Infiltration Of Lymphoplasmacytic Cells Research Articles

A 35-year-old male with Waldenstrom Macroglobulinemia

Waldenstrom macroglobulinemia is one type of non- Hodgkin lymphoma that presents with a wide range of clinical features mainly due to secretion of the IgM paraprotein and infiltration of lymphoplasmacytic cells. Patients with Waldenstrom macroglobulinemia may experience fatigue, unexplained weight loss, enlarged lymph nodes, abdominal swelling, weakness, nose bleeding, blurring of vision etc. Waldenstrom macroglobulinemia is a disease of elderly individuals. The median age at diagnosis is 65 years, with a slight male predominance. We reported a case of a 35-yearold male who presented with fever, abdominal pain, loose motion and epistaxis. Examination revealed cervical and inguinal lymphadenopathy with hepatosplenomegaly. Investigation revealed diffuse lymphocyte infiltration on bone marrow study . Protein electrophoresis showed monoclonal gammopathy with high serum IgM level. On the basis of clinical features and investigation reports finally we diagnosed that, this was a rare case of Waldenstrom macroglobulinemia in a young patient. The patient was transfused 5 units of whole blood and was treated following standard treatment protocol with dexamethasone, Bortezomib and G-CSF. Unfortunately, patient died 3 weeks after initiation of treatment. J Bangladesh Coll Phys Surg 2023; 41(4): 325-328

Determination of Helicobacter heilmannii in cats by real time polymerase chain reaction and histopathology

Helicobacter species such as Helicobacter heilmannii, Helicobacter pylori, Helicobacter felis, Helicobacter bizzozeronii, Helicobacter salomonis have been identified in cats and dogs, and research suggesting that these species may be zoonotic agents have increased in recent years. The aim of this study is to reveal the presence of H. heilmannii in stomach and liver tissues taken during necropsies of domestic, stray and shelter cats by real-time PCR and histopathology and to compare histopathological findings and real-time PCR results. The material of the study consisted of stomach and liver tissues taken from 41 cats who died from different causes and were necropsied. DNA of H. heilmannii was determined in the stomach tissues of 36 cats and the liver tissues of 24 cats in the study conducted with real-time PCR using specific primers of H. heilmannii. Epithelial degeneration in the stomach, fibrosis and oedema in the lamina propria and lymphoplasmacytic cell infiltration were detected histopathologically in the cats diagnosed with gastritis. Eight cats were positive in Hematoxylin-Eosin staining and nineteen cats were positive in the staining with Warthin Starry of the sections, in terms of Helicobacter-like microorganisms. Microscopically, dissociation of the remark cords and hydropic degeneration in hepatocytes and mononuclear cell infiltrations in some sections were determined in the livers. In conclusion, with this study, it was understood that real-time PCR analyses are very useful in determining H. heilmannii in cats. However, histopathological examinations are necessary to associate the presence of bacteria with the development of the disease.

Investigación con PCR en Tiempo Real e Histopatología sobre la presencia de H. felis, H. heilmannii y H. pylori en Perros

Helicobacter species such as H. heilmannii, H. pylori, H. felis, H. bizzozeronii and H. salomonis have been identified in cats and dogs, and research suggesting that these species may be zoonotic agents and has been studied intensified in recent years. The aim of this study was to reveal the presence, comparing the histopathological findings and Real-time PCR results of H. felis, H. heilmannii, and H. pylori in the stomach and liver tissues taken during the necropsies of owned, stray or shelter dogs. The material of the study consisted of stomach and liver tissues taken from 35 dogs that died for different reasons and were brought for necropsy. DNA copies of H. heilmannii were detected by Real-time PCR in the liver samples of 30 dogs using H. heilmannii-specific primers. In the case of gastric samples, Real-time PCR detected H. heilmannii in 13 cases, H. pylori in 3 cases, both H. heilmannii and H. pylori in 13 cases, and H. felis, H. heilmannii and H. pylori in 3 cases. Microscopically, neutrophil leukocyte infiltration, epithelial degeneration, fibrosis and oedema in the lamina propia, and lymphoplasmacytic cell infiltration were determined in the stomachs. In the Hemotoxylin Eosin staining of the sections, 5 cases and 14 cases in the Warthin–Starry staining were found positive for Helicobacter-like microorganisms. Microscopically, dissociation of the remark cords and hydropic degeneration in hepatocytes, and focal mononuclear cell infiltrations in some sections were determined in the livers. In conclusion, with this study, it was understood that Real-time PCR analyzes are very useful in the diagnosis of H. felis, H. heilmannii, and H. pylori. However, histopathological examinations are necessary to associate the presence of bacteria with the development of the disease.

