Atypical Lymphoid Proliferation Research Articles

Highly atypical lymphoid proliferation after COVID-19 vaccine, with spontaneous regression.

Highly atypical lymphoid proliferation after COVID-19 vaccine, with spontaneous regression.

Successful Management of Malakoplakia in a Patient Combined Variable Immunodeficiency

Malakoplakia is a rare tumor-like inflammatory condition often affecting the urogenital and gastrointestinal tract of immunocompromised individuals. Malakoplakia is posited to be secondary to impaired macrophages with an inability to phagocyte bacteria completely. The resulting inflammation may trigger CVID-associated enteropathy. Treatment is challenging, often requiring prolonged antibiotics, and reducing immunosuppression.Our patient is a 45-year-old female with heterozygous TNFRSF13B c.310T>C common variable immunodeficiency (CVID). She has granulomatous lymphocytic interstitial lung disease (GILD), CVID-enteropathy in remission, immune thrombocytopenia requiring splenectomy, and chronic norovirus shedding. She was on mycophenolate mofetil, immunoglobulin replacement, and trimethoprim/sulfamethoxazole prophylaxis.She presented with two months history of progressive diarrhea, weight loss, and abdominal pain. Stool calprotectin was 131 uG/G. GI pathogen panel was only positive for Norovirus. Endoscopy showed intraepithelial lymphocytosis and villus blunting (seen on previous scopes). Colonoscopy demonstrated rectal polyps, submucosal atypical lymphoid proliferation, abundant histiocytes with Michaelis-Gutman bodies, positive calcium deposits with Von-Kossa stain, consistent with malakoplakia, with no evidence of malignancy.We started ciprofloxacin 500 milligrams twice daily, Vitamin C, oral immunoglobulin, and probiotics. We continued mycophenolate mofetil to avoid GLILD flares. Diarrhea resolved several weeks after starting ciprofloxacin. However, attempts to wean ciprofloxacin led to recurrence of diarrhea. She completed nine months of ciprofloxacin, after which calprotectin normalized, and repeat scopes showed no evidence of malakoplakia; diarrhea did not recur after stopping ciprofloxacin.We highlight a rare CVID GI complication that successfully responded to a prolonged course of antibiotics. Balancing immune suppression with worsening malakoplakia requires vigilant diagnostic and therapeutic decision-making. Malignancy surveillance will be warranted.

Primary vitreoretinal lymphoma: a case report

Primary vitreoretinal lymphoma (PVRL) is a rare entity and is considered a part of central nervous system lymphoma. PVRL most commonly occurs as diffuse large B cell lymphoma with a high rate of CNS recurrence. We present here a case of a 61-year-old male who initially presented with progressive visual loss. Imaging demonstrated an intraocular soft tissue mass with no other foci. The biopsy was suggestive but not conclusive of B cell lymphoma. The patient proceeded to enucleation. Microscopy showed an atypical lymphoid proliferation with lymphoid follicles and distorted germinal centres.

ODP508 PRIMARY DIFFUSE LARGE B-CELL THYROID LYMPHOMA: A CASE REPORT

Abstract Background Primary thyroid lymphoma (PTL) is a rare disease, accounting for <5% of all thyroid malignancies. Diffuse B-cell lymphoma (DBCL) is the most common type of PTL. The diagnosis and management of PTL depends on the histopathologic results. Clinical Case A 77-year-old female (with T2DM, Hypertension and CAD) had sudden onset of rapidly enlarging neck mass associated with compressive symptoms such as dyspnea, dysphagia and hoarseness of voice. She underwent neck CT scan with contrast revealing a heterogenous mass on right thyroid lobe 9.7×6.9×5.5 cm, with encasement and invasion of adjacent portions of trachea and right common carotid artery . Thyroid function tests were normal and anti-TPO was elevated at 1891.76 IU/ml (NV: < 5.6 IU/mL). Patient underwent tracheostomy and excision biopsy of the mass. Histopathologic report showed atypical lymphoid proliferation and Hashimoto's thyroiditis. Immunohistochemical staining of the neoplastic cells were reactive to CD20, CD3; and Ki67 was positive in 70-90%. Fluorescence in situ hybridization test for c-myc, BCL 6 rearrangement and IGH/BCL 2 fusion for ruling out aggressive double-hit lymphomas came out negative, hence the final diagnosis of Non-Hodgkin's Diffuse Large B-cell Lymphoma. Most PTL originate from B cells, especially DBCL, which accounts for 50–80%. A standard management in PTL is controversial due to limited clinical trials and due to its heterogenous nature. A multidisciplinary approach is the best management technique for PTL. Chemotherapy can cause remission but recent studies showed better response with chemoradiotherapy. The patient was started on R-CVP (rituximab, cyclophosphamide, vincristine, prednisolone). One year after, FDG-PET showed resolution of right thyroid lobe mass with mild diffuse metabolic activity in the rest of thyroid gland, and absence of hypermetabolic lymph nodes. Conclusion In older patients presenting with a rapidly enlarging goiter, especially in a setting of Hashimoto's thyroiditis, the diagnosis of PTL should be considered and differentiated from other types of thyroid disease. The prognosis and management of PTL differ from secondary thyroid lymphoma and other types of neoplasm. It can have a good outcome when recognized in earlier stages and if with less aggressive histopathologic type. Presentation: No date and time listed

