www.nature.com/cddis REVIEW ARTICLE OPEN To be or not to be a fat burner, that is the question for cpt1c in cancer cells ✉ Rut Fadó1,2, Sebastian Zagmutt1, Laura Herrero 3,4,5 , Helena Muley1, Rosalía Rodríguez-Rodríguez1,5, Huichang Bi6 , ✉ Dolors Serra 3,4,5 and Núria Casals 1,5 © The Author(s) 2023 There is an urgent need to identify reliable genetic biomarkers for accurate diagnosis, prognosis, and treatment of different tumor types. Described as a prognostic marker for many tumors is the neuronal protein carnitine palmitoyltransferase 1 C (CPT1C). Several studies report that CPT1C is involved in cancer cell adaptation to nutrient depletion and hypoxia. However, the molecular role played by CPT1C in cancer cells is controversial. Most published studies assume that, like canonical CPT1 isoforms, CPT1C is a mediator of fatty acid transport to mitochondria for beta-oxidation, despite the fact that CPT1C has inefficient catalytic activity and is located in the endoplasmic reticulum. In this review, we collate existing evidence on CPT1C in neurons, showing that CPT1C is a sensor of nutrients that interacts with and regulates other proteins involved in lipid metabolism and transport, lysosome motility, and the secretory pathway. We argue, therefore, that CPT1C expression in cancer cells is not a direct regulator of fat burn, but rather 1234567890();,: is a regulator of lipid metabolic reprograming and cell adaptation to environmental stressors. We also review the clinical relevance of CPT1C as a prognostic indicator and its contribution to tumor growth, cancer invasiveness, and cell senescence. This new and integrated vision of CPT1C function can help better understand the metabolic plasticity of cancer cells and improve the design of therapeutic strategies. Cell Death and Disease (2023)14:57 ; https://doi.org/10.1038/s41419-023-05599-1 FACTS INTRODUCTION Metabolic reprograming is a signature of cancer cells that allows CPT1C favors tumor survival in conditions of hypoxia and them to adapt to environmental stresses such as hypoxia and nutrient deprivation and is a prognostic marker in several nutrient scarcity [1]. Currently, there is a search for surrogate human tumors. genetic markers involved in metabolic reprograming that would In the cancer field, CPT1C is considered a marker of fatty acid lead to more accurate diagnosis and prognosis, but also optimized oxidation, which is controversial because it has inefficient catalytic targeted therapy [1]. Studies have shown that lipid metabolism activity, and is located in the endoplasmic reticulum of cells. alterations are associated with tumor growth and progression Evidence from the field of neuroscience allow us to deduce that [2, 3]. Indeed, some cancer cells exhibit a high rate of fatty acid CPT1C is a nutrient sensor that regulates the function of other oxidation (FAO) in producing energy [4, 5], and identified as proteins involved in lipid metabolic reprogramming. emerging therapeutic targets are key genes involved in FAO, such as carnitine palmitoyltransferase 1 (CPT1) [6, 7]. CPT1 enzymes catalyze the conversion of long-chain acyl-CoAs to OPEN QUESTIONS acyl-carnitines to facilitate fatty acid (FA) entry to the mitochondrial matrix for their beta-oxidation [8, 9]. The canonical members of the Identify the molecular mechanism by which CPT1C modulates CPT1 family are CPT1A, a ubiquitous enzyme with greater expression mitochondrial function. in liver, pancreas, and kidney, and CPT1B, expressed mainly in Only 3 proteins are known to date to interact with CPT1C and muscle, heart, and brown fat tissue. Canonical CPT1 enzymes control be regulated by CPT1C sensing of nutrients. What other the limiting step in FAO, and their activity is regulated by malonyl- proteins relevant to tumor growth bind CPT1C? CoA, a metabolite mainly derived from glucose and a precursor of The crystal structure of CPT1 proteins, and the design of new FA synthesis. In the feeding state, CPT1A and CPT1B are inhibited by drugs that specifically target the CPT1C isoform. malonyl-CoA, whose levels increase when the energy balance is positive. By contrast, in conditions of glucose deprivation, malonyl- 1 Basic Sciences Department, Faculty of Medicine and Health Sciences, Universitat Internacional de Catalunya, E-08195 Sant Cugat del Vallès, Spain. 2Institut de Neurociències, Universitat Autònoma de Barcelona, E-08193Bellaterra, Cerdanyola del Vallès, Spain. 3Department of Biochemistry and Physiology, School of Pharmacy and Food Sciences, Universitat de Barcelona, E-08028 Barcelona, Spain. 