Genetic and Functional Drivers of Diffuse Large B Cell Lymphoma.

Diffuse large B cell lymphoma (DLBCL) is the most common form of blood cancer and is characterized by a striking degree of genetic and clinical heterogeneity. This heterogeneity poses a major barrier to understanding the genetic basis of the disease and its response to therapy. Here, we performed an integrative analysis of whole-exome sequencing and transcriptome sequencing in a cohort of 1,001 DLBCL patients to comprehensively define the landscape of 150 genetic drivers of the disease. We characterized the functional impact of these genes using an unbiased CRISPR screen of DLBCL cell lines to define oncogenes that promote cell growth. A prognostic model comprising these genetic alterations outperformed current established methods: cell of origin, the International Prognostic Index comprising clinical variables, and dual MYC and BCL2 expression. These results comprehensively define the genetic drivers and their functional roles in DLBCL to identify new therapeutic opportunities in the disease.

Postmortem study of organ-specific toxicity in glioblastoma patients treated with a combination of temozolomide, irinotecan and bevacizumab.

PURPOSE: Systemic chemotherapy including monotherapy with temozolomide (TMZ) or bevacizumab (BEV); two-drug combinations, such as irinotecan (IRI) and BEV, TMZ and BEV and a three-drug combination with TMZ, IRI and BEV (TIB) have been used in treating patients with progressive high-grade gliomas including glioblastoma (GBM). Most patients tolerated these regimens well with known side effects of hypertension, proteinuria, and reversible clinical myelosuppression (CM). However, organ- or system- specific toxicities from chemotherapy agents have never been examined by postmortem study. This is the largest cohort used to address this issue in glioma patients. METHODS: Postmortem tissues (from all major systems and organs) were prospectively collected and examined by standard institution autopsy and neuropathological procedures from 76 subjects, including gliomas (N = 68, 44/M, and 24/F) and brain metastases (N = 8, 5/M, and 3/F) between 2009 and 2019. Standard hematoxylin and eosin (H&E) were performed on all major organs including brain specimens. Electronic microscopic (EM) study was carried out on 14 selected subject's kidney samples per standard EM protocol. Medical records were reviewed with adverse events (AEs) analyzed and graded according to the Common Terminology Criteria for Adverse Events (CTCAE), version 4.03. A swimmer plot was utilized to visualize the timelines of patient history by treatment group. The binary logistic regression models were performed to explore any associations between treatment strategies and incident myelosuppression. RESULTS: Twenty-four glioma subjects were treated with TIB [median: 5.5 (range: 1-25) cycles] at tumor recurrence. Exposure to IRI significantly increased the frequency of CM (p = 0.05). No unexpected adverse events clinically, or permanent end-organ damage during postmortem examination was identified in glioma subjects who had received standard or prolonged duration of BEV, TMZ or TIB regimen-based chemotherapies except rare events of bone marrow suppression. The most common causes of death (COD) were tumor progression (63.2%, N = 43) followed by aspiration pneumonia (48.5%, N = 33) in glioma subjects. No COD was attributed to acute toxicity from TIB. The study also demonstrated that postmortem kidney specimen is unsuitable for studying renal ultrastructural pathological changes due to autolysis. CONCLUSION: There is no organ or system toxicity by postmortem examinations among glioma subjects who received BEV, TMZ or TIB regimen-based chemotherapies regardless of durations except for occasional bone marrow suppression and reversible myelosuppression clinically. IRI, but not the extended use of TMZ, significantly increased CM in recurrent glioma patients. COD most commonly resulted from glioma tumor progression with infiltration to brain stem and aspiration pneumonia.

Immunohistochemical expression of vitamin D receptor and forkhead box P3 in classic Hodgkin lymphoma: correlation with clinical and pathologic findings.

