Glutamoptosis is the induction of apoptotic cell death as a consequence of the aberrant activation of glutaminolysis and mTORC1 signaling during nutritional imbalance in proliferating cells. The role of the bioenergetic sensor AMPK during glutamoptosis is not defined yet. Here, we show that AMPK reactivation blocks both the glutamine-dependent activation of mTORC1 and glutamoptosis in vitro and in vivo. We also show that glutamine is used for asparagine synthesis and the GABA shunt to produce ATP and to inhibit AMPK, independently of glutaminolysis. Overall, our results indicate that glutamine metabolism is connected with mTORC1 activation through two parallel pathways: an acute alpha-ketoglutarate-dependent pathway; and a secondary ATP/AMPK-dependent pathway. This dual metabolic connection between glutamine and mTORC1 must be considered for the future design of therapeutic strategies to prevent cell growth in diseases such as cancer.

Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest tumors, partly due to its intrinsic aggressiveness, metastatic potential, and chemoresistance of the contained cancer stem cells (CSCs). Pancreatic CSCs strongly rely on mitochondrial metabolism to maintain their stemness, therefore representing a putative target for their elimination. Since mitochondrial homeostasis depends on the tightly controlled balance between fusion and fission processes, namely mitochondrial dynamics, we aim to study this mechanism in the context of stemness. In human PDAC tissues, the mitochondrial fission gene DNM1L (DRP1) was overexpressed and positively correlated with the stemness signature. Moreover, we observe that primary human CSCs display smaller mitochondria and a higher DRP1/MFN2 expression ratio, indicating the activation of the mitochondrial fission. Interestingly, treatment with the DRP1 inhibitor mDivi-1 induced dose-dependent apoptosis, especially in CD133⁺ CSCs, due to the accumulation of dysfunctional mitochondria and the subsequent energy crisis in this subpopulation. Mechanistically, mDivi-1 inhibited stemness-related features, such as self-renewal, tumorigenicity, and invasiveness and chemosensitized the cells to the cytotoxic effects of Gemcitabine. In summary, mitochondrial fission is an essential process for pancreatic CSCs and represents an attractive target for designing novel multimodal treatments that will more efficiently eliminate cells with high tumorigenic potential.

Pancreatic ductal adenocarcinoma (PDAC), the most common form of pancreatic cancer, is an extremely lethal disease due to late diagnosis, aggressiveness and lack of effective therapies. Considering its intrinsic heterogeneity, patient stratification models based on transcriptomic and genomic signatures, with partially overlapping subgroups, have been established. Besides molecular alterations, PDAC tumours show a strong desmoplastic response, resulting in profound metabolic reprogramming involving increased glucose and amino acid consumption, as well as lipid scavenging and biosynthesis. Interestingly, recent works have also revealed the existence of metabolic subtypes with differential prognosis within PDAC, which correlated to defined molecular subclasses in patients: lipogenic subtype correlated with a classical/progenitor signature, while glycolytic tumours associated with the highly aggressive basal/squamous profile. Bioinformatic analyses have demonstrated that the representative genes of each metabolic subtype are up-regulated in PDAC samples and predict patient survival. This suggests a relationship between the genetic signature, metabolic profile, and aggressiveness of the tumour. Considering all this, defining metabolic subtypes represents a clear opportunity for patient stratification considering tumour functional behaviour independently of their mutational background.

Renal cell carcinoma (RCC) represents the main renal tumors and are highly metastatic. They are heterogeneous tumors and are subdivided in 12 different subtypes where clear cell RCC (ccRCC) represents the main subtype. Tumor extracellular matrix (ECM) is composed, in RCC, mainly of different fibrillar collagens, fibronectin, and components of the basement membrane such as laminin, collagen IV, and heparan sulfate proteoglycan. Little is known about the role of these ECM components on RCC cell behavior. Analysis from The Human Protein Atlas dataset shows that high collagen 1 or 4A2, fibronectin, entactin, or syndecan 3 expression is associated with poor prognosis whereas high collagen 4A3, syndecan 4, or glypican 4 expression is associated with increased patient survival. We then analyzed the impact of collagen 1, fibronectin 1 or Matrigel on three different RCC cell lines (Renca, 786-O and Caki-2) in vitro. We found that all the different matrices have little effect on RCC cell proliferation. The three cell lines adhere differently on the three matrices, suggesting the involvement of a different set of integrins. Among the 3 matrices tested, collagen 1 is the only component able to increase migration in the three cell lines as well as MMP-2 and 9 activity. Moreover, collagen 1 induces MMP-2 mRNA expression and is implicated in the epithelial to mesenchymal transition of two RCC cell lines via Zeb2 (Renca) or Snail 2 (Caki-2) mRNA expression. Taken together, our results show that collagen 1 is the main component of the ECM that enhances tumor cell invasion in RCC, which is important for the metastasic process.