From the identification of actionable molecular targets to the generation of faithful neuroblastoma patient-derived preclinical models.

BACKGROUND: Neuroblastoma (NB) represents the most frequent and aggressive form of extracranial solid tumor of infants. Although the overall survival of patients with NB has improved in the last years, more than 50% of high-risk patients still undergo a relapse. Thus, in the era of precision/personalized medicine, the need for high-risk NB patient-specific therapies is urgent. METHODS: Within the PeRsonalizEd Medicine (PREME) program, patient-derived NB tumors and bone marrow (BM)-infiltrating NB cells, derived from either iliac crests or tumor bone lesions, underwent to histological and to flow cytometry immunophenotyping, respectively. BM samples containing a NB cells infiltration from 1 to 50 percent, underwent to a subsequent NB cells enrichment using immune-magnetic manipulation. Then, NB samples were used for the identification of actionable targets and for the generation of 3D/tumor-spheres and Patient-Derived Xenografts (PDX) and Cell PDX (CPDX) preclinical models. RESULTS: Eighty-four percent of NB-patients showed potentially therapeutically targetable somatic alterations (including point mutations, copy number variations and mRNA over-expression). Sixty-six percent of samples showed alterations, graded as "very high priority", that are validated to be directly targetable by an approved drug or an investigational agent. A molecular targeted therapy was applied for four patients, while a genetic counseling was suggested to two patients having one pathogenic germline variant in known cancer predisposition genes. Out of eleven samples implanted in mice, five gave rise to (C)PDX, all preserved in a local PDX Bio-bank. Interestingly, comparing all molecular alterations and histological and immunophenotypic features among the original patient's tumors and PDX/CPDX up to second generation, a high grade of similarity was observed. Notably, also 3D models conserved immunophenotypic features and molecular alterations of the original tumors. CONCLUSIONS: PREME confirms the possibility of identifying targetable genomic alterations in NB, indeed, a molecular targeted therapy was applied to four NB patients. PREME paves the way to the creation of clinically relevant repositories of faithful patient-derived (C)PDX and 3D models, on which testing precision, NB standard-of-care and experimental medicines.

Rathke's cyst with xanthogranulomatous change or chronic cystic craniopharingioma? A rare case.

A 9-year-old girl presented to our neurosurgery clinic complaining of visual disturbances for a week. Magnetic resonance imaging showed an extensive sellar lesion with suprasellar involvement and compression of the optic chiasm. Based on these findings, a cystic craniopharyngioma, a pituitary macroadenoma and - less likely - a Rathke's cleft cyst were considered as differential diagnoses. In view of the progressive loss of vision, the parents agreed to resection of the lesion through an endoscopic endonasal transtubercular approach, with the aim of a gross total resection. Microscopic examination revealed that the lesion was cystic, surrounded by an epithelium that was partly composed of columnar ciliated cells with interspersed mucous cells and partly had a flattened appearance. The observed findings were complex to interpret: if, on the one hand, the clinical-surgical and neuroradiologic data suggested a craniopharyngioma, this hypothesis was not supported by the microscopic data, because the presence of columnar ciliated epithelium associated with mucous cells was a microscopic feature inconsistent with a craniopharyngioma and was instead consistent with a Rathke's cleft cyst, a histologic diagnosis that was made. The incidence of Rathke's cleft cyst, which mimics clinical and neuroradiologic aspects of craniopharyngiomas, is extremely unusual, as only 2 cases have been described in the literature.

SA-ß-Gal in Kidney Tubules as a Predictor of Renal Outcome in Patients with Chronic Kidney Disease.

Cellular senescence has emerged as an important driver of aging and age-related disease in the kidney. The activity of ß-galactosidase at pH 6 (SA-ß-Gal) is a classic maker of senescence in cellular biology; however, the predictive role of kidney tissue SA-ß-Gal on eGFR loss in chronic kidney disease (CKD) is still not understood. We retrospectively studied the expression of SA-ß-Gal in kidney biopsies obtained in a cohort [n = 22] of incident patients who were followed up for 3 years as standard of care. SA-ß-Gal staining was approximately fourfold higher in the tubular compartment of patients with CKD vs. controls [26.0 ± 9 vs. 7.4 ± 6% positive tubuli in patients vs. controls; p < 0.025]. Tubular expressions of SA-ß-Gal, but not proteinuria, at the time of biopsy correlated with eGFR loss at the follow up; moreover, SA-ß-Gal expression in more than 30% of kidney tubules was associated with fast progressive kidney disease. In conclusion, our study shows that SA-ß-Gal is upregulated in the kidney tubular compartment of adult patients affected by CKD and suggests that tubular SA-ß-Gal is associated with accelerated loss of renal function.