A case of B Cell Lymphoma, NOS – A Manifestation in the Labial Mucosa

<h3>Introduction</h3> Non-Hodgkin's lymphoma of B cell lineage is the third most common malignancy of the maxillofacial region. DLBCL, NOS constitutes 25% - 35% of the adult non-Hodgkin's lymphomas and is more common in elderly individuals, as well as more prevalent in males than in females. The etiology of DLBCL, NOS remains unknown. Underlying immunodeficiency is a significant risk factor. <h3>Materials and Methods</h3> We report a case of a 78-year old female patient who presented with a lump in the upper labial mucosa since two weeks, there were no symptoms prior to the stated time duration. The soft tissue mass was located approximately 4mm from the vermillion border, adjacent to the incisal edges of teeth #10 and 11 (left maxillary lateral incisor and canine), measured around 1cm in diameter and was firm on palpation. The clinical differential included benign salivary gland neoplasms, lipoma and a mucocele. The patient had a past medical history of diffuse large B cell lymphoma three years ago. <h3>Results</h3> Microscopic examination showed an atypical lymphoid proliferation involving the soft tissue with infiltration into the adjacent skeletal muscles. A few residual ductular/glandular elements were also noted. The biopsy was composed of a monotonous infiltrate of medium sized cells with round to irregular nuclei, one or more prominent nucleoli and scant cytoplasm. A polymorphous inflammatory background was not observed, and the cells diffusely infiltrated in to the soft tissue. Immunohistochemistry revealed that the atypical cells were positive for CD20 and BCL2 while negative for CD10, BCL6 and c-Myc. Ki-67 proliferation index was moderate to high. <h3>Conclusions</h3> In summary, the overall features were consistent with a B-cell lymphoma that was difficult to further classify due to limited sampling. Based on the relatively high proliferative rate, the lesion was likely consistent with recurrence of the previously diagnosed diffuse large B-cell lymphoma.

EBV-POSITIVE B-CELL ULCERATIVE PROLIFERATION IN THE ORAL CAVITY

<h3>Objective</h3> Lymphoproliferative disorders (LPDs) are a group of conditions characterized by excessive production of lymphocytes manifested by lymphadenopathy, monoclonal lymphocytosis, and bone marrow infiltration. LPDs are often associated with Epstein-Barr virus (EBV) infection. EBV-associated LPDs most commonly appear in the setting of immunocompromised status such as HIV and in transplantation with the extended use of immunosuppressive medications. Individuals with primary immunodeficiency disorders, such as common variable immunodeficiency disease (CVID) or severe combined immunodeficiency, are also at risk of LPD. <h3>Case Summary</h3> A 67-year-old woman complained of mouth ulcerations for 3 months before presentation that were progressively getting worse. Past medical history included CVID, CD4 lymphopenia, rhinosinusitis, eosinophilic asthma, chronic back pain, anxiety, and depression. Medications included immunoglobulin replacement therapy, mepolizumab, montelukast, albuterol, cetirizine, bupropion, clonazepam, fluoxetine, and gabapentin. The patient was allergic to penicillin and azithromycin. Past surgical history was notable for removal of skin melanoma and total knee replacement. Review of systems included a 3-month, 25-lb weight loss, odynophagia, constipation, weakness, and fatigue. Extraoral examination was unremarkable. Intraoral examination revealed 3 × 2 cm² necrotizing palatal ulcerations bilaterally. Necrotizing ulceration was also noted on multiple areas of the gingiva. Differential diagnosis included acute necrotizing ulcerative periodontitis, deep fungal infection (aspergillosis), and lymphoma/LPD. Complete blood count with differential, EBV polymerase chain reaction (PCR), cytomegalovirus PCR and maxillofacial computed tomography (CT) were ordered. The CT scan was unremarkable and laboratory evaluation was negative except for positive EBV PCR. Oral biopsy demonstrated EBV positive B-cell ulcerative proliferation. Positron emission tomography/CT showed widespread fluorodeoxyglucose (FDG)-avid lymphadenopathy. An axillary lymph node biopsy showed atypical lymphoid proliferation in a background of atypical and reactive follicular hyperplasia most consistent with EBV-negative follicular lymphoma, which appeared histologically distinct from the oral EBV-positive LPD. The patient is currently being treated with single-agent rituximab therapy. <h3>Conclusions</h3> EBV may cause oral diseases including infectious mononucleosis, oral hairy leukoplakia, necrotizing ulcerative mucositis, non-Hodgkin's lymphoma, and EBV-related LPD. Persistent oral lesions in patients with CVID could be related to underlying systemic disease. This case is unique because the patient seemingly had follicular lymphoma unrelated to her EBV-positive LPD oral disease, yet in the setting of severe immunosuppression from CVID.

Atypical lymphoid proliferation of the orbit.