4Institut de Biomedicina de la Universitat de Barcelona (IBUB), Universitat de Barcelona, E-08028 Barcelona, Spain. 5Centro de Investigación Biomédica en Red de Fisiopatología de la Obesidad y la Nutrición (CIBEROBN), Instituto de Salud Carlos III, Madrid, Spain. 6School of Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515, China. ✉email: bihchang@smu.edu.cn; ncasals@uic.es Edited by Wei Jia Received: 8 September 2022 Revised: 13 January 2023 Accepted: 16 January 2023 Official journal of CDDpress R. Fadó et al. 2 Fig. 1 Kaplan–Meier analysis. Overall survival rate for patients expressing low or high CPT1C mRNA levels (614 and 1144 patients with ovarian cancer and lung cancer, respectively) analyzed with Kaplan–Meier plotter. Dataset and analysis system from [62]. CoA levels decrease, leading to the activation of canonical CPT1 mRNA was upregulated in 81% of cases of tumorous tissues enzymes, and driving FAO and ATP production [8, 9]. Therefore, FAO compared to normal adjacent tissue; interestingly, they also found is finely regulated, depending on nutritional state, through the that CPT1C was expressed in a broad range of human malonyl-CoA-CPT1A/B axis. malignancies and suggested that CPT1C was involved in a parallel CPT1C is an intriguing isoform: despite its high sequence pathway to mTORC-enhanced glycolysis [23]. similarity with CPT1A and CPT1B, it has hardly any catalytic activity Other authors subsequently reported higher CPT1C expression and is mainly localized in the endoplasmic reticulum (ER) rather (mRNA or protein levels) in other types of tumors, e.g., papillary than in mitochondria [10–12], indicating a cellular role different thyroid carcinomas [24] and hepatocellular carcinomas [21] than in from that of the canonical CPT1 enzymes. CPT1C is expressed normal tissues. Moreover, patients with gastric cancer, hepatocarci- almost exclusively in neurons, stem cells, and cancer cells [12–14]. noma, and basal-like breast cancer with high CPT1C expression were Interestingly, CPT1C maintains the ability to bind malonyl-CoA, reported to experience significantly shorter overall survival com- and therefore, to sense the nutritional status of the cells [10, 11]. pared to patients with low CPT1C expression [20–22]. Kaplan-Meier Our group has recently demonstrated that the main role of CPT1C analysis confirms the same poor survival in patients with ovarian or in neurons is to regulate the activity of other proteins, with which lung cancer with high CPT1C expression (Fig. 1), pointing to CPT1C it forms complexes, depending on malonyl-CoA levels [15–17]. as a prognostic marker for several tumor types. Additionally, a study Specifically, those other proteins are involved in the regulation of using The Cancer Genome Atlas (TCGA) dataset showed that CPT1C lipid metabolism, Golgi-mediated secretory transport, and the is part of the 5-gene metabolic signature associated with epithelial- endolysosomal system, impacting on synaptic transmission and mesenchymal transition, metastatic progression, and poor prognosis axon growth [15–19]. Therefore, in neurons, CPT1C is a nutrient across multiple cancers [25]. sensor that regulates neuron metabolism and function. However, not all tumor types benefit from CPT1C expression, However, in the cancer field, most authors assume that CPT1C since human bladder cancer samples compared to normal has the same catalytic function as the other canonical members of adjacent tissues show decreased levels of CPT1C. [26]. its enzyme family, and that CPT1C is a specific marker of FAO. This assumption comes from studies demonstrating that CPT1C CPT1C favors tumor survival and growth in response to stress overexpression and silencing regulates FAO [14, 20–22]. Here, Numerous reports demonstrate the key role of CPT1C in human- we exhaustively review the literature and question the role of derived cancer cell lines in response to metabolic stress. CPT1C CPT1C as a facilitator of FA entry in the mitochondria of cancer overexpression in breast cancer (MCF7) and papillary thyroid cells. Instead, we propose that CPT1C is a sensor of cell nutritional carcinoma (KTC-1) cell lines has been reported to promote survival status and a hallmark of lipid metabolic adaptation. in response to hypoxia or glucose depletion [14, 24]. Accordingly, CPT1C downregulation by siRNAs in colon cancer (HCT116), breast CPT1C in human tumors cancer (MCF7, Hs578T), lung carcinoma (A549), pancreatic The first time that CPT1C was