Microtubule-associated protein MAP1LC3C regulates lysosomal exocytosis and induces zinc reprogramming in renal cancer cells.

Microtubule-associated protein 1 light chain 3 gamma (MAP1LC3C or LC3C) is a member of the microtubule-associated family of proteins that are essential in the formation of autophagosomes and lysosomal degradation of cargo. LC3C has tumor-suppressing activity, and its expression is dependent on kidney cancer tumor suppressors, such as von Hippel-Lindau protein and folliculin. Recently, we demonstrated that LC3C autophagy is regulated by noncanonical upstream regulatory complexes and targets for degradation postdivision midbody rings associated with cancer cell stemness. Here, we show that loss of LC3C leads to peripheral positioning of the lysosomes and lysosomal exocytosis (LE). This process is independent of the autophagic activity of LC3C. Analysis of isogenic cells with low and high LE shows substantial transcriptomic reprogramming with altered expression of zinc (Zn)-related genes and activity of polycomb repressor complex 2, accompanied by a robust decrease in intracellular Zn. In addition, metabolomic analysis revealed alterations in amino acid steady-state levels. Cells with augmented LE show increased tumor initiation properties and form aggressive tumors in xenograft models. Immunocytochemistry identified high levels of lysosomal-associated membrane protein 1 on the plasma membrane of cancer cells in human clear cell renal cell carcinoma and reduced levels of Zn, suggesting that LE occurs in clear cell renal cell carcinoma, potentially contributing to the loss of Zn. These data indicate that the reprogramming of lysosomal localization and Zn metabolism with implication for epigenetic remodeling in a subpopulation of tumor-propagating cancer cells is an important aspect of tumor-suppressing activity of LC3C.

A novel RYR1 variant in an infant with a unique fetal presentation of central core disease.

Ryanodine receptor type 1-related disorder (RYR1-RD) is the most common subgroup of congenital myopathies with a wide phenotypic spectrum ranging from mild hypotonia to lethal fetal akinesia. Genetic testing for myopathies is imperative as the diagnosis informs counseling regarding prognosis and recurrence risk, treatment options, monitoring, and clinical management. However, diagnostic challenges exist as current options are limited to clinical suspicion prompting testing including: single gene sequencing or familial variant testing, multi-gene panels, exome, genome sequencing, and invasive testing including muscle biopsy. The timing of diagnosis is of great importance due to the association of RYR1-RD with malignant hyperthermia (MH). MH is a hypermetabolic crisis that occurs secondary to excessive calcium release in muscles, leading to systemic effects that can progress to shock and death if unrecognized. Given the association of MH with pathogenic variants in RYR1, a diagnosis of RYR1-RD necessitates an awareness of medical team to avoid potentially triggering agents. We describe a case of a unique fetal presentation with bilateral diaphragmatic eventrations who had respiratory failure, dysmorphic facial features, and profound global hypotonia in the neonatal period. The diagnosis was made at several months of age, had direct implications on her clinical care related to anticipated need to long-term ventilator support, and ultimately death secondary an arrhythmia as a result of suspected MH. Our report reinforces the importance of having high suspicion for a genetic syndrome and pursuing early, rapid exome or genome sequencing as first line testing in critically ill neonatal intensive care unit patients and further evaluating the pathogenicity of a variant of uncertain significance in the setting of a myopathic phenotype.

Carfilzomib-based antibody mediated rejection therapy in pediatric kidney transplant recipients.

