GLUT1 overexpression enhances CAR T cell metabolic fitness and anti-tumor efficacy.

The tumor microenvironment presents many obstacles to effective chimeric antigen receptor (CAR) T cell therapy, including glucose competition from tumor and myeloid cells. Using mouse models of acute lymphoblastic leukemia (ALL), renal cell carcinoma (RCC), and glioblastoma (GBM), we show that enforced expression of the glucose transporter GLUT1 enhances anti-tumor efficacy and promotes favorable CAR-T cell phenotypes for two clinically relevant CAR designs, 19-28z and IL13Rα2-BBz. In the NALM6 ALL model, 19-28z-GLUT1 promotes T stem cell-like memory formation and prolongs survival. RNA sequencing of these CAR-T cells reveals that the overexpression of GLUT1, but not GLUT3, enriches for genes involved in glycolysis, mitochondrial respiration, and memory precursor phenotypes. Extending these data, 19-28z-GLUT1 CAR-T cells improve tumor control and response to rechallenge in an RCC patient-derived xenograft model. Furthermore, IL13Rα2-BBz CAR-T cells overexpressing GLUT1 prolong the survival of mice bearing orthotopic GBMs and exhibit decreased exhaustion markers. This novel engineering approach can offer a competitive advantage to CAR-T cells in harsh tumor environments where glucose is limiting.

Quantitative assessment of circulating tumor cells in cerebrospinal fluid as a clinical tool to predict survival in leptomeningeal metastases.

PURPOSE: Circulating tumor cells in cerebrospinal fluid are a quantitative diagnostic tool for leptomeningeal metastases from solid tumors, but their prognostic significance is unclear. Our objective was to evaluate CSF-CTC quantification in predicting outcomes in LM. METHODS: This is a single institution retrospective study of patients with solid tumors who underwent CSF-CTC quantification using the CellSearch® platform between 04/2016 and 06/2019. Information on neuroaxis imaging, CSF results, and survival was collected. LM was diagnosed by MRI and/or CSF cytology. Survival analyses were performed using multivariable Cox proportional hazards modeling, and CSF-CTC splits associated with survival were identified through recursive partitioning analysis. RESULTS: Out of 290 patients with CNS metastases, we identified a cohort of 101 patients with newly diagnosed LM. In this group, CSF-CTC count (median 200 CTCs/3 ml) predicted survival continuously (HR = 1.005, 95% CI: 1.002-1.009, p = 0.0027), and the risk of mortality doubled (HR = 2.84, 95% CI: 1.45-5.56, p = 0.0023) at the optimal cutoff of ≥ 61 CSF-CTCs/3 ml. Neuroimaging findings of LM (assessed by 3 independent neuroradiologists) were associated with a higher CSF-CTC count (median CSF-CTCs range 1.5-4 for patients without radiographic LM vs 200 for patients with radiographic LM, p < 0.001), but did not predict survival. CONCLUSION: Our data shows that CSF-CTCs quantification predicts survival in newly diagnosed LM, and outperforms neuroimaging. CSF-CTC analysis can be used as a prognostic tool in patients with LM and provides quantitative assessment of disease burden in the CNS compartment.

A Sox2:miR-486-5p Axis Regulates Survival of GBM Cells by Inhibiting Tumor Suppressor Networks.

Glioblastoma multiforme (GBM) and other solid malignancies are heterogeneous and contain subpopulations of tumor cells that exhibit stem-like features. Our recent findings point to a dedifferentiation mechanism by which reprogramming transcription factors Oct4 and Sox2 drive the stem-like phenotype in glioblastoma, in part, by differentially regulating subsets of miRNAs. Currently, the molecular mechanisms by which reprogramming transcription factors and miRNAs coordinate cancer stem cell tumor-propagating capacity are unclear. In this study, we identified miR-486-5p as a Sox2-induced miRNA that targets the tumor suppressor genes PTEN and FoxO1 and regulates the GBM stem-like cells. miR-486-5p associated with the GBM stem cell phenotype and Sox2 expression and was directly induced by Sox2 in glioma cell lines and patient-derived neurospheres. Forced expression of miR-486-5p enhanced the self-renewal capacity of GBM neurospheres, and inhibition of endogenous miR-486-5p activated PTEN and FoxO1 and induced cell death by upregulating proapoptotic protein BIM via a PTEN-dependent mechanism. Furthermore, delivery of miR-486-5p antagomirs to preestablished orthotopic GBM neurosphere-derived xenografts using advanced nanoparticle formulations reduced tumor sizes in vivo and enhanced the cytotoxic response to ionizing radiation. These results define a previously unrecognized and therapeutically targetable Sox2:miR-486-5p axis that enhances the survival of GBM stem cells by repressing tumor suppressor pathways. SIGNIFICANCE: This study identifies a novel axis that links core transcriptional drivers of cancer cell stemness to miR-486-5p-dependent modulation of tumor suppressor genes that feeds back to regulate glioma stem cell survival.