associated with tumor growth was in carcinoma (PANC-1), and thyroid carcinoma (KTC-1 and B-CPAP) 2011 [14], 9 years after its discovery as a new CPT1 isoform [10]. cell lines increases cell death in response to different stressors, Searching for transcripts involved in tumor cell metabolic such as hypoxia, glucose depletion, glycolysis inhibition, and transformation, Zaugg and collaborators [14] found that 2 rapamycin treatment (which mimics amino acid-like starvation) independent screening methods unexpectedly concurred in [14, 24, 27]. Overall, those data indicate that CPT1C expression pointing to the Cpt1c gene. Cpt1c was defined as a new cancer provides some metabolic advantage that favors tumor growth in gene that both conferred rapamycin resistance and was activated the face of different metabolic challenges. This feature may be of by p53. The same authors then analyzed CPT1C expression in high relevance for cells in the central zone of solid tumors, which patients with non-small cell lung carcinoma, finding that, CPT1C are usually poorly irrigated and have low access to nutrients and Cell Death and Disease (2023)14:57 R. Fadó et al. 3 Fig. 2 Regulation of CPT1C expression. Metabolic challenges such as hypoxia and nutrient deprivation increase CPT1C expression in most cancer cells, promoting metabolic adaptation and survival in response to these environmental stressors. On the one hand, low oxygen levels promote accumulation of the HIF1α transcription factor, which is translocated to the nucleus to bind the Cpt1c promoter and activate gene transcription. On the other hand, glucose depletion and glycolysis inhibition increase the AMP/ATP ratio leading to activation of AMPK, which in turn, phosphorylates and activates the CREB transcription factor. Phosphorylated CREB increases expression of the PGC-1β and ERRα transcription factors, ultimately promoting Cpt1c transcription. PPARα activates CPT1C expression by binding directly to the promoter region. p53 is activated by DNA-damaging stimuli and binds to the first intron of the Cpt1c, thereby enhancing CPT1C expression. Interestingly, 2 different miRNAs have been described that downregulate this signaling pathway, one acting on ERRα expression, and the other on CPT1C expression. oxygen. The fact that blood tumor cell lines are those with less tumor progression, also activates CPT1C expression by binding CPT1C expression may reflect their easy availability to nutrients in directly to the promoter region of CPT1C, indicating that CPT1C is a the bloodstream [23]. novel downstream target gene of PPARα [31]. Drug treatments that Interestingly, and in accordance with its role in cell metabolism, affect cellular metabolism, such as metformin (an AMPK activator) CPT1C transcription is precisely regulated in response to distinct and imatinib (a glycolysis inhibitor), also upregulate CPT1C metabolic challenges (Fig. 2). On the one hand, oxygen restriction expression [14, 32]. Finally, note that CPT1C is a bona fide target (0.2%) increases CPT1C transcription through binding of the hypoxia- of the tumor suppressor p53, and therefore, CPT1C expression is inducible factor HIF1α to the Cpt1c promoter [20]. Moreover, tumor- enhanced by DNA-damaging stimuli, such as irradiation and bearing mice subjected to chronic hypoxia showed an increase in certain chemotherapeutic drugs [33]. Interestingly p53 is also CPT1C expression in the tumor mass [14]. Interestingly, this increase in activated by stress stimuli such as hypoxia and starvation, having a CPT1C expression was not found in the normal adjacent tissue, key role in metabolic reprogramming [33]. It has recently been suggesting that hypoxic regulation of CPT1C depends on the unique demonstrated that the mutated form of p53, which is prevalent in circumstances of the tumor microenvironment. human cancer, maintain the ability of activating CPT1C expression On the other hand, glucose depletion or glycolysis inhibition by in basal-like breast cancers through the miR-200c-ZEB2 axis [22]. 