Christian A. Kunder

Noninvasive monitoring of diffuse large B-cell lymphoma by immunoglobulin high-throughput sequencing

Recent studies have shown limited utility of routine surveillance imaging for diffuse large B-cell lymphoma (DLBCL) patients achieving remission. Detection of molecular disease by immunoglobulin high-throughput sequencing (Ig-HTS) from peripheral blood provides an alternate strategy for surveillance. We prospectively evaluated the utility of Ig-HTS within 311 blood and 105 tumor samples from 75 patients with DLBCL, comparing Ig-HTS from the cellular (circulating leukocytes) and acellular (plasma cell-free DNA) compartments of peripheral blood to clinical outcomes and (18)fluoro-deoxyglucose positron emission tomography combined with computed tomography (PET/CT; n = 173). Clonotypic immunoglobulin rearrangements were detected in 83% of patients with adequate tumor samples to enable subsequent monitoring in peripheral blood. Molecular disease measured from plasma, compared with circulating leukocytes, was more abundant and better correlated with radiographic disease burden. Before treatment, molecular disease was detected in the plasma of 82% of patients compared with 71% in circulating cells (P = .68). However, molecular disease was detected significantly more frequently in the plasma at time of relapse (100% vs 30%; P = .001). Detection of molecular disease in the plasma often preceded PET/CT detection of relapse in patients initially achieving remission. During surveillance time points before relapse, plasma Ig-HTS demonstrated improved specificity (100% vs 56%, P < .0001) and similar sensitivity (31% vs 55%, P = .4) compared with PET/CT. Given its high specificity, Ig-HTS from plasma has potential clinical utility for surveillance after complete remission.

Distinct biological subtypes and patterns of genome evolution in lymphoma revealed by circulating tumor DNA

Circulating tumor DNA reveals patterns of clonal evolution and allows classification of tumor subtypes in lymphoma. The telltale DNA in lymphoma Diffuse large B cell lymphoma is a relatively common type of tumor that can exhibit a wide range of behaviors, from indolent and curable cancers to ones that are very aggressive and difficult to treat. By analyzing DNA in tumor samples and blood of lymphoma patients, Scherer et al. have shown that specific genetic characteristics can determine each tumor’s cell of origin and identify tumors that are going to transform into more aggressive subtypes and may require more intensive treatment. The authors also demonstrated that circulating tumor DNA in the patients’ blood is suitable for this analysis, allowing for periodic monitoring of each patient without repeated invasive biopsies. Patients with diffuse large B cell lymphoma (DLBCL) exhibit marked diversity in tumor behavior and outcomes, yet the identification of poor-risk groups remains challenging. In addition, the biology underlying these differences is incompletely understood. We hypothesized that characterization of mutational heterogeneity and genomic evolution using circulating tumor DNA (ctDNA) profiling could reveal molecular determinants of adverse outcomes. To address this hypothesis, we applied cancer personalized profiling by deep sequencing (CAPP-Seq) analysis to tumor biopsies and cell-free DNA samples from 92 lymphoma patients and 24 healthy subjects. At diagnosis, the amount of ctDNA was found to strongly correlate with clinical indices and was independently predictive of patient outcomes. We demonstrate that ctDNA genotyping can classify transcriptionally defined tumor subtypes, including DLBCL cell of origin, directly from plasma. By simultaneously tracking multiple somatic mutations in ctDNA, our approach outperformed immunoglobulin sequencing and radiographic imaging for the detection of minimal residual disease and facilitated noninvasive identification of emergent resistance mutations to targeted therapies. In addition, we identified distinct patterns of clonal evolution distinguishing indolent follicular lymphomas from those that transformed into DLBCL, allowing for potential noninvasive prediction of histological transformation. Collectively, our results demonstrate that ctDNA analysis reveals biological factors that underlie lymphoma clinical outcomes and could facilitate individualized therapy.

Germinal centre protein HGAL promotes lymphoid hyperplasia and amyloidosis via BCR-mediated Syk activation

The human germinal centre associated lymphoma (HGAL) gene is specifically expressed in germinal centre B-lymphocytes and germinal centre-derived B-cell lymphomas, but its function is largely unknown. Here we demonstrate that HGAL directly binds Syk in B-cells, increases its kinase activity upon B-cell receptor stimulation and leads to enhanced activation of Syk downstream effectors. To further investigate these findings in vivo, HGAL transgenic mice were generated. Starting from 12 months of age these mice developed polyclonal B-cell lymphoid hyperplasia, hypergammaglobulinemia and systemic reactive AA amyloidosis, leading to shortened survival. The lymphoid hyperplasia in the HGAL transgenic mice are likely attributable to enhanced B-cell receptor signalling as shown by increased Syk phosphorylation, ex vivo B-cell proliferation and increased RhoA activation. Overall, our study shows for the first time that the germinal centre protein HGAL regulates B-cell receptor signalling in B-lymphocytes which, without appropriate control, may lead to B-cell lymphoproliferation.

