Biology subjects

Lazaris, A.

Publications and source records attributed to Lazaris, A..

8 recordsLinked to original sources

bioRxiv2026

Iterative Extracellular Vesicle Protein Co-Expression Biomarker Refinement for Preoperative Classification of Histopathological Growth Patterns in Colorectal Liver Metastasis Patients

Preoperative triage of colorectal liver metastases (CRCLM) by histopathological growth pattern (HGP)--angiogenesis-dependent desmoplastic (dHGP) and vessel co-opting replacement (rHGP)--could guide anti-angiogenic therapy, yet HGP scoring requires resected tissue. We present an extracellular vesicle (EV) inner and outer protein (EVPio) co-expression assay and iterative biomarker refinement for plasma-based HGP classification. We established a minimal, high-throughput plasma pre-processing workflow (low-speed centrifugation and 0.45 m filtration) with comparable EVPio assay performance to size-exclusion chromatography. We established an EV biomarker selection template with growing cohorts--feasibility (n = 3), pilot (n = 9), discovery (n = 67)--ranking candidate protein pairs by signal quality (SNR, CV), redundancy (inter-correlation/orthogonality), and HGP separation (effect size, significance, ROC). This process reduced an initial 19x18 capture/detection set to a focused 9x9 panel (81 co-expression pairs). In a 58-patient CRCLM subset (22 dHGP, 14 rHGP, 22 mixed), three pairs achieved high signal quality with significant differential expression across HGPs. A three-feature linear discriminant model yielded 75.9% cross-validated accuracy (AUC 0.77) for classifying pure dHGP vs. non-dHGP. The results show that co-expression signatures capture defining features of HGP biology while revealing heterogeneity. The proposed EV biomarker refinement template is generalizable and our results show that co-expression signatures capture defining features of HGP biology supporting efforts towards clinically actionable, HGP-driven therapeutic guidance.

bioengineering↗

bioRxiv2025

Glucagon-induced PGC-1α4/PPARγ promotes hepatic lipid storage and macrosteatosis in fasting and MASLD

The liver plays a central role in regulating the transition between fasting and feeding states, coordinating glycogen breakdown, gluconeogenesis, fatty acid catabolism, and lipid storage. Disruptions in this balance contribute to metabolic disorders, including Metabolic dysfunction Associated Steatotic Liver Disease (MASLD). Here, we identify a novel biological pathway downstream of glucagon that promotes the hepatic lipid accumulation associated with fasting. Using gain- and loss-of-function in vivo and in vitro models, we found that glucagon induces sustained expression of PGC-14, promoting lipid uptake and storage in hepatocytes by increasing PPAR{gamma} activity. Increased PPAR{gamma}/PGC-14 promotes hepatic Fsp27/Cidec expression, leading to lipid droplet expansion and triglyceride trapping in liver. Activity of the PPARGC1A alternative promoter and PGC-14 expression were higher in livers of patients with MASLD, and PGC-14 expression correlated with macrosteatosis. Consistently, persistent expression of hepatic PGC-14 in mice fed a western diet promoted macrosteatosis, exacerbated oxidative stress and altered hepatic lipid composition to resemble worsening Metabolic dysfunction Associated Steatohepatitis (MASH) in humans. Our findings demonstrate that glucagon-induced hepatic PGC-14/PPAR{gamma} activity facilitates efficient uptake and storage of lipids during fasting, but over-activation of this coordinated metabolic pathway leads to lipid accumulation and worsening of steatosis in MASLD.

physiology↗

bioRxiv2023

Inhibition of proprotein convertase subtilisin-like kexin type 9 (PCSK9) potentiates anti-angiogenic therapy in colorectal cancer liver metastases