BACKGROUND: To date, the evidence for proteasome-inhibitor (PI) based antibody mediated rejection (AMR) therapy has been with the first-generation PI bortezomib. Results have demonstrated encouraging efficacy for early AMR with lesser efficacy for late AMR. Unfortunately, bortezomib is associated with dose-limiting adverse effects in some patients. We report use of the second generation proteosome inhibitor carfilzomib for AMR treatment in two pediatric patients with a kidney transplant. METHODS: The clinical data on two patients who experienced dose limiting toxicities from bortezomib were collected along with their short- and long-term outcomes. RESULTS: A two-year-old female with simultaneous AMR, multiple de novo DSAs (DR53 MFI 3900, DQ9 MFI 6600, DR15 2200, DR51 MFI 1900) and T-cell mediated rejection (TCMR) completed three carfilzomib cycles and experienced stage 1 acute kidney injury after the first two cycles. At 1 year follow up, all DSAs resolved, and her kidney function returned to baseline without recurrence. A 17-year-old female also developed AMR with multiple de novo DSAs (DQ5 MFI 9900, DQ6 MFI 9800, DQA*01 MFI 9900). She completed two carfilzomib cycles, which were associated with acute kidney injury. She had resolution of rejection on biopsy and decreased but persistent DSAs on follow-up. CONCLUSIONS: Carfilzomib treatment for bortezomib-refractory rejection and/or bortezomib toxicity may provide DSA elimination or reduction, but also appears to be associated with nephrotoxicity. Clinical development of carfilzomib for AMR will require a better understanding of efficacy and development of approaches to mitigate nephrotoxicity.

Primary hyperoxaluria diagnosed after kidney transplant: A review of the literature and case report of aggressive renal replacement therapy and lumasiran to prevent allograft loss.

Primary hyperoxaluria type 1 is a rare inherited disorder caused by abnormal liver glyoxalate metabolism leading to overproduction of oxalate, progressive kidney disease, and systemic oxalosis. While the disorder typically presents with nephrocalcinosis, recurrent nephrolithiasis, and/or early chronic kidney disease, the diagnosis is occasionally missed until it recurs after kidney transplant. Allograft outcomes in these cases are typically very poor, often with early graft loss. Here we present the case of a child diagnosed with primary hyperoxaluria type 1 after kidney transplant who was able to maintain kidney function, thanks to aggressive renal replacement therapy as well as initiation of a new targeted therapy for this disease. This case highlights the importance of having a high index of suspicion for primary hyperoxaluria in patients with chronic kidney disease and nephrocalcinosis/nephrolithiasis or with end stage kidney disease of uncertain etiology, as initiating therapies early on may prevent poor outcomes.

Metastatic neuroblastoma masquerading as infantile hemangioma in a 4-month-old child.

INTRODUCTION: Diffuse liver lesions in an infant have a differential diagnosis including infantile hemangioma (IH), which is common in the first year of life, and neuroblastoma (NBL) which presents at a median age of 18 months. RESULTS: We describe the case of a 4-month-old girl with a known superficial/deep IH who presented with new axillary nodules and hepatosplenomegaly, initially suspected to reflect IH but later determined to be widely metastatic NBL. CONCLUSION: Hepatic IH and metastatic NBL can present similarly. Clinicians must maintain a broad differential when evaluating new findings in a patient with previously diagnosed IH.

Human Nephrogenesis can Persist Beyond 40 Postnatal Days in Preterm Infants.

Introduction: Human nephrogenesis is typically completed by 36 weeks gestation; however, it is impacted by preterm birth. Early studies suggested that nephrogenesis persisted for ≤40 postnatal days in preterm infants. However, the postmenstrual age (PMA) of the preterm infants who survived >40 days was uncertain. In this study, we sought to reexamine postnatal kidney development in preterm infants surviving >40 days. Methods: Human kidney samples were obtained from an institutional biobank. Samples were considered controls if survival was ≤4 days after birth with PMA of 30 to ≤36 weeks. Kidneys from preterm neonates with postnatal survival >40 days and PMA of 30 to ≤36 weeks were compared to controls. We counted glomerular generations, measured nephrogenic zone widths (NZW), and performed immunofluorescence (IF) with SIX1 and RET. We compared kidney weights and quantified the cross-sectional area of proximal (lotus tetragonolobus lectin [LTL], SL22A2), distal (SLC12A3, KCNJ10), and glomerular (nephrin) markers using IF. Results: Seven preterm infants surviving >40 days and 8 controls were analyzed. Four of 7 preterm infants had histologic and molecular evidence of nephrogenesis. Cessation of nephrogenesis in preterm infants occurred 2 weeks earlier than PMA-matched controls with attenuated expression of both SIX1 and RET. We found increased kidney weight-to-body weight ratio, increased distal tubular cross-sectional staining in the superficial nephrons, and distal tubular hypertrophy and hyperplasia in the preterm infant kidneys. Conclusion: Our study supports that nephrogenesis in preterm infants persists longer than previously thought with evidence of early nephron stress, placing importance on the neonatal environment.

