RNA aggregates harness the danger response for potent cancer immunotherapy.

Cancer immunotherapy remains limited by poor antigenicity and a regulatory tumor microenvironment (TME). Here, we create "onion-like" multi-lamellar RNA lipid particle aggregates (LPAs) to substantially enhance the payload packaging and immunogenicity of tumor mRNA antigens. Unlike current mRNA vaccine designs that rely on payload packaging into nanoparticle cores for Toll-like receptor engagement in immune cells, systemically administered RNA-LPAs activate RIG-I in stromal cells, eliciting massive cytokine/chemokine response and dendritic cell/lymphocyte trafficking that provokes cancer immunogenicity and mediates rejection of both early- and late-stage murine tumor models. In client-owned canines with terminal gliomas, RNA-LPAs improved survivorship and reprogrammed the TME, which became "hot" within days of a single infusion. In a first-in-human trial, RNA-LPAs elicited rapid cytokine/chemokine release, immune activation/trafficking, tissue-confirmed pseudoprogression, and glioma-specific immune responses in glioblastoma patients. These data support RNA-LPAs as a new technology that simultaneously reprograms the TME while eliciting rapid and enduring cancer immunotherapy.

Midline brain hamartomatous lesions in fibrodysplasia ossificans progressiva with ACVR1 mutations.

Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disorder characterized by extensive heterotopic ossification of soft tissue structures leading to severe limitations in movement. FOP is caused by a germline mutation in the activating receptor type IA (ACVR1) gene. Worrisome is the fact that up to a third of diffuse intrinsic pontine gliomas (DIPG) also harbor the same point mutation in ACVR1. Radiological reports of central nervous system (CNS) involvement by FOP have described brainstem masses; however, the literature on the histopathology or pathogenesis of these lesions is scant. Here we present detailed neuropathologic findings of a brainstem mass in a patient with FOP and suggest that the tumor is hamartomatous in nature. This report, along with a literature review of radiographic and laboratory data, offers support for the idea that the ACVR1 mutation may incite CNS proliferation, predominantly in the brainstem, but is probably not an oncologic driver. These lesions may be seen at autopsy and are likely noncontributory to death.

Low Frequency of Lymph Node Metastases in Patients in the United States With Early-stage Gastric Cancers That Fulfill Japanese Endoscopic Resection Criteria.

BACKGROUND & AIMS: In the West, early gastric cancer is increasingly managed with endoscopic resection (ER). This is, however, based on the assumption that the low prevalence and risk of lymph node metastases observed in Asian patients is applicable to patients in the United States. We sought to evaluate the frequency of and factors associated with metastasis of early gastric cancers to lymph nodes, and whether the Japanese ER criteria are applicable to patients in the US. METHODS: We performed a retrospective study of 176 patients (mean age 68.5 years; 59.1% male; 58.5% white) who underwent surgical resection with lymph node dissection of T1 and Tis gastric adenocarcinomas, staged by pathologists, at 7 tertiary care centers in the US from January 1, 1999, through December 31, 2016. The frequency of lymph node metastases and associated risk factors were determined. RESULTS: The mean size of gastric adenocarcinomas was 23.0 ± 16.6 mm-most were located in the lower-third of the stomach (67.0%), invading the submucosa (55.1%), and moderately differentiated (31.3%). Lymphovascular invasion was observed in 18.2% of lesions. Overall, 20.5% of patients had lymph node metastases. Submucosal invasion (odds ratio, 3.9; 95% CI, 1.4-10.7) and lymphovascular invasion (odds ratio, 4.6; 95% CI, 1.8-12.0) were independently associated with increased risk of metastasis to lymph nodes. The frequency of lymph node metastases among patients fulfilling standard and expanded Japanese criteria for ER were 0 and 7.5%, respectively. CONCLUSIONS: The frequency of lymph node metastases among patients with early gastric cancer in a US population is higher than that of published Asian series. However, early gastric cancer lesions that meet the Japanese standard criteria for ER are associated with negligible risk of metastasis to lymph nodes, so ER can be recommended for definitive therapy. Expanded criteria cancers appear to have a higher risk of metastasis to lymph nodes, so ER may be considered for select cases.