Objective: Lymphoproliferative disorders are a group of lesions characterized by abnormal proliferation of lymphocytes. In the orbit, they can occur in the ocular adnexae. These neoplasms have defined clinical and pathologic characteristics and account for more than 20% of all orbital tumors. Several types of lymphoproliferative lesions have been described in the orbit. One example is lymphoid hyperplasia, which commonly involves the lacrimal gland. A benign lesion like lymphoid hyperplasia will show a general normal archetype of the tissues-involved lacrimal gland. We expect a polyclonal group of cells with more or less normal architecture of a follicle. On the other hand, lymphoma will show less organized arrangement of cells, and we expect them to be of monoclonal lineage.Methods: This is a case report of a 55-year-old Filipino female who came in for blurring of vision of both eyes. During her assessment, there was an incidental finding of bilateral upper eyelid swelling, and a 30x15 mm palpable firm mass under the right superior orbital rim and a 30x10 mm mass under the left were noted. The right globe was displaced inferiorly, but no proptosis was seen on exophthalmometry. On plain CT scan, we noted a homogenous mass with molding or contouring around the orbital structures. On coronal view, we noted homogenous masses that mold around the globe and recti, and this also confirmed on axial cuts. A section biopsy was done via anterior orbitotomy.Results: Our patient’s histopath had features of both. On scanning magnification, we noted a very cellular round cell tumor. The round cell lesion seemed to be reminiscent of a germinal center of a lymph node. Around it we noticed the glandular structures, which were expected, since this specimen was from the lacrimal gland. A closer view of the lesion on high power showed these lymphocytes within a germinal center. These are large macrophages that actively phagocytose apoptotic lymphoid cells in germinal centers. We expect to see a lot of them in benign hyperplasia, but only a few will be present in malignancies. In summary, our patient had both benign and malignant features, resulting in a histopath result of atypical lymphoid proliferation.Conclusion: Atypical lymphoid proliferation is a rare orbital tumor with benign and malignant features. There is no standard protocol for treatment, and proper multi-specialty coordination is important. External beam radiation therapy with linear accelerator (LINAC) appears to be an effective treatment, with no recurrence in our patient after 5 months.

Flow Cytometry Analysis of Recurrent or Persistent Lymphadenopathy in Patients with Nodular Lymphocyte-Predominant Hodgkin Lymphoma

Objectives: We recently examined the utility of flow cytometric analysis in the diagnosis of nodular lymphocyte predominant Hodgkin lymphoma (NLPHL) by examining reactive T-cell features. This study aims to compare these features in sequential biopsies of persistent or recurrent lymphadenopathy in patients with NLPHL. Methods: We reanalysed the histopathology and flow cytometry findings of 9 patients with multiple biopsies for persistent or recurrent lymphadenopathy and either initial or recurrent NLPHL. A flow cytometry signature was considered suggestive of NLPHL if ≥12% of T-cells expressed CD57 or ≥3% of T-cells co-expressed CD4 and CD8. Results: A flow cytometry signature considered suggestive of NLPHL was seen in 18 of 20 specimens. Based on histopathology, 11 were diagnosed as NLPHL, 3 were initially underdiagnosed as atypical lymphoid proliferation, and 4 were initially incorrectly diagnosed as negative or progressive transformation of germinal centers. Flow cytometry showed similar expression patterns of CD57 and CD4/CD8 in T-cells between initial and subsequent biopsies. The remaining 2 specimens lacked the flow cytometry signature suggestive of NLPHL and were histopathologically diagnosed as reactive hyperplasia. Conclusion: Flow cytometry analysis based on our criteria is highly sensitive in detecting NLPHL. Correlation with the cytospin cytology may increase the diagnostic specificity. A negative flow essentially ruled out the possibility of NHLPHL.

Allogeneic Stem Cell Transplantation for the Treatment of Lymphoproliferative Disorders in Patients Affected By Primary Immunodeficiencies: The UK Experience