Anatomic and molecular development of corticostriatal projection neurons in mice.

Corticostriatal projection neurons (CStrPN) project from the neocortex to ipsilateral and contralateral striata to control and coordinate motor programs and movement. They are clinically important as the predominant cortical population that degenerates in Huntington's disease and corticobasal ganglionic degeneration, and their injury contributes to multiple forms of cerebral palsy. Together with their well-studied functions in motor control, these clinical connections make them a functionally, behaviorally, and clinically important population of neocortical neurons. Little is known about their development. "Intratelencephalic" CStrPN (CStrPNi), projecting to the contralateral striatum, with their axons fully within the telencephalon (intratelencephalic), are a major population of CStrPN. CStrPNi are of particular interest developmentally because they share hodological and axon guidance characteristics of both callosal projection neurons (CPN) and corticofugal projection neurons (CFuPN); CStrPNi send axons contralaterally before descending into the contralateral striatum. The relationship of CStrPNi development to that of broader CPN and CFuPN populations remains unclear; evidence suggests that CStrPNi might be evolutionary "hybrids" between CFuPN and deep layer CPN-in a sense "chimeric" with both callosal and corticofugal features. Here, we investigated the development of CStrPNi in mice-their birth, maturation, projections, and expression of molecular developmental controls over projection neuron subtype identity.

Arteriovenous malformation-associated aneurysms in the pediatric population.

OBJECT: Conventional cerebral angiography and treatment for ruptured arteriovenous malformations (AVMs) in children are often performed in a delayed fashion. In adults, current literature suggests that AVM-associated aneurysms may be more likely to hemorrhage than isolated AVMs, which often leads to earlier angiography and endovascular treatment of associated aneurysms. The nature of AVM-associated aneurysms in the pediatric population is virtually unknown. In this report, the authors investigate the relationship of associated aneurysms in a large group of children with AVMs. METHODS: Seventy-seven pediatric patients (≤ 21 years old) with AVMs were treated at the Columbia University Medical Center between 1991 and 2010. Medical records and imaging studies were retrospectively reviewed, and associated aneurysms were classified as arterial, intranidal, or venous in location. Clinical presentation and outcome variables were compared between children with and without AVM-associated aneurysms. RESULTS: A total of 30 AVM-associated aneurysms were found in 22 children (29% incidence). Eleven were arterial, 9 intranidal, and 10 were venous in location. There was no significant difference in the rate of hemorrhage (p = 0.91) between children with isolated AVMs (35 of 55 [64%]) and children with AVM-associated aneurysms (13 of 22 [59%]). However, of the 11 children with AVM-associated aneurysms in an arterial location, 10 presented with hemorrhage (91%). An association with hemorrhage was significant in univariate analysis (p = 0.045) but not in multivariate analysis (p = 0.37). CONCLUSIONS: Associated aneurysms are present in nearly a third of children with AVMs, and when arterially located, are more likely to present with hemorrhage. These data suggest that early angiography with endovascular treatment of arterial-based aneurysms in children with AVMs may be indicated.

Role of fever in ventriculoperitoneal shunt placement after aneurysmal subarachnoid hemorrhage.

BACKGROUND: Central fever is common after aneurysmal subarachnoid hemorrhage (aSAH) and may delay ventriculoperitoneal shunt (VPS) placement. OBJECTIVE: We hypothesize that drain-dependent aSAH patients with central fever or persistent fever after treatment of an identifiable cause are not at an increased risk of infectious VPS failure. METHODS: Patient demographics, radiographic characteristics, temperature, incidence of infection, and shunt failure were prospectively recorded in a consecutive cohort of aSAH patients. Central fever was defined as temperature higher than 38.3°C with no identifiable cause. RESULTS: Of 580 patients, 61 (11%) were drain dependent. Central fever developed in 18, 35 had fever of known etiology, and 8 remained afebrile. There was no shunt failure at discharge, and 2 failures (3.2%) at follow-up were attributed to infection. One patient with central fever (6%), none with fever of identifiable etiology, and 1 (13%) with no fever had infectious shunt failures at a median follow-up of 10.2 ± 3.6 months (P > .05). Nine patients with central fever (50%) and 6 (17%) who were treated for fever of known etiologies had persistent fever at shunt placement. Patients who were febrile on the day of surgery had similar infectious shunt failure rates at discharge compared with those who were afebrile (0% vs 0%; P = 1.0). Similarly, febrile and afebrile patients at VPS insertion had comparable rates of infectious shunt failure at follow-up (7% vs 2%; P = .43). CONCLUSION: aSAH patients with central fever or persistent fever after treatment of fever of identifiable etiology are not at an increased risk of infectious VPS failure.