Gastric Plexiform Fibromyxoma: A Great Mimic of Gastrointestinal Stromal Tumor (GIST) and Diagnostic Pitfalls.

Through a multicenter study, we collected seven cases of gastric plexiform fibromyxoma including four females and three males, 21 to 79 y old (46.1 ± 10.1). All cases showed a unilocular lesion measuring 0.3 to 17 cm (5.3 ± 2.4), arising from antrum (5/7) or body (2/7). Six of the seven cases had intraoperative frozen sections and/or endoscopic ultrasound fine needle aspiration (EUS-FNA), and all of them were preoperatively or intraoperatively diagnosed as gastrointestinal stromal tumor (GIST). EUS-FNA material showed markedly elongated spindle cells with streaming oval to elongated nuclei with rounded ends. Histologically, the tumors exhibited a plexiform growth pattern and were composed of a rich myxoid stroma and cytologically bland uniform spindle cells without mitotic figures, with the exception of one case which displayed nuclear pleomorphism and increased mitosis. Immunostains showed the tumor cells to be focally positive for SMA (6/6), focally and weakly positive for desmin (3/6) and caldesmon (2/3), negative for CD117 (0/7), CD34 (0/7), DOG1 (0/4), and S100 (0/5). No mutations were identified on Next-Generation Sequencing test, and no loss of SDHB immunoreactivity was identified in the tumor with nuclear pleomorphism. One case was treated with Gleevec because of the initial diagnosis of GIST. All patients had a follow-up for up to 11 y, with no tumor recurrence or metastasis reported. Our results suggest that gastric plexiform fibromyxoma is rare and may be underrecognized and misinterpreted as GIST during intraoperative frozen section or preoperative EUS-FNA diagnosis without immunostains leading to inappropriate treatment.

Analysis of immunobiologic markers in primary and recurrent glioblastoma.

Glioblastoma (GBM) generates a varied immune response and understanding the immune microenvironment may lead to novel immunotherapy treatments modalities. The goal of this study was to evaluate the expression of immunologic markers of potential clinical significance in primary versus recurrent GBM and assess the relationship between these markers and molecular characteristics of GBM. Human GBM samples were evaluated and analyzed with immunohistochemistry for multiple immunobiologic markers (CD3, CD8, FoxP3, CD68, CD163, PD1, PDL1, CTLA4, CD70). Immunoreactivity was analyzed using Aperio software. Degree of strong positive immunoreactivity within the tumor was compared to patient and tumor characteristics including age, gender, MGMT promoter methylation status, and ATRX, p53, and IDH1 mutation status. Additionally, the TCGA database was used to perform similar analysis of these factors in GBM using RNA-seq by expectation-maximization. Using odds ratios, IDH1 mutated GBM had statistically significant decreased expression of CD163 and CD70 and a trend for decreased PD1, CTLA4, and Foxp3. ATRX-mutated GBMs exhibited statistically significant increased CD3 immunoreactivity, while those with p53 mutations were found to have significantly increased CTLA4 immunoreactivity. The odds of having strong CD8 and CD68 reactivity was significantly less in MGMT methylated tumors. No significant difference was identified in any immune marker between the primary and recurrent GBM, nor was a significant change in immunoreactivity identified among age intervals. TCGA analysis corroborated findings related to the differential immune profile of IDH1 mutant, p53 mutant, and MGMT unmethylated tumors. Immunobiologic markers have greater association with the molecular characteristics of the tumor than with primary/recurrent status or age.

Increased proportion of FoxP3+ regulatory T cells in tumor infiltrating lymphocytes is associated with tumor recurrence and reduced survival in patients with glioblastoma.