▪Malignancies represent a significant morbidity for patients (pts) affected by primary immunodeficiency (PID). 4-25% of pts with PID will develop a malignancy during the course of their life, lymphoproliferative disorders being the most common. Dysfunctional T-cell surveillance, particularly against EBV, and impaired DNA repair mechanisms lie at the basis of this susceptibility. Malignancies in PID pts are difficult to treat as these pts often harbor co-morbidities that increase the risk of treatment-related toxicity. Allogeneic stem cell transplantation (SCT) offers a treatment for both PID and LPD. Data on the outcome of pts referred to SCT for LPD in PID are scant, as PID are rare and sometimes under diagnosed.We retrospectively collected data on pts 0-18 years referred to SCT for LPD in PID in the 12 pediatric SCT centres in the UK from 2000 to 2015. Centralized review for histopathology specimens and radiological imaging was performed to confirm the LPD diagnosis and the disease staging/remission status at SCT. Ethical approval was obtained.40 patients affected by 13 different PIDs were identified, X-linked lymphoproliferative disease being the most common (15% of the pts).Median age at LPD diagnosis was 6.1 years (range 0.2-17.2). Pts were diagnosed with a B-cell lymphoma (60%), Hodgkin lymphoma (15%), polymorphic LPD (10%), plasmacytic hyperplasia (7%), T-cell lymphoma (5%) or atypical lymphoid proliferation (3%). 83% of the pts presented with stage III/IV disease, 4% had CNS involvement. LPD proved EBER positive in 66% of the tested samples. Pts received a median of 2 lines of chemo/immunotherapy to treat LPD, which proved resistant to front line treatment in 64% of the pts. 7 pts were referred for SCT but did not receive one due to disease progression (3), donor unavailability (1) or medical decision (3). Among the non-transplanted pts, 1 died from sepsis during chemotherapy, 4 died from disease progression; the OS of this group is 28% at a median follow up of 10 months from LPD diagnosis. For the 33 pts who underwent SCT, median age at SCT was 8.4 yrs (range 0.6-17.7). 58% of the pts had 1 or more comorbidities at SCT (chronic lung disease being most common). Of these, 67% of the pts receiving SCT were in complete remission (CR), 24% in partial remission, and 10% had active disease when starting their conditioning regimen.7 different types of conditioning regime were used, as per participating center local policies. In 94% of the cases a reduced intensity conditioning (RIC) was used. 74% of treated pts received in vivo T cell depletion with Alemtuzumab (71%) or Anti-Thymocyte Globulin (3%). 27% of the pts received the graft from a matched related, 39% matched unrelated and 33% mismatched donor. Bone marrow and peripheral blood were the most common sources of stem cells (46% and 42% of the cases, respectively).All pts achieved neutrophil engraftment and secondary graft loss was experienced by 2 pts (6%). 4 pts (12%) died from SCT related toxicity (3 infections, 1 pulmonary VOD/hemorrhage). Acute GVHD grade I-IV developed in 42% of the pts. LPD recurred in 2 cases (1 after secondary graft loss) and was successfully treated with surgery/chemotherapy. With a median follow up of 48 months, 88.0% patients are alive, in CR post SCT. 76% of the evaluable patients exhibited a full donor engraftment at the latest follow up.Disease stage, histology, underlying PID and remission status at SCT did not impact on the patients´ outcomes. Overall survival was significantly higher in SCT recipients as compared to patients who did not receive SCT (88 vs 28%, p= 0.03). Even among the 3 pts who underwent SCT without achieving LPD remission, OS was 67%, thus underlying that restoring competent immunity might play a role in this context, allowing for LPD control post SCT via the newly grafted T-cells.This cohort of pts is the largest described of its kind. We conclude that SCT with RIC is safe and efficacious for pts affected by LPD in PID. While larger cohorts of pts are required to confirm these data, our study suggests that remission status at SCT plays a partial role in LPD control. As a consequence, limited pre-SCT chemotherapy might be required and SCT indication might be plausible for pts not achieving CR. DisclosuresLucchini:Alexion: Membership on an entity's Board of Directors or advisory committees.

S1618 Finding the Needle in the Haystack: PTLD Occurring in the Background of GVHD and Infection

INTRODUCTION: Post-transplant lymphoproliferative disorder (PTLD) is an uncommon complication after hematopoietic stem cell transplant and is associated with Epstein-Barr virus infection and more aggressive immunosuppressive regimens. PTLD can coincide with graft versus host disease (GVHD) and mimic other disorders making endoscopic tissue sampling central to diagnosis and treatment. CASE DESCRIPTION/METHODS: A 27-year-old male was admitted for worsening diarrhea after undergoing umbilical cord blood transplant for Blastic plasmacytoid dendritric cell neoplasm 5 months prior. His transplant course was complicated by several hospitalizations for GVHD of the gut. A colonoscopy 4 months earlier was endoscopically normal with biopsies consistent with GVHD (Figure 1). At this time, the patient presented with 10 days of non-bloody diarrhea in the setting of tapering prednisone dose, similar to prior flares of GVHD. He reported mild chronic abdominal pain but was otherwise asymptomatic. Flexible sigmoidoscopy showed deep linear ulcerations from the proximal rectum to the sigmoid colon with edema, friability, and loss of vascular pattern (Figure 2) which were biopsied. GI PCR returned positive for salmonella and C difficile. A few days later he developed massive hematochezia necessitating ICU level care and massive blood transfusion protocol. Repeat colonoscopy showed interval progression of ulcerations with diffuse bleeding. Biopsies from original flexible sigmoidoscopy showed CD20/EBV + immunoblasts with intermixed CD3+ T cells and polytypic plasma cells (Figure 3) and serum EBV viral load was elevated confirming the diagnosis of PTLD. Patient was started on rituximab and achieved negative viremia after 8 weeks though his course was complicated by recurrent hematochezia. DISCUSSION: The gastrointestinal tract is a common site for PTLD. As patients can present with nonspecific symptoms and confounding diagnoses, identification requires a high index of suspicion. Endoscopic diagnosis through biopsy is critical given the associated high mortality. The characteristic lesion of PTLD in the gastrointestinal tract has been described as a raised, rubbery and erythematous lesion with central ulceration. Although this patient did not have the classic endoscopic finding and had several competing GI diagnoses, he had several risk factors for developing PTLD and once diagnosed responded appropriately to therapy.Figure 1.: First colonoscopy after umbilical cord transplant showed normal endoscopic appearance, random colonic biopsies were consistent with GVHD.Figure 2.: Subsequent flexible sigmoidoscopy showed deep linear ulcerations with edematous mucosa, friability, and loss of vascular pattern in the sigmoid colon just four months later.Figure 3.: H&E stain; 40× field: Atypical lymphoid proliferation as exhibited by numerous large, irregular immunoblasts with prominent nucleoli (examples: black arrows).