A rare case of Riedel's thyroiditis: Immunoglobulin G4-related thyroid disease

Abstract Introduction Riedel's thyroiditis is a rare thyroiditis in which normal thyroid tissue is replaced by dense fibrous connective tissue. Its etiology is not fully known. An autoimmune process is thought to exist. In this disease group, lymphoplasmocytic tissue infiltration, mostly consisting of IgG4 (+) plasma cells and small lymphocytes, is seen accompanying fibrosis. In our case, we presented a very rare case of Reidel's thyroiditis. Clinical Case We report the case of a 46-year-old woman who came to the general surgery department with an enlargement at the base of her neck. Among the patient's complaints were dysphonia, a feeling of pressure and pain in the neck. She had no history of thyroid disease. Laboratory findings; TSH 0.06 uIU/ml (0.27–4.2), fT3 2.69 pg/mL (2.3–4.5), fT4 1.81 ng/dl (0.93–1.70), crp 73.6 mg/l (0.0–5.0), sedimentation 90 mm/h (0–20), leukocytes 11.74 10^3/ul (4.49–12.68), absolute neutrophil count 9.22 10 ^3/ul (1.9–7.9). Trucut biopsy was performed with the preliminary diagnosis of malignancy due to the history of a mass growing in a short time on the neck of the patient. Pathology result; “Intense fibrosis increase, severe lymphoplasmacytic cell infiltration rich in IgG4 positive cells. Thyroid follicle epithelial cells with ck7, panck, pax8, thyroglobulin, ttf1 positive and cea, ck20, alk negative Intense plasma cells with CD38 have polyclonal staining with kappa and lambda, and IgG4/IgG ratio > 50%. In the background, cd3, pax5 and bcl2 and T cell-rich reactive inflammatory cells are accompanied. Obliterative vascular structures were noted in rust and elastic histochemical stains, and storiform fibrosis was observed in places on the ground. When histopathological and immunohistochemical findings are evaluated together, it is primarily compatible with IgG4-related disease/reidel thyroiditis.'' She was then referred to the endocrinology department. The serum IgG4 level was found to be above normal. She was started on steroid treatment with 64 mg daily methylprednisolone. A weekly control plan was made. In these controls, the steroid dose was gradually reduced. In the follow-up one week later, it was observed that the patient's CRP and sedimentation returned to normal values. There was also a decrease in enlargement in the neck area. In the third month of the treatment, the patient didn't want to continue the treatment. She did not visit our clinic for the next 3 months. Then, she came for the control after three months without treatment. Biochemistry and thyroid function tests were normal. The swelling in the neck area of the patient was completely gone. Conclusion Reidel's thyroiditis is a very rare disease with publications on the basis of case reports in the literature. Therefore, there are no broad-spectrum clinical studies. Clarity on treatment approaches was also not provided. We also benefited from case reports while managing our patient's treatment. We believe that our case will contribute to the literature for the same purpose.

Phaeohyphomycotic cyst.

Phaeohyphomycosis is a term used for a rare opportunistic infection caused by a group of dematiaceous fungi which contains melanin in their cell walls. In 1974, the term phaeohyphomycosis was first coined by Ajello for an entity caused by pigmented fungi.1 Four clinical forms of phaeohyphomycosis exist: i) cutaneous, ii) subcutaneous, iii) systemic, and iv) cerebral. Among these, the subcutaneous form (phaeohyphomycotic cyst) is the most common subtype which usually presents as nodular swelling mainly over the distal extremities, which may be misdiagnosed as epidermal inclusion cyst, ganglion or lipoma. These fungi are present in the soil, where they infect mostly farmers and persons working in fields and farms. It was also highlighted that these infections are seen mostly in immunocompromised individuals and are byproducts of antimicrobial, steroid, and immunosuppressive therapy for various illnesses, including cancer, autoimmune diseases, and transplant cases.1 The pigment giving the characteristic brownish-black appearance to the fungi is melanin, which prevents phagocytosis and hence acts as a virulence factor.2 This group has more than 120 species and 70 genera.3 They have a broad spectrum of clinical manifestations, including superficial and deep fungal infections, sinus involvement, and disseminated forms, including lung and brain abscesses. The incidence ranges from 1-3.1 per 100,000 patients.4 The gold standard method for diagnosis is histopathological examination and culture. Fontana-Masson stain is of immense help in identifying these dematiaceous fungi in tissue as it highlights the melanin pigment in the cell walls. Figure 1 refers to a 59-year-old farmer man who presented with nodular swelling over the dorsum of the left hand for the last 9 months. Initially, the swelling was of peanut size, which gradually progressed to the present size of 5x3x2cm. The lesion was painless, well-defined, and freely mobile. The joint was not affected. He is on medication for type II diabetes mellitus and hypertension. He underwent Whipple's surgery for periampullary carcinoma. He was treated for proximal sensory-motor axonal neuropathy and tuberculosis three years back. Because of isoniazid-induced hepatitis, the patient received a modified anti-tuberculous regime for 9 months, after which he developed gastric ulceration with hematemesis and melena. On endoscopic biopsy, he was found to have chronic active gastritis with Helicobacter pylori infection. In addition, the patient had severe iron deficiency and hypoalbuminemia due to malabsorption. After treatment, he completely recovered at the time of hospital discharge. Figure 1 Phaeohyphomycotic cyst. A - A well-encapsulated and unilocular cyst measuring 4.5x2.2x1.8cm in size (scale bar = 2.5cm); The cyst lumen contains homogenous and translucent gelatinous soft material; B - Multinucleated giant cells showing fungal profile with septate, branching and globose swelling (H&E; x200); C - Periodic Acid-Schiff stain showing bright magenta positivity (PAS stain; x200); D - Fontana Masson stain giving brownish black color due to melanin in the fungal cell walls (x200).: Thus, the nodular swelling was wholly excised and sent for histopathological examination. Grossly, a well-encapsulated mass measuring 4.5x2.2x1.8 cm was submitted for histological analysis (Figure 1A). The external aspect appeared intact, yellowish-white, and congested. A unilocular cyst was identified on serial slicing with a capsular thickness of 0.1-0.2cm. The cut surface was soft in consistency with homogenous and gelatinous translucent material within the lumen. On light microscopy, an outer thick fibrous capsule layer was identified. Just beneath this capsular layer was the vascularized granulation tissue, proliferating fibroblasts, numerous multinucleated foreign and Langhan’s giant cells, and variable lymphoplasmacytic cell infiltrate. Also, abundant basophilic mucoid material with a background of many degenerated cells. These giant cells engulfed pigmented fungal profiles that depicted branching, septate and globose swelling (Figure 1B). Periodic Schiff-Acid stain gave bright magenta color to these fungi (Figure 1C), whereas Fontana Masson stain gave brownish black color due to melanin in the fungal cell walls (Figure 1D). Given the morphology, a diagnosis of a phaeohyphomycotic cyst was rendered. On follow-up, the patient is doing well, and has not received any antifungal agent. The leading treatment choice in non-invasive subcutaneous phaeohyphomycosis is local excision.