Glioblastoma multiforme (GBM) is an aggressive malignancy associated with profound host immunosuppression mediated in part by FoxP3 expressing regulatory CD4+ T lymphocytes (Tregs) that down-regulate anti-tumor immunity. In order to assess whether FoxP3 was an independent driver differentially expressed in primary versus recurrent GBMs, we stained resected primary and recurrent GBM tumors for CD3, CD4, CD8 and FoxP3 expression using standard immunohistochemistry. Slides were scanned with a high-resolution scanner (ScanScope CS; Aperio), and image analysis software (Aperio ScanScope) was used to enumerate lymphocyte subpopulations allowing for high-throughput analysis and bypassing manual selection bias. As shown in previous studies, enumeration of individual lymphocyte populations did not correlate with clinical outcomes in patients with GBM. However, the CD4+ to regulatory FoxP3+ T cell ratio was diminished in recurrent disease, and increased CD3 and CD8+ to regulatory T cell ratios showed a positive correlation with survival outcomes in primary GBM. These results suggest that while absolute numbers of tumor infiltrating lymphocytes may not be informative for predicting clinical outcomes in patients with GBM, the effective balance of CD3, CD4 and CD8+ T cells to immunosuppressive FoxP3+ regulatory cells may influence clinical outcomes in this patient population.

Validation and implementation of telecytology at an academic medical center using digital cameras and Microsoft Teams software.

INTRODUCTION: Rapid On-Site Evaluation of cytological samples obtained through fine needle aspiration for adequacy is a critical component of a cytology service; however, it imposes a significant time and cost burden for the practicing pathologist and the cytology service. Telecytology enables adequacy assessment by a pathologist remotely, greatly saving time. Telecytology also allows slide preparation and manipulation at the procedure site by an employee with less training requirements, liberating the cytotechnologist to screen cases and perform other laboratory duties - an important aspect to consider during times of cytotechnologist shortages. We propose a telecytology system with a simple setup of a microscope, microscope camera, laptop, and Microsoft Teams software. MATERIALS AND METHODS: We designed a system consisting of a mobile cart, backup battery, microscope, digital camera, and a laptop computer with microscope imaging software and Microsoft Teams software for image transmission. Validation was performed by 4 pathologists making adequacy assessments on randomly selected previously signed out cases using the telecytology system. RESULTS: Our validation of this system demonstrated a greater than 90% concurrence rate between the original adequacy call and the call made by pathologists using the telecytology system - a benchmark used by most, if not all, published validations of similar telecytology systems. In addition, the adequacy assessment concordance rate between select pathologists exceeded 90%. CONCLUSIONS: In summary, our telecytology system provides excellent adequacy services for the clinicians and patients we serve. The Microsoft Teams software is a great tool for transmission of video microscopy. This system will be used with the goal of saving time and increasing efficiency for the cytopathology department.

Cecal Metastasis of Clear Cell Renal Cell Carcinoma After Previous Nephrectomy.

Metastasis of renal cell carcinoma (RCC) to the gastrointestinal (GI) tract is exceedingly rare. We present a case of a man in his 40s with a history of RCC that had metastasized to his abdominal wall and brain who then presented with abdominal pain and melena. On presentation, imaging showed new bone metastases and a colonic mass in the ascending colon. The biopsy of the mass from colonoscopy demonstrated RCC primary. Although rare, this case report highlights the importance of a thorough evaluation of patients with a history of RCC and considers GI tract involvement in those presenting with GI bleeding.

KR158 Spheres Harboring Slow-Cycling Cells Recapitulate High-Grade Glioma Features in an Immunocompetent System.

Glioblastoma (GBM) poses a significant challenge in clinical oncology due to its aggressive nature, heterogeneity, and resistance to therapies. Cancer stem cells (CSCs) play a critical role in GBM, particularly in treatment resistance and tumor relapse, emphasizing the need to comprehend the mechanisms regulating these cells. Also, their multifaceted contributions to the tumor microenvironment (TME) underline their significance, driven by their unique properties. This study aimed to characterize glioblastoma stem cells (GSCs), specifically slow-cycling cells (SCCs), in an immunocompetent murine GBM model to explore their similarities with their human counterparts. Using the KR158 mouse model, we confirmed that SCCs isolated from this model exhibited key traits and functional properties akin to human SCCs. KR158 murine SCCs, expanded in the gliomasphere assay, demonstrated sphere forming ability, self-renewing capacity, positive tumorigenicity, enhanced stemness and resistance to chemotherapy. Together, our findings validate the KR158 murine model as a framework to investigate GSCs and SCCs in GBM pathology, and explore specifically the SCC-immune system communications, understand their role in disease progression, and evaluate the effect of therapeutic strategies targeting these specific connections.

