Molecular landscape of glucose metabolism in glioblastoma and the normal human brain: A narrative review

Glioblastoma (GBM) is a highly aggressive and lethal brain tumor with limited treatment options, such as the chemotherapeutic agent, temozolomide (TMZ). However, many GBM tumors develop resistance to TMZ, which is a major obstacle to effective therapy. Recently, dysregulated lipid metabolism has emerged as an important factor contributing to TMZ resistance in GBM. The dysregulation of lipid metabolism is a hallmark of cancer and alterations in lipid metabolism have been linked to multiple aspects of tumor biology, including proliferation, migration, and resistance to therapy. In this review, we aimed to summarize current knowledge on lipid metabolism in TMZ-resistant GBM, including key metabolites and proteins involved in lipid synthesis, uptake, and utilization, and recent advances in the application of metabolomics to study lipid metabolism in GBM. We also discussed the potential of lipid metabolism as a target for novel therapeutic interventions. Finally, we highlighted the challenges and opportunities associated with developing these interventions for clinical use, and the need for further research to fully understand the role of lipid metabolism in TMZ resistance in GBM. Our review suggests that targeting dysregulated lipid metabolism may be a promising approach to overcome TMZ resistance and improve outcomes in patients with GBM.

A positive feedback loop of HK3-mediated activation of NF-κB modulates glucose metabolism and promotes proliferation and metastasis in glioblastoma.

Glioblastoma multiforme is a common primary brain tumor, which exhibits an imbalance between glioma cell growth and glucose metabolism. Recent discoveries have found that the multiple pathways and downstream genes involved in the dysregulated metabolic pathway allow tumor to manifest and progress, which is critical to patients with glioblastoma associated with significant systemic and immunosuppression. Moreover, immune microenvironment is considered a major obstacle to generating an effective antitumor immune response. Therefore, identification of patient-specific tumor antigens through highly personalized approach, and effective combination with other therapeutic modalities such as molecular agents targeting tumor metabolic oncogene addiction and potent host immune modulators, may provide targets for more effective therapeutic strategies for glioblastoma. In this review, we aim to highlight the most recent findings regarding glucose uptake and proliferation, cell mobility and to expand our investigations and more comprehensively examine different aspects of glucose metabolism in glioblastoma, such as pentose phosphate pathway (PPP) and its enzymes, metabolic modulation of genetics and epigenetics and key metabolic regulators, importantly, tumor cell-induced glucose deprivation inhibits T-cell glycolysis and immunogenic functions. Furthermore, this review will concentrate on how to discover effective drug targets to regulate glucose metabolism in tumor and T cell growth for future glioblastoma therapies, and the challenges faced by the field of metabolism in tumor immune microenviroment.

Glioblastoma (GBM) is a primary malignant brain tumor with a dismal prognosis, partially due to our inability to completely remove and kill all GBM cells. Rapid tumor recurrence contributes to a median survival of only 15 months with the current standard of care which includes maximal surgical resection, radiation, and temozolomide (TMZ), a blood–brain barrier (BBB) penetrant chemotherapy. Radiation and TMZ cause sphingomyelinases (SMase) to hydrolyze sphingomyelins to generate ceramides, which induce apoptosis. However, cells can evade apoptosis by converting ceramides to sphingosine-1-phosphate (S1P). S1P has been implicated in a wide range of cancers including GBM. Upregulation of S1P has been linked to the proliferation and invasion of GBM and other cancers that display a propensity for brain metastasis. To mediate their biological effects, SMases and S1P modulate signaling via phospholipase C (PLC) and phospholipase D (PLD). In addition, both SMase and S1P may alter the integrity of the BBB leading to infiltration of tumor-promoting immune populations. SMase activity has been associated with tumor evasion of the immune system, while S1P creates a gradient for trafficking of innate and adaptive immune cells. This review will explore the role of sphingolipid metabolism and pharmacological interventions in GBM and metastatic brain tumors with a focus on SMase and S1P.