Pathogenesis and treatment of a giant occipital bone defect with meningoencephalocele in an NF1 child: case report and review of the literature.

Autosomal dominantly inherited neurofibromatosis type I (NF1) is a systemic disorder caused by a mutation of a gene on chromosome 17q11.2 and characterized by multiple café-au-lait spots, lentiginous macules, Lisch nodules of the iris, and tumors of the nervous system. Bony manifestations such as scoliosis, dysplasia of the greater sphenoidal wing, tibial pseudoarthrosis, short stature, and macrocephaly have been reported in approximately 50% of patients. However, calvarial bone defects are rare. After screening 324 articles, 23 cases (12 adult and 11 pediatric patients) of occipital bone defects in NF1 patients were selected. All patients had a single/multiple bone defect over the lambdoid suture. Adjacent benign plexiform neurofibromas were observed in 14 patients (60.8%, 7 adults and 7 children); one adult patient was diagnosed with neurofibrosarcoma. Meningoencephalocele over the occipital defect was noted in 8 cases (34.78%, all adults). Cranioplasty was performed in only 17.39% of patients. Histologic examination was performed in 7 of the 15 patients with associated neurofibromas/neurofibrosarcomas. Biopsy of the bone margins surrounding the defect was performed in only one case. Pathologic examination of the herniated parieto-occipital or cerebellar tissue was not performed in any of the patients studied. We report the case of a 9-year-old girl with NF1 and a significant occipital bone defect and performed a systematic review of the relevant literature to highlight the challenges in treating this condition and to investigate the underlying mechanisms contributing to bone defects or dysplasia in NF1.

Characterization of Tumor and Immune Tumor Microenvironment of Primary Tumors and Metastatic Sites in Advanced Renal Cell Carcinoma Patients Based on Response to Nivolumab Immunotherapy: Preliminary Results from the Meet-URO 18 Study.

BACKGROUND: Prognostic and predictive factors for patients with metastatic renal cell carcinoma (mRCC) treated with immunotherapy are highly warranted, and the immune tumor microenvironment (I-TME) is under investigation. METHODS: The Meet-URO 18 was a multicentric retrospective study assessing the I-TME in mRCC patients treated with ≥2nd-line nivolumab, dichotomized into responders and non-responders according to progression-free survival (≥12 months and ≤3 months, respectively). The primary objective was to identify differential immunohistochemical (IHC) patterns between the two groups. Lymphocyte infiltration and the expressions of different proteins on tumor cells (CD56, CD15, CD68, and ph-mTOR) were analyzed. The expression of PD-L1 was also assessed. RESULTS: A total of 116 tumor tissue samples from 84 patients (59% were primary tumors and 41% were metastases) were evaluated. Samples from responders (N = 55) were significantly associated with lower expression of CD4+ T lymphocytes and higher levels of ph-mTOR and CD56+ compared with samples from non-responders (N = 61). Responders also showed a higher CD3+ expression (p = 0.059) and CD8+/CD4+ ratio (p = 0.084). Non-responders were significantly associated with a higher percentage of clear cell histology and grading. CONCLUSIONS: Differential IHC patterns between the tumors in patients who were responders and non-responders to nivolumab were identified. Further investigation with genomic analyses is planned.

Stem-like signatures in human meningioma cells are under the control of CXCL11/CXCL12 chemokine activity.

BACKGROUND: Meningiomas are mainly benign brain tumors, although about 20% of histologically benign cases are clinically aggressive and recur after resection. We hypothesize that meningioma brain invasiveness and recurrence may be related to the presence of cancer stem cells and their high responsiveness to the CXCL12-CXCR4/CXCR7 chemokine axis. The aim of this study was to isolate meningioma stem cells from human samples, characterize them for biological features related to malignant behavior, and to identify the role of CXCR4/CXCR7 in these processes. METHODS: Meningioma stem cells were isolated from patient-derived primary cultures in stem cell-permissive conditions, and characterized for phenotype, self-renewal, proliferation and migration rates, vasculogenic mimicry (VM), and in vivo tumorigenesis, in comparison with differentiated meningioma cells and stem-like cells isolated from normal meninges. These cell populations were challenged with CXCL12 and CXCL11 and receptor antagonists to define the chemokine role in stem cell-related functions. RESULTS: Stem-like cells isolated from meningioma cultures display higher proliferation and migration rates, and VM, as compared to meningioma non-stem cells or cells isolated from normal meninges and were the only tumorigenic population in vivo. In meningioma cells, these stem-like functions were under the control of the CXCR4/CXCR7 chemokine axis. CONCLUSIONS: We report a role for CXCL11 and CXCL12 in the control of malignant features in stem-like cells isolated from human meningioma, providing a possible basis for the aggressive clinical behavior observed in subsets of these tumors. CXCR4/CXCR7 antagonists might represent a useful approach for meningioma at high risk of recurrence and malignant progression.