Diagnosis of prostate cancer by desorption electrospray ionization mass spectrometric imaging of small metabolites and lipids

Significance Desorption electrospray ionization mass spectrometry imaging (DESI-MSI) is a label-free molecular imaging technique that provides a window into the biochemical processes present in benign and malignant prostate tissue. This is important both in improving the understanding of tissue biology and in achieving rapid cancer diagnosis. We applied DESI-MSI to record lipid, carbohydrate, and most importantly, small metabolite images from 54 normal and malignant prostate tissue specimens. Several Krebs cycle intermediates were present at different concentrations in prostate cancer compared with normal tissue. Statistical calculations identified panels of metabolites that could readily distinguish prostate cancer from normal tissue with nearly 90% accuracy in a validation set. The results also indicated that the ratio of glucose to citrate ion signals could be used to accurately identify prostate cancer. Accurate identification of prostate cancer in frozen sections at the time of surgery can be challenging, limiting the surgeon’s ability to best determine resection margins during prostatectomy. We performed desorption electrospray ionization mass spectrometry imaging (DESI-MSI) on 54 banked human cancerous and normal prostate tissue specimens to investigate the spatial distribution of a wide variety of small metabolites, carbohydrates, and lipids. In contrast to several previous studies, our method included Krebs cycle intermediates (m/z <200), which we found to be highly informative in distinguishing cancer from benign tissue. Malignant prostate cells showed marked metabolic derangements compared with their benign counterparts. Using the “Least absolute shrinkage and selection operator” (Lasso), we analyzed all metabolites from the DESI-MS data and identified parsimonious sets of metabolic profiles for distinguishing between cancer and normal tissue. In an independent set of samples, we could use these models to classify prostate cancer from benign specimens with nearly 90% accuracy per patient. Based on previous work in prostate cancer showing that glucose levels are high while citrate is low, we found that measurement of the glucose/citrate ion signal ratio accurately predicted cancer when this ratio exceeds 1.0 and normal prostate when the ratio is less than 0.5. After brief tissue preparation, the glucose/citrate ratio can be recorded on a tissue sample in 1 min or less, which is in sharp contrast to the 20 min or more required by histopathological examination of frozen tissue specimens.

Pediatric Plastic Bronchitis: Case Report and Retrospective Comparative Analysis of Epidemiology and Pathology

Plastic bronchitis (PB) is a pathologic condition in which airway casts develop in the tracheobronchial tree causing airway obstruction. There is no standard treatment strategy for this uncommon condition. We report an index patient treated using an emerging multimodal strategy of directly instilled and inhaled tissue plasminogen activator (t-PA) as well as 13 other cases of PB at our institution between 2000 and 2012. The majority of cases (n = 8) occurred in patients with congenital heart disease. Clinical presentations, treatments used, histopathology of the casts, and patient outcomes are reviewed. Further discussion is focused on the epidemiology of plastic bronchitis and a systematic approach to the histologic classification of casts. Comorbid conditions identified in this study included congenital heart disease (8), pneumonia (3), and asthma (2). Our institutional prevalence rate was 6.8 per 100,000 patients, and our case fatality rate was 7%.

Anterolateral congenital diaphragmatic hernia with omphalocele: A case report and literature review

The combination of congenital diaphragmatic hernia (CDH) and omphalocele is quite rare but can be seen in several syndromes. We report on a female newborn with this combination that had not been diagnosed prenatally. The patient suffered respiratory failure that persisted despite intensive care support, suggesting severe secondary pulmonary hypoplasia. Autopsy revealed the combination of an anterolateral CDH and omphalocele in the absence of other anomalies. We believe this to be the first such case to be reported in the literature.

Presence of Even a Small Ground-Glass Component in Lung Adenocarcinoma Predicts Better Survival