Diabetes mellitus (DM)-mediated impaired glucose metabolism increase in the glioblastoma (GB) risk by inducing hyperglycemia and hyperinsulinemia. An integral membrane transport protein, glucose transporter 3 (GLUT3) facilitates glucose transport into GB tumor cells. We aimed to explore the regulation of GLUT3 in GB tumors of patients who were concurrently diagnosed with DM. Formalin-fixed paraffin-embedded (FFPE) tumor samples were collected from 93 GB patients and retrospectively analyzed. Of the total, 15 patients were concurrently diagnosed with DM (GB-DM). The role of GLUT3 in tumor aggressiveness was evaluated by analyzing its correlation with Ki67, P53 expression, MALAT1 expression, and peripheral blood hemoglobin A1C (HbA1c) level. T98G cells were treated with empagliflozin and metformin to modulate GLUT3. The RNA expression of GLUT3, SOX2, and MALAT1 was analyzed by real-time qPCR. The lactate levels of T98G cells were measured by Cobas c502 analyzer. A scratch wound assay was performed to investigate the migration rate of T98G cells. GLUT3 expression was lower in GB-DM tumors than in GB-only tumors. In GB-DM, the expression of tumoral GLUT3 and peripheral blood glycated hemoglobin (HbA1c) levels were negatively correlated with P53 and Ki67. A decreased GLUT3 shortened the disease-free survival duration in GB-DM patients. Empagliflozin reduced GLUT3, while metformin-induced GLUT3 in T98G cells. The empagliflozin-mediated GLUT3 suppression induced SOX2 and MALAT1 expressions and influenced the migration capacity of T98G cells. Our findings suggest that the low GLUT3 expression of the tumors of GB-DM patients may induce the production of adenosine triphosphate (ATP) from cellular energy sources other than glucose metabolism. However, further studies are warranted to confirm these results.

RPP25, a 25 kDa protein subunit of ribonuclease P (RNase P), is a protein-coding gene. Disorders associated with RPP25 include chromosome 15Q24 deletion syndrome and diffuse scleroderma, while systemic sclerosis can be complicated by malignancy. However, the functional role of RPP25 expression in glioblastoma multiforme (GBM) is unclear. In this study, comprehensive bioinformatics analysis was used to evaluate the impact of RPP25 on GBM occurrence and prognosis. Differential analysis of multiple databases showed that RPP25 was commonly highly expressed in multiple cancers but lowly expressed in GBM. Survival prognostic results showed that RPP25 was prognostically relevant in six tumors (CESC, GBM, LAML, LUAD, SKCM, and UVM), but high RPP25 expression was significantly associated with poor patient prognosis except for CESC. Analysis of RPP25 expression in GBM alone revealed that RPP25 was significantly downregulated in GBM compared with normal tissue. Receiver operating characteristic (ROC) combined with Kaplan-Meier (KM) analysis and Cox regression analysis showed that high RPP25 expression was a prognostic risk factor for GBM and had a predictive value for the 1-year, 2-year, and 3-year survival of GBM patients. In addition, the expression of RPP25 was correlated with the level of immune cell infiltration. The gene set enrichment analysis (GSEA) results showed that RPP25 was mainly associated with signalling pathways related to tumor progression and tumor metabolism.

The metabolism of glioblastoma (GBM), the most aggressive and lethal primary brain tumor, is flexible and adaptable to different adverse conditions, such as nutrient deprivation. Beyond glycolysis, altered lipid metabolism is implicated in GBM progression. Indeed, metabolic subtypes were recently identified based on divergent glucose and lipid metabolism. GBM is also characterized by an immunosuppressive microenvironment in which myeloid-derived suppressor cells (MDSCs) are a powerful ally of tumor cells. Increasing evidence supports the interconnection between GBM and MDSC metabolic pathways. GBM cells exert a crucial contribution to MDSC recruitment and maturation within the tumor microenvironment, where the needs of tumor-infiltrating lymphocytes (TILs) with antitumor function are completely neglected. In this review, we will discuss the unique or alternative source of energy exploited by GBM and MDSCs, exploring how deprivation of specific nutrients and accumulation of toxic byproducts can induce T-cell dysfunction. Understanding the metabolic programs of these cell components and how they impact fitness or dysfunction will be useful to improve treatment modalities, including immunotherapeutic strategies.