Hyaline Protoplasmic Astrocytopathy in the Setting of Epilepsy.

OBJECTIVES: Cerebral hyaline protoplasmic astrocytopathy (HPA) is a clinicopathologic entity characterized by eosinophilic cytoplasmic inclusions within astrocytes. It has been observed in a subset of patients with early-onset epilepsy, brain malformations, and developmental delay. The exact association of this entity with epilepsy is still unknown. This report, with its review of the literature, aims to summarize HPA features to raise awareness regarding this entity. METHODS: We report on 2 HPA cases and critically review the literature. RESULTS: Approximately 42 cases of HPA have been reported, including the 2 cases presented here, consisting of 23 female and 19 male patients. Patient age ranged from 3 to 39 years. All patients had early-onset seizures (3-20 months of age), ranging from partial to generalized, that were refractory despite treatment with antiepileptic drugs. Postoperative follow-up intervals ranged from 2 to 93 months, and the clinical outcome was graded according to the Engel classification, showing variable results. CONCLUSIONS: Clinicians should consider HPA in differential diagnosis in patients with intractable seizures, especially when they are associated with developmental delay and brain malformations. Increasing awareness of this entity among pathologists may promote better understanding of this condition as well as better diagnosis and treatment for these patients.

Early molecular diagnosis of BRAF status drives the neurosurgical management in BRAF V600E-mutant pediatric low-grade gliomas: a case report.

BACKGROUND: To date, this is the only report showing with close and consecutive magnetic resonance images the extremely rapid response of two types of pediatric low-grade gliomas (PLGG) to vemurafenib and its impact on the surgical approach. CASES PRESENTATION: We report two cases of symptomatic PLGG treated with vemurafenib, a BRAF inhibitor: in a 12-year-old girl it was used as first-line medical treatment, reducing the tumor by 45% within a month and stabilizing to 76% after a year; in a 3-year-old boy with no improvement after SIOP LGG 2004 Protocol, vemurafenib induced in only one week a 34% shrinkage and solved the hydrocephalus, avoiding surgical operation. DISCUSSION AND CONCLUSIONS: Our cases demonstrate how an early molecular diagnosis of BRAF mutations through the neurosurgical biopsy is essential to promptly start targeted therapies., whose effect can influence both therapeutic and surgical decisions, hopefully reducing the occurrence of second neurosurgery with associated risks of neurological sequelae.

Analysis of A Disintegrin and Metalloprotease 17 (ADAM17) Expression as a Prognostic Marker in Ovarian Cancer Patients Undergoing First-Line Treatment Plus Bevacizumab.

To find prognostic factors for advanced ovarian cancer patients undergoing first-line therapy with carboplatin, paclitaxel and bevacizumab, we investigated the expression of a disintegrin and metalloprotease 17 (ADAM17) in cancer tissues. ADAM17 has been involved in ovarian cancer development, progression and cell resistance to cisplatin. Tissue microarrays from 309 ovarian cancer patients enrolled in the MITO16A/MANGO-OV2 clinical trial were analyzed by immunohistochemistry for ADAM17 protein expression. Intensity and extent of staining were combined into a semi-quantitative visual grading system (H score) which was related to clinicopathological characteristics of cases and the clinical outcome of patients by univariate and multivariate Cox regression models. ADAM17 immunostaining was detected in most samples, mainly localized in the tumor cells, with variable intensity across the cohort. Kaplan-Meier survival curves, generated according to the best cut-off value for the ADAM17 H score, showed that high ADAM17 expression was associated with worse prognosis for PFS and OS. However, after the application of a shrinkage procedure to adjust for overfitting hazard ratio estimates, the ADAM17 value as prognostic factor was lost. As subgroup analysis suggested that ADAM17 expression could be prognostically relevant in cases with no residual disease at baseline, further studies in this patient category may be worth planning.

Lymphoplasmacyte-rich meningioma with hematologic signs and PD-L1 over-expression.