OA16 A great imitator of scleroderma

Introduction/BackgroundEosinophilic fasciitis is a rare disease of idiopathic aetiology, which can often mimic scleroderma in presentation and has been associated with haematological diseases. We present a case of eosinophilic fasciitis where the diagnosis was delayed due to the attribution of skin manifestations to scleroderma. This case highlights the importance of careful history and examination to recognize these differential diagnoses, prevent delay and improve outcome.Description/MethodA 69-year-old gentleman was referred to the Rheumatology clinic with suspected diffuse cutaneous systemic sclerosis. He presented with a 9-month history of skin tightening, pain and swelling affecting his legs, finding it difficult to fully extend at the knees, to drive or walk. His symptoms progressed overtime with skin changes spreading to all four extremities, abdomen and back. He reported weight loss of 12Kg associated with early satiety. He also experienced intermittent numbness of his hands. There were no respiratory symptoms, dysphagia or Raynaud’s phenomenon reported. His medical history includes atrial fibrillation and asbestos exposure. Currently on Apixaban 5mg/daily and Bisoprolol 2.5mg/daily.Physical examination revealed diffuse brown hyperpigmentation, erythema and thickened skin on his upper and lower limbs, abdomen and back, it was difficult to crease the skin and there was ‘peau d’orange’ appearance in his forearms. His face and hands were spared. Phalen's and Tinel’s tests were negative.Prior to clinical review he was investigated for paraneoplastic disease: a CT chest, abdomen and pelvis showed an incidental cyst in the thymus and emphysema with no underlying malignancy. Laboratory tests showed normocytic normochromic anaemia with haemoglobin 112 g/L and eosinophilia 1.5 x 109/L, elevated erythrocyte sedimentation rate 62mm/h, CRP 57mg/L, polyclonal hypergammaglobulinaemia IgA 4.3g/L and IgG 24.85g/L. The myeloma screen were negative.The absence of Raynaud’s, and the distribution of skin changes sparing his face, hands and feet, eosinophilia and raised inflammatory markers, made the diagnosis of eosinophilic fasciitis. Histological analysis of a full thickness skin biopsy confirmed patchy lymphoplasmacytic inflammatory cell infiltrate within the fascia and spilling into subcutis, with eosinophils.Subsequently, he was commenced on prednisolone 40mg/daily reducing regimen and Methotrexate 20mg/weekly, with a good response and regression of the swelling and pain. He is now able to walk without difficulty.Discussion/ResultsEosinophilic fasciitis is rare. Symptoms can be similar to those of scleroderma, sometimes considered a variant. The exact aetiology is unknown; however, the literature mentions a potential paraneoplastic, inflammatory, infectious or physical trigger, leading to an abnormal response with accumulation of eosinophils in different tissues. Consequently, this may release transforming growth factor beta to activate fibroblasts, increasing expression of other factors in the connective tissue, resulting in fibrosis.This case highlights that eosinophilic fasciitis present with hardening and thickening of the skin and surrounding tissue similar to that of scleroderma, however, lacks Raynaud’s phenomenon, sclerodactyly. Another striking features from his examination which helped us distinguishing from scleroderma was that his skin changes spare his face, hands and feet.Carpal tunnel syndrome is described in the literature which can progress if untreated.Typical blood tests findings documented in the literature include eosinophilia in 80% of the cases, elevated ESR and C-reactive protein (CRP), polyclonal hypergammaglobulinaemia, are usually present, however, eosinophilia may be transient, therefore normal eosinophil count does not exclude the disease. ANA and Rheumatoid factor can be positive in up to 10 % of the cases.MRI of the extremities to look for thickening of the fascia has been used in other cases, however, it was not required in our case because our Orthopaedics colleagues helped us with performing an urgent full thickness biopsy.Recommended treatment includes high dose steroids and immunosuppressive medication such Methotrexate is most often used for severe or resistant cases.The prognosis is good, however if treatment is delayed patients can develop joint contractures in 85% of patients and skin hardness may persist.In conclusion, careful history and examination is essential in differentiating scleroderma from the scleroderma-mimics especially when there is an atypical distribution, no sclerodactyly and no autoantibodies.Key learning points/Conclusion1. This case suggest that clinicians should be aware that eosinophilic fasciitis typically spare the hands, feet and face.2. Eosinophilia may be transient, therefore normal count does not exclude the disease.3. Recognizing this rare but great imitator of scleroderma set the stage for prompt therapy in either condition and reduce morbidity.

Otologic Manifestations of IgG4-Related Disease: Literature Review and Report of Two Cases

Background: IgG4-related disease (IgG4-RD) is an inflammatory process. The literature on IgG4-RD is rather limited, with mastoid involvement being uncommon. In such cases, presentation may mimic other middle ear and mastoid cavity pathologies. Objective: To summarize findings of patients with IgG4-RD involving the mastoid. Methods: Description of two new cases and summary of findings with previous reports. Results: Nineteen cases of IgG4-RD with mastoid bone involvement were reported in the literature, the earliest appearing at the beginning of the previous decade. Most frequent symptoms included hearing deterioration, tinnitus and otalgia. In 58% of the cases, the process was restricted to otologic manifestations. In 7 out of 19 cases, tissue IgG4 levels were elevated. In all histopathologic samples taken from the operative sites, a dense lymphoplasmacytic cell infiltration was observed. Following definitive pathological diagnosis, the most common treatment was corticosteroids. Generally, the time from onset to final diagnosis was usually more than six months. The treatment is corticosteroids, followed by immunosuppressive agents such as rituximab, cyclophosphamide, and methotrexate. Conclusions: IgG4-RD involving the mastoid is a challenging condition, both in diagnosis and treatment. Hence, IgG4-RD should be included in the differential diagnosis of middle ear pathologies and include a multi-disciplinary team for treatment.