Proliferating embryonic and tumor tissues rely on aerobic glycolysis, or the metabolism of glucose to lactate under oxygenated conditions, to assist in the synthesis of biosynthetic precursors necessary for proliferation. The reliance on aerobic glycolysis may be mediated by the expression of specific metabolic enzymes. Mammalian Hexokinases HK1 and HK2 are 100kDa proteins that phosphorylate glucose to glucose-6-phosphate as the first step of the glycolytic pathway. In normal adult tissues, HK1 is ubiquitously expressed but is particularly prominent in the brain and kidney. HK2 is generally expressed at low levels within adipose tissue and skeletal tissue and negligently in normal brain. In this study, we wished to determine the ontogeny of these isoforms within the developing brain and determine how their expression relates to the extent of aerobic glycolysis in Glioblastoma Multiforme (GBM), a highly aggressive malignant brain tumor. Mining of existing published microarray data found strong expression of HK1 in 1, 2, 4 cell stage but a switch to stronger expression of HK2 in the blastocyst stage, previously reported to rely heavily on aerobic glycolysis. We performed HK2 immunohistochemistry in mice from gestational age E12, E16, post natal 1 month and 2 months. Our results demonstrated an age and cell specific HK1 and HK2 immunoreactivity in embryonic brain tissue with decreased expression of HK2 post-natally. Subsequently, we investigated the expression of HK1 and HK2 isoforms in a panel of GBM cell lines with varying levels of dependence on aerobic glycolysis, as measured by lactate levels and O2 consumption. HK2 but not HK1 expression was higher in GBM cells that had low O2 consumption and high extracellular lactate levels, supporting an association of HK2 with aerobic glycolysis. As HK2 expression is nearly silent in adult brain but expressed in fetal tissue and GBM cells, we hypothesized that DNA methylation/demethylation events may be playing important in its regulation. Adult normal human brain and GBM cell lines that had no HK2 expression were found to be methylated at CpG islands within intron 1 by bisulfite sequencing in contrast to fetal tissue and HK2-expressing GBM cells. The degree of methylation correlated with transcript expression in GBM cell lines. Treatment of U343 and A172 cell lines with 5-azacytidine restored HK2 transcript expression, supporting that HK2 maybe epigenetically regulated. Overall, our results demonstrate that the expression of the HK2 isoform, in contrast to HK1, may be particularly important in tissues relying on aerobic glycolysis for proliferation including embryonic tissue and GBMs. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 40.