Lymphoplasmacyte-rich meningioma (LPRM) is one of the rarest variants of grade I meningiomas. It can be clinically associated with prominent peripheral blood abnormalities, anemia, and/or various gammopathy, which usually disappear after surgical removal of the tumor. We document a case of right frontal LPRM in a 72-year-old male who presented general cognitive decadence. The patient suffered from mild anemia. The LPRM is a rare variant of meningioma, with only a few cases globally reported in the literature. It has been categorized as a grade I tumor in the 2021 World Health Organization (WHO) classification central nervous system. Due to the rarity, this meningioma variant origin and biological behavior are still not clear. Immunohistochemistry profile showed prominent PD-L1 expression, leading to additional interrogation on LPRM immunomorphological characteristics, the significance of the inflammatory tumoral microenvironment and its correlation with the immune-checkpoints.

Dyslipidemia in Children Treated with a BRAF Inhibitor for Low-Grade Gliomas: A New Side Effect?

BRAF inhibitors, in recent years, have played a central role in the disease control of unresectable BRAF-mutated pediatric low-grade gliomas (LGGs). The aim of the study was to investigate the acute and long-term effects of vemurafenib on the lipid metabolism in children treated for an LGG. In our cohort, children treated with vemurafenib (n = 6) exhibited alterations in lipid metabolism a few weeks after starting, as was demonstrated after 1 month (n = 4) by the high plasma levels of the total cholesterol (TC = 221.5 ± 42.1 mg/dL), triglycerides (TG = 107.8 ± 44.4 mg/dL), and low-density lipoprotein (LDL = 139.5 ± 51.5 mg/dL). Despite dietary recommendations, the dyslipidemia persisted over time. The mean lipid levels of the TC (222.3 ± 34.7 mg/dL), TG (134.8 ± 83.6 mg/dL), and LDL (139.8 ± 46.9 mg/dL) were confirmed abnormal at the last follow-up (45 ± 27 months, n = 6). Vemurafenib could be associated with an increased risk of dyslipidemia. An accurate screening strategy in new clinical trials, and a multidisciplinary team, are required for the optimal management of unexpected adverse events, including dyslipidemia.

Calcifications in diffuse leptomeningeal glioneuronal tumors: a case series.

Diffuse leptomeningeal glioneuronal tumor (DLGNT) is a new rare entity, typically seen in the pediatric population. Classical neuroimaging features at clinical onset include marked subarachnoid/leptomeningeal enhancement and tiny pseudo-cystic lesions along the subpial surface of the neuroaxis, frequently associated with communicating hydrocephalus. However, data on the long-term appearance of this tumor on computed tomography (CT) and magnetic resonance imaging (MRI) are still lacking. We describe a peculiar pattern of progressive leptomeningeal calcifications in three young patients with DLGNT. The calcifications, mainly located in the basal cisterns, sylvian fissures and posterior surface of the thalami, were present at clinical onset in the older subject and appeared about 2 years after clinical onset in the other two. Patients underwent different schemes of chemotherapy, variably associated with craniospinal irradiation and/or bevacizumab. In all cases, calcifications were present before starting craniospinal irradiation and/or treatment with bevacizumab. This novel peculiar pattern of progressive leptomeningeal calcifications expands the imaging phenotype of DLGNT and should be carefully sought, especially in later phases of the disease. Taking into consideration the onset, evolution, and absence of direct relationship with treatments, we hypothesize that calcifications in DLGNTs might be the effect of natural changes in the tumor and its environment.

Validation of a Novel Three-Dimensional (3D Fusion) Gross Sampling Protocol for Clear Cell Renal Cell Carcinoma to Overcome Intratumoral Heterogeneity: The Meet-Uro 18 Study.

We aimed to overcome intratumoral heterogeneity in clear cell renal cell carcinoma (clearRCC). One hundred cases of clearRCC were sampled. First, usual standard sampling was applied (1 block/cm of tumor); second, the whole tumor was sampled, and 0.6 mm cores were taken from each block to construct a tissue microarray; third, the residual tissue, mapped by taking pieces 0.5 × 0.5 cm, reconstructed the entire tumor mass. Precisely, six randomly derived pieces of tissues were placed in each cassette, with the number of cassettes being based on the diameter of the tumor (called multisite 3D fusion). Angiogenic and immune markers were tested. Routine 5231 tissue blocks were obtained. Multisite 3D fusion sections showed pattern A, homogeneous high vascular density (10%), pattern B, homogeneous low vascular density (8%) and pattern C, heterogeneous angiogenic signatures (82%). PD-L1 expression was seen as diffuse (7%), low (33%) and absent (60%). Tumor-infiltrating CD8 scored high in 25% (pattern hot), low in 65% (pattern weak) and zero in 10% of cases (pattern desert). Grading was upgraded in 26% of cases (G3-G4), necrosis and sarcomatoid/rhabdoid characters were observed in, respectively, 11 and 7% of cases after 3D fusion (p = 0.03). CD8 and PD-L1 immune expressions were higher in the undifferentiated G4/rhabdoid/sarcomatoid clearRCC subtypes (p = 0.03). Again, 22% of cases were set to intermediate to high risk of clinical recurrence due to new morphological findings of all aggressive G4, sarcomatoid/rhabdoid features by using 3D fusion compared to standard methods (p = 0.04). In conclusion, we propose an easy-to-apply multisite 3D fusion sampling that negates bias due to tumor heterogeneity.