Immunoglobulin-G4-related disease

After years of confusion about apparently distinct clinical disease symptoms, the term IgG4-related disease (IgG4-RD) has been coined in 2001, uniting these fibroinflammatory clinical entities with atendency for tumorous enlargement and tissue fibrosis. Over the past two decades, experimental and clinical studies could make astounding progress in the understanding of this elusive disease. By now, we have areasonable idea of the pathophysiological mechanisms, which opens up new avenues for therapeutic approaches. It seems like adense lymphoplasmacytic cell infiltrate, consisting of B‑cells, IgG4+ plasma cells, follicular T‑helper cells, CD4+ cytotoxic T‑cells and M2 macrophages induces a smoldering inflammatory reaction with afibrogenic cytokine milieu. This stimulates fibroblasts to secrete extracellular matrix components, leading to the histopathologically characteristic storiform fibrosis and obliterative phlebitis. Macroscopically, this reaction results in diffuse organ swelling and tumorous lesions. The macroscopic and histological differentiation from conditions mimicking IgG4-RD can be challenging. This is especially true for granulomatous diseases, such as antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). The situation is further complicated by the fact that ANCAs can be positive in IgG4-RD and, vice versa IgG4 antibodies can be elevated in numerous differential diagnoses, such as infections, AAV, sarcoidosis, and malignancies. This article provides an overview of the multifaceted clinical condition of IgG4-RD with respect to the pathophysiology, diagnostic steps and treatment. Furthermore, an overview of the differential diagnoses is discussed especially with respect to granulomatous diseases.

Calcifying fibrous tumor of the ileum resected by single-port laparoscopic surgery: a case report

BackgroundCalcifying fibrous tumors (CFTs) are rare benign tumors. Because CFTs sometimes relapse, radical resection with adequate margins is necessary. We report a case of ileal CFT resected using single-port laparoscopic surgery.Case presentationA 33-year-old man presented with chief complaints of abdominal pain and vomiting. Computed tomography demonstrated a 45-mm-sized pelvic mass with partial calcification in the ileum. The patient was diagnosed with an ileal tumor, and partial resection of the ileum was performed using the single-port laparoscopic technique. Pathologic findings revealed hypocellular spindle cells with dense hyalinized collagen, interspersed calcification, and infiltration of lymphoplasmacytic cells. Immunohistochemical analysis showed that the factor XIIIa was positive and other tumor-specific markers were negative. Based on these findings, the tumor was finally diagnosed as a CFT.ConclusionsAlthough CFT is benign, multifocal and recurrent CFTs have been reported. Therefore, careful intraperitoneal observation and curative resection are necessary. Single-port laparoscopic surgery is acceptable, both in terms of curability and minimal invasiveness.

Clinicopathological characteristics of IgG4-related lung disease

BackgroundImmunoglobulin G4-related lung disease (IgG4-RLD) is a rare entity. We retrospectively analyzed the clinical and histopathological characteristics of patients with pathologically confirmed IgG4-RLD to improve the diagnosis rate and reduce the risk of misdiagnosis.MethodsWe screened the pathological reports of 4838 patients with pulmonary surgery and/or biopsy specimens from April 2017 to April 2021 at Sun Yat-Sen Memorial Hospital affiliated with Sun Yat-Sen University, and specimens from 65 patients with suspected IgG4-RLD were subjected to immunohistochemical staining for IgG4 and IgG. Finally, 10 patients with definite IgG4-RLD that was pathologically confirmed were enrolled and analyzed.ResultsThe incidence of pathologically confirmed IgG4-RLD was 0.2% (10/4838). The ten patients had an average age of 59.7 years at diagnosis, and the male-to-female ratio was 9:1. The initial clinical manifestations were nonspecific, and cough was the most common symptom (4/10). More than one organ was involved in most patients (8/10), and mediastinal/hilar lymph node involvement was often observed (7/10). Serum IgG4 was analyzed in 6 patients and found to be elevated. Serum tumor marker levels were within the normal range or were slightly elevated. Computed tomography (CT) of the chest and/or 18F-fluorodeoxyglucose positron emission tomography-computed tomography (18F-FDG PET-CT) imaging revealed that 5 patients had a mixed type, 3 patients had the solid nodular type, and 2 patients had the bronchovascular type. All pulmonary masses and large nodules with solid patterns had spiculated margins and inhomogeneous enhancement with or without pleural indentation and a lobulated appearance. Abundant lymphoplasmacytic cell infiltration and fibrosis were observed in all patients. The expression of IgG4 and IgG was upregulated in the pulmonary sections. Seven patients were treated with glucocorticoids with or without additional immunosuppressants and responded well.ConclusionsThe results of our study suggest that multiple imaging findings, an elevated serum IgG4 concentration, and no significant increase in serum tumor biomarkers could provide diagnostic support for IgG4-RLD, especially for isolated IgG4-RLD or IgG4-RLD that includes other organ involvement that does not aid in establishing the diagnosis.

Overexpression of HIF1α in Hunner Lesions of Interstitial Cystitis: Pathophysiological Implications.