BackgroundNovel insights into the pathophysiology of schizophrenia are needed to move the field forward by providing the conceptual framework to facilitate development of new treatment strategies. It is well established that glutamatergic systems are disrupted in schizophrenia, which are intimately linked to metabolic function. While there are many promising new directions, accumulating evidence suggests that bioenergetic function is impaired in the brain in schizophrenia. There are multiple mechanisms in the brain to meet neuronal energy demands, including glycolysis, lactate uptake, and oxidative phosphorylation. In normal brain, neurons and astrocytes are coupled through the astrocyte-neuron lactate shuttle, where astrocytes metabolize glucose to lactate and pyruvate, primary energy substrates that are transported to neurons via monocarboxylate transporters (MCTs). Lactate generated by glycolysis in glial cells constitutively supports synaptic transmission even under conditions in which a sufficient supply of glucose and intracellular adenosine triphosphate (ATP) are present. Interestingly, working memory and other cognitive domains are dependent on the shuttling of lactate from astrocytes to neurons. This process highlights the bioenergetic coupling between astrocytes and neurons that develops as the brain matures, forming a critical biological process in the mature adult brain. We assessed elements of these systems in postmortem brain, testing the hypothesis that there are cell-subtype defects in bioenergetics function in the frontal cortex in schizophrenia.MethodsWell-validated assays were used to assess the activity of three glycolytic enzymes in postmortem dorsolateral prefrontal cortex (DLPFC) samples (n=16/group): lactate dehydrogenase (LDH), hexokinase (HXK), and phosphofructokinase (PFK). Each sample was assayed with and without a specific inhibitor (in duplicate) and normalized to protein loaded into the assay. We also probed for differences in protein expression using western blot analysis. Western blot analyses were run in duplicate using the following antibodies optimized for postmortem brain: MCT1, LDH, LDHA, LDHB, HXK1, glucose transporter 3 (GLUT3). We performed real time quantitative polymerase chain reaction (RT-qPCR) using TaqMan PCR assays (MCT1, MCT4, HXK1, HXK2, LDHA, LDHB, PFK1, GLUT1, and GLUT3) in duplicate on cDNA samples in 96-well optical plates on a Stratagene MX3000P (Stratagene, La Jolla, California). We also coupled laser capture microdissection (LCM) with RT-qPCR from superficial and deep layers of DLPFC using the Veritas Microdissection instrument and CapSure Macro LCM caps (Life Technologies, formerly Arcturus, Mountain View, CA, USA). Similar studies were performed in haloperidol-decanoate or vehicle (sesame oil) treated rats (intramuscular injection every 3 weeks for 9 months).ResultsWe found a 24% decrease in PFK1 mRNA expression in the dorsolateral prefrontal cortex in schizophrenia (p=0.039). We also found decreases in HXK (26%, p=0.002) and PFK (16%, p<0.001) activity in the dorsolateral prefrontal cortex. These changes were not present in haloperidol treated rats. At the cell-level, in pyramidal neurons we found an increase in MCT1 mRNA expression (22%, p= 0.038), and decreases in HXK1 (19%, p= 0.023), PFK1 (22%, p=0.003), GLUT1 (20%, p=0.008), and GLUT3 (20%, p=0.023) mRNA expression. We found increases in MCT1 (17%, p<0.05) and GLUT3 (20%, p<0.05), but not HXK1, PFK1, or GLUT1, mRNA expression in enriched pyramidal neuron samples of antipsychotic treated rats.DiscussionAs the brain develops, bioenergetic organization and the formation of synapses occur simultaneously, creating a fundamentally interdependent system. There is accumulating evidence of implicating a number of abnormalities associated with glucose metabolism, the lactate shuttle, and bioenergetic coupling in schizophrenia, suggesting energy storage and usage deficits in the brain. Bioenergetic deficits and genetic risk for synaptic dysfunction in schizophrenia could contribute to the pathophysiology of this illness. In normal brain, glucose enters cells through GLUTs and is processed by glycolytic enzymes resulting in bioenergetic substrates such as pyruvate. Pyruvate can then be converted to lactate and transported between cells or intracellularly by MCTs to be oxidized in the TCA cycle when neuronal energy demand is high. Our findings of decreased glycolytic enzyme and lactate transporter mRNA expression suggests a decrease in the capacity of pyramidal neurons to generate bioenergetic substrates from glucose via glycolytic pathways. Additionally, if neurons were unable to take up adequate amounts of glucose for glycolysis, the intracellular pool of available pyruvate/lactate for transport into mitochondria may be diminished, ultimately impacting energy supply. It is also possible that there is attenuated glycolysis in pyramidal neurons, with a shift towards pathways that boost protection from oxidative stress (pentose phosphate pathway). Other studies also report region and cell-subtype specific changes in the expression of genes encoding proteins involved in metabolism in this illness. Importantly, the above changes were not attributable to antipsychotic treatment. Both synaptic function and meeting of energetic demands are essential for cognition, and failure of either could contribute to the cognitive symptoms seen in schizophrenia. Augmenting affected systems such as glucose utilization pathways could offer a novel approach to restoring cognitive function in schizophrenia. This could include targeting pro-metabolic substrates pharmacologically.