2-deoxy-D-glucose also increase CPT1C mRNA levels in different However, in human non-small-cell lung carcinoma, CPT1C pro- cancer cell lines [24, 28]. Wu and collaborators [28] moreover, vided a growth advantage that did not correlate with p53 elegantly demonstrated that this regulation is mediated by the expression or mutation status, indicating that CPT1C is also energy sensor AMPK (which is phosphorylated and activated when involved in a p53-independent metabolic adaptation [14]. ATP levels decrease) through the downstream signaling axis CREB- In summary, it seems that CPT1C upregulation is involved in PGC-1β-ERRα [28]. The proteins PGC-1β and ERRα form functional enabling cellular adaptation to ever-changing metabolic condi- complexes to regulate the expression of genes involved in long- tions in tumor cells (Fig. 2). term and sustained adaptive responses to environmental stressors, such as nutrient deprivation [29]. The Cpt1c promoter has multiple CPT1C involvement in migration, invasion, and cancer cell ERRα binding sites that effectively promote Cpt1c transcriptional senescence activation, as demonstrated by dual luciferase reporter gene assays In addition to its role in meeting metabolic challenges, CPT1C also and ChIP assays [30]. Interestingly, mouse embryonic fibroblasts protects cells from senescence (Fig. 3). Late PANC-1 cells (with deficient for AMPK fail to upregulate CPT1C when exposed to low- many passages) or PANC-1 cells stably transfected with an empty glucose and/or hypoxic conditions [14], confirming that CPT1C vector show a myriad of characteristics that reflect cell senes- expression in response to energy stress is controlled by AMPK. cence, such as increased beta-galactosidase staining, cell mor- Recently, Bi’s group [31] has discovered that PPARα, a ligand- phology changes, cell cycle arrest, decreased telomere elongation, activated transcription factor that regulates lipid metabolism and and reduced gene expression of the senescence-associated Cell Death and Disease (2023)14:57 R. Fadó et al. 4 Fig. 3 Effects of CPT1C silencing in cancer cell metabolism. Downregulation of CPT1C favors cancer cell senescence, as demonstrated by the increased b-galactosidase marker, diminished telomere length, cell cycle arrest, and activation of the senescence-associated secretory phenotype. CPT1C downregulation, as revealed by lipidomic analysis, results in great lipid remodeling, including impaired expression of lipid metabolic genes, decreased cardiolipin levels, diminished phosphatidylcholine/phosphatidylethanolamine (PC/PE) ratio, increased levels of FAs, triacylglycerol, and sphingolipids, and lipid droplet accumulation. CPT1C downregulation impairs mitochondrial morphology, function (decreased mitochondrial respiration, decreased FAO, and altered electrochemical gradient) and biogenesis. All those changes result in decreased ATP levels and oxidative stress. secretory phenotype [27, 34]. The senescence phenotype includes are associated with an increase in ROS formation [27, 35, 38] and a impairment of mitochondrial function, reduced cellular ATP levels, decrease in ATP production [14, 21, 27, 35]. Moreover, genes increased ROS accumulation, and IL-8 inflammation marker involved in the electron transport chain and mitochondrial expression. Interestingly, CPT1C overexpression in senescent cells biogenesis are downregulated [27, 35, 38] (Fig. 3). Interestingly, can reverse the phenotype, while CPT1C downregulation recapi- CPT1C has also been involved in neuron redox homeostasis tulates senescence in diverse cancer cell lines, leading to lower system. Specifically, an increase in the glutathione oxidized form proliferation, migration, and invasion rates, and reduced in vivo (GSSG) was detected in CPT1C KO mouse brains resulting in an tumor growth [14, 20, 21, 27]. Interestingly, of all the members of increased oxidative environment and a reduced capacity to the CPT family, CPT1C exhibits the most substantial effect on cell protect cells from oxidative damage [39]. Accordingly, CPT1C senescence and mitochondrial dysfunction [35]. overexpression in a hippocampal cell line downregulated ROS and The effect of CPT1C on cancer cell senescence described above malondialdehyde (a final product of polyunsaturated fatty acid is also present in normal diploid fibroblast MRC-5 cells [36]. peroxidation) and increased superoxide dismutase (an important Moreover, CPT1C expression in a non-tumorigenic epithelial cell antioxidant mechanism) in response to cell damage [40]. All that line (MCF12A) enhances epithelial-mesenchymal transition phe- evidence points to the idea that CPT1C is necessary to maintain notypes, such as migration, invasion, and anchorage-independent normal mitochondrial performance in cancer cells. growth [22]. The lipidomic profile is also altered by CPT1C silencing (Fig. 3), It is worth mentioning that 2 miRNAs with antitumor roles have as an increase in total FA, triacylglycerol, and sphingomyelin been described that downregulate CPT1C expression