BACKGROUND: Expression of forkhead box P3 (FOXP3), a key regulator of T-cell function, in the tumor immune microenvironment is related to survival in classic Hodgkin lymphoma (CHL). Vitamin D receptor (VDR), a transcription factor agonists have been shown to induce FOXP3 expression in T-cells and enhance recruitment of these cells to the inflammatory sites. VDR expression is CHL has been described. However, there is no data on expression of VDR in context of quantity of FOXP3 positive cells in CHL. METHODS: We examined and correlated immunohistochemical expression of VDR and FOXP3 along with clinical and pathology findings in 29 cases of CHL. RESULTS: VDR was expressed in Hodgkin Reed-Sternberg (HRS) cells and background lymphocytes and FOXP3 was expressed in background lymphocytes. 82% of CHL cases, regardless of the subtype, expressed VDR and in majority of the cases, VDR expression was directly proportional to the quantity of FOXP3 expressing lymphocytes in the tumor microenvironment. In cases with higher clinical stage (III/IV), only 28.5% of cases diffusely expressed VDR and FOXP3 compared to 71.4% showing focal positivity. Whereas in cases with lower clinical stages (I/II), the expression pattern of VDR and FOXP3 was almost similar (41.6% diffuse versus 33.3% focal). Interestingly, focal VDR and FOXP3 expression pattern was significantly higher among males. Mixed cellularity cases showed predilection for focal VDR and FOXP3 expression (80% cases); whereas nodular sclerosis subtype had focal and diffuse VDR and FOXP3 expression patterns in similar proportion. Cases with diffuse VDR and FOXP3 expression were less likely to have bone marrow involvement. Epstein Barr virus- encoded small RNA (EBER) positive cases were predominantly focally positive (80%) for VDR and FOXP3. CONCLUSIONS: In summary, quantity of FOXP3 positive T-cells in CHL microenvironment seems to correlate with VDR expression. Clinical stage show a trend of inverse correlation with expression of VDR and quantity of FOXP3 positive T-cells. These findings suggest that VDR could be a possible prognostic and therapeutic target in CHL.

Implantable chemotherapy-loaded silk protein materials for neuroblastoma treatment.

Neuroblastoma is the most common extracranial childhood solid tumor. Treatment of high risk tumors require intense multicycle chemotherapies, resulting in short- and long-term toxicities. Here, we present treatment of an orthotopic neuroblastoma mouse model, with silk fibroin materials loaded with vincristine, doxorubicin or the combination as a intratumoral, sustained release system. The materials, loaded with vincristine with or without doxorubicin, significantly decreased neuroblastoma tumor growth compared to materials loaded without drug or doxorubicin only as well as intravenous (IV) drug treatment. The intratumoral drug concentration was significantly higher with intratumoral delivery versus IV. Furthermore, intratumor delivery decreased the maximum plasma concentration compared to IV delivery, reducing systemic exposure and possibly reduing long-term side effects of chemotherapy exposure. Histopathologically, tumors with remission periods >25 days before recurrence transformed from a "small-round-blue cell" (SBRC) to predominantly "large cell" neuroblastoma (LCN) histopathology, a more aggressive tumor subtype with unfavorable clinical outcomes. These results show that intratumoral chemotherapy delivery may be a treatment strategy for pediatric neuroblastoma, potentially translatable to other focal tumors types. Furthermore, this treatment modality allows for a clinically relevant mouse model of tumor transformation that may be used for studying the phenotypical tumor recurrence and developing more effective treatment strategies for recurrent tumors.

How I manage patients with grey zone lymphoma.

Since grey zone lymphoma (GZL) was originally included in the 2008 World Health Organization classification as a B-cell lymphoma unclassifiable with features intermediate between diffuse large B-cell lymphoma (DLBCL) and classical Hodgkin lymphoma (cHL), new biological and clinical knowledge have been learned. It is important to highlight that diagnosis of this entity is complex and involvement by haematopathologists with expertise in this disease is recommended. It is recognized now that patients with GZL may present clinically with primary mediastinal localization or systemic disease without mediastinal involvement. Regardless of clinical presentation, patients with GZL have relatively high relapse rates, especially compared with primary mediastinal DLBCL or cHL. Interestingly, relapsed/refractory GZL patients appear to be salvaged fairly successfully, especially with haematopoietic stem cell transplantation (HSCT). Off of a clinical trial, we recommend R-CHOP (rituximab, cyclophosphamide, doxorubicin, oncovin, prednisolone) or dose-adjusted EPOCH-R (etoposide, prednisolone, oncovin, cyclophosphamide, doxorubicin, rituximab) for frontline treatment of GZL. Additionally, we advocate use of consolidative radiotherapy for localized and/or bulky disease. For patients with relapsed/refractory GZL, salvage chemotherapy followed by consolidative autologous HSCT should be considered. Finally, continued biological and pathologic examination of this unique disease entity is warranted as well as exploration towards the integration of targeted therapeutic agents (e.g., brentuximab vedotin, programmed cell death 1inhibitors, B-cell receptor inhibitors, proteasome inhibitors, etc.) into the treatment paradigm of GZL.