Redox rewiring in glioblastoma: the thioredoxin system as a precision therapeutic target.

The cellular form of the prion protein (PrPc) is a glycoprotein anchored to the cell membrane by a glycosylphosphatidylinositol moiety. An aberrant form of PrPc that is partially resistant to proteases, PrPres, is a hallmark of prion diseases, which in humans include Cruetzfeldt-Jakob disease (CJD), Gerstmann-Sträussler-Scheinker syndrome, and fatal familial insomnia. We have characterized the major forms of PrP in normal and pathological human brains. A COOH-terminal fragment of PrPc, designated C1, is abundant in normal and CJD brains as well as in human neuroblastoma cells. Sequence analysis revealed that C1 contains alternative NH2 termini starting at His-111 or Met-112. Like PrPc, C1 is glycosylated, anchored to the cell membrane, and is heat-stable. Consistent with the lack of the NH2-terminal region of PrPc, C1 is more acidic than PrPc and does not bind heparin. An additional fragment longer than C1, designated C2, is present in substantial amounts in CJD brains. Like PrPres, C2 is resistant to proteases and is detergent-insoluble. Our data indicate that C1 is a major product of normal PrPc metabolism, generated by a cleavage that disrupts the neurotoxic and amyloidogenic region of PrP comprising residues 106-126. This region remains intact in C2, suggesting a role for C2 in prion diseases.

Dysregulated sphingolipid metabolism is associated with many cancers; allowing cells to evade apoptosis through increases in sphingosine-1-phosphate (S1P) and decreases in ceramides. Ceramides can be hydrolyzed by ceramidases to sphingosine, which can then be phosphorylated by sphingosine kinases to S1P. S1P allows cells to evade apoptosis and increase migration, while shifts toward ceramides favor cell death. Glioblastoma (GBM) exhibits shifts in the sphingolipid balance towards S1P, contributing to chemoresistance and migration. Understanding of sphingolipid metabolism in GBM is still limited, and currently, there are no approved treatments to target the dysregulation. Acid ceramidase (ASAH1), a key enzyme in the production of S1P, is highly expressed in GBM and is associated with worse survival of GBM patients, as per The Cancer Genome Atlas data. To address the altered sphingolipid metabolism and therapeutic resistance in GBM, we explored the efficacy of pharmacologic and genetic inhibition of ASAH1 in both parental and temozolomide (TMZ)-resistant patient-derived xenografts. Cells were infected with ASAH1 shRNA or treated with ASAH1 inhibitors and assessed for cell growth and migration. Our work suggests that pharmacologic inhibition of ASAH1 induces cell death and that this effect is maintained in TMZ-resistant cells. Furthermore, we find a novel role for carmofur, an ASAH1 inhibitor, in the inhibition of GBM migration. Together, these data suggest the potential utility of normalizing the sphingolipid balance in the context of GBM TMZ resistance.

