avelumab 10 mg/kg (1 h IV) Q2W. Response was assessed every 6 wks by independent review per RECIST v1.1. Endpoints included objective response rate (ORR), duration of response (DOR), progression-free survival (PFS), overall survival (OS), safety (NCI CTCAE v4.0), and tumor PD-L1 expression (clone 73-10). Results: As of Mar 19, 2016, 241 pts received avelumab for a median of 8 wks (range 2-80); median follow-up was 7.3 mos (range 0-18.2). Primary tumor sites were upper tract (renal pelvis/ureter [23.7%]) and lower tract (bladder/urethra [76.3%]). 95.4% of pts had progressed on prior platinum therapy, and 63.4% had received ≥ 2 prior lines for advanced disease (range 0-6). In 153 pts with ≥ 6 mos follow-up, confirmed ORR was 17.6% (95% CI 12.0-24.6) with 9 complete responses and 18 partial responses; 24/27 (88.9%) were ongoing. Median DOR was not reached, and the 24-wk DOR rate was 92.0% (95% CI 71.6, 97.9). 36 pts had stable disease as best response (disease control rate 41.2%). Median PFS was 6.4 wks (95% CI 6.1-11.4), and median OS was 7.0 mos (95% CI 5.6-11.1). Based on a ≥ 5% PD-L1 staining cutoff in evaluable pts with PD-L1+ (n = 56) and PD-L1– (n = 75) tumors, ORR was 25.0% (95% CI 14.4-38.4) vs 14.7% (95% CI 7.6-24.7; p = 0.178). Treatment-related adverse events (TRAE) of any grade occurring in ≥ 10% of pts were infusion-related reaction (22.8%) and fatigue (12.0%). 7.5% had a grade ≥ 3 TRAE; fatigue (1.2%)/asthenia (0.8%) occurred in > 1 pt. 28 pts (11.6%) had an immune-related TRAE (grade ≥ 3 in 2.5%). There was 1 treatment-related death (pneumonitis). Conclusions: Avelumab is well tolerated and shows promising clinical activity, including durable responses, in pts with mUC, regardless of tumor PD-L1 expression status. Follow-up is ongoing. Clinical trial information: NCT01772004.

Purpose: To identify the genetic alterations associated with renal adenomas.Materials and Methods: We analyzed 37 renal adenomas obtained at autopsy (23 papillary and 14 non-papillary) by comparative genomic hybridization.Results: In papillary tumors, the median number of gains and losses of genetic material per tumor was 2.0 and 1.0, respectively. Papillary tumors were characterized predominantly by gains of genetic material on chromosomes 7 (57%), 17 (35%), 16 (26%), 12 (26%), 3 (22%), 20 (22%) and loss of a sex chromosome (83%). In 6 papillary tumors less than or equal to 5 mm. in diameter, gain of chromosome 7 occurred in 4 specimens. Initiating events for papillary renal adenomas include gain of chromosome 7 and loss of a sex chromosome. In non-papillary tumors, the median number of gains and losses of genetic material per tumor was 1.0 and 1.0, respectively. Non-papillary tumors were characterized by loss of genetic material on chromosome 3p (50%), loss of a sex chromosome (36%) and a gain o...

BACKGROUND The prognostic values of intratumoral microvessel density (iMVD), tumor cell proliferation rate, DNA content (ploidy), and p53 protein expression are controversial or have not been well studied in patients with renal cell carcinoma (RCC) confined to the kidney. METHODS A uniform group of 52 clear cell (conventional) RCCs confined to the kidney (classified as T1N0M0 or T2N0M0) were analyzed for iMVD, MIB-1 score, DNA content, S-phase fraction, and p53 protein expression by immunohistochemical methods or flow cytometry. iMVD was evaluated in a single area (X200, 1.15 mm2) representative of the highest MVD (neovascular "hot spot") after independently highlighting endothelial cells with antibodies specific for factor VIII-related antigen (F8/86) and CD31 (JC/70A). The MIB-1 antibody (Ki-67 antigen) score was used as a marker for the tumor cell proliferation rate. DNA content and S-phase fraction were determined by flow cytometry using paraffin embedded tissue. p53 expression was assessed using the D07 antibody. RESULTS The median time of clinical follow-up was > 9 years. Eleven patients died of disease; the median time to death was 26 months. iMVD counts using antifactor VIII and anti-CD31 were tightly correlated (correlation coefficient = 0.89). S-phase fraction was higher in aneuploid tumors than in diploid tumors (mean, 12.4% vs. 4.3%; P = 0.01). Using univariate survival analyses, tumor size (stage classification pT1 vs. PT2; P = 0.01) and nuclear grade (P = 0.04) were associated with shortened survival. No statistically significant differences in survival were found for iMVD, MIB-1 score, DNA content, S-phase fraction, or p53 expression. Only two cases strongly expressed p53 protein; both tumors were of high nuclear grade. Using multivariate survival analyses, nuclear grade and tumor size were the only independent prognostic factors (best model P = 0.002). CONCLUSIONS In this study, nuclear grade and tumor size were found to be independent predictors of survival in locally confined clear cell (conventional) RCC, as has been shown previously for locally confined RCC in general. MIB-1 score, iMVD counts, DNA content, S-phase fraction, and p53 expression did not contribute additional prognostic information. Cancer 1997; 80:1768-75. © 1997 American Cancer Society.