and cause content and an accumulation of lipid droplets have been reported cell senescence (Fig. 2). First, miR-377-3p directly targets the [21, 35, 38]. Moreover, an increase in cell membrane stiffness and CPT1C 3’-untranslated region and decreases proliferation, migra- permeability is suggested by a reduced phosphatidylcholine/ tion, and invasion of hepatocarcinoma cells [21]. Accordingly, in phosphatidylethanolamine (PC/PE) ratio [38]. Also reduced is the patient hepatocarcinoma samples, miR-377-3p levels are inversely expression of lipid anabolic enzymes, such as DGAT1/2, and FASN, correlated with CPT1C expression and predict clinical outcomes. and the transcription factor SREBF1. Interestingly, CPT1C silencing Second, miR-1291 downregulates CPT1C expression by targeting in MDA-MB-231 and PANC-1 cells exerts more significant effects the transcription factor ERRα [30]. Consequently, cell proliferation on the mitochondrial function and lipidome than downregulation is decreased, and lipid metabolism is impaired. Targeting these of CPT1A and CPT1B [35]. Overall, those data point to an important miRNAs has been proposed as a therapeutic strategy to combat role for CPT1C in mitochondrial function and in lipid metabolism malignant cancers with high CPT1C expression [21, 23, 30, 37]. in cancer cells that goes beyond FAO modulation (Fig. 3). What metabolic pathways are regulated by CPT1C? The main function of CPT1C is not to burn fat CPT1C silencing in different cancer cell lines leads to important CPT1C can exert a myriad of metabolic effects, but what is the alterations in mitochondrial function and morphology: loss of molecular function of this protein? In the cancer field it is assumed mitochondrial electrochemical gradient [27, 35], reduced mito- that the function of CPT1C—like that of the other canonical CPT1 chondrial respiration (basal oxygen consumption rate and proteins—is to facilitate FA transport to the mitochondrial matrix maximal respiration using glucose as a substrate) [27, 35], reduced to boost FAO. This idea arises from the very first study linking levels of the mitochondrial-specific lipid cardiolipin [35, 38], CPT1C to cancer, which reported that CPT1C overexpression swollen mitochondria with loss of internal crista density, and increases FAO in MCF7 [14]. Subsequent studies have reported abnormal membrane structures [14, 27] (Fig. 3). All these changes FAO changes resulting from CPT1C overexpression or silencing in Cell Death and Disease (2023)14:57 R. Fadó et al. 5 other cancer cell lines [21, 22, 30], even though none of those studies measured the catalytic activity of the protein. The following evidence leads us to reason that CPT1C is not a direct regulator of FA entry in mitochondria: CPT1C overexpression in yeast and some mammalian tumor and embryonic cell lines (PC12 and HEK293) demonstrates that CPT1C does not have efficient CPT1 activity; specifically, it cannot efficiently convert acyl-CoAs into acyl-carnitines, as the limiting step in FAO [10, 11]. The assumption that CPT1C is located in the mitochondrial external membrane of cancer cells is still a controversial assumption. We accordingly analyzed CPT1C localization in MCF7 cells by cell sub-fractionation and by confocal microscopy using specific markers for ER and mitochondria. On the one hand, we found that endogenous CPT1C is present in microsomes and mitochondria-associated membranes but is hardly observed in crude mitochondria (Fig. 4A and Supple- mental data). On the other hand, we found that CPT1C fused to a fluorescent protein colocalizes mainly with ER and, to a lesser extent, with mitochondria (Fig. 4B). Those findings would indicate that CPT1C is an ER-resident protein present on the ER side of mitochondria–ER contacts, corroborating published data regarding neurons and stem cells [12, 13]. The fact that CPT1C expression in cancer cells increases with oxygen depletion [14, 20, 24] does not fit with the idea that CPT1C exclusively upregulates FAO. In conditions of hypoxia, cells enhance anaerobic glycolysis over FAO [41]. In fact, ATP generation derived from FADH2 and NADH formed during beta-oxidation requires oxygen. Accordingly, enhanced levels of HIF1α, which plays a fundamental role in cancer hypoxia adaptation driving transition from oxidative to glycolytic meta- bolism [41–43] correlate with lower expression of FAO-related genes [44], but activates CPT1C expression [20]. All these data suggest that enhanced expression of CPT1C by HIF-1α might support tumor progression