Dosage of X-linked Toll-like receptor 8 determines gender differences in the development of systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune disease with a high incidence in females and a complex phenotype. Using 564Igi mice, a model of SLE with knock-in genes encoding an autoreactive anti-RNA Ab, we investigated how expression of Toll-like receptors (TLRs) in B cells and neutrophils affects pathogenesis. We established that TLR signaling through MyD88 is necessary for disease. Autoantibody was produced in mice with single deletions of Tlr7, Tlr8, or Tlr9 or combined deletions of Tlr7 and Tlr9. Autoantibody was not produced in the combined absence of Tlr7 and Tlr8, indicating that TLR8 contributes to the break in tolerance. Furthermore, TLR8 was sufficient for the loss of B-cell tolerance, the production of class-switched autoantibody, heightened granulopoiesis, and increased production of type I IFN by neutrophils as well as glomerulonephritis and death. We show that dosage of X-linked Tlr8 plays a major role in the high incidence of disease in females. In addition, we show that the negative regulation of disease by TLR9 is exerted primarily on granulopoiesis and type I IFN production by neutrophils. Collectively, we suggest that individual TLRs play unique roles in the pathogenesis of systemic lupus erythematosus, suggesting new targets for treatment.

BK Virus Nephropathy in Heart Transplant Recipients.

Polyomavirus-associated nephropathy (PVAN) has become an important cause of kidney failure in kidney transplant recipients. PVAN is reported to affect 1% to 7% of kidney transplant recipients, leading to premature transplant loss in approximately 30% to 50% of diagnosed cases. PVAN occurring in the native kidneys of solid-organ transplant recipients other than kidney only recently has been noted. We report 2 cases of PVAN in heart transplant recipients, which brings the total of reported cases to 7. We briefly review the literature on the hypothesized causes of PVAN in kidney transplant recipients and comment on whether these same mechanisms also may cause PVAN in other solid-organ transplant recipients. PVAN should be considered in the differential diagnosis when evaluating worsening kidney function. BK viremia surveillance studies of nonkidney solid-organ recipients should be conducted to provide data to assist the transplantation community in deciding whether regular monitoring of nonkidney transplant recipients for BK viremia is indicated.

High male chimerism in the female breast shows quantitative links with cancer.

Clinical observations suggest that pregnancy provides protection against cancer. The mechanisms involved, however, remain unclear. Fetal cells are known to enter the mother's circulation during pregnancy and establish microchimerism. We investigated if pregnancy-related embryonic/fetal stem cell integration plays a role in breast cancer. A high-sensitivity Y-chromosome assay was developed to trace male allogeneic cells (from male fetus) in females. Fixed-embedded samples (n = 206) from both normal and breast cancer patients were screened for microchimerism. The results were combined with matching clinicopathological and histological parameters and processed statistically. The results show that in our samples (182 informative) more than half of healthy women (56%) carried male cells in their breast tissue for decades (n = 68), while only one out of five in the cancer sample pool (21%) (n = 114) (odds ratio = 4.75, CI at 95% 2.34-9.69; p = 0.0001). The data support the notion that a biological link may exist between chimerism and tissue-integrity. The correlation, however, is non-linear, since male microchimerism in excess ("hyperchimerism") is also involved in cancer. The data suggest a link between hyperchimerism and HER2-type cancers, while decreased chimerism ("hypochimerism") associates with ER/PR-positive (luminal-type) breast cancers. Chimerism levels that correlate with protection appear to be non-random and share densities with the mammary progenitor components of the stem cell lineage in the breast. The results suggest that protection may involve stem/progenitor level interactions and implicate novel quantitative mechanisms in chimerism biology.

Validation of Linear Range HER2/Estrogen Receptor/Progesterone Receptor IHControls for Daily Quality Assurance.

OBJECTIVES: To evaluate a new US Food and Drug Administration (FDA)-cleared immunohistochemistry (IHC) control (IHControls [Boston Cell Standards]) comprising peptide epitopes for HER2, estrogen receptor (ER), and progesterone receptor (PR) attached to cell-sized microspheres and to compare its performance against conventional tissue controls. METHODS: IHControls and tissue/cell line controls for HER2, ER, and PR were compared side by side daily at 5 clinical IHC laboratories for 1 to 2 months. Separately, the sensitivity of the 2 types of controls was evaluated in simulated IHC assay failure experiments by diluting the primary antibody. Additional evaluations included lot-to-lot manufacturing reproducibility of 3 independent lots and specificity against 26 antigenically irrelevant IHC stains. RESULTS: Side-by-side testing revealed a 99.6% concordance between IHControls and tissue controls across 5 IHC laboratories and 766 individual evaluations. Three discordant quality control events were the result of operator error. Simulated assay failure data showed that both IHControls and tissue controls are similarly capable of detecting IHC staining errors. Manufacturing reproducibility of IHControls showed less than 10% variability (coefficient of variation). No cross-reactions were detected from 26 antigenically irrelevant IHC stains. CONCLUSIONS: IHControls, the first FDA-cleared IHC controls, can sensitively and accurately detect IHC assay problems, similar to tissue controls.