Atypical Presentation of Granulosa Cell Tumor in an Adolescent: A Case Report.

BACKGROUND: Granulosa cell tumors (GCTs) frequently present with elevated levels of estrogen and inhibin. Most diagnoses in the pediatric and adolescent population are juvenile-type GCTs; adult-type GCTs in this population are rare. CASE: We describe a 14-year-old female who presented with a large adnexal mass and clinical hyperandrogenism. Laboratory evaluation revealed elevated levels of free and total testosterone, low-normal estradiol, and mildly elevated alpha-fetoprotein (AFP). Other tumor markers, including inhibin, were within normal limits. Intraoperative assessment with unilateral oophorectomy, pathology, and imaging resulted in a diagnosis of a stage IA adult-type GCT. SUMMARY AND CONCLUSION: GCTs often result in elevated estrogen and inhibin B levels; however, this case demonstrates that non-classic elevations in testosterone and normal inhibin levels should not eliminate concern for a GCT, particularly in the setting of a large ovarian mass.

Multiparametric quantitative renal MRI in children and young adults: comparison between healthy individuals and patients with chronic kidney disease.

PURPOSE: Multiparametric quantitative renal MRI may provide noninvasive radiologic biomarkers of chronic kidney disease (CKD) based on investigations in animal models and adults. We aimed to (1) obtain normative multiparametric quantitative MRI data from the kidneys of healthy children and young adults, (2) compare MRI measurements between healthy control participants and patients with CKD, and (3) determine if MRI measurements correlate with clinical and laboratory data as well as histology. METHODS: This was a prospective, case-control study of 20 healthy controls and 12 CKD patients who underwent percutaneous renal biopsy ranging from 12 to 23 years of age between October 2018 and March 2020. Kidney function was documented and pathology assessed for fibrosis/inflammation. Utilizing a field strength of 1.5T, we examined renal T1, T2, and T2* relaxation mapping, MR elastography (MRE), and diffusion-weighted imaging (DWI). A single analyst made all manual measurements for quantitative MRI pulse sequences. Independent measurements from cortex, medulla, and whole kidney were obtained by drawing regions of interest on single slices from the upper, mid, and lower kidney. A weighted average was calculated for each kidney; if two kidneys, the right and left were averaged. Continuous variables were compared with Mann-Whitney U test; bivariate relationships were assessed using Spearman rank-order correlation. RESULTS: Median estimated glomerular filtration rate (eGFR) was 112.3 ml/min/1.73 m2 in controls (n = 20, 10 females) and 55.0 ml/min/m2 in CKD patients (n = 12, 2 females) (p < 0.0001). Whole kidney (1333 vs. 1291 ms; p = 0.018) and cortical (1212 vs 1137 ms; p < 0.0001) T1 values were higher in CKD patients. Cortical T1 values correlated with eGFR (rho = - 0.62; p = 0.0003) and cystatin C (rho = 0.58; p = 0.0007). Whole kidney (1.87 vs. 2.02 10-3 mm2/s; p = 0.007), cortical (1.89 vs. 2.04 10-3 mm2/s; p = 0.008), and medullary (1.87 vs. 1.98 10-3 mm2/s; p = 0.0095) DWI apparent diffusion coefficients (ADC) were lower in CKD patients. Whole kidney ADC correlated with eGFR (rho = 0.45; p = 0.012) and cystatin C (rho = - 0.46; p = 0.009). Cortical histologic inflammation correlated with DWI ADC (rho = - 0.71; p = 0.011). CONCLUSION: Renal T1 relaxation and DWI ADC measurements differ between pediatric healthy controls and CKD patients, correlate with laboratory markers of CKD, and may have histologic correlates.

Case Report: Atypical HUS Presenting With Acute Rhabdomyolysis Highlights the Need for Individualized Eculizumab Dosing.