S1704 Mantle Cell Lymphoma Presenting as Colonic Polypoid Nodules in a Crohn’s Disease Patient With History of Azathioprine Use

INTRODUCTION: The gastrointestinal (GI) tract is the most common extra nodal site affected by lymphoma, accounting for 5–20% of all cases. Of all GI lymphoma subtypes, mantle cell lymphoma (MCL) is more frequently noted in the ileum and colon. We present a case of MCL presenting as mucosal polypoid nodules in a Crohn's disease patient with history of azathioprine use. CASE DESCRIPTION/METHODS: A 71-year- old male presented to the clinic for management of Crohn’s disease (CD). He was diagnosed at the age of 25 when he developed acute small bowel obstruction secondary to a terminal ileum (TI) stricture and required resection and ileo-colonic anastomosis. He had treatment failure with infliximab due to antibody formation and was managed with azathioprine for over 30 years. Surveillance colonoscopy a year ago showed active ileo-colonic disease and ustekinumab was initiated. Past medical history was remarkable for generalized lymphadenopathy involving the supraclavicular, mediastinal and mesenteric regions. Previously, patient had undergone an excisional lymph node biopsy that was negative for lymphoma. Laboratory testing showed leukocyte count of 5.5 × 109/L with normal differential, hemoglobin of 14.1 g/dL, and a platelet count of 106 L K/uL. Colonoscopy showed scattered erosions and friable mucosa in the entire examined colon and TI. Interestingly, a 2 cm submucosal nodule was noted in the proximal colon that appeared to be a lipoma with mucosal inflammation. Similar small submucosal nodules were noted in the rectum and biopsies were obtained. Pathology of the biopsied nodules revealed atypical lymphoid proliferation. On immunohistochemical staining, B cells were identified, positive for CD 20, CD5, BCL-1, BCL-2, CD43 but negative for CD10 and CD23. The findings were consistent with mantle cell lymphoma. PET scan and bone marrow biopsy revealed disseminated disease with bone marrow involvement. The patient’s immunosuppression was discontinued, and he was referred to oncology for chemotherapy. DISCUSSION: MCL is an aggressive subtypes of B-cell non-Hodgkin lymphoma and most patients have advanced-stage disease at diagnosis. Patients with MCL and GI involvement could be asymptomatic or may present with abdominal pain, obstruction, diarrhea, or overt bleeding. Endoscopic findings may include normal mucosa, nodules, polyps, ulcers, and masses. It is important to biopsy seemingly benign lesions as in our case, if index of suspicion is high. Further history of azathioprine is known to increase lymphoma risk.Figure 1.: A 2 cm smooth submucosal nodule with erythematous mucosal changes noted in the proximal colon.

COMPLETE REMISSION OF BURKITT LYMPHOMA IN A CHILD: A CASE REPORT

Burkitt lymphoma (BL) is a type of non-Hodgkin lymphoma that originates from the malignant transformation of B lymphocytes. A 5-year-old boy presented bilateral swelling between the middle and lower third of the face, with limitation of mouth opening and headache. The computed tomography revealed areas suggestive of a tumor with invasion of the maxillary sinuses and initially of the orbital cavities. Under general anesthesia, an intraoral incisional biopsy was performed. Microscopic analysis revealed atypical lymphoid proliferation, and the immunohistochemical labeling was positive for CD10, CD20, and BCL-6 and in 100% for Ki-67 and negative for BCL-2. The final diagnosis of BL was established. The patient underwent treatment at a referral hospital following the protocol of the Brazilian Society of Pediatric Oncology and remains free of the disease for 24 months. BL is highly aggressive, and the dentist plays an important role in early diagnosis. Acknowledgments: CNPq.

Abstract A23: A second site KrasG12D mutation that impairs PI3K binding rescues embryonic lethality, abrogates myeloproliferative disease, and delays lung tumorigenesis