Sphingolipid metabolism is dysregulated in many cancers, allowing cells to evade apoptosis through increased sphingosine-1-phosphate (S1P) and decreased ceramides. Ceramidases hydrolyze ceramides to sphingosine, which is phosphorylated by sphingosine kinases to generate S1P. The S1P allows cells to evade apoptosis by shifting the equilibrium away from ceramides, which favor cell death. One tumor type that exhibits a shift in the sphingolipid balance towards S1P is glioblastoma (GBM), a highly aggressive brain tumor. GBMs almost always recur despite surgical resection, radiotherapy, and chemotherapy with temozolomide (TMZ). Understanding sphingolipid metabolism in GBM is still limited, and currently, there are no approved treatments to target dysregulation of sphingolipid metabolism in GBM. Carmofur, a derivative of 5-fluorouracil, inhibits acid ceramidase (ASAH1), a key enzyme in the production of S1P, and is in use outside the USA to treat colorectal cancer. We find that the mRNA for ASAH1, but not other ceramidases, is elevated in recurrent GBM. When TMZ-resistant GBM cells were treated with carmofur, decreased cell growth and increased apoptosis were observed along with cell cycle perturbations. RNA-sequencing identified decreases in cell cycle control pathways that were specific to TMZ-resistant cells. Furthermore, the transcription factor and G1 to S phase regulator, E2F8, was upregulated in TMZ-resistant versus parental GBM cells and inhibited by carmofur treatment in TMZ-resistant GBM cells, specifically. These data suggest a possible role for E2F8 as a mediator of carmofur effects in the context of TMZ resistance. These data suggest the potential utility of normalizing the sphingolipid balance in the context of recurrent GBM.

Previously, we have demonstrated that IL-13 receptor alpha2 (IL-13Rα2), a high affinity receptor for Th2 cytokine IL-13, is overexpressed in most glioblastoma multiforme (GBM) cell lines and approximately 78% of the patient-derived samples. We have also demonstrated that IL-13Rα2 can be targeted by a number of immunotherapeutic agents including chimeric antigen receptor modified T (CAR-T) cells, targeted lentivirus and adenovirus vectors, and a chimeric fusion immunotoxin consisting of IL-13 and truncated Pseudomonas exotoxin (IL-13-PE). However, the signal transduction initiated by IL-13 in GBM tumor is not known to occur through the IL13Rα2, which has a high affinity for IL-13. We have recently observed that IL-13 can signal through IL-13Rα2 by activating AP-1 transcription factors in human brain tumor cell lines. In this study, we have examined IL-13Rα2 expression in human brain tumor and normal brain specimens, and the subsequent signaling through the AP-1 pathway in situ. Using six human glioblastoma and three astrocytoma specimens, we evaluated the expression of AP-1 transcription factors by immunohistochemistry (IHC) and compared the extent of immunostaining and percent positive fields with three normal brain specimens. Six GBM specimens examined showed high degree of immunostaining for c-Fos, c-Jun, Jun D and Fra-1 (AP-1 family of transcription factors) and a high percentage of positive fields. These specimens also showed strong immunostaining for IL-13Rα2 (4+) in &amp;gt;70% fields (P&amp;lt;0.001 compared to normal brain). Three astrocytoma specimens showed staining for IL-13Rα2 (&amp;gt;2+ and 32% fields, P&amp;lt;0.01 compared to normal brain), but the extent of staining was lower compared to GBM. Similar to IL-13Rα2 expression, the extent of staining and percentage of positive fields for AP-1 transcription factors were highly statistically significant between tumors and normal brain (P&amp;lt;0.001 for GBM compared to normal brain and P&amp;lt;0.01 for astrocytoma compared to normal brain). The extent of immunostaining in GBM was highest for c-Fos (4+, 78% fields) followed by c-Jun (3+, 57% fields), Fra-1 (2+, 70% fields) and Jun-D (2+, 28% fields). Jun-B expression was the lowest among the AP-1 transcription factors (&amp;lt;1+, 7% fields) in GBM specimens. Astrocytoma specimens showed lesser extent of immunostaining for AP-1 members compared to GBM; c-Fos showed 2+ staining and 42% positive fields followed by c-Jun (2+, 12% fields), Fra-1 (2+, 48% fields) and Jun-D (&amp;lt;1+, 18% fields). Jun-B staining intensity was &amp;lt;1+ in only 6% fields. Normal brain specimens showed no immunostaining for AP-1 family members. Our results generally corroborate with data obtained from GBM cell lines and confirm that IL-13 can signal in IL-13Rα2 positive GBM tumors in-situ through the AP-1 pathway, thus indicating that this pathway may be an important target for therapeutic intervention of GBM in addition to targeting IL-13Rα2. Citation Format: Rukmini Bhardwaj, Akiko Suzuki, Pamela Leland, Bharat H. Joshi, Raj K. Puri. Analysis of IL-13 signaling through IL-13Rα2 in human brain tumor specimens in situ [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2438. doi:10.1158/1538-7445.AM2017-2438