The aim of our study was to elucidate biological changes in Hunner lesions, which underlie the pathophysiology of Hunner-type interstitial cystitis, by characterizing their whole transcriptome and immunopathological profiles. Paired bladder mucosal biopsies, 1 sample each from the Hunner lesion and nonlesion area, were obtained from 25 patients with Hunner-type interstitial cystitis. The samples were subjected to whole-transcriptome profiling; immunohistochemical quantification of CD3, CD4, CD8, CD20, CD138, mast cell tryptase, cytokeratin and HIF1α; and quantitative polymerase chain reaction for IFN-α, IFN-β, IFN-γ, TNF, TGF-β1, HIF1α, IL-2, IL-4, IL-6, IL-10 and IL-12A. The results were compared between the lesion and nonlesion areas. RNA sequencing identified 109 differentially expressed genes and 30 significantly enriched biological pathways in Hunner lesions. Up-regulated pathways (24) included HIF1α signaling pathway, PI3K-Akt signaling pathway, RAS signaling pathway and MAPK signaling pathway. By contrast, down-regulated pathways (6) included basal cell carcinoma and protein digestion and absorption. The mRNA levels of HIF1α, IFN-γ and IL-2, and the HIF1α protein level were significantly higher in lesion areas. Otherwise, there were no significant differences between the lesion and nonlesion samples in terms of mRNA levels of inflammatory cytokines or histological features such as lymphoplasmacytic and mast cell infiltration, epithelial denudation and CD4/CD8 T-lymphocyte ratio. Our findings demonstrate significant overexpression of HIF1α and up-regulation of its related biological pathways in Hunner lesions. The results indicate that ischemia, in conjunction with inflammation, plays a pathophysiological role in this subtype of interstitial cystitis/bladder pain syndrome, particularly in Hunner lesions.

Soluble PD-1 Ligands Regulate T-Cell Function in Waldenstrom's Macroglobulinemia

While immune checkpoint molecules contribute to the malignant development of several cancers, including certain types of lymphomas, their potential significance in malignant progression of Waldenstrom's Macroglobulinemia (WM), a low-grade B-cell lymphoma, has not fully been described. The main characteristic of WM is infiltration of lymphoplasmacytic cells in the bone marrow (BM) and increased monoclonal IgM production by malignant cells within BM microenvironment. While next generation sequencing analysis has identified mutations in the MYD88 gene as the driver of WM development, studies by our group have also highlighted a role for the BM microenvironment in pathophysiology of WM and shown that elevated levels of certain cytokines in the BM, such as IL-6 and IL-21, promote both cell proliferation and IgM secretion in WM.In this study, we investigated the impact of cytokines on the immune checkpoint receptor Programmed Death-1 (PD-1) and its ligands PD-L1 and PD-L2 and evaluated how these molecules contribute to malignant progression in WM.Using RT-PCR analysis we have shown that treatment of WM cell lines with IL-21 and IL-6 significantly induce PD-L1 and PD-L2 gene expression, implying that elevated levels of these cytokines within the BM microenvironment could be associated with increased PD-L1 and PD-L2 expression by malignant cells in WM patients. We therefore performed RT-PCR analysis on cells isolated from patients' BM and confirmed increased PD-L1 and PD-L2 expression by both WM and BM cells when compared normal BM cells. In contrast, flow cytometry analysis showed minimal expression of these ligands on the surface of the WM cells, with no significant difference between WM and normal BM cells, indicating that there might be a regulatory mechanism controlling surface expression of these ligands. Further analysis using ELISA and Western Blotting showed that PD-L1 and PD-L2 are found as secreted forms in both BM plasma and peripheral blood serum of the WM patients and their level is significantly higher in WM as compared to normal subjects. Interestingly, in vitro treatment of the WM cell lines by IL-21 and IL-6 increased secreted PD-L1 and PD-L2 in the culture media of the cells, providing an additional line of evidence for elevated PD-L1 and PD-L2 levels in WM patients. Subsequent analysis showed that the secreted form of the PD-L1 and PD-L2 binds to T-cells and induces signals that reduce T-cell proliferation and cell cycle progression, the effects that are associated with a reduction in p-ERK and cyclin A levels.In summary, our data identifies that secreted form of PD-1 ligands is elevated in WM and play an important role in regulating T-cell function. Given the capability of secreted form to be bioactive at distant sites, soluble PD-1 ligands may have potential to promote disease progression. DisclosuresAnsell:Seattle Genetics: Research Funding; Bristol-Myers Squibb: Research Funding; Affimed: Research Funding; Merck: Research Funding; Celldex: Research Funding.

The Bendamustine Plus Rituximab Regimen Is Active and Safe in Previously Untreated Patients with Waldenström Macroglobulinemia, a Study on Behalf of the French Innovative Leukemia Organization (FILO)