KR158 spheres harboring slow-cycling cells recapitulate GBM features in an immunocompetent system.

Glioblastoma (GBM) poses a significant challenge in clinical oncology due to its aggressive nature, heterogeneity, and resistance to therapies. Cancer stem cells (CSCs) play a critical role in GBM, particularly in treatment-resistance and tumor relapse, emphasizing the need to comprehend the mechanisms regulating these cells. Also, their multifaceted contributions to the tumor-microenvironment (TME) underline their significance, driven by their unique properties. This study aimed to characterize glioblastoma stem cells (GSCs), specifically slow-cycling cells (SCCs), in an immunocompetent murine GBM model to explore their similarities with their human counterparts. Using the KR158 mouse model, we confirmed that SCCs isolated from this model exhibited key traits and functional properties akin to human SCCs. KR158 murine SCCs, expanded in the gliomasphere assay, demonstrated sphere forming ability, self-renewing capacity, positive tumorigenicity, enhanced stemness and resistance to chemotherapy. Together, our findings validate the KR158 murine model as a framework to investigate GSCs and SCCs in GBM-pathology, and explore specifically the SCC-immune system communications, understand their role in disease progression, and evaluate the effect of therapeutic strategies targeting these specific connections.

Primary Hodgkin lymphoma of the central nervous system: two case reports and review of the literature.

CNS involvement by systemic Hodgkin lymphoma (HL) is quite rare, but the disease limited to the CNS is an exceptionally rare entity. The incidence of CNS-HL has been estimated at 0.2-0.5% of cases, but a more recent study has modified that figure to less than 0.02%. Like the conventional form, the diagnosis of primary CNS-HL rests upon distinct morphological and immunohistochemical characteristics, including diagnostic Reed-Sternberg cells, in addition to staging studies demonstrating a lack of disease elsewhere. The paucity of cases in the literature precludes reliable clinical and demographic data, as well as a consensus on treatment and prognosis. We present two cases of primary cerebellar HL, one with 10-year follow-up, and a relevant review of the literature.

Increasing medical student awareness of pathology through a virtual, asynchronous pathology elective.

Faculty from the University of Florida Department of Pathology, Immunology, and Laboratory Medicine developed an asynchronous, fully virtual pathology elective for medical students that emphasizes both foundational pathology concepts as well as the role of pathologists in the broader health system. The program includes ten core modules as well as several selective modules which allows students to tailor their coursework to better align with their desired specialty. After completing each module, students were required to concisely summarize the topic in the form of a 280-character tweet. Students were surveyed immediately after finishing the course and again one year after finishing the course to assess the effectiveness of the course in teaching students about pathology. Survey results showed significant improvement in student knowledge about the field of pathology and high levels of satisfaction with the course content and delivery. The tweet summaries required in the course provided a unique challenge for students. Although the course was initially developed in response to the COVID-19 pandemic, enrollment has continued steadily through 2023. Our study suggests that online pathology electives can be an effective way to increase medical student exposure to pathology and facilitate learning about the field of pathology, especially among students who may not ultimately pursue a career in pathology.

A Case of Statin-Associated Autoimmune Myopathy: Management and Treatment.

Elevated lipid panels are associated with an increased risk of cardiovascular disease. Management of heart disease with lipid lowering agents play a vital role in medicine. Statins are one group of medications that are widely utilized in the medical field to decrease the risk of atherosclerotic disease. Statins work by inhibiting the hepatic enzyme 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR). Although statins are one of the most effective drugs for secondary and primary prevention of heart disease, they are not without risks and side effects such as hepatotoxicity and myopathy. We present a case of a male patient who developed progressively worsening muscle weakness and elevated muscle enzyme markers upon initiation of a statin. His symptoms persisted despite a trial of an alternative statin and subsequent discontinuation of all statin medications. A multitude of possible etiologies were considered and ranged from infectious, autoimmune, cancerous, to congenital in nature. Environmental factors, such as exposure to medications or toxins, were also considered as one of the possible precipitating factors. The association between his statin consumption and muscle weakness were not easily apparent at first. He required further workup including physical examination, electromyography, panel of myositis antibodies, and muscle biopsy. After clinical suspicion and elevated antibodies to HMGCR beyond the normal limit, he was discovered to have statin-associated autoimmune myopathy. The patient improved with the treatment of immunosuppressive agent's prednisone and methotrexate.