BACKGROUND. Renal cell carcinomas include several distinct entities with a range of biologic and clinical behavior from relatively indolent to extremely aggressive. Although conventional prognostic factors such as stage and grade are quite useful, other clinical, laboratory, and pathologic findings are now believed to have additional predictive value. METHODS. A review of the literature on the evaluation of prognostic factors in general and on the current status of prognostic factors in renal cell carcinoma in particular was undertaken. A working classification of prognostic factors, as recommended by the College of American Pathologists, was used. For clarity, the prognostic indicators were grouped according to whether each was a patient-related or tumor-related factor. RESULTS. Patient-related prognostic factors include symptomatic presentation, significant weight loss, poor performance status, anemia, hypercalcemia, elevated alkaline phosphatase and, perhaps, elevated serum ferritin. The most widely used tumor-related prognostic factors include stage, grade, and histologic type. Recently proposed biomarkers still under investigation include DNA content, as well as markers of cellular proliferation, apoptosis, and angiogenesis, among others. CONCLUSIONS. Current prognostic factors for renal cell carcinoma yield considerable information for assisting with patient management and predicting clinical outcome. Traditional prognostic factors remain the most valuable, even though a variety of other patient-related and tumor-related factors may significantly contribute to prognostic information. A number of recently described biomarkers show great promise but the current data are insufficient to recommend their use.

Colorectal carcinoma remains a leading cause of cancer morbidity and mortality. Various clinical signs and pathologic factors have been shown to have a bearing on a patient’s prognosis. Some of these factors, such as extent of disease (stage) and histologic grade, are generally accepted, while others, primarily biologic and molecular markers, have been proposed recently and remain controversial. The authors describe both more established and newly proposed variables, reviewing multivariate analyses to examine their relative importance. The recommendations of the Association of Directors of Anatomic and Surgical Pathology for the reporting of colorectal carcinomas are presented.

Abstract.The following is the editor's summary of the Pre-symposium workshop on Contemporary Assessment of Technologies presented at the Symposium on Molecular Approaches to Laboratory Diagnosis at San Francisco in February 1995. This workshop was moderated by Dr Joel M. Palefsky, and Dr Michael P. Busch. We have briefly summarized the presentations by: (1) Dr Indira Hewlett of the Center for Biologics Evaluation and Research, Food and Drug Administration entitled «Technology overview»; (2) Dr John J. Sninsky of Roche Molecular Systems Inc. entitled «Polymerase Chain Reaction»; (3) Dr Terrance Walker of Becton Dickinson Research Center entitled «Strand Displacement Amplification»;(4) Dr Mickey Urdea of Chiron Corporation entitled «bDNA assay» and (5) Dr Robert H. Singer of University of Massachussets Medical Center entitled «Image analysis of in situ hybridization». Although it was not possible to list all the references to the primary literature, we have attempted to provide the key references as far as possible.

BACKGROUND The natural histories of Hodgkin's disease (HD) and non-Hodgkin's lymphomas (NHL) during pregnancy are not well understood. METHODS All cases of HD and NHL diagnosed during pregnancy at Stanford University Medical Center since 1987 were reviewed and clinical follow-up was obtained. Various immunohistochemical studies and in situ hybridization for Epstein–Barr virus (EBV) encoded RNA were performed in a subset of cases. RESULTS Seventeen cases of HD and 12 cases of NHL were accessioned (median age; 27 yrs). The HD cases were classified as 13 nodular sclerosis type, 3 mixed cellularity type, and 1 unclassified. Clinical follow-up revealed most of the patients had Stage II to III disease and were diagnosed on average at 22 weeks gestation. Most of the patients deferred therapy until after delivery and had no evidence of disease at the last follow-up except for one death with disease but not from it. NHL were classified according to the working formulation as high or intermediate grade lymphomas of various types, including both nodal and extranodal sites. Clinical follow-up revealed most had Stage II to IV disease and were diagnosed on average at 23 weeks gestation. Patients with HD tended to survive longer than those with NHL (raw mortality, P = 0.04). In situ hybridization failed to provide support for the presence of EBV in a subset of patients with NHL. CONCLUSIONS The clinical behavior of these neoplasms during pregnancy does not appear to be significantly different from that outside of the setting of pregnancy. Treatment of selected HD patients apparently may be safely deferred until after delivery. Patients with NHL present with higher stage disease and have a poorer prognosis than those with HD. Cancer 1996;78:304-10.