BackgroundWaldenström macroglobulinemia (WM) is a rare low-grade B-cell malignancy characterized by the infiltration of IgM-secreting lymphoplasmacytic cells in the bone marrow. There is no standard therapy for WM, but several drugs have been shown to be effective either alone or in combination. Bendamustine (B)-Rituximab (R) has been found to be effective in salvage (Tedeschi , Leukemia Lymphoma 2015) or first setting in MW (Treon, ASH2016).BR- regimen has however reported to be associated with an increase in secondary cancer and the development of secondary myelodysplastic syndrome (MDS) or acute myeloblastic leukemia (AML) in patients with previously treated non-Hodgkin lymphoma. Prolonged neutropenia after R-chemotherapy treatment of lymphoma or CLL has also been associated with an increase of MDS/AML, and a shorter interval between the occurrence of the two malignancies. However, this has not been evaluated in previously untreated WM, nor the possible relationship with secondary cytopenia to WM treatment.MethodsForty-four symptomatic previously untreated patients with WM based on the second WM Workshop recommendations for diagnostic criteria were enrolled in this retrospective multi-center study between January 2012 and January 2017 in ten French centers. MYD88L265P and CXCR4 mutations were searched using q PCR and or next generation sequencing. Del (17) p and/ or TP53 mutation were investigated for 22 patients. All patients received a BR regimen consisting of 375 mg/m2 of rituximab on day 1 and 90 mg/m2 of bendamustine on days 1 and 2, repeated every 4 weeks, with a maximum of 6 cycles. Long-term complications of BR, 6 months or more after treatment discontinuation were analyzed. PFS and OS were plotted using the log rank test and Kaplan-Meier comparison.ResultsThe median age at WM diagnosis was 69 years old (range 45-88) and the median age at initiation of therapy was 70 years old (range 46-89). The MYD88L265Pmutation was present in 24/30 (80%) of the cases. CXCR4 mutations were observed in 2/14 cases, alteration of TP53 in 1/22 cases.The median time from diagnosis to therapy was 2 months (range 0-123 months). The median serum IgM level was 3,870mg/dl (range 650-12000). The criteria for initiating BR treatment were: anemia (54%), B-symptoms (45%), hyper viscosity (25%), adenopathy and/or splenomegaly (18%), thrombocytopenia (16%), neuropathy (11%), and amyloidosis (4%). According to the IPSSMW score, 22% of patients were low risk, 35% intermediate risk, and 43% high risk. Overall 82% of patients completed the planned 6 courses.Using consensus response criteria, the ORR was 95% (CR15%, VGPR 40% PR38% MR 2%) and the major response rate (CR+VGPR+PR) 93%.Toxicities attributed to BR led to dose reduction and/ or incomplete regimen of intended therapy in 18% of patients. Their median age was 78 years old. Toxicities were related to excessive or prolonged myelosuppression in 5 patients and infection in 3.Most of the adverse events were hematological and infectious toxic effects. Grade 3-4 neutropenia occurred in 4 (11%) cases and grade 3-4 thrombocytopenia in one (3%). Minor infections developed in eight cases: three cutaneous infections, two herpes zosters, one upper respiratory tract infection, one sinusitis, and one oral-herpes. Grade 3-4 pneumonia was observed in two cases and sepsis in one.Among 38 evaluable patients for toxicities after BR therapy, 16(42%) developed one or more long-lasting cytopenias: neutropenia (34%), anemia (10%), and thrombocytopenia (8%), all grade I and II.The median duration for prolonged neutropenia, anemia, and thrombocytopenia were 6 months (range: 6-24 months), 7.5 months (range: 6-9 months) and 12 months (6-30 months), respectively.After a median observation time of 27 months (range 5-51 months) after BR-regimen, the PFS and OS were 92% and 100%, respectively. Two patients relapsed at 24 and 45 months. One patient with a history of breast cancer prior to WM treatment relapsed from this solid tumor at 23 months. None of the patients developed aggressive lymphoma or secondary MDS/AML at the last follow-up.ConclusionThe BR regimen is efficient in patients with previously untreated WM. In elderly patients, the dose of bendamustine needs to be lowered. Long-lasting cytopenia occurred in half of patients and resolved in most cases within one year after the end of treatment. Longer follow-up is needed to assess the long-term risks of BR treatment. DisclosuresLaribi:MUNDIPHARMA: Research Funding; NOVARTIS: Honoraria, Research Funding; ROCHE: Research Funding; TEVA: Research Funding; HOSPIRA: Research Funding; AMGEN: Honoraria; TAKEDA: Honoraria, Research Funding. Leblond:SERVIER: Honoraria, Membership on an entity's Board of Directors or advisory committees; GILEAD: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; JANSSEN: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; BMS: Honoraria; ABBVIE: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; ROCHE: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; NOVARTIS: Honoraria, Membership on an entity's Board of Directors or advisory committees.

Glomerulonephritis with severe nephrotic syndrome induced by immune complexes composed of galactose-deficient IgA1 in primary Sj\xf6gren\u2019s syndrome: a case report

BackgroundPrimary Sjögren’s syndrome (pSS) is an auto-immune disease characterized by sialadenitis and dacryoadenitis with lymphoplasmacytic cell infiltration. In pSS, not only sicca symptoms, but also extra-glandular involvement induced by immune abnormalities based on pSS occurs. Renal involvement is one such important life-threatening extra-glandular involvement. Although the aberrant glycosylated IgA in pSS as a product of over-activated B cells is a risk factor of renal involvement, its association has not been clarified. Here we report a case of glomerulonephritis (GN) induced by immune complexes (IC) composed of galactose-deficient IgA1 (Gd-IgA1) in a patient with pSS.Case presentationA 48-year-old Japanese woman with pSS was admitted to our hospital because of a two-month history of nephrotic syndrome. Seven years before she had been diagnosed with pSS from keratoconjunctivitis sicca, elevation of serum anti-Ro/SSA antibody titer and lymphoplasmacytic cell infiltration around salivary ducts of the small salivary glands. Renal biopsy revealed diffuse bubbling appearance in glomerular basement membrane (GBM) with scarce mesangial proliferation. Immunofluorescence showed granular IgA, C3 and Gd-IgA1 staining of GBM. Light chain staining showed no monoclonality. Electron microscopy showed electron dense deposits mainly in the intra-membranous and paramesangial areas and slightly in the subepithelial area. Additional serum analysis confirmed elevation of Gd-IgA1 (13.5 μg/mL), which was comparable with that seen in IgA nephropathy, and qualitative enzyme-linked immunosorbent assay of IgA-containing circulating immune complex (IgA-CIC) was positive. Thus, we diagnosed GN induced by IC composed of Gd-IgA1. Furthermore, retrospectively performed immunofluorescence of the small salivary gland evaluated at the diagnosis of pSS showed positive Gd-IgA1 staining of infiltrating lymphoplasmacytic cells. Therefore, we concluded that Gd-IgA1 produced by over-activated B cells in pSS formed circulating IC and thereby induced GN. After induction therapy with high dose prednisolone and mycophenolate mofetil, the nephrotic syndrome remitted within 3 weeks, the serum Gd-IgA1 level decreased to the normal range (3.8 μg/mL), and serum IgA-CIC disappeared in the 6th month after induction therapy.ConclusionsOur findings clearly demonstrate an association between aberrant glycosylated IgA and the renal involvement seen in pSS, thereby helping to clarify the renal significance of aberrant glycosylated IgA in pSS.