Abstract Phosphatidylinositol 3-kinase (PI3K) signaling is essential for RAS-driven transformation. To directly investigate the role of oncogenic K-Ras binding to PI3K in development and tumorigenesis, we generated KrasG12D/+,Y64G/+ mice that express, from the endogenous locus, a K-Ras oncoprotein that also contains a “second site” amino acid substitution at tyrosine 64 (Y64G) that disrupts the interaction between oncogenic K-Ras and PI3K. Surprisingly, KrasG12D,Y64G mice are viable, fertile, and phenotypically unremarkable, although they are born at a lower than expected Mendelian frequency (26% versus the expected 50% on a C57BL/6 strain background). As opposed to the enhanced proliferative rate observed in mouse embryonic fibroblasts (MEFs) from K-RasG12D mice, KrasG12D,Y64G MEFs exhibit wild-type rates of proliferation. Detailed analysis of the hematopoietic compartment in KrasG12D,Y64G mice reveals a reduced proportion of long-term hematopoietic stem cells, but no evidence of the aggressive myeloproliferative neoplasm observed when a conditional mutant KrasG12D is activated in hematopoietic cells. Consistent with these observations, bone marrow cells isolated from K-RasG12D,Y64G mice exhibit a normal pattern of myeloid progenitor colony growth in response to cytokine stimulation. In contrast to the lack of observed hematologic malignancy, 100% of KrasG12D,Y64G mice we examined at 1 year of age showed lung lesions, ultimately succumbing to lung tumors with a median survival of 496 days. Despite the ubiquitous KrasG12D,Y64G expression, these mice survive longer than models with mosaic, adenoviral-Cre recombinase-controlled KrasG12D (median survival of 185 days). The majority of the lung lesions that arise in KrasG12D, Y64G mice are low grade, classified pathologically as atypical lymphoid proliferation or papillary adenomas; a few adenocarcinomas are also observed. These studies reinforce the importance of oncogenic KRas-mediated activation of PI3K for transformation and demonstrate that expressing a Y64G amino acid substitution in the context of oncogenic KrasG12D normalizes cell proliferation, rescues embryonic lethality, abrogates myeloid disease, and attenuates lung tumorigenesis. Beyond the bone marrow and lung, this mutant strain is a potent genetic tool for dissecting the role of aberrant PI3K signaling in pancreatic, colon, and other tissues characterized by tumors driven by somatic KRAS mutations, and also have implications for treating human cancers with KRAS mutations. Citation Format: Jasmine C. Wong, Pedro A. Perez-Mancera, Jangkyung Kim, Kuang-Yu Jen, Scott C. Kogan, Ari J. Firestone, Eric A. Collisson, David A. Tuveson, Kevin Shannon. A second site KrasG12D mutation that impairs PI3K binding rescues embryonic lethality, abrogates myeloproliferative disease, and delays lung tumorigenesis [abstract]. In: Proceedings of the AACR Special Conference on Targeting RAS-Driven Cancers; 2018 Dec 9-12; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(5_Suppl):Abstract nr A23.

Abstract IA31: Genetic approaches for testing candidate drug targets for Ras-induced tumorigenesis in vivo

Abstract Despite remarkable progress over the past few years, developing chemical inhibitors that directly and selectively target mutant Ras proteins remains a fundamental challenge. Given this, inhibitors of proteins that function upstream or downstream of oncogenic Ras are a promising alternative therapeutic strategy for RAS mutant tumors. However, the complexity of Ras signaling networks and adaptive mechanisms that result in pathway “rewiring” complicates identifying genes and proteins that are essential for the growth and survival of these cancers. Genetically engineered mice that express mutant Ras alleles from the endogenous locus are a robust system for uncovering pathway dependencies and potential therapeutic vulnerabilities. We have generated novel strains of mice to address two specific questions. First, we collaborated with Kevin Haigis to generate a “second site” C181S substitution within a conditional NrasLSL-G12D mutant allele (Nat Genet 2008;40:600) to ask if palmitoylation is essential for myeloid transformation in vivo. We intercrossed mice carrying this mutation with the Mx1-Cre strain and induced Cre recombinase expression in the hematopoietic compartment by injecting compound mutant pups with polyI:polyC. As previously reported, homozygous NrasLSL-G12D mice uniformly developed an aggressive myeloproliferative disorder (MPD) characterized by markedly elevated blood leukocyte counts, splenomegaly, and death by age 9 months. By contrast, homozygous NrasLSL-G12D,C181S mice remained healthy with no evidence of hematologic disease despite constitutively elevated levels of RasGTP in bone marrow cells that were similar to those observed in NrasLSL-G12D mice. Whereas normal NRas and N-RasG12D proteins are predominantly in the plasma membrane, N-RasG12D, C181S was largely mis-localized in the cytosol. We also generated Mx1-Cre, NrasG12D,C181S/G12D compound heterozygous mutant mice to investigate potential interactions between oncogenic NrasG12D alleles with and without the C181S substitution. Over half of these mice developed aggressive hematologic malignancies, albeit with prolonged latency compared to homozygous NrasG12D mice. Importantly, genetic analysis of diseased tissues revealed reduced frequency or complete absence of the NrasG12D,C181S allele due to secondary somatic genetic events, which indicates strong selective pressure to overcome the growth-suppressive properties of unpalmitoylated N-Ras. Together, these studies validate the palmitoylation/depalmitoylation cycle as a therapeutic target in NRAS mutant hematologic malignancies and establish a novel model system for addressing this question in other cancers. In another ongoing project, we collaborated with Pedro Perez-Macera and David Tuveson to modify a LSL-KrasG12D knock-in allele to introduce a second-site amino acid Y64G substitution that severely impairs binding to PI3 kinase. Surprisingly, mice expressing KrasG12D,Y64G in the germline are viable and phenotypically normal, although they are born at a lower than expected Mendelian frequency. Whereas Mx1-Cre, KrasG12D/+ mice uniformly develop aggressive MPD and die by 4 months of age, KrasG12D,Y64G animals have normal blood counts at 12-18 months of age. However, 100% of KrasG12D,Y64G mice develop lung lesions by 1 year of age, and ultimately succumb from lung tumors with a median survival of 496 days. The majority of these lesions are classified pathologically as atypical lymphoid proliferation or papillary adenomas, with a few mice developing adenocarcinomas. These studies demonstrating that expressing a Y64G amino acid substitution in the context of oncogenic KrasG12D rescues embryonic lethality, abrogates myeloid disease, and attenuates lung tumorigenesis are generally consistent with and also extend elegant studies of Pi3kca mutant mice from the Downward lab (Cell 2007;129:957; Cancer Cell 2013;24:617). Beyond the bone marrow and lung, KrasG12D,Y64G mice are a potent genetic tool for dissecting the role of aberrant PI3 kinase signaling in lung, pancreatic, colon, and other tissues characterized by tumors driven by somatic KRAS mutations, and these data have implications for treating human cancers with KRAS mutations. Citation Format: Kevin M. Shannon. Genetic approaches for testing candidate drug targets for Ras-induced tumorigenesis in vivo [abstract]. In: Proceedings of the AACR Special Conference on Targeting RAS-Driven Cancers; 2018 Dec 9-12; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(5_Suppl):Abstract nr IA31.