Introduction: Hyaluronan (HA) is an extracellular matrix glycosaminoglycan that has been shown to accumulate at high levels in several cancers, including non-small cell lung cancer (NSCLC). The accumulation of HA in NSCLC is associated with a more aggressive phenotype and shortened overall survival. In a prevalence analysis using tissue microarrays of archival human specimens (N = 194) stained by affinity histochemistry, we demonstrated that 55% of adenocarcinomas, 74% of squamous cell carcinomas and 83% of large cell carcinomas accumulated HA. The engineered enzyme PEGylated recombinant human hyaluronidase PH20 (PEGPH20) removes HA from the tumor microenvironment. PEGPH20-based therapies are being evaluated in clinical studies of several solid tumors, including NSCLC (NCT02346370, NCT02563548). Herein, we evaluated the hypothesis that PEGPH20 combination therapy could enhance the efficacy of pemetrexed (PEM), a folate antimetabolite approved by FDA as a first-line treatment in combination with cisplatin, against locally advanced and metastatic NSCLC in patients with non-squamous histology. Methods: We evaluated PEGPH20 plus PEM in a NSCLC xenograft model. First, the human NSCLC adenocarinoma cell line A549 was transduced with hyaluronan synthase-3 (HAS3) to generate the A549/HAS3 cell line. Next, A549/HAS3 cells were inoculated adjacent to the tibial periosteum of nude mice and tumor growth was monitored via ultrasonography. When tumors size reached ∼250 mm3, mice were staged into six groups: (1) vehicle; (2) PEGPH20, (3) PEM daily, (4) PEM weekly, (5) PEGPH20+PEM daily or (6) PEGPH20+PEM weekly. Vascular volume, hypoxia and tumor growth inhibition (TGI) were assessed using conventional methods. Affinity histochemical staining for HA was performed on formalin-fixed paraffin-embedded tissue sections after harvest. Results: A549/HAS3 tumors grew faster than parental A549 tumors in vivo. PEGPH20 monotherapy significantly reduced tumoral HA, increased vascular volume and reduced hypoxia in tumors. Further, TGI with PEGPH20+PEM, weekly or daily, was 29.7 or 31.1%, respectively, whereas in either PEGPH20 alone or PEM alone, both were Conclusion: These data suggest that addition of PEGPH20 to pemetrexed increases the efficacy of PEM and support further investigation of PEGPH20 plus PEM treatment in NSCLC tumors of non-squamous histology that accumulate HA. Citation Format: Renee Clift, Jessica A. Cowell, Susan J. Zimmerman, Mathieu Marella, Ping Jiang, Arnold B. Gelb, Michael J. LaBarre, Daniel C. Maneval, Curtis B. Thompson, Xiaoming Li. PEGylated recombinant hyaluronidase PH20 (PEGPH20) enhances pemetrexed antitumor efficacy in a human nonsquamous NSCLC xenograft model. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 283.

We describe the first case of an Epstein-Barr virus (EBV)-associated natural killer-large granular lymphocyte (NK-LGL) leukemia in the United States to the best of our knowledge. A 29-year-old woman of Japanese descent developed EBV infection after a blood transfusion as indicated by a rise in serum antibody titers. Peripheral blood and bone marrow aspirate smears demonstrated increased LGLs. Flow cytometry showed that these cells expressed NK-associated surface antigens. Cytogenetic analysis of the bone marrow aspirate showed two distinct but related clones with multiple copies of a modified 7 marker chromosome. Death followed colonic perforation. Findings at necropsy included bone marrow lymphocytosis and erythrophagocytosis, a mononucleosis-like lymphadenitis, atypical hepatitis with a mixed, predominantly T-cell infiltrate, interstitial pneumonitis, and multiorgan system vasculitis with perforation of the transverse colon. Epstein-Barr virus transcripts were identified in lymphocytes infiltrating liver and peripheral nerve by in situ hybridization. In addition, Southern blot analyses showed monoclonal bands superimposed on oligoclonal ladders of EBV termini in liver and lymph node. The identical episomal form of EBV was found in the bone marrow, lymph node, and liver. No immunoglobulin (Ig), T-cell receptor beta, or T-cell receptor gamma chain gene rearrangements were identified. These studies support the hypothesis that the LGL population was a neoplastic EBV-related clonal proliferation of NK cells.