The type of interstitial inflammatory cell infiltrate and the severity of glomerular injury; an underestimated morphologic correlation

Introduction: The interrelation between the type of interstitial inflammatory cells and the severity of glomerulonephritis was not considered in most of the relevant medical literature. Objectives: To investigate the relationship between the type of interstitial cell infiltrate and the morphological severity of glomerular injury in different types of proliferative glomerulonephritides. Patients and Methods: We retrospectively reviewed 138 native kidney biopsies and assessed the relationship between the type of interstitial inflammatory cell infiltrate and the severity of glomerular injury in the form of cellular crescents and fibrinoid necrosis. Results: The predominant type of interstitial inflammatory cell infiltrate was lymphocytic, noted in more than half of the cases. Lymphoplasmacytic inflammatory cell infiltrate was the second most common type which observed. Fifty-five of patients had inflammation in areas of fibrosis. Cellular/fibrocellular crescents were observed in 44% of cases, and fibrinoid necrosis in 30% of cases. As compared to the ‘lymphocytic’ group, patients in the ‘lymphoplasmacytic’ group had ~3 times higher probability of presenting with crescents and fibrinoid necrosis. Conclusion: Our study highlights the significance of morphological correlations that may predict the severity of glomerular injury. Such findings would be helpful in limited or inadequate renal biopsy samples where the pathologist can alert the clinician, in the appropriate clinical context, to the possibility of having crescents and/or necrotizing lesions in the unsampled glomeruli.

18F-FDG PET/CT in Hyalinized Cholecystitis.

Hyalinizing cholecystitis is a rare type of chronic cholecystitis. Moderate patchy transmural especially perivascular lymphoplasmacytic inflammatory cell infiltration is observed in the gallbladder wall. We present the 18F-FDG PET/CT and MRI findings of this rare subtype of chronic cholecystitis. Hyalinizing cholecystitis should be kept in mind in the differential diagnosis of gallbladder wall thickening with intense 18F-FDG uptake.

Mass Cytometry Identifies a Novel Signature for Myeloid-Derived Suppressor-Cells in Waldenstrom's Macroglobulinemia

Myeloid derived suppressor cells (MDSC) are a heterogeneous population of undifferentiated myeloid cells that are expanded and activated in pathological conditions and have the ability to potently suppress T-cell function and thereby contribute to immunosuppression and tumor progression. While there have been studies showing a role for MDSC in a variety of hematological malignancies, no data is available indicating that MDSCs contribute to the tumor progression in Waldenstrom's Macroglobulinemia (WM), an indolent lymphoma characterized by bone marrow (BM) infiltration of lymphoplasmacytic (LPL) cells and increased secretion of monoclonal IgM. In previous work, we have found increased GM-CSF and reduced arginine and cysteine in the BM microenvironment in WM. We hypothesized that this was due to the presence and activity of MDSCs in WM. BM aspirates from WM patients (n=17) were therefore processed to isolate LPL (CD19+/CD138+) cells from the rest of the BM cells (CD19-/CD138-). Sorted (CD19-/CD138-) cells from BM of patients with WM were studied with flow cytometry. Using a sequential gating strategy (lack of lineage markers, low levels of HLADR, CD33+, CD11b+) we identified a population of MDSCs that were then subdivided using CD14 and CD15 expression into total-, monocytic-, or granulocytic- MDSCs (m-MDSC, g-MDSC). We also analyzed unsorted BM cells using cytometry by time-of-flight (CyTOF) in order to further identify and phenotypically characterize the BM MDSC population in a group of WM patients with smoldering (asymptomatic) disease, symptomatic disease, or in remission post-treatment. BM samples from normal subjects were used as a control. Flow cytometry data showed significant higher numbers of MDSC subsets expressing PD-L1 and Arginase1 in WM patients when compared to the normal samples. BM cells from WM patients (n=18) then were compared to controls (n=11), and the absolute number of the total MDSC (p=0.05), m-MDSC (p=0.002), g-MDSC (p=0.02) was increased in WM specimens. When MDSCs from WM or normal monocytes from healthy controls were co-cultured with activated T-cells, the proliferation of activated T-cells in the presence of MDSCs from WM patients was impaired compared to controls, confirming the suppressive role of MDSCs. We then performed high dimensional analysis of the total BM MDSC cells using t-SNEand identified phenotypically distinct MDSC cell populations in the BM that were differentially present when healthy controls were compared to patients with smoldering WM or those with WM needing treatment. Specifically, WM patients needing treatment had increased numbers of a distinct MDSC population that was highly positive for CD163, and CD138. Moreover, conventional markers denoting m-MDSC and g-MDSC, such as CD14 and CD15, were highly expressed in all populations and their pattern of expression did not specifically define the MDSC subtypes, indicating that high dimensional phenotyping further details the MDSC sub-compartments beyond the conventional categorization of MDSC using conventional cytometry. In summary, we find that MDSCs are increased in the BM of WM patients compared to controls. MDSCs expressing CD163 and CD138 increase when WM patients become symptomatic and require therapy. Furthermore, MDSCs in the BM of WM patients suppress T-cell function and likely contribute to progression of the disease. MDSCs in the BM therefore present a therapeutic target that should be explored in WM patients. Disclosures Ansell: Bristol Myers Squibb: Other: research funding for clinical trials; Merck: Other: research funding for clinical trials; AI Therapeutics: Other: research funding for clinical trials; Affimed: Other: research funding for clinical trials; Takeda: Other: research funding for clinical trials; Pfizer: Other: research funding for clinical trials; Regeneron: Other: research funding for clinical trials; Seattle Genetics: Other: research funding for clinical trials.

Immunoglobulin G4-related sclerosing cholangitis.