Gain-of-function polymorphisms of cystathionine ß-synthase and delayed cerebral ischemia following aneurysmal subarachnoid hemorrhage.

OBJECT: Cystathionine ß-synthase (CBS) is an enzyme that metabolizes homocysteine to form H(2)S in the brain. Hydrogen sulfide functions as a vasodilator as well as a regulator of neuronal ion channels and multiple intracellular signaling pathways. Given the myriad effects of H(2)S, the authors hypothesized that patients possessing gain-of-function polymorphisms of the CBS gene will experience a decreased incidence of delayed cerebral ischemia (DCI) following aneurysmal subarachnoid hemorrhage (aSAH). METHODS: Patients were enrolled in a prospective observational database of aSAH outcomes. DNA was extracted from buccal swabs and sequenced for 3 functional polymorphisms of the CBS gene (699C→T, 844ins68, and 1080C→T) by polymerase chain reaction. Serum homocysteine levels (µmol/L) were assayed. Multivariate analysis was used to determine the relationship between CBS genotype and occurrence of both angiographic vasospasm and DCI. RESULTS: There were 87 patients included in the study. None of the polymorphisms investigated were significantly associated with the incidence of angiographic vasospasm. However, after controlling for admission hypertension, patients with the gain-of-function 844 WT/ins genotypes were less likely to experience DCI relative to those with the 844 WT/WT genotype (86 patients, p = 0.050), while the decrease-in-function genotype 1080 TT was more likely to experience DCI relative to those with 1080 CC and CT genotypes (84 patients, p = 0.042). Serum homocysteine levels did not correlate with the extent of either angiographic vasospasm or DCI in this analysis. CONCLUSIONS: Polymorphisms of the CBS gene that impart gain-of-function may be associated with a reduced risk of DCI after aSAH, independent of serum homocysteine. Signaling through H(2)S may mediate protection from DCI following aSAH through a mechanism that does not involve macrovascular vasodilation.

Polymorphisms in complement component 3 (C3F) and complement factor H (Y402H) increase the risk of postoperative neurocognitive dysfunction following carotid endarterectomy.

BACKGROUND: Up to 28% of patients undergoing carotid endarterectomy (CEA) are estimated to experience neurocognitive dysfunction following surgery. The complement cascade plays a central role in ischaemia-reperfusion injury. The authors investigated the effect of common polymorphisms in the complement component 3 (C3F) and complement factor H (CFH Y402H) genes on incidence of neurocognitive dysfunction post-CEA. METHODS: This study examined a nested cohort of prospectively recruited patients receiving elective CEA, who were genotyped for the C3F or Y402H polymorphisms. Each patient underwent a standard battery of eight neuropsychometric tests before, and 1 day and 30 days after, surgery. RESULTS: 57 of 142 (40%) CEA patients had at least one copy of the C3F allele (C3F+), and 17 of 137 (12%) patients had two copies of the CFH Y402H allele (Y402H++). At postoperative day 1, patients were three times (OR 3.05, p=0.045) or six times (OR 6.41, p=0.006) more likely to experience moderate-to-severe neurocognitive dysfunction if they carried the C3F+ or Y402H++ genotype, respectively. Patients with both risk genotypes had an almost eightfold risk of dysfunction (OR 7.67, p=0.046). Right-hand-dominant C3F+ subjects undergoing right-side CEA performed significantly worse on tests of visuospatial function than C3F- subjects. At day 30, C3F+ and Y402H++ genotypes trended towards significance as predictors of dysfunction (p=0.07 and p=0.22, respectively). CONCLUSION: The C3F and Y402H polymorphisms are strong independent predictors of moderate-to-severe neurocognitive dysfunction at 1 day following CEA. Furthermore, patients undergoing right-sided CEA are predisposed to deficits associated with cortex ipsilateral to the operative carotid artery.