by other ways nonrelated to FAO. As explained above, CPT1C expression affects the whole mitochondrial function in cancer cells (glucose-mediated respira- tion, morphology, electrochemical gradient of the membrane, etc) [27, 35, 38]. Therefore, FAO alterations observed in CPT1C silencing may be a consequence of global mitochondrial function changes rather than a direct effect on FA transport to mitochondria. CPT1C is involved in proliferation, migration, and invasion of cancer cells in conditions of full glucose availability [14, 20–22, 27]. Altogether, those findings lead us to conclude that CPT1C, more than just an enzyme that facilitates FA transport to the mitochondria for FAO, plays a more general role in cancer cells (see Fig. 3). CPT1C is a nutrient sensor Before rethinking the role of CPT1C in cancer cells, it is worthwhile reviewing its function in neurons. Our group has recently demonstrated that CPT1C in neurons acts as a nutrient sensor that regulates the function of other proteins involved in cell metabolism [9]. Specifically, CPT1C regulates the activity of its interacting proteins depending on malonyl-CoA, whose levels recapitulate nutrient availability. Importantly, malonyl-CoA levels SAC1 is a phosphatidylinositol (PI4P) phosphatase that reg- increase when the energy balance is positive, and decrease when ulates PI4P levels in the Golgi and is involved in cholesterol and nutrients are scarce [45]. MNR studies demonstrate that the 5‘ tail sphingolipid transport from the trans-Golgi network (TGN) to of CPT1 switches between 2 conformations—open and closed— the plasma membrane. SAC1 is also a negative regulator of the depending on malonyl-CoA binding [46, 47]. Therefore, we secretory pathway [48]. Dysregulation of SAC1 and Golgi PI4P hypothesize that malonyl-CoA binding triggers a conformational levels, which features in many tumors, is often associated with change in CPT1C that affects the activity of its interacting proteins. the release of proteins that remodel the extracellular matrix, To date, 3 proteins that form complexes with CPT1C and are promoting angiogenesis and cell motility, and is also associated regulated by malonyl-CoA have been described: with poor tumor prognosis [49, 50]. Our group has Cell Death and Disease (2023)14:57 R. Fadó et al. 6 Fig. 4 Human CPT1C is mainly localized in ER and other mitochondrial proteins, thereby regulating their activity. mitochondria–ER contact sites in cancer cells. A MCF7 cells were Overall, we propose that CPT1C does not directly regulate FAO by seeded, collected and isolated by cellular subfractionation and mediating FA entry to mitochondria, but indirectly, whether by Percol gradient in order to separate cytoplasm (Cyto), crude buffering malonyl-CoA locally at mitochondria–ER contact sites, or mitochondria (CMito, mitochondria and other contaminating membranes), microsomes (fraction containing the ER), and by interacting with other proteins involved in mitochondrial mitochondria-associated membranes (MAMs, fraction containing function (Fig. 5). mitochondria–ER contacts). Also kept was whole lysate (input). All In summary, we are of the opinion that CPT1C in cancer cells fractions were prepared for the same total protein concentration behaves as a nutrient sensor that facilitates tumor progression and and processed by Western blotting, and equal volumes of each metabolic adaptation to environmental stressors through regulation fraction were loaded. Endogenous CPT1C was mainly localized at ER of SAC1, protrudin, ABHD6, and possibly other proteins (Fig. 5). and MAMs. B MCF7 cells were transfected with human CPT1C or CPT1A tagged with mTurquoise2 (mTurq2); 48 h later, Mitotracker Orange (200 nM) was added, and incubation was 30 min. The cells CONCLUSIONS AND FUTURE PERSPECTIVES were then fixed. and calnexin (used as an ER marker) was detected by immunocytochemistry. CPT1C mainly colocalized with the ER CPT1C is an ER transmembrane protein with inefficient catalytic (orange spots), while CPT1A colocalized with mitochondria (light activity, and as such, is not able to directly mediate FA transport to blue spots). The colocalization of CPT1C with mitochondria (the few mitochondria for FAO. Instead, CPT1C acts as a malonyl-CoA light blue spots) may correspond to ER–mitochondria contacts. The sensor that, in regulating the function of other proteins such as scale bar in B is 10 µm. SAC1, ABHD6, and protrudin, also regulates key cancer cell processes such as lipid metabolism, mitochondrial function, secretory