Paraneoplastic pemphigus and Castleman's disease in the setting of herpes simplex virus infection.

A 14-year-old girl presented with a 3-week history of mucosal erosions, injected conjunctiva, dehydration, and respiratory distress. She had been treated with intravenous acyclovir for herpes simplex infection with positive herpes simplex virus immunoglobulin M and immunoglobulin G. Physical examination and imaging revealed a large abdominal mass. Incisional biopsy was obtained, and pathology demonstrated angiofollicular hyperplasia with hyalinized germinal centers and Castleman's syndrome-like features. Based on the mucosal erosions, herpes simplex virus serology and positive herpes simplex virus-1 direct fluorescent antibody, Castleman's disease secondary to overwhelming herpes simplex virus infection was the initial impression. The poor response to antivirals and subsequent development of a bullous eruption on the hands resulted in dermatology consultation. Skin biopsy was obtained from a bullae and revealed suprabasilar acantholysis with necrosis as well as upper dermal, perivascular, and interface infiltrate of lymphocytes and eosinophils. No viropathic changes were present. Direct immunofluorescence was significant for immunoglobulin G deposition intercellularly and along the dermoepidermal junction and focal trace C3 deposition along the dermoepidermal junction consistent with paraneoplastic pemphigus, later confirmed by indirect immunofluorescence. We report this case of paraneoplastic pemphigus secondary to Castleman's syndrome confounded by herpes simplex virus-1 positive mucosal erosions.

A Rare Case of Waldenstrom Macroglobulinemia of the Rectosigmoid Colon.

Waldenstrom macroglobulinemia is an uncommon mature B-cell lymphoma characterized by monoclonal immunoglobulin M protein in peripheral blood and lymphoplasmacytic cells in bone marrow and/or extramedullary sites. The gastrointestinal tract is a rare site of involvement. The diagnosis is based on clinicopathologic findings, although somatic mutations, such as MYD88, can aid in the diagnosis. We present a patient with irregular stools diagnosed with Waldenstrom macroglobulinemia involving the rectosigmoid colon by histopathology and immunohistochemistry on colonic biopsies, immunoglobulin M protein in serum, clonal plasma cells in bone marrow, and MYD88 mutation in colonic and bone marrow specimens.

Association between spinal stenosis and wild-type ATTR amyloidosis.

Age-related cardiac amyloidosis results from deposits of wild-type tranthyretin amyloid (ATTRwt) in cardiac tissue. ATTR may play a role in carpal tunnel syndrome (CTS) and in spinal stenosis (SS), indicating or presaging systemic amyloidosis. We investigated consecutive patients undergoing surgery for SS for ATTR deposition in the resected ligamentum flavum (LF) and concomitant risk of cardiac amyloidosis. Each surgical specimen (LF) was stained with Congo red, and if positive, the amyloid deposits were typed by mass spectrometry. Patients with positive specimens underwent standard of care evaluation with fat pad aspirates, serum and urine protein electrophoresis with immunofixation, free light-chain assay, TTR gene sequencing and technetium 99 m-pyrophosphate-scintigraphy. In 2018-2019, 324 patients underwent surgery for SS and 43 patients (13%) had ATTR in the LF with wild-type TTR gene sequences. Two cases of ATTRwt cardiac amyloidosis were diagnosed and received treatment. In this large series, ATTRwt was identified in 13% of the patients undergoing laminectomy for SS. Patients with amyloid in the ligamentum flavum were older and had a higher prevalence of CTS, suggesting a systemic form of ATTR amyloidosis involving connective tissue. Further prospective study of patients with SS at risk for systemic amyloidosis is warranted.

Use of Eculizumab in Transplant-Associated Thrombotic Microangiopathy in a Patient With Polycystic Kidney Disease Immediately Post-Kidney Transplant: A Case Report.

Transplant-associated thrombotic microangiopathy (TMA) in the post-organ transplantation setting occurs from a number of potential inciting factors, such as the use of calcineurin inhibitors, ischemic injury, infections, or antibody-mediated rejection leading to unchecked complement activation and end-organ damage. Delayed recognition of this condition can result in allograft loss. In this case description, we describe the first case of de novo TMA in a patient with polycystic kidney disease that occurred immediately after kidney transplantation. The diagnosis was made promptly on the basis of clinical and laboratory characteristics by a multidisciplinary team and confirmed through kidney biopsy, which showed acute TMA. The patient was successfully managed by replacing tacrolimus with belatacept, which targets cytotoxic T lymphocyte antigen 4, and use of eculizumab, a C5 inhibitor. Eculizumab treatment was discontinued after 3 months of complement inhibition on the patient's request, and relapse of TMA has not been encountered after more than 1 year of follow-up.