Accelerating development of high-risk neuroblastoma patient-derived xenograft models for preclinical testing and personalised therapy

BackgroundPredictive preclinical models play an important role in the assessment of new treatment strategies and as avatar models for personalised medicine; however, reliable and timely model generation is challenging. We investigated the feasibility of establishing patient-derived xenograft (PDX) models of high-risk neuroblastoma from a range of tumour-bearing patient materials and assessed approaches to improve engraftment efficiency.MethodsPDX model development was attempted in NSG mice by using tumour materials from 12 patients, including primary and metastatic solid tumour samples, bone marrow, pleural fluid and residual cells from cytogenetic analysis. Subcutaneous, intramuscular and orthotopic engraftment were directly compared for three patients.ResultsPDX models were established for 44% (4/9) of patients at diagnosis and 100% (5/5) at relapse. In one case, attempted engraftment from pleural fluid resulted in an EBV-associated atypical lymphoid proliferation. Xenogeneic graft versus host disease was observed with attempted engraftment from lymph node and bone marrow tumour samples but could be prevented by T-cell depletion. Orthotopic engraftment was more efficient than subcutaneous or intramuscular engraftment.ConclusionsHigh-risk neuroblastoma PDX models can be reliably established from diverse sample types. Orthotopic implantation allows more rapid model development, increasing the likelihood of developing an avatar model within a clinically useful timeframe.

Rapidly fatal encephalitis associated with atypical lymphoid proliferations of the basal ganglia subsequent to aneurysmal subarachnoid hemorrhage.

Rapidly fatal encephalitis associated with atypical lymphoid proliferations after intracranial aneurysm rupture has not been reported. Here, we describe a 52-year-old female who presented to the emergency department with a severe headache. Imaging demonstrated aneurysmal subarachnoid hemorrhage due to a ruptured left posterior inferior cerebellar artery aneurysm, which was treated with endovascular embolization and subsequent external ventricular drain. She recovered without neurologic sequelae by day seven; however, five weeks later she represented with a severe headache associated with nausea and fever. Initial repeat imaging was unremarkable. She deteriorated quickly and was empirically treated for meningitis despite negative cerebrospinal fluid studies. Magnetic resonance imaging revealed diffuse cerebral edema within the basal ganglia and thalamus. Biopsy of the caudate nuclei revealed atypical lymphoid proliferations. She was treated accordingly with no significant improvement. This case highlights the necessity for a better understanding of the etiology, chronology, and natural history of atypical lymphoid proliferations.

Atypical Lymphoid Proliferations and Clonality in Helicobacter-associated Inflammatory Infiltrates in Children.

Helicobacter infection is considered the major predisposing factor for gastric mucosa-associated lymphoid tissue (MALT) lymphoma with initial infection likely occurring in childhood. Primary gastric MALT lymphoma most commonly occurs in patients older than 50 years which is attributed to the lengthy chronic infection time required before the development of MALT lymphoma. Our study analyzes the histologic features and presence of immunoglobulin heavy chain (IGH) clonality in Helicobacter-associated chronic gastritis (62 cases) and Helicobacter-negative chronic gastritis (17 cases) biopsies within the pediatric population, diagnosed between 1996 and 2018. Helicobacter-associated gastritis was more likely to show active inflammation (P=0.01), with no significant difference in number of germinal centers or the strength, linear property, or depth of the inflammatory infiltrate. In total, 47% (29/62) of the Helicobacter-associated cases had at least 1 lymphoepithelial lesion, equivocal or definitive (a modified Wotherspoon score of 3 to 5), compared with 24% (4/17) of the Helicobacter-negative cases (P=0.5). All cases with lymphoepithelial lesions were assessed for IGH clonality, showing the presence of monoclonality in 27% (8/30) of evaluable cases. None of our patients were diagnosed with gastric lymphoma within available follow-up data. Although 4% of our cases could be considered MALT lymphoma in an adult patient based on prominent lymphoepithelial lesions and IGH monoclonality, caution is advised when diagnosing lymphoma in the pediatric population given the good prognosis of Helicobacter-associated gastritis in this age group. It is unclear if these monoclonal lymphoid proliferations require close follow-up.