pathway, and protrusion growth. Since malonyl-CoA levels fluctuate depending on nutrient availability, CPT1C could be demonstrated that CPT1C downregulates SAC1 activity and said to be a nutrient sensor involved in metabolic reprograming of increases PI4P levels in the ER-TGN contact sites depending on cancer cells to be able to adapt to constantly changing conditions malonyl-CoA [16]. In this way, CPT1C may drive lipid membrane in the tumor microenvironment. remodeling and favor the secretory pathway, therefore enhan- In relation to FAO, since our results show that CPT1C is present cing cell migration and invasion (Fig. 5). at mitochondria–ER contact sites, we hypothesize that CPT1C may Protrudin, which is present in ER-late endosome contact sites, locally buffer malonyl-CoA and indirectly regulate CPT1A/B activity regulates the anterograde transport of late endosomes and FAO. Identifying novel mitochondrial proteins that interact necessary for axon growth in neurons [51]. In tumors, protrudin with CPT1C would throw light on the wide spectrum of CPT1C orchestrates invadopodia maturation, resulting in tumor cell effects on mitochondrial function. invasion and metastasis [52, 53]. Interestingly, Palomo–Guerrero Interestingly, in metabolism, neurons and cancer cells are and collaborators (17) demonstrate that CPT1C regulates similar in that they do not usually rely on fats for energy fuel. In protrudin function depending on malonyl-CoA levels, suggest- neurons, lipids are reserved for the constant membrane remodel- ing that CPT1C can modulate tumor invasion depending on ing that takes place during spinogenesis, synaptic plasticity, and nutrient availability (Fig. 5). interconnectivity; in cancer cells, lipids are reserved for continuous ABHD6, a hydrolase of monoacylglycerols, regulates levels of membrane synthesis during cell division and growth. Therefore, it the most abundant endocannabinoid, 2-araquidonoylglycerol, is not unreasonable to think that cancer cells may take advantage which is a modulator of neurotransmitter release through its of the neuronal protein CPT1C for their own benefit, i.e., to act as a binding to the endocannabinoid receptor CB1 at the neuron nutritional sensor that regulates lipid metabolism in such a presynaptic membrane [54]. Importantly, CB1 has also been restricted context. found in the external mitochondrial membrane, where it Future research should deepen understanding of the CPT1C- directly controls mitochondrial respiration and energy produc- regulated molecular pathways that drive cell metabolic repro- tion [55]. ABHD6 also hydrolyzes bis(monoacylglycero)phos- gramming. How does CPT1C drive survival in conditions of phate a key lipid for endolysosomal lipid sorting [56]. hypoxia and nutrient depletion? Which CPT1C-interacting proteins Interestingly, ABHD6 has been associated with cancer growth are involved? More specifically, how does CPT1C regulate and metastasis in different tumor types [57–59]. Since CPT1C mitochondrial function? Is it only by buffering the malonyl-CoA, upregulates ABHD6 activity when malonyl-CoA levels decrease or does it regulate the function of a specific mitochondrial [17], we postulate that 2-araquidonoylglycerol (2-AG) and protein? Are high CPT1C-expressing cancers more dependent on bis(monoacylglycero)phosphate (BMP) levels will decrease with lipids as fuel? A better understanding of CPT1C downstream nutrient scarcity, resulting in enhanced mitochondrial function pathways would undoubtedly help us draw the complex and and the endolysosomal-mediated lipid transport changes dynamic picture of cancer cell metabolic plasticity. necessary for the metabolic adaptation of cancer cells (Fig. 5). An important issue that needs to be addressed in the near future is the search for drugs that target CPT1C, and so prevent Other interactors have been described for CPT1C, including the metabolic adaptation of cancer cells to hypoxia or nutrient hydrolase ABHD12 [60]. ABHD12 silencing suppresses breast depletion, as supportive treatment to conventional therapy. In cancer cell proliferation, migration, and invasion [61]. Therefore, xenograft mouse models, CPT1C depletion has been shown to even though we still do not know how CPT1C regulates ABHD12 reduce tumor growth and development, and, in several types of activity, we cannot rule out a role for the CPT1C-ABHD12 complex human tumors, low CPT1C expression is associated with increased in cancer cell progression. overall survival. Because CPT1C is expressed only