Immunoglobulin (Ig)G4-related sclerosing cholangitis (IgG4-SC) is a biliary tract manifestation of IgG4-related diseases (IgG4-RD); a subgroup of SC defined as a condition with progressive stenosis and destruction of the bile ducts due to diffuse inflammation and fibrosis. IgG4-SC is clinically characterized by the (a) chronic elevation of cholestatic enzyme levels, (b) significant elevation of serum IgG4 levels, (c) diffuse or segmental narrowing of intra and/or extra hepatic bile ducts with thickening of the bile duct wall in imaging studies, (d) marked lymphoplasmacytic and IgG4-positive plasma cell infiltration and fibrosis in histology, (e) presence of IgG4-RD in other organs, mainly involving autoimmune pancreatitis, and (f) excellent response to corticosteroids. The diagnosis of IgG4-SC is based on a combination of these findings. Although the IgG4-SC diagnosis is different from that of primary sclerosing cholangitis (PSC) or biliary and pancreatic malignancies, it is extremely important to diagnose or suspect IgG4-SC appropriately; as the incorrect diagnosis of PSC or malignant diseases may lead to the progression of fibrosis in patients due to untreated chronic cholestasis, or to unnecessary major surgical resections. Although its etiology remains unclear, recent studies of IgG4-SC have attempted to clarify the roles of the IgG4 molecule and novel autoantibodies detected in patients with IgG4-SC.

PS1305 CHARACTERIZATION OF ENDOGENOUS CXCR4 INHIBITORY PEPTIDES TO TARGET WALDENSTRÖM'S MACROGLOBULINEMIA

Background:Waldenströḿs Macroglobulinemia (WM) is an indolent B‐cell lymphoma with elevated monoclonal IgM serum levels and bone marrow infiltration of lymphoplasmacytic cells. Whole‐genome sequencing has led to identification of two prevalent mutations affecting MYD88 and CXCR4 that allow the classification into 3 genotypes. It was shown that CXCR4 mutations contribute to malignant growth and are associated with Ibrutinib resistance in WM. So far, naturally occurring mechanisms counteracting CXCR4 driven proliferation in cancer are largely unknown. We screened a human blood‐derived peptide library in order to identify novel anti‐cancer peptides that target CXCR4. This approach succeeded in identifying an as‐yet‐unknown CXCR4 blocking peptide called EPI‐X4 (endogenous peptide inhibitor of CXCR4) that is generated naturally in the human body. The 16‐mer peptide is cleaved from its precursor albumin under acidic conditions and was shown to specifically interfere in the cross‐talk between CXCR4 positive cancer cells and their environment. Successful development of highly active and more stable EPI‐X4 derivatives (WSC02 and JM#21) could be of clinical relevance in the future.Aims:Our goal is to analyse the functional role of the naturally occurring EPI‐X4 and to test optimized EPI‐X4 derivatives as potential therapeutic tool in WM.Methods:We performed multiple cell line based functional assays such as apoptosis and migration as well as experiments with EPI‐X4 treated WM cells in vitro and in vivo using NSG mice. Overexpression of CXCR4 mutations was achieved via lentivirus‐mediated infection of WM cells. Whole transcriptome analysis based on next‐generation sequencing was carried out using the Illumina platform and validated via Real‐Time PCR. Proteomic analysis was done using highly sensitive tandem mass spectrometry.Results:We have investigated the effect of EPI‐X4 and its optimized derivatives on blocking the CXCR4 receptor, WM cell migration and inhibition of cell proliferation in presence or absence of different CXCR4 mutations. Efficient and dose depending blocking of CXCR4 by the peptides was documented by two different clones of CXCR4 antibodies, recognizing different epitopes. Both optimized forms of EPI‐X4 were able to abrogate growth and efficiently induced apoptosis in WM cells (up to 90% compared to the control). Migration of WM cells with or without activating CXCR4 mutations was inhibited by 50% for the naturally occurring EPI‐X4 and 95% for the optimized peptides compared to the control. Mice injected with WSC02 treated WM cells showed a prolonged survival of 26 days (median survival control 54,5 days versus 80,5 days for WSC02, respectively). To better understand the impact of EPI‐X4 on the transcriptome, we profiled EPI‐X4 treated cells at the mRNA level using RNA‐seq technique. By performing gene enrichment analysis, genes related to carbohydrate and lipid metabolism pathways as well as the RAF/RAS pathway, hypoxia signal cascades and p53 were enriched. Proteomic changes upon EPI‐X4 treatment on different WM cells are currently being analyzed.Summary/Conclusion:Collectively, our data indicate that naturally occurring mechanisms exist that counteract CXCR4 induced lymphoma growth and that this can be used to develop innovative treatment concepts which target CXCR4 mutations in WM.

Ibrutinib for the treatment of Bing-Neel syndrome, a complication of Waldenström macroglobulinemia: Patient case report

Bing-Neel syndrome is a rare complication of Waldenström macroglobulinemia, characterized by infiltration of lymphoplasmacytic cells to the central nervous system. Multiple treatment modalities exist including purine analogs, bendamustine, high-dose methotrexate, or high-dose cytarabine. Of interest, ibrutinib, a Bruton tyrosine kinase inhibitor has also displayed efficacy in Bing-Neel syndrome. Current literature is limited for the treatment of Bing-Neel syndrome considering its rarity, and while ibrutinib is indicated for the treatment of Waldenström macroglobulinemia, it is utilized off-label for treatment of Bing-Neel syndrome. Additionally, debate exists regarding the recommended dosing strategy for ibrutinib for this indication with disease remission demonstrated at 560 mg and 420 mg. We present a case report that provides additional evidence for this debate with a patient who received 560 mg of ibrutinib initially and maintained disease control despite a dose reduction to 420 mg for tolerability. Ultimately, more data are needed to develop standardized Bing-Neel syndrome treatment strategies with specific consideration to the use of ibrutinib in this condition.