Colorectal cancer liver metastatic (CRCLM) tumours present as two main histopathological growth patterns (HGPs) including desmoplastic HGP (DHGP) and replacement HGP (RHGP). The DHGP tumours obtain their blood supply by sprouting angiogenesis, whereas the RHGP tumours utilize an alternative vascularisation known as vessel co-option. In vessel co-option, the cancer cells hijack the mature sinusoidal vessels to obtain blood supply. Vessel co-option has been reported as an acquired mechanism of resistance to anti-angiogenic treatment in CRCLM. Herein, we showed that inhibiting proprotein convertase subtilisin-like kexin type 9 (PCSK9) via clinically approved PCSK9-neutralizing antibody (Evolocumab) can boost the response of vessel co-opting tumours to anti-angiogenic therapy. Mechanistically, we found that PCSK9 inhibition downregulates runt related transcription factor-1 (RUNX1) expression levels in CRCLM cancer cells in vivo, which its expression positively correlates with the development of vessel co-option. Collectively, these results suggest that inhibiting PCSK9 is a promising way to improve the efficacy of anti-angiogenic therapy against vessel co-opting tumours in CRCLM.

molecular biology↗

bioRxiv2022

Isolation of Extracellular Vesicles from human plasma samples: The importance of controls

There are a number of methods for the isolation of extracellular vesicles (EV) which include the traditional ultracentrifugation to column-based kits available from different companies. Isolation of EVs from complex fluids, such as blood, has several challenges as the detection of low abundance molecules can easily be masked by more abundant proteins, when performing mass spectrometry. For this reason, several commercially available kits contain Thromboplastin D (TP-D) to promote clotting, thus removing clotting factors and abundant proteins resulting in increased detection of proteins. Our study demonstrates that plasma pretreated with Rabbit brain derived TP-D (the most common additive), generated a dynamic range of proteins compared to plasma alone, however, most of these proteins were contaminants introduced from the TP-D (99.1% purity). As an alternative, we tested recombinant TP and demonstrated that although it did not introduce any significant contaminants, we did not see any difference in the detection of proteins. Thus TP-D is not required, and any protein additives must be carefully screened.

cell biology↗

bioRxiv2022

Disruption of integrin alpha-5/beta-1-dependent transforming growth factor beta-1signaling pathway attenuates vessel co-option in colorectal cancer liver metastases

Colorectal cancer liver metastases (CRCLM) have two major histopathological growth patterns (HGPs) including angiogenic desmoplastic HGP (DHGP) and non-angiogenic replacement HGP (RHGP). The RHGP lesions obtain their blood supply through vessel co-option, where the cancer cells hijack the pre-existing blood vessels of the surrounding liver tissue. Consequently, anti-angiogenic therapies are less efficacious in CRCLM patients with RHGP lesions. Recently, we identified a positive correlation between the expression of Angiopoietin1 (Ang1) and the development of vessel co-opted CRCLM lesions in vivo. However, the mechanisms underlying Ang1 upregulation in vessel co-opting CRCLM lesions are unclear. Herein, we demonstrated that transforming growth factor {beta}1 (TGF{beta}1) modulates the expression of Ang1 in hepatocytes in vitro. Significantly, pharmaceutical inhibition of integrin alpha-5/beta-1 (ITG5{beta}1) through ATN-161 impaired TGF{beta}1-dependent Ang1 expression in vitro and in vivo. Moreover, blocking ITG5{beta}1 attenuated the formation of vessel co-opting lesions. Furthermore, treatment with ATN-161 significantly improved survival in tumour-bearing mice. Taken together, our results suggest the molecular mechanism of Ang1 upregulation in vessel co-opting CRCLM and targeting this pathway may serve as promising therapeutic strategy to overcome the development of vessel co-option in CRCLM.

cancer biology↗

bioRxiv2021

Cancer cells induce hepatocytes apoptosis in co-opted colorectal cancer liver metastatic lesions