in the brain, In this context, how can we explain the observation that CPT1C while CPT1A and CPT1B are widely expressed in peripheral tissues, expression modulates FAO? Although it may well be through the a promising anticancer strategy—as suggested by the group of interaction with ABHD6, as explained above, we propose 2 Tak W. Mak [23, 37]—is the development of small drugs or sh- additional putative explanations. First, CPT1C may indirectly RNA-carrying nanomedicines that specifically target CPT1C but are regulate CPT1A/B activity by locally buffering malonyl-CoA at unable to penetrate the blood–brain–barrier [23, 37]. Since CPT1C mitochondria–ER contact sites. If so, CPT1C overexpression would shows high homology with canonical isoforms, it is essential to sequester malonyl-CoA, leading to an increase in CPT1A/B activity understand small conformational variations between isoforms that and in FAO. Second, we cannot rule out that CPT1C at may be druggable. Unfortunately, CPT1 protein structure predic- mitochondria–ER contact sites may interact with CPT1A/B or tions as provided by the AlphaFold database are not sufficiently Cell Death and Disease (2023)14:57 R. Fadó et al. 7 Fig. 5 CPT1C regulates the function of key proteins involved in cancer growth and metastasis. CPT1C has inefficient catalytic activity but is able to interact with and regulate the function of other proteins. At the membrane contact sites between the ER and the trans-Golgi network (TGN), CPT1C downregulates the activity of the phosphatase SAC1, favoring the increased PI4P levels necessary for the secretory pathway, and consequently, for remodeling of the extracellular matrix, angiogenesis, and cancer cell migration and invasion. Moreover, PI4P at TGN is necessary for efficient transport of cholesterol and sphingolipids to the plasma membrane, and so can contribute to those processes. At the ER-late endosome (LE) contacts, CPT1C enhances the function of protrudin, which is necessary for anterograde LE transport and for protrusion growth, and therefore, for tumor invasion and metastasis. CPT1C also interacts with the hydrolase ABHD6 and downregulates its activity. This downregulation of ABHD6 increases levels of the endocannabinoid 2-araquidonoylglicerol (2-AG), which, through the CB1 receptor, possibly regulates mitochondrial function. ABHD6 also controls levels of the bis(monoacylglycero)phosphate (BMP) lipid, which is enriched in the intraluminal vesicles of LE and is involved in lipid metabolism and transport. ABHD6 has been associated with tumor growth and metastasis. 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Lipidomics reveals ACKNOWLEDGEMENTS carnitine palmitoyltransferase 1C protects cancer cells from lipotoxicity and This study was supported by the Spanish Ministry of Science and Innovation (MCIN) senescence. J Pharm Anal. 2021;11. https://doi.org/10.1016/j.jpha.2020.04.004. (PID2020-114953RB-C21 to LH and DS, and PID2020-114953RB-C22 to NC and RR co- 39. Lee J, Wolfgang MJ. Metabolomic profiling reveals a role for CPT1c in neuronal funded by the European Regional Development Fund), the Biomedical Research oxidative metabolism. BMC Biochem. 2012;13:23. Center in Pathophysiology of Obesity and Nutrition (CIBEROBN) (Grant CB06/03/0001 40. Ding Y, Zhang H, Liu Z, Li Q, Guo Y, Chen Y, et al. Carnitine palmitoyltransferase 1 to LH), the Merck Health Foundation (to LH), the Government of Catalonia (CPT1) alleviates oxidative stress and apoptosis of hippocampal neuron in response (2017SGR278 to DS), and the Marató de TV3 Foundation (201627-30 to DS and to beta-Amyloid peptide fragment Aβ25-35. Bioengineered. 2021;12:5440–9. NC). Figures were created with BioRender.com. Cell Death and Disease (2023)14:57 R. Fadó et al. 9 AUTHOR CONTRIBUTIONS Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims NC, RF, RR-R, and DS performed study design; RF, SZ, LH, and HM performed in published maps and institutional affiliations. development of methodology and analysis and interpretation of data; RF, SZ, LH, HM, RR-R, HB, DS, and NC performed writing, review and revision of the paper. All authors read and approved the final paper. 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To view a copy of this license, visit http:// Correspondence and requests for materials should be addressed to Huichang Bi or creativecommons.org/licenses/by/4.0/. Núria Casals. Reprints and permission information is available at http://www.nature.com/ © The Author(s) 2023 reprints Cell Death and Disease (2023)14:57