Clinicopathologic characteristics of secondary squamous cell carcinoma of head and neck in survivors of allogeneic hematopoietic stem cell transplantation for hematologic malignancies.

The risk of late complications including secondary malignancies is increased in long-term survivors of allogeneic hematopoietic stem cell transplants (HSCT). There is limited literature on the biological behavior and clinical features of squamous cell carcinoma (SCC) of head and neck post-HSCT. We present the clinical and pathologic characteristics on six patients who were diagnosed with SCC while in remission following an allogeneic HSCT. Median follow-up was 8 years. Five patients (83%) developed SCC of tongue and one developed esophageal SCC. Five patients had oral chronic graft-versus-host disease (cGvHD). The conventional risk factors of alcohol, tobacco, and human papillomavirus were absent. The most common presenting finding was the new-onset focal oral pain and ulcerated plaques clinically indistinguishable from a flare of their oral cGvHD lesions. We demonstrated that the SCC in three patients was of donor origin.

Correction: Comparative oncology DNA sequencing of canine T cell lymphoma via human hotspot panel.

[This corrects the article DOI: 10.18632/oncotarget.25209.].

Comparative oncology DNA sequencing of canine T cell lymphoma via human hotspot panel.

T-cell lymphoma (TCL) is an uncommon and aggressive form of human cancer. Lymphoma is the most common hematopoietic tumor in canines (companion animals), with TCL representing approximately 30% of diagnoses. Collectively, the canine is an appealing model for cancer research given the spontaneous occurrence of cancer, intact immune system, and phytogenetic proximity to humans. We sought to establish mutational congruence of the canine with known human TCL mutations in order to identify potential actionable oncogenic pathways. Following pathologic confirmation, DNA was sequenced in 16 canine TCL (cTCL) cases using a custom Human Cancer Hotspot Panel of 68 genes commonly mutated in human TCL. Sequencing identified 4,527,638 total reads with average length of 229 bases containing 346 unique variants and 1,474 total variants; each sample had an average of 92 variants. Among these, there were 258 germline and 32 somatic variants. Among the 32 somatic variants there were 8 missense variants, 1 splice junction variant and the remaining were intron or synonymous variants. A frequency of 4 somatic mutations per sample were noted with >7 mutations detected in MET, KDR, STK11 and BRAF. Expression of these associated proteins were also detected via Western blot analyses. In addition, Sanger sequencing confirmed three variants of high quality (MYC, MET, and TP53 missense mutation). Taken together, the mutational spectrum and protein analyses showed mutations in signaling pathways similar to human TCL and also identified novel mutations that may serve as drug targets as well as potential biomarkers.

CD4+/CD56+ hematodermic neoplasm ("blastic natural killer cell lymphoma"): neoplastic cells express the immature dendritic cell marker BDCA-2 and produce interferon.

CD4+/CD56+ hematodermic neoplasm ("blastic natural killer [NK]-cell lymphoma") is a rare highly aggressive neoplasm associated with cutaneous manifestations followed by dissemination to blood, bone marrow, and other tissues. Neoplastic cells exhibit a lineage-negative CD4+/CD56+/CD43+/HLA-DR+ immunophenotype, initially suggesting an NK-cell derivation. The recent discovery of CD123 antigen expression by tumor cells has provided evidence for a relationship to immature dendritic cells (DCs), a group of myeloid and lymphoid early-committed progenitors capable of differentiating into antigen-presenting DCs. Based on flow cytometric analysis, myeloid DCs represent the majority of human peripheral blood DCs and are positive for blood dendritic cell antigen (BDCA)-1 (or CD1c), CD13, CD11c(high), CD33, and CD123(low). Plasmacytoid DCs are BDCA-2+/ CD123(high)+/CD13-/CD33- and produce interferon (IFN)-alpha when triggered by antigens. IFN-alpha production may be detected in tissue sections using antibodies for myxovirus A (MxA) protein, a surrogate marker. This report describes the clinical, histologic, immunophenotypic, cytogenetic, and molecular genetic findings for 3 cases of CD4+/CD56+ hematodermic neoplasms. In all cases, neoplastic cells were reactive for CD123, BDCA-2, and MxA protein, providing strong evidence for an immature plasmacytoid DC derivation for this rare neoplasm.