3049 Mantle Cell Lymphoma in Routine Screening Colonoscopy

INTRODUCTION: The incidence of colorectal cancer has been on the rise in young and middle aged adults in the United States. While adenocarcinoma remains the most common type of colorectal cancer, lymphoma, particularly mantle cell lymphoma (MCL) is a rare type of lymphoma that can occur in those between the ages of 35-85. We present a case of MCL seen during routine colonoscopy. CASE DESCRIPTION/METHODS: A 51 year old male with recent diagnosis of MCL by oncology was referred to gastroenterology for a screening colonoscopy. During the procedure innumerable large polypoid lesions were visualized in the cecum and ascending colon with similar findings scattered throughout the colon. Biopsies taken from the ascending colon revealed dense atypical lymphoid proliferation with immunohistochemistry confirming MCL. DISCUSSION: MCL comprises about 7% of non-Hodgkin lymphomas. Extranodal manifestations of MCL occurs in 25% of those that are newly diagnosed, with GI involvement estimated to be 15-60%. At the time of diagnosis, most patients present in the advanced stages of MCL. It is associated with poor long term survival and high relapse rates. However, recent advances in potential new therapies for MCL have shown promise. Our case is unusual in that the typical signs of intestinal malaise (abdominal pain, diarrhea, anorexia, and hematochezia) and/or systemic B symptoms were not present. With remission rates being dismal, and treatment strategies based on patient fitness and symptoms, early detection is imperative. It is important to be cognizant to the complaints patients have as it may warrant earlier colonoscopy and therefore diagnosis of colorectal cancer.

Utility of Flow Cytometry in the Diagnostic Evaluation of Recurrent or Persistent Nodular Lymphocyte-Predominant Hodgkin Lymphoma

Abstract Objectives We previously reported that lymph nodes involved by nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) had unique flow cytometric features. To further validate their diagnostic utility, we have examined the flow cytometry features in cases of persistent lymphadenopathy (LAD) prior to the diagnosis of NLPHL or recurrent LAD following diagnosis and treatment for NLPHL. Methods We retrospectively identified nine patients (2-3 specimens per patient for a total of 20 specimens) with persistent or recurrent lymphadenopathy before or after they were diagnosed with NLPHL between the ages of 13 and 76 years. Their histopathology diagnoses were reviewed and flow cytometry data were reanalyzed. Results Based on our published criteria (Am J Clin Pathol 2016;145:107-115), positive flow cytometry findings (at least 12% of T cells expressing CD57 or at least 3% of T cells coexpressing CD4 and CD8) were seen in 18 specimens. Based on histopathology, 11 of them were correctly diagnosed as NLPHL, 3 of them initially underdiagnosed as atypical lymphoid proliferation, and 4 of them initially incorrectly diagnosed as negative or progressive transformation of germinal centers (PTGCs). The flow cytometry studies showed similar expression patterns of CD57, CD4, and CD8 in T cells and very similar high percentages of CD57+ T cells and CD4+CD8+ T cells between initial biopsies and subsequent biopsies in these patients. Negative flow cytometry findings were seen in two specimens pathologically confirmed as negative for NLPHL in two patients after treatment. Their initial diagnostic specimens showed positive flow cytometry findings, different from that seen in the subsequent specimens posttreatment. Conclusion Flow cytometry appears to be a useful adjunct in detecting early or relapsed NLPHL, especially in atypical lesions. The presence of positive flow cytometry features is very sensitive in detecting recurrent or persistent NLPHL, while absence of positive flow cytometry features helps rule out NLPHL.

Spectrum of cytomorphological features of extranodal NK/T-cell lymphoma, nasal type.

Extranodal natural killer/T-cell lymphoma, nasal type (ENKTL) is an aggressive extranodal lymphoma of NK-cell or T-cell lineage. Its clinical features overlap with those of several sinonasal mass lesions. While the histopathological features are well described, diagnosis is often difficult, owing to presence of extensive coagulative necrosis, so that repeated biopsies may sometimes be necessary for correct diagnosis. Literature on cytological findings of ENKTL is limited. Cytomorphological features of cases of histologically confirmed ENKTL having corresponding cytology samples were reviewed retrospectively, to identify distinctive features that could possibly suggest this entity. Aspirates from five patients were studied: four from cervical nodes, one from cheek swelling and one from pleural fluid. Two aspirates were reported as positive for malignancy, two as atypical lymphoid proliferation and one was non-diagnostic. Pleural fluid was reported as malignant, favouring a diagnosis of carcinoma. On cytology, aspirates showed medium to large cells with folded, indented nuclei and abundant pale cytoplasm, some with tongue-like cytoplasmic protrusions. A distinctive feature was presence of large loose clusters of tumour cells with arborising capillaries running through them. Interestingly, necrosis was consistently absent. Subsequent biopsies from palate (three cases) and nasal masses (two cases) confirmed the diagnosis of ENKTL. Suspicion of ENKTL on cytology is crucial for timely diagnosis to avoid diagnostic delay, especially when only highly necrotic biopsy samples are available. Awareness of distinctive cytomorphological features is required to make fine needle aspiration an effective diagnostic tool for initial diagnosis and for evaluation of possible recurrences.