Background While lepidic‐predominant lung adenocarcinomas are known to have better outcomes than similarly sized solid tumors, the impact of smaller noninvasive foci within predominantly solid tumors is less clearly characterized. We tested the hypothesis that lung adenocarcinomas with even a small ground‐glass opacity (GGO) component have a better prognosis than otherwise similar pure solid (PS) adenocarcinomas. Patients and Methods The maximum total and solid‐component diameters were determined by preoperative computed tomography in patients who underwent lobar or sublobar resection of clinical N0 adenocarcinomas without induction therapy between May 2003 and August 2013. Survival between patients with PS tumors (0% GGO) or tumors with a minor ground‐glass (MGG) component (1%‐25% GGO) was compared by Kaplan‐Meier and Cox analyses. Results A total of 123 patients met the inclusion criteria, comprising 54 PS (44%) and 69 MGG (56%) whose mean ground‐glass component was 18 ± 7%. The solid component tumor diameter was not significantly different between the groups (2.3 ± 1.2 cm vs. 2.5 ± 1.3 cm, P = .2). Upstaging to pN1‐2 was more common for the PS group (13% [7/54] vs. 3% [2/69], P = .04), but the distribution of pathologic stage was not significantly different between the groups (PS 76% stage I [41/54] vs. MGG 80% stage I [55/69], P = .1). Having a MGG component was associated with markedly better survival in both univariate analysis (MGG 5‐year overall survival 86.7% vs. PS 64.5%, P = .001) and multivariable survival analysis (hazard ratio, 0.30, P = .01). Conclusion Patients with resected cN0 lung adenocarcinoma who have even a small GGO component have markedly better survival than patients with PS tumors, which may have implications for both treatment and surveillance strategies. Micro‐Abstract The impact of having a small ground‐glass opacity component in an otherwise mostly solid lung adenocarcinoma at prognosis was evaluated using Kaplan‐Meier and Cox analyses of 123 patients who underwent lobar or sublobar resection of clinical N0 lung adenocarcinomas without induction therapy. Patients with tumors that had a minor ground‐glass component that comprised less than 25% of the overall tumor size had less pathologic upstaging and far better 5‐year overall survival compared to patients who had a 100% solid cancer.

Indolent thyroid cancer: knowns and unknowns

Thyroid cancer incidence is rapidly increasing due to increased detection and diagnosis of indolent thyroid cancer, i.e. cancer that is likely to be clinically insignificant. Clinical, radiologic, and pathologic features predicting indolent behavior of thyroid cancer are still largely unknown and unstudied. Existing clinicopathologic staging systems are useful for providing prognosis in the context of treated thyroid cancer but are not designed for and are inadequate for predicting indolent behavior. Ultrasound studies have primarily focused on discrimination between malignant and benign nodules; some studies show promising data on using sonographic features for predicting indolence but are still in their early stages. Similarly, molecular studies are being developed to better characterize thyroid cancer and improve the yield of fine needle aspiration biopsy, but definite markers of indolent thyroid cancer have yet to be identified. Nonetheless, active surveillance has been introduced as an alternative to surgery in the case of indolent thyroid microcarcinoma, and protocols for safe surveillance are in development. As increased detection of thyroid cancer is all but inevitable, increased research on predicting indolent behavior is needed to avoid an epidemic of overtreatment.

Predominance of CD4+ T Cells in T-Cell/Histiocyte-Rich Large B-Cell Lymphoma and Identification of a Subset of Patients With Peripheral B-Cell Lymphopenia

Objectives T-cell/histiocyte-rich large B-cell lymphoma (THRLBCL) is a morphologic variant of large B-cell lymphoma whose flow cytometry findings are not well characterized. Methods Nineteen cases with flow cytometric immunophenotyping were identified from the case records of four institutions between 2001 and 2016. Results In most cases, neoplastic B cells were not detected by flow cytometry. Overall, cases showed a predominance of CD4+ T cells, which in some cases was marked. Significant coexpression of CD57 was seen on CD4+ T cells where this marker was analyzed, which correlated with PD-1 expression. Two cases also showed a profound systemic B-cell lymphopenia, which was associated in one case with hypogammaglobulinemia. Conclusions Overall, our work challenges previous findings that cases of THRLBCL are rich in CD8+ T cells and highlights parallels between THRLBCL and nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL). Also, an association of THRLBCL with systemic B-cell lymphopenia has not been previously reported but may represent an underrecognized manifestation.

KLHL6 Is Preferentially Expressed in Germinal Center-Derived B-Cell Lymphomas

Objectives KLHL6 is a recently described BTB-Kelch protein with selective expression in lymphoid tissues and is most strongly expressed in germinal center B cells. Methods Using gene expression profiling as well as immunohistochemistry with an anti-KLHL6 monoclonal antibody, we have characterized the expression of this molecule in normal and neoplastic tissues. Protein expression was evaluated in 1,058 hematopoietic neoplasms. Results Consistent with its discovery as a germinal center marker, KLHL6 was positive mainly in B-cell neoplasms of germinal center derivation, including 95% of follicular lymphomas (106/112). B-cell lymphomas of non-germinal center derivation were generally negative (0/33 chronic lymphocytic leukemias/small lymphocytic lymphomas, 3/49 marginal zone lymphomas, and 2/66 mantle cell lymphomas). Conclusions In addition to other germinal center markers, including BCL6, CD10, HGAL, and LMO2, KLHL6 immunohistochemistry may prove a useful adjunct in the diagnosis and future classification of B-cell lymphomas.