Recent literature suggests that usual Hodgkin's disease (nodular sclerosing and mixed cellularity types or UHD) and nodular lymphocyte predominance Hodgkin's disease (NLPHD) may be distinct clinical and pathologic entities. Thus, coexistence of NLPHD and UHD in the same patient is expected to be rare. We undertook a review of cases accessioned as NLPHD and UHD in the Laboratory of Surgical Pathology at Stanford University Hospital between January 1980 and May 1992 and found five patients with UHD that predated, followed, or coexisted with lesions histologically typical of NLPHD. All of the patients were male with ages ranging from 10 to 30 years at presentation (median, 22 years; mean, 22.2 years). The sites initially involved by disease were primarily peripheral lymph nodes in the region of the head and neck: cervical (three), supraclavicular (one), submandibular (one). One patient presented with mixed-cellularity Hodgkin's disease (MCHD), two with nodular sclerosis Hodgkin's disease including the cellular phase, one with NLPHD, and the remaining patient presented with a composite malignancy comprising MCHD and NLPHD. Development of the second lymphoma was associated with a somewhat more variable distribution of nodal involvement. The morphologic features in each biopsy specimen resembled either typical NLPHD or UHD, except for one case in which cells with features of both Reed-Sternberg cells and lymphocytic and histiocytes cells were identified. However, the immunophenotypic profiles obtained with a panel of monoclonal antibodies remained distinct for all cases studied. None of the cases showed reactivity with antibodies against the Epstein-Barr-virus latent membrane protein. Thus, NLPHD and UHD maintain a distinct phenotype, even when occurring in the same patient. A second conclusion is that the utility of Leu-7 (CD57) reactivity in distinguishing NLPHD applies to problematic as well as classic cases. Finally, Epstein-Barr virus is not implicated in NLPHD cases associated with UHD.

A 48-year-old man with a history of recurrent urolithiasis and chronic renal failure underwent a nephrectomy for a renal mass. At surgery the mass proved to be a calculus impacted in a dilated calyx. Gross examination of the kidney revealed chalky white deposits in the deep medulla and papillary tips. Histologic examination revealed chronic interstitial nephritis with brown spicules within some tubular epithelial cells and larger deposits of brown crystals within tubular lumina, the interstitium of the medulla, and papillary tips. Polarization microscopy revealed individual crystals scattered throughout the renal parenchyma. Although the arrangement of the crystals was reminiscent of uric acid, and, in fact, a clinical diagnosis of gouty nephropathy was made, x-ray diffraction analysis demonstrated crystals of 2,8-dihydroxyadenine. Enzymatic studies confirmed the complete absence of adenine phosphoribosyltransferase activity in erythrocyte lysates.

Recombinant interleukin-2 (rIL-2) is a pleiotropic cytokine that activates select immune effector cell responses associated with antitumor activity, including antibody-dependent cellular cytotoxicity (ADCC). Rituximab is an anti-CD20 monoclonal antibody that activates ADCC in non-Hodgkin lymphoma (NHL). The ability of rIL-2 to augment rituximab-dependent tumor responses was investigated. The efficacy of rIL-2 in combination with rituximab was evaluated in 2 NHL tumor xenograft models: the CD20hi, rituximab-sensitive, low-grade Daudi model and the CD20lo, aggressive, rituximab-resistant Namalwa model. Combination of rIL-2 plus rituximab was synergistic in a rituximab-sensitive Daudi tumor model, as evidenced by significant tumor regressions and increased time to tumor progression, compared with rIL-2 and rituximab single agents. In contrast, rituximab-resistant Namalwa tumors were responsive to single-agent rIL-2 and showed an increased response when combined with rituximab. Using in vitro killing assays, rIL-2 was shown to enhance activity of rituximab by activating ADCC and lymphokine-activated killer activity. Additionally, the activity of rIL-2 plus rituximab F(ab')2 was similar to that of rIL-2 alone, indicating a critical role for immunoglobulin G1 Fc-FcgammaR-effector responses in mediating ADCC. Antiproliferative and apoptotic tumor responses, along with an influx of immune effector cells, were observed by immunohistochemistry. Collectively, the data suggest that rIL-2 mediates potent tumoricidal activity against NHL tumors, in part, through activation and trafficking of monocytes and natural killer cells to tumors. These data support the mechanistic and therapeutic rationale for combination of rIL-2 with rituximab in NHL clinical trials and for single-agent rIL-2 in rituximab-resistant NHL patients.