Background: Atypical hemolytic uremic syndrome (aHUS) is an ultra-rare orphan disease caused by dysregulated complement activation resulting in thrombotic microangiopathy. Although complement-mediated endothelial injury predominantly affects the renal microvasculature, extra-renal manifestations are present in a significant proportion of patients. While eculizumab has significantly improved the morbidity and mortality of this rare disease, optimizing therapeutic regimens of this highly expensive drug remains an active area of research in the treatment of aHUS. Case Presentation: This report describes the case of a previously healthy 4 year-old male who presented with rhabdomyolysis preceding the development of aHUS with anuric kidney injury requiring dialysis. Clinical stabilization required increased and more frequent eculizumab doses compared with the standardized weight-based guidelines. In the maintenance phase of his disease, pharmacokinetic analysis indicated adequate eculizumab levels could be maintained with an individualized dosing regimen every 3 weeks, as opposed to standard 2 week dosing, confirmed in this patient over a 4 year follow up period. Cost analyses show that weight-based maintenance dosing costs $312,000 per year, while extending the dosing interval to every 3 weeks would cost $208,000, a savings of $104,000 per year, relative to the cost of $72,000 from more frequent eculizumab dosing during his initial hospitalization to suppress his acute disease. Conclusion: This case exemplifies the potential of severe, multisystem involvement of aHUS presenting with extra-renal manifestations, including rhabdomyolysis as in this case, and highlights the possibility for improved clinical outcomes and higher value care with individualized eculizumab dosing in patients over the course of their disease.

Olfactomedin 4 as a novel loop of Henle-specific acute kidney injury biomarker.

Acute kidney injury (AKI) is associated with morbidity and mortality. Urinary biomarkers may disentangle its clinical heterogeneity. Olfactomedin 4 (OLFM4) is a secreted glycoprotein expressed in stressed neutrophils and epithelial cells. In septic mice, OLFM4 expression localized to the kidney's loop of Henle (LOH) and was detectable in the urine. We hypothesized that urine OLFM4 (uOLFM4) will be increased in patients with AKI and sepsis. Urine from critically ill pediatric patients was obtained from a prospective study based on AKI and sepsis status. uOLFM4 was quantified with a Luminex immunoassay. AKI was defined by KDIGO severe criteria. Sepsis status was extracted from the medical record based on admission diagnosis. Immunofluorescence on pediatric kidney biopsies was performed with NKCC2, uromodulin and OLFM4 specific antibodies. Eight patients had no sepsis, no AKI; 7 had no sepsis but did have AKI; 10 had sepsis, no AKI; 11 had sepsis and AKI. Patients with AKI had increased uOLFM4 compared to no/stage 1 AKI (p = 0.044). Those with sepsis had increased uOLFM4 compared to no sepsis (p = 0.026). uOLFM4 and NGAL were correlated (r2 0.59, 95% CI 0.304-0.773, p = 0.002), but some patients had high uOLFM4 and low NGAL, and vice versa. Immunofluorescence on kidney biopsies demonstrated OLFM4 colocalization with NKCC2 and uromodulin, suggesting expression in the thick ascending LOH (TALH). We conclude that AKI and sepsis are associated with increased uOLFM4. uOLFM4 and NGAL correlated in many patients, but was poor in others, suggesting these markers may differentiate AKI subgroups. Given OLFM4 colocalization to human TALH, we propose OLFM4 may be a LOH-specific AKI biomarker.

A mutant fibrinogen that is unable to form fibrin can improve renal phenotype in mice with sickle cell anemia.

Sickle cell anemia (SCA) causes nephropathy which may progress to kidney failure. To determine if soluble fibrinogen (FibAEK) can prevent kidney damage in mice with SCA, we performed bone marrow transplantation (BMT) of Berkeley sickle mice into wild-type fibrinogen (FibWT), and FibAEK mice that bear a germ-line mutation in fibrinogen Aα chain at thrombin cleavage site which prevents fibrin formation. We found improved albuminuria in SS FibAEK mice compared with SS FibWT mice at 12 months post-BMT due to the reduced kidney fibrosis, ischemic lesions, and increased survival of podocytes in the glomeruli, but did not improve urine concentrating defect. Therefore, our study clarifies the distinct role of fibrinogen and fibrin in the renal pathology of SCA.

Elimination of the fibrinogen integrin αMß2-binding motif improves renal pathology in mice with sickle cell anemia.