Vessel co-option in colorectal cancer liver metastases (CRCLM) has been recognized as one of the mechanistic pathways that contribute to resistance against anti-angiogenic therapy. In vessel co-opted CRCLM lesions, the cancer cells are highly motile that move toward and along the pre-existing sinusoidal vessels and hijack them to gain access to nutrient. The movement of cancer cells is accompanied by replacement of the hepatocytes. However, the molecular mechanisms by which this replacement occurs are unclear yet. To examine the involvement of apoptosis in hepatocytes replacement by cancer cells in co-opted lesions, we conducted immunohistochemical staining for chemonaive CRCLM specimens using pro-apoptotic markers antibody, such as cleaved caspase-3 and cleaved poly (ADP-ribose) polymerase-1 (PARP-1). The results suggested overexpression of pro-apoptotic markers in liver parenchyma of co-opted lesions compared to angiogenic lesions, specifically the hepatocytes that are in close proximity to the cancer cells. Importantly, co-culturing hepatocytes with colorectal cancer cells induced overexpression of pro-apoptotic markers in the hepatocytes. Altogether, these results propose that cancer cells could exploit apoptosis to replace the hepatocytes and establish vessel co-option in CRCLM.

cancer biology↗

bioRxiv2021

Ectopic FVIII expression and misfolding in hepatocytes as a potential cause of human hepatocellular carcinoma

Hemophilia A gene therapy targets hepatocytes to express B domain deleted-(BDD) clotting factor VIII (FVIII) to permit viral encapsidation. Since BDD is prone to misfolding in the endoplasmic reticulum (ER) and ER protein misfolding in hepatocytes followed by high fat diet (HFD) can cause hepatocellular carcinoma (HCC), we studied how FVIII misfolding impacts HCC development using hepatocyte DNA delivery to express three proteins from the same parental vector: 1) well-folded cytosolic dihydrofolate reductase (DHFR); 2) BDD-FVIII, which is prone to misfolding in the ER; and 3) N6-FVIII which folds more efficiently than BDD-FVIII. One week after DNA delivery, when FVIII expression was undetectable, mice were fed HFD for 65 weeks. Remarkably, all mice that received BDD-FVIII vector developed liver tumors, whereas only 58% of mice that received N6 and no mice that received DHFR vector developed liver tumors, suggesting that the degree of protein misfolding in the ER increases predisposition to HCC in the context of a HFD and in the absence of viral transduction. Our findings raise concerns of ectopic BDD-FVIII expression in hepatocytes in the clinic, which poses risks independent of viral vector integration. Limited expression per hepatocyte and/or use of proteins that avoid misfolding may enhance safety. Graphical abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

cancer biology↗

bioRxiv2019

PGC-1α isoforms coordinate to balance hepatic metabolism and apoptosis in inflammatory environments

Liver is regularly exposed to changing metabolic and inflammatory environments. It must sense and adapt to metabolic need while balancing resources required to protect itself from insult. PGC-1 is a transcriptional coactivator that both coordinates metabolic adaptation to diverse stimuli and protects against inflammation. However, it is not known how PGC-1 integrates extracellular signals to balance metabolic and anti-inflammatory outcomes. PGC-1 exists as multiple, alternatively spliced variants expressed from different promoters. Primary mouse hepatocytes were used to evaluate the role(s) of different PGC-1 proteins in regulating hepatic metabolism and inflammatory signaling downstream of TNF. PGC-11 and PGC-14 were expressed in hepatocytes and expression analysis uncovered shared and isoform-specific roles for these variants linked to metabolism and inflammation. PGC-11 primarily impacted gene programs of nutrient and mitochondrial metabolism, while TNF signaling revealed that PGC-14 influenced several pathways related to innate immunity and cell death. Gain- and loss-of-function models illustrated that PGC-14 uniquely enhanced expression of anti-apoptotic gene programs and attenuated hepatocyte apoptosis in response to TNF or LPS. This was in contrast to PGC-11, which reduced the expression of a wide inflammatory gene network, but did not prevent liver cell death in response to the cytokine. We conclude that PGC-1 variants have distinct, yet complimentary roles in hepatic responses to metabolism and inflammation and identify PGC-14 as an important mitigator of apoptosis.

molecular biology↗