Glioblastoma (GBM) is the most malignant brain tumor, characterized by frequent resistance to standard therapy, Temozolomide (TMZ), which is highly related to GBM metabolic rewiring. The transmembrane protein CD47 is known for its role in innate immune suppression by binding to SIRPα on myeloid cells. However, non-canonical CD47 pathway activated by thrombospondin 1 (TSP1) suggests a different role for CD47 in tumor progression, and whether this signaling axis modulates GBM metabolism remains unclear. Our patient biopsy data revealed strong CD47 immunoreactivity in GBM compared with adjacent normal tissue (n=21, p &amp;lt; 0.05). Paired (recurrent/primary) GBM patient RNA-Seq (n=14) shows that CD47 is upregulated in recurrent tumors, and high CD47 levels, along with high TSP1 expression in TCGA GBM database, are associated with poor survival, suggesting the importance of the non-canonical CD47 pathway in GBM. In CD47-overexpressing GBM cells, RNA-Seq revealed enrichment of OXPHOS, fatty acid metabolism, and phospholipid synthesis. Lipidomics confirmed CD47-driven upregulation of cardiolipin (CL), suggesting metabolic rewiring via non-canonical CD47 signaling. Patient tumors and TMZ-resistant GBM cells similarly showed induction of CL biosynthesis and remodeling enzymes, which regulate mitochondrial function by incorporating fatty acids. The key enzyme of monounsaturated FA (MUFA) synthesis, Stearoyl-CoA Desaturase, is increased in recurrent tumors. MUFA-CL is studied to promote stable and efficient mitochondrial respiration, whereas polyunsaturated FA (PUFA)-CL is prone to damage by mitochondrial stress. We found that GBM cells pretreated with MUFA oleate increased the mitochondrial spare capacity compared to PUFA linoleic acid incubation. Further, we showed that TSP1 activates CL processing in GBM cells and suggested that the non-canonical CD47 pathway may promote TMZ resistance by increasing MUFA-CL to prevent TMZ-induced mitochondrial stress. Interestingly, CD47 knockdown using different shRNAs showed distinct gene enrichments between metabolism and immune phagocytosis. Also, anti-sense morpholinos targeting CD47 (CD47M) reduced FA-dependent mitochondrial respiration and the activity of key enzymes involved in CL processing in GBM cells. Moreover, TMZ combined with CD47M significantly reduced the GBM viability. On the other hand, we found that targeting non-canonical CD47 on microglia produced the opposite effect to that in cancer cells, with increased OXPHOS and FA metabolism. This alteration significantly increases microglia-mediated killing of GBM. Further, human GBM organoids showed that CD47M pre-treated microglia increased organoid transparency, indicating increased microglial cytotoxicity toward patient-derived GBM cells. Taking together, our data suggests that targeting the non-canonical CD47 pathway may be a potential approach for TMZ-resistant GBM and enhance microglia-mediated cytotoxicity. Citation Format: Yu-Ting Tsai, Mitra Kooshki, Jamie Sagastume, Valerie Payne, Ashley Szymonski, Pin-Yuan Chen, Jian-Ying Chuang, Tsung-I Hsu, Shay Soker, Glenn Lesser, David Soto-Pantoja. Non-canonical CD47 enhances cardiolipin biosynthesis and remodeling in Temozolomide-resistant establishment and reduces microglia-mediated cytotoxicity in glioblastoma progression [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Brain Cancer; 2026 Mar 23-25; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2026;86(6_Suppl):Abstract nr A017.