Background: Hyaluronan (HA), a glycosaminoglycan distributed in the extracellular matrix, is frequently upregulated in cancer and correlates with disease progression. Administration of a first-in-class PEGylated recombinant human hyaluronidase (PEGPH20) depletes HA, unmasks HA around tumor cells and decreases tumor interstitial fluid pressure, increasing accessibility to systemically administered agents in preclinical models. PEGPH20 recently received Orphan Drug designation in pancreatic ductal adenocarcinoma (PDA) in the USA. We report the development and analytical validation of a novel prototype assay for a companion diagnostic to select patients with PDA for molecularly targeted therapies with PEGPH20. Design: A recently described biotinylated recombinant immunoadhesin (HTI-601, Jadin 2014) was adapted for use in an immunohistochemistry-based assay on formalin-fixed paraffin-embedded tissue. Sensitivity, specificity, and within-laboratory precision studies were performed in a research mode and then at a central laboratory on a validation set of approximately 200 tumor and normal tissues under GCP conditions. Both pathologist scoring and operator-assisted image analysis (positive pixel count for strong positive pixels) were evaluated. Results: Analytical sensitivity studies identified an optimal probe dilution of 0.417 μg/mL on an immunostainer based on dynamic range in 4 human tumor xenografts containing differential levels of HA. The frequency of high HA observed was 62.7% of archival PDA (N = 75). During analytical specificity, 5.1% of 75 PDA and 3 normal adjacent tissue samples had faint or focal staining after pre-digestion with recombinant PH20 hyaluronidase. No cross-reactivity was identified in 99 normal human tissues across a panel of histotypes; only 1 (1.0%) colon sample had staining of macrophages. Within-laboratory precision among three observers’ annotations had average%CVs as follows: between day 4.8%, between run 12.2%, and repeatability 16.6%. Conclusion: The sensitivity, specificity and within-laboratory precision of the prototype assay using image analysis are acceptable. The frequency of high HA expression is similar to other studies in the literature. Cross-reactivity was focal and faint so is unlikely to interfere. Subset analysis showed contributions to%CV to be higher for HAlow than HAmedium or HAhigh samples as expected. These findings warrant further evaluation of HTI-601 staining on biopsies for PEGPH20-associated therapies in clinical studies. Citation Format: Arnold B. Gelb, Ping Jiang, Laurence Jadin, Daniel C. Maneval, H. Michael Shepard. Development and analytical validation of a novel assay for tissue detection of hyaluronan in the tumor microenvironment to select patients for molecularly targeted pancreatic cancer therapies. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 576. doi:10.1158/1538-7445.AM2015-576

Approximately 50% of de novo acute leukemias have distinctive molecular abnormalities, most frequently chromosomal translocations (1,2). These translocations typically involve genes that are involved in transcription and differentiation (3). As a result of the translocation, these genes are disrupted and the 5’-segment of one gene is joined to the 3’-end of a second gene to form a novel fusion gene, from which chimeric mRNA is transcribed and protein is translated. Inversions occur as well, which also create fusion genes from which novel proteins are generated (1,3). In contrast, many of the known translocations that occur in chronic leukemias and malignant lymphomas affect proto-oncogenes, which are involved in cell proliferation or survival, and an antigen-receptor gene. As a result of the translocation, the proto-oncogene is brought into proximity with the involved antigen receptor gene locus, typically without disrupting the coding region of the proto-oncogene. Under the influence of the antigen-receptor gene enhancers or promoters, the proto-oncogene is constitutively expressed, resulting in increased expression (overexpression) of the normal (nonmutated) protein (1,3). A common theme in such translocations are errors in the normal recombinational mechanisms involving the antigen-receptor genes.

Objectives.—To study the clinicopathologic and molecular genetic findings in posttransplantation lymphoproliferative disorders (PTLDs) following pediatric liver transplantation and to determine the applicability of a recently proposed consensus classification system.Design.—The clinical, pathologic, and molecular genetic findings of 11 PTLDs that occurred in 10 patients are presented. These 10 patients were derived from a group of 121 pediatric patients who underwent liver transplantation at the University of California, San Francisco. The PTLDs were classified using the proposed Society for Hematopathology scheme. Clonality was determined by immunohistochemical detection of monotypic immunoglobulin or by using polymerase chain reaction–based methods to detect monoclonal immunoglobulin heavy-chain gene rearrangements. Epstein-Barr virus (EBV) was detected by immunohistochemistry, in situ hybridization, or polymerase chain reaction. Epstein-Barr virus typing and the presence of LMP1 gene deletions were also analyzed by polymerase chain reaction.Results.—There were 3 early lesions, 4 polymorphic PTLDs, and 4 monomorphic PTLDs. Monoclonality was demonstrated in 8 of 9 cases assessed. Epstein-Barr virus was present in all cases; of 9 cases assessed by polymerase chain reaction, the virus was type A in 8 and type B in 1. No EBV LMP1 gene deletions were identified. The corresponding liver explants were negative for EBV in 8 cases and positive in 1 case. Greater than 3 foci of disease and monomorphic PTLD were associated with decreased actuarial survival (P < .05).Conclusions.—The prognosis of pediatric patients with PTLD is favorable for early lesions and polymorphous PTLD, particularly in patients with localized disease. Multifocal disease and monomorphic PTLD are associated with an unfavorable prognosis.