Metabolomic and Lipidomic Characterization of Meningioma Grades Using LC-HRMS and Machine Learning.

Meningiomas are among the most common brain tumors that arise from the leptomeningeal cover of the brain and spinal cord and account for around 37% of all central nervous system tumors. According to the World Health Organization, meningiomas are classified into three histological subtypes: benign, atypical, and anaplastic. Sometimes, meningiomas with a histological diagnosis of benign tumors show clinical characteristics and behavior of aggressive tumors. In this study, we examined the metabolomic and lipidomic profiles of meningioma tumors, focusing on comparing low-grade and high-grade tumors and identifying potential markers that can discriminate between benign and malignant tumors. High-resolution mass spectrometry coupled to liquid chromatography was used for untargeted metabolomics and lipidomics analyses of 85 tumor biopsy samples with different meningioma grades. We then applied feature selection and machine learning techniques to find the features with the highest information to aid in the diagnosis of meningioma grades. Three biomarkers were identified to differentiate low- and high-grade meningioma brain tumors. The use of mass-spectrometry-based metabolomics and lipidomics combined with machine learning analyses to prospect and characterize biomarkers associated with meningioma grades may pave the way for elucidating potential therapeutic and prognostic targets.

Association of Suppressive Myeloid Cell Enrichment with Aggressive Oropharynx Squamous Cell Carcinoma.

BACKGROUND: While immune-cell infiltrated tumors, such as human papillomavirus positive (HPV+) ororpharyngeal squamous cell carcinomas (OPSCC) have been associated with an improved clinical prognosis, there is evidence to suggest that OPSCCs are also subjected to increased immunoregulatory influence. The objective of this study was to assess whether patients with clinically aggressive OPSCC have a distinct immunosuppressive immune signature in the primary tumor. METHODS: This retrospective case-control study analyzed 37 pre-treatment tissue samples from HPV+ and HPV-negative OPSCC patients treated at a single institution. The cases were patients with known disease recurrence and the controls were patients without disease recurrence. An mRNA-expression immune-pathway profiling was performed, and correlated to clinical outcomes. The TCGA head and neck cancer database was utilized to make comparisons with the institutional cohort. RESULTS: In our cohort, HPV-negative and HPV+ patients with known disease recurrence both had significantly increased suppressive monoctyte/macrophage and granulocyte cell-expression-profile enrichment. Similar findings were found in the TCGA cohort when comparing HPV-negative to positive patients. CONCLUSIONS: our study demonstrates that patients with recurrent HPV+ OPSCC had suppressive monocyte/macrophage and granulocyte immune-cell enrichment, similar to those seen in the more aggressive HPV-negative OPSCC.

Evaluation of multinodular goiter and primary hyperparathyroidism leads to a diagnosis of AL amyloidosis.