A microcystic/reticular schwannoma in an unusual site: description of a retroperitoneal location and review of the literature.

Microcystic/reticular (MRV) schwannoma has been described since 2008, but remains a rarely encountered entity. MRV has a predilection for visceral locations and has variable histologic appareances. Given its rarity and anatomic variability, this entity could raise differential diagnostic issues with other tumours and malignancies.We describe the case of a 69-year-old male followed at IRCCS Ospedale Policlinico San Martino of Genoa for his previous history of non-Hodgkin lymphoma. A para-aortic mass was discovered during follow-up, which -due to its stability, also after chemotherapy- had been hypothesized to be a non-lymphomatous lesion; given the dimensions and the site, the mass was removed. Histological evaluation showed a nodule limited by a slight fibrous capsule and characterized by a proliferation of medium-sized fusiform cells, with elongated nuclei and scarce eosinophilic cytoplasm. Given the lack of malignant signs and the strong expression of protein S-100, a diagnosis of mesenchymal neoplasia with expression of neural markers compatible with reticular schwannoma was made. The neoplasm has not recurred since its removal.The case we present is, at our best knowledge, the first described in the retroperitoneum, a site where the exclusion of other mesenchymal malignancies is mandatory. The rarity and variability of presentations could create problems of differential diagnosis both with mucinous-producing carcinomas or with other soft tissue tumours, with myxoid or reticular structure. The description of this case could help raise information on this rare neoplasm and help distinguish it from other malignancies, especially in unusual sites.

Mind the gap: IgG4-related disease mimicking infectious cerebral mass lesions.

BACKGROUND: Cerebral intraparenchymal masses represent usually a neoplastic, or infectious differential diagnostic workup in neurology or infectious disease units. CASE PRESENTATION: Our patient was an 82-year-old male presenting with seizures, cerebral masses and a history of past treated pulmonary tuberculosis. Initial workup included a differential diagnosis of an infectious mass/multiple abscess. After exclusion of infectious or primary neoplastic origins by negative HIV serology, the absence of immune suppression, endocarditic lesions, negative results of blood cultures and bronchoalveolar lavage, negative cerebrospinal fluid workout on spinal tap led to exclusion of infectious causes. A surgical procedure was performed to access one of the lesions. This yielded a firm, cyst-like mass of histiocytic granulomatous tissue with a conspicuous plasmacellular component and a relevant IgG4 plasmacellular component consistent with IgG4-related disease. Steroid treatment determined conspicuous improvement and led to discharge of the patient. CONCLUSION: Parenchymal IgG4-related disease may be included as a new entity in the differential diagnosis of single or multiple cerebral masses in addition to infectious or neoplastic etiology.

Secondary Membranous Nephropathy Due to Benign Tumors in 2 Young Women: A Case Report.

Membranous nephropathy (MN) is one of the most common causes of adult-onset nephrotic syndrome. We describe the cases of 2 young women in their 20s presenting with nephrotic syndrome due to antiphospholipase A2 receptor (anti-PLA2R)-negative MN, that was found to be associated with benign tumors. Both women had no extrarenal symptoms of a connective tissue disease, infection, or malignancy. They both had been previously healthy and were not receiving treatment with any drugs. Both had MN on kidney biopsy. Biopsies were negative for PLA2R antigen, and their serum did not demonstrate the presence of anti-PLA2R antibodies. Both were investigated for a secondary cause on the basis of negative anti-PLA2R serology and biopsy features supportive of secondary MN and were found to have benign tumors on radioimaging: a uterine leiomyoma and mesenteric fibromatosis, respectively. In both instances, the nephrotic syndrome remitted following resection of the tumors. To our knowledge, uterine leiomyoma and mesenteric fibromatosis have not previously been described in association with MN. These cases highlight the importance of pursuing a secondary cause of MN in patients without anti-PLA2R antibodies in serum or PLA2R antigen on kidney biopsy.