A 61-year-old man with acquired immunodeficiency syndrome (AIDS) sought care because of the onset of progressive dysphagia. He was found to have a perforated, fungating esophageal mass. The combined histologic and immunologic findings were diagnostic of Hodgkin's disease, nodular sclerosis type, lymphocyte-depleted variant, arising in the esophagus. The Reed-Sternberg cells and mononuclear variants were positive for Epstein-Barr virus (EBV) latent membrane protein (LMP1) and EBV RNA. Occasional small lymphoid cells were also positive for EBV RNA. Polymerase chain reaction studies demonstrated the presence of EBV type A without deletion of the EBV LMP1 gene. Other authors have reported an increased frequency of type B EBV and deletion of the EBV LMP1 gene in cases of human immunodeficiency virus-associated Hodgkin's disease. Hodgkin's disease arising in the esophagus is rare in immunocompetent patients. However, in the presence of AIDS, Hodgkin's disease should be considered in the differential diagnosis of patients with signs or symptoms of esophageal disease.

Solitary fibrous tumors are spindle-cell neoplasms that originally were described in the pleura but that can occur in a large variety of sites. We report a well-circumscribed tumor, apparently involving the renal capsule, clinically thought to be a renal-cell carcinoma or oncocytoma. It was composed of bland spindle-shaped cells with a patchy lymphoplasmacytic infiltrate, suggesting sarcomatoid renal-cell carcinoma, inflammatory myofibroblastic tumor, or solitary fibrous tumor; however, immunohistochemical stains were negative for keratin, alpha-smooth-muscle actin, and desmin but strongly positive for CD34. Ultrastructural examination revealed fibroblast-like cells without myofibroblastic or epithelial differentiation. The combined findings favor a diagnosis of a solitary fibrous tumor involving the renal capsule. To our knowledge, this lesion has not been reported in this location.

The authors analyzed the frequency of immunophenotypic abnormalities in 1,474 cases of routinely fixed, paraffin-embedded B-lineage non-Hodgkin's lymphomas. B-lineage was determined by immunoreactivity for CD20 (L26, 92%); CD45RA (4KB5, an additional 3%) or immunoglobulin (Ig) light chain restriction (remaining 5%). CD45RA was found to be especially helpful on Bouin's-fixed or decalcified tissue and Ig staining was most helpful in plasmacytoid lesions. Coexpression of the T-cell marker CD43 (Leu-22) was the most common immunophenotypic abnormality, seen in 60% of mantle cell lymphomas (MCL), 39% of CLL/small lymphocytic lymphomas, 16% of diffuse large cell lymphomas (DLCL), but only 5% of follicular lymphomas (FL). Antibodies to CD45RO (A6 and UCHL1) and CD3 (polyclonal) were useful in distinguishing infiltrating T cells from B cells coexpressing CD43. Ig light chain restriction was the next commonest immunophenotypic abnormality, which was identified in 67% of plasmacytoid diffuse small cell lymphomas, 43% of MCLs, 35% of monocytoid B-cell lymphomas and 28% of FLs. Overexpression of bcl-2 oncogenic protein was observed in 71% of FLs (n = 96), but not in a control group of reactive follicular hyperplasias (n = 34). Combining two criteria increased the sensitivity of immunodiagnosis in certain circumstances.

Helicobacter pylori infection is a risk factor for gastric adenocarcinoma. We examined whether this infection is also a risk factor for primary gastric non-Hodgkin's lymphoma.This nested case-control study involved two large cohorts (230,593 participants). Serum had been collected from cohort members and stored, and all subjects were followed for cancer. Thirty-three patients with gastric non-Hodgkin's lymphoma were identified, and each was matched to four controls according to cohort, age, sex, and date of serum collection. For comparison, 31 patients with nongastric non-Hodgkin's lymphoma from one of the cohorts were evaluated, each of whom had been previously matched to 2 controls. Pathological reports and specimens were reviewed to confirm the histologic type of the tumor. Serum samples from all subjects were tested for H. pylori IgG by an enzyme-linked immunosorbent assay.Thirty-three cases of gastric non-Hodgkin's lymphoma occurred a median of 14 years after serum collection. Patients with gastric lymphoma were significantly more likely than matched controls to have evidence of previous H. pylori infection (matched odds ratio, 6.3; 95 percent confidence interval, 2.0 to 19.9). The results were similar in both cohorts. Among the 31 patients with nongastric lymphoma, a median of six years had elapsed between serum collection and the development of disease. No association was found between nongastric non-Hodgkin's lymphoma and previous H. pylori infection (matched odds ratio, 1.2; 95 percent confidence interval, 0.5 to 3.0).Non-Hodgkin's lymphoma affecting the stomach, but not other sites, is associated with previous H. pylori infection. A causative role for the organism is plausible, but remains unproved.