BACKGROUND: Amyloid goiter, defined as excess amyloid within the thyroid gland in such quantities that it produces a clinically apparent goiter, is a very rare manifestation of systemic amyloidosis with cases commonly seen in the setting of Amyloid A (AA) amyloidosis. Amyloid goiter as the primary clinical manifestation secondary to Amyloid light chain (AL) amyloidosis is very rare. We present a case of AL amyloidosis with initial manifestation as goiter with amyloid deposition in the thyroid and the parathyroid gland. CASE PRESENTATION: A 73 year old male presented with goiter and compressive symptoms of dysphagia and hoarseness. Laboratory workup revealed normal thyroid function, nephrotic range proteinuria, elevated serum calcium level with an elevated parathyroid hormone level (PTH) consistent with primary hyperparathyroidism. Thyroid ultrasound showed an asymmetric goiter with three dominant nodules. Cervical computed tomography revealed a goiter with substernal extension and deviation of the trachea. Fine needle aspiration was unsatisfactory. There was also evidence of osteoporosis and hypercalciuria with negative Sestamibi scan for parathyroid adenoma. The patient underwent a total thyroidectomy and one gland parathyroidectomy. Pathology revealed benign thyroid parenchyma with diffuse amyloid deposition in the thyroid and parathyroid gland that stained apple green birefringence under polarized light on Congo Red stain. Immunochemical staining detected AL amyloid deposition of the lambda type. Bone marrow biopsy revealed an excess monoclonal lambda light chain of plasma cells consistent with a diagnosis of AL amyloidosis secondary to multiple myeloma affecting the kidney, thyroid, parathyroid gland, and heart. He was treated with 4 cycles of chemotherapy with a decrease in the M spike and light chains with a plan to pursue a bone marrow transplant. CONCLUSION: Amyloid goiter as the primary clinical manifestation secondary to AL amyloidosis with deposition in the thyroid and parathyroid gland is rare. The top differential for amyloid deposits in the thyroid includes systemic amyloidosis or medullary thyroid carcinoma. The definitive diagnosis lies in the histopathology of the thyroid tissue. To diagnose systemic amyloidosis as the etiology for a goiter, a solid understanding of the causes of systemic amyloidosis coupled with a thorough evaluation of the patient's history and laboratory data is necessary.

Histologic Findings at the Time of Repeat Resection Predicts Survival in Patients with Glioblastoma.

OBJECTIVE: Radiographic worsening in patients with glioblastoma undergoing treatment may be due to tumor recurrence or treatment effect. The overall prognosis of these patients based on histologic findings at the time of repeat resection is not well established. METHODS: Patients with glioblastoma at our institution were identified. Patients who only had 1 surgery were excluded. Demographic and clinical data were recorded. The histologic diagnosis at the time of repeat surgery was recorded as either tumor recurrence or pseudoprogression. For this study, pseudoprogression was defined as absence of tumor histologic features and could show coagulative necrosis, reactive gliosis, and/or inflammatory infiltration. RESULTS: A total of 115 patients were identified, 106 with tumor recurrence and 9 with pseudoprogression. The pseudoprogression group had a greater percentage of patients with O6-methylguanine DNA-methyltransferase (MGMT) methylation (37.7% vs. 66.7%), but these results did not reach statistical significance. The overall median survival was 23.1 months. The overall median survival was 22.0 months in the tumor recurrence group and 33.3 months in the pseudoprogression group (P = 0.0814). The overall median survival from the time of repeat surgery was 8.4 months for the entire cohort, 8.3 months for the tumor recurrence group and 18.4 months for the pseudoprogression group (P = 0.0063). In multivariable analysis, presence of tumor features was predictive of worse overall survival from the time of second surgery (hazards ratio 3.96, 95% confidence interval: 1.30-12.06, P = 0.0156). CONCLUSIONS: In patients with worsening imaging, the absence of tumor on histologic diagnosis is associated with improved survival from the time of second surgery.

Glioblastoma with a primitive neuroectodermal component: two cases with implications for glioblastoma cell-of-origin.

BACKGROUND: Glioblastoma (GBM) is the most common primary brain malignancy, but much remains unknown about the histogenesis of these tumors. In the great majority of cases, GBM is a purely glial tumor but in rare cases the classic-appearing high-grade glioma component is admixed with regions of small round blue cells with neuronal immunophenotype, and these tumors have been defined in the WHO 2016 Classification as "glioblastoma with a primitive neuronal component." METHODS: In this paper, we present two cases of GBM-PNC with highly divergent clinical courses, and review current theories for the GBM cell-of-origin. RESULTS AND CONCLUSIONS: GBM-PNC likely arises from a cell type competent to give rise to glial or neuronal lineages. The thesis that GBM recapitulates to some extent normal neurodevelopmental cellular pathways is supported by molecular and clinical features of our two cases of GBM-PNC, but more work is needed to determine which cellular precursor gives rise to specific cases of GBM. GBM-PNC may have a dramatically altered clinical course compared to standard GBM and may benefit from specific lines of treatment.