Sickle cell anemia (SCA) is caused by a point mutation in the ß-globin gene that leads to devastating downstream consequences including chronic hemolytic anemia, episodic vascular occlusion, and cumulative organ damage resulting in death. SCA patients show coagulation activation and inflammation even in the absence of vascular occlusion. The coagulation factor fibrinogen is not only central to hemostasis but also plays important roles in pathologic inflammatory processes, in part by engaging neutrophils/macrophages through the αMß2 integrin receptor. To determine whether fibrin(ogen)-mediated inflammation is a driver of SCA-associated pathologies, hematopoietic stem cells from Berkeley sickle mice were transplanted into homozygous Fibγ390-396A mice that express normal levels of a mutant form of fibrin(ogen) that does not engage αMß2 Fibγ390-396A mice with SCA displayed an impressive reduction of reactive oxygen species (ROS) in white blood cells (WBCs), decreased circulating inflammatory cytokines/chemokines, and significantly improved SCA-associated glomerular pathology highlighted by reduced glomerulosclerosis, inflammatory cell infiltration, ischemic lesions, mesangial thickening, mesangial hypercellularity, and glomerular enlargement. In addition, Fibγ390-396A mice with SCA had improved glomerular protective responses and podocyte/mesangial transcriptional signatures that resulted in reduced albuminuria. Interestingly, the fibrinogen γ390-396A mutation had a negligible effect on cardiac, lung, and liver functions and pathologies in the context of SCA over a year-long observation period. Taken together, our data support that fibrinogen significantly contributes to WBC-driven inflammation and ROS production, which is a key driver of SCA-associated glomerulopathy, and may represent a novel therapeutic target against irreversible kidney damage in SCA.

Primary intracranial Ewing's sarcoma with unusual features.

Pediatric primary "small round blue cell" tumors in the CNS represent several entities, some more common than others. Ewing sarcoma/peripheral primitive neuroectodermal tumor (ES/pPNET) is rare and must be distinguished from other tumors such as medulloblastoma [1, 2], atypical rhabdoid/teratoid tumor, ependymomal tumors, metastatic sarcomas, hematologic malignancies, and other mimics. Although therapy for ES/pPNET is effective, it brings severe side effects, including cardiac toxicity, making correct recognition important [3]. As small blue cell tumors look similar, diagnosis often depends on special stains, immunohistochemistry, and molecular techniques. While the combination of membranous immunohistochemical reactivity for CD99 with cytoplasmic glycogen provides effective screening, demonstration of characteristic translocations of EWSR1 (chromosome 22) or FUS (chromosome 16) by fluorescent in situ hybridization (FISH) can confirm the diagnosis. We are reporting three primary ES/pPNET of the CNS, two of which occurred in children. While the adult case demonstrates the classic histopathology, the two pediatric cases have histopathology that significantly deviates from the usual. One is suggestive of a primary sarcoma, and the other mimics an ependymoma, but all three cases are confirmed with FISH. These observations suggest that primary ES in the CNS may have histology different from the classic morphology and a high index of suspicion should be maintained in order to make the correct diagnosis. A search of the literature suggests that these tumors are most frequently seen in children and young adults. Imaging often shows a supratentorial enhancing mass that touches the leptomeninges. Survival over three years is good but long term prognosis is unknown [3, 4].

Ectomesenchymoma with embryonal rhabdomyosarcoma and ganglioneuroma, arising in association with benign triton tumor of the tongue.

Soft-tissue tumors known as "triton" tumors are rare lesions containing neural tissue and skeletal muscle at varying levels of maturity and malignant potential. Benign triton tumors, also called "neuromuscular choristomas" or "neuromuscular hamartomas," consist of neural tissue containing mature skeletal muscle in intimate relationship with peripheral nerve. These tumors are rare in the head and neck in children. Ectomesenchymomas are similar tumors consisting of a malignant mesenchymal component, usually embryonal rhabdomyosarcoma, and a neuroectodermal component represented by mature ganglion cells or primitive neuroblastic/neuroectodermal foci (primitive ectomesenchymoma). Benign triton tumors have been regarded as benign, whereas ectomesenchymomas have been operationally considered to be variants of rhabdomyosarcoma. We present here a unique case that combines features of these 2 entities in a recurrent lesion on the tongue of a 35-month-old girl. This lesion raises questions about the "benign" nature of benign triton tumor and its possible relationship to ectomesenchymoma.