Primary myocardial diseases in the pediatric age group encompass a variety of metabolic, infectious, congenital, and acquired disorders, one of which is "histiocytoid cardiomyopathy." We describe clinical and pathologic features in two infants. Autopsy findings in the first case were consistent with sudden cardiac death. The second infant has survived for 2 years on antiarrhythmic therapy with amiodarone. Microscopically, cells with vacuolated to granular cytoplasm were grouped in fascicles, imparting a pseudonodular appearance, but following a distribution reminiscent of conduction fibers. Ultrastructurally, the cells lack a T-tubule system, possess scattered lipid droplets and desmosomes rather than side-to-side junctions, and contain leptomeric fibrils that predominantly marginate to the cell periphery without sarcomeres. Immunostaining of paraffin-embedded tissue reveals perimembranous immunoreactivity for muscle-specific actin, but not for the histiocytic markers CD68 (KP1) and lysozyme. Immunohistochemistry may be a practical alternative when tissue is not saved for electron microscopy. The clinical and pathologic features of this disorder in light of the current literature suggest that it may be hamartoma, possibly of conduction system origin.

Stage has been established as the most important predictor of prognosis in renal cell carcinoma. The predictive value of other morphologic features is less well established. Therefore, in this study we assessed morphologic parameters in 82 Robson stage I renal cell carcinomas. Insufficient numbers of T1 lesions were present in this series to permit a confident comparison of Robson staging with the Union Internationale Contre le Cancer-American Joint Committee on Cancer TNM (tumor, node, metastasis) staging. Morphologic parameters--including size, character of cytoplasm, nuclear grade, and pelvic invasion--were studied. Both univariate survival analysis and multivariate or tree-structured survival analysis (TSSA) were employed, with disease-free survival and actuarial survival as end points. Nuclear grade was the most important predictor of prognosis in stage I neoplasms by TSSA. Nuclear grade greater than 2 correlated with significantly shorter survival (p = 0.018). Stage I tumors measuring less than 5.0 cm by survival analysis (or 6.0 cm by TSSA) were associated with improved disease-free survival (p = 0.040), although TSSA indicated that the effect was pronounced only in low-grade neoplasms. The character of cell cytoplasm was not independent of nuclear grade (Kruskal-Wallis test, p = 0.028). The contingency table indicated disproportionate numbers of grade 4 mixed cell tumors, grade 3 granular cell tumors, and low-grade clear cell tumors. By TSSA, younger patients with low-grade but larger tumors had a poor clinical outcome. Elderly patients with high-grade tumors had the worst overall survival. None of the other clinical parameters or architectural pattern correlated with survival or disease-free survival. When nuclear grade was combined with tumor size and age at diagnosis in a decision tree, patients with stage I neoplasms were separated into favorable, intermediate, and poor prognosis groups.

A case of Langerhans' cell histiocytosis confined to the mediastinum and presenting with de novo superior cava syndrome is reported. The causes of superior vena cava syndrome in childhood are discussed as is the importance of obtaining pathologic diagnosis prior to initiating therapy. © 1993 Wiley-Liss, Inc.

Background: The phosphatidylinositol 3-kinase (PI3K) family of enzymes plays key roles in the transduction of cell signals, and several inhibitors of this pathway are currently in clinical development for the treatment of breast and other cancers. PI3K signaling is activated by growth factor receptors and regulated and terminated by multiple factors including dephosphorylation of the 3′ phosphate of PIP3 by the phosphatase PTEN. Reduced PTEN expression in breast cancer confers susceptibility to inhibitors of the PI3 kinase/Akt pathway. In a subset of breast cancer cases, PTEN protein has been found to be expressed at lower levels (e.g., as compared with normal breast epithelium), however, human breast cancer cells rarely have PTEN mutations. On the other hand PI3K gene mutations have been found in a majority of breast tumors. Methods: To examine the relationship between PTEN protein loss and PI3K gene mutation, PTEN immunoreactivity was assessed semi-quantitatively using histoscores in freshly-cut sections of paraffin blocks from 26 anonymized breast cancer specimens. Sections from the same tumor blocks were used for PI3K mutation detection. One H&E-stained section was used to mark the area of tumor and 2-4 additional unstained sections were macrodissected to enrich for tumor cells, which then underwent DNA extraction. The DNA was analyzed with a commercial PI3K mutation test kit (DxS PI3K Mutation Test Kit, Manchester, UK) that detects 4 somatic mutations in exons 9 and 20 of PIK3CA in three discrete mutation groups or “buckets.” Results: Of the 26 breast cancer specimens examined, 7 (27%) had absence of PTEN protein expression (histoscore=0) and an additional 4 (15%) had a reduction in PTEN expression (histoscore>0 to G), 1 (4%) of E542K (1624 G>A), and 3 (12%) of E545D (1635 G>T) or E545K (1633 G>A). Conclusions: There was a statistically significant correlation between PTEN loss (histoscore

To assess the impact of prostatic involvement with transitional cell carcinoma we reviewed the clinical outcome of 49 patients with transitional cell carcinoma of the prostate. In addition, 115 step-sectioned cystoprostatectomy specimens removed for bladder transitional cell carcinoma were studied to determine the true incidence of secondary prostatic involvement by transitional cell carcinoma. Specimens from 300 prostates removed for prostatic adenocarcinoma also were reviewed to investigate the presence of incidental transitional cell carcinoma arising within the prostate. Transitional cell carcinoma was found in 29% of the step-sectioned specimens and in none of the radical prostatectomy specimens. The presence of prostatic invasion either into the stroma or involving prostatic ducts and acini only had no adverse effect on outcome. Lymph node status and bladder stage, and not prostatic invasion were the determining factors of survival. The presence of seminal vesicle involvement or prostatic stromal invasion appeared to predict for lymph node involvement. With a mean followup of more than 3 years 75% of our patients who had negative lymph nodes and low stage bladder lesions are alive without evidence of disease. In our series prostatic involvement by transitional cell carcinoma did not impact on survival when patients were treated aggressively with radical cystoprostatectomy.

Homozygous adenine phosphoribosyltransferase deficiency is a genetic defect that is associated with 2,8-dihydroxyadenine urolithiasis. Since the prevalence of the heterozygous state is found in 0.4% to 1.2% of the population, it is surprising that more cases of 2,8-dihydroxyadenine urolithiasis have not been reported. Herein we describe a patient with complete adenine phosphoribosyltransferase deficiency with 2,8-dihydroxyadenine urolithiasis leading to chronic renal failure. Gene sequencing revealed that the patient is a compound heterozygote. One of the mutations (a T insertion between bases 346 and 347) has been encountered before, but the second (a G-to-A substitution at base 1356) has not been previously reported. Possible explanations for the unexpected rarity of 2,8-dihydroxyadenine urolithiasis are discussed. (Arch Intern Med. 1993;153:767-770)

Fifty-three low-grade renal cell carcinomas and oncocytomas were reviewed to assess the prognostic significance of nuclear grade (grade 1 versus grade 2), character of cell cytoplasm, size, and stage. Twelve of 46 patients (27%) with follow-up died of disease. Stage 4 disease (most often, extensive local spread) best predicted decreased survival. The survival of patients with stage 1, 2, and 3 tumors was similar; and renal vein invasion was not an adverse prognostic indicator. Size also appeared to predict patient survival; tumors greater than 10 cm metastasized at a higher rate than neoplasms 5 cm or less. We were unable to demonstrate statistically significant differences in survival between individuals with grade 1 and 2 neoplasms, nor were we able to demonstrate a statistically significant difference in survival on the basis of the character of the cell cytoplasm. However, there was a trend toward better survival for patients with granular cell carcinomas and oncocytomas as opposed to neoplasms with clear or mixed cell cytoplasm. Although the number of patients with granular cell carcinomas and oncocytomas is small, the data suggest that the distinction between low-grade granular cell carcinomas and oncocytomas adds little prognostic information.

Morphologic parameters were correlated with survival in 121 renal cortical neoplasms including 116 carcinomas and five oncocytomas. An increasing nuclear grade was generally correlated with a significant decrease in disease-free survival although no statistical difference was found between nuclear Grade 1 and 2 tumors. Similarly, a higher stage at diagnosis predicted a shorter disease-free survival. Renal vein invasion adversely affected prognosis only for high nuclear grade carcinomas. Papillary and spindled carcinomas, independent of nuclear grade, were associated with a significant decrease in disease-free survival compared to tumors with a solid pattern. Patients with large neoplasms (greater than 10 cm) had a significantly worse disease-free survival than patients with tumors 10 cm or less. The prognostic significance of tumor cell type is less clear. Patients with oncocytomas had the best disease-free survival compared with patients with tumors of other cell types. However, the difference in survival was not statistically significant for low-grade tumors, suggesting that nuclear grade rather than cell type may be the more important determinant.

Diffuse small-cell lymphomas of B-lineage comprise a group of immunophenotypically related lymphoid malignancies that display variable clinical aggressiveness. We compared a variety of clinical, pathologic, and immunologic characteristics of 64 B-lineage diffuse small-cell lymphomas to patient survival in an effort to define prognostically relevant subtypes of these neoplasms. Neither clinical parameters nor histological subclassification correlated with patient outcome. In contrast, three immunologic features of these lymphomas showed a statistically significant relationship with actuarial survival. Neoplasms that manifested greater than or equal to 25% Ki-67+ cells (proliferation-associated antigen), less than 25% Leu 4+ cells (pan-T antigen), or less than 15% Leu 3+ cells (helper/inducer T-subset antigen) were associated with significantly decreased patient survival as compared to neoplasms with the reverse phenotype (P = .02, P = .003, P = .0005, respectively). Leu 3 findings were of particular importance in initial biopsies (P = .0007), while the Ki-67 findings were significant regardless of time of biopsy (P = .01 for biopsies at diagnosis and P = .004 for other biopsies). These data indicate that immunologic analysis can demonstrate subsets of diffuse small-cell lymphoma with different biologic potential, and suggest that such analysis be included in the routine work-up of patients with this type of neoplasm.