Biology subjects

Karcini, A.

Publications and source records attributed to Karcini, A..

4 recordsLinked to original sources

bioRxiv2024

Proteomic Assessment of SKBR3/HER2+ Breast Cancer Cellular Response to Lapatinib and Investigational Ipatasertib Kinase Inhibitors

Modern cancer treatment approaches aim at achieving cancer remission by using targeted and personalized therapies, as well as harnessing the power of the immune system to recognize and eliminate the cancer cells. To overcome a relatively short-lived response due to the development of resistance to the administered drugs, combination therapies have been pursued, as well. To expand the outlook of combination therapies, the objective of this study was to use high-throughput data generation technologies such as mass spectrometry and proteomics to investigate the response of HER2+ breast cancer cells to a mixture of two kinase inhibitors that has not been adopted yet as a standard treatment regime. The broader landscape of biological processes that are affected by inhibiting two major pathways that sustain the growth and survival of cancer cells, i.e., EGFR and PI3K/AKT, was investigated by treating SKBR3/HER2+ breast cancer cells with Lapatinib or a mixture of Lapatinib/Ipatasertib small molecule drugs. Changes in protein expression and/or activity in response to the drug treatments were assessed by using two complementary quantitative proteomic approaches based on peak area and peptide spectrum match measurements. Over 900 proteins matched by three unique peptide sequences (FDR<0.05) were affected by the exposure of cells to the drugs. The work corroborated the anti-proliferative activity of Lapatinib and Ipatasertib, and, in addition to cell cycle and growth arrest processes enabled the identification of several multi-functional proteins with roles in cancer-supportive hallmark processes. Among these, immune response, adhesion and migration emerged as particularly relevant to the ability to effectively suppress the proliferation and dissemination of cancer cells. The supplementation of Lapatinib with Ipatasertib further affected the expression or activity of additional transcription factors and proteins involved in gene expression, trafficking, DNA repair, and development of multidrug resistance. Furthermore, over fifty of the affected proteins represented approved or investigational targets in the DrugBank database, which through their protein-protein interaction networks can inform the selection of effective therapeutic partners. Altogether, our findings exposed a broad plethora of yet untapped opportunities that can be further explored for enhancing the anti-cancer effects of each drug as well as of many other multi-drug therapies that target the EGFR/ERBB2 and PI3K/AKT pathways. The data are available via ProteomeXchange with identifier PXD051094.

pharmacology and toxicology↗

bioRxiv2022

Mapping the Cell-Membrane Proteome to the Cancer Hallmarks

The hallmarks of biological processes that underlie the development of cancer have been long recognized, yet, existing therapeutic treatments cannot prevent cancer from continuing to be one of the leading causes of death worldwide. This work was aimed at exploring the extent to which the cell-membrane proteins are implicated in triggering cancer hallmark processes, and assessing the ability to pinpoint novel therapeutic targets through a combined membrane proteome/cancer hallmark perspective. By using GO annotations, a database of human proteins associated broadly with ten cancer hallmarks was created. Cell-membrane cellular subfractions of SKBR3/HER2+ breast cancer cells used as a model system were analyzed by high resolution mass spectrometry, and high-quality proteins (FDR<3 %) identified by at least two unique peptides were mapped to the cancer hallmark database. Over 1100 experimentally detected cell-membrane or cell-membrane associated proteins, representing ~8 % of the human cell-membrane proteome, were mapped to the hallmark database. Representative membrane constituents such as receptors, CDs, adhesion and transport proteins were distributed over the entire genome and present in every hallmark category. Sustained proliferative signaling/cell cycle, adhesion/tissue invasion, and evasion of immune destruction emerged as prevalent hallmarks represented by the membrane proteins. Construction of protein-protein interaction networks uncovered a high level of connectivity between the hallmark members, with some receptor (EGFR, ERBB2, FGFR, MTOR, CSF1R), antigen (CD44), and adhesion (MUC1) proteins being implicated in most hallmark categories. An illustrative subset of 116 hallmark proteins that included 44 oncogenes, 28 tumor suppressors, and 38 approved drug targets was subjected to a more in-depth analysis. The existing drug targets were implicated mainly in signaling processes. Network centrality analysis revealed that nodes with high degree, rather than betweenness, represent a good resource for informing the selection of putative novel drug targets. Through heavy involvement in supporting cancer hallmark processes, we show that the functionally diverse and networked landscape of cancer cell-membrane proteins fosters unique opportunities for guiding the development of novel therapeutic interventions, including multi-agent, immuno-oncology and precision medicine applications.

cancer biology↗

bioRxiv2021

The SKBR3 Cell-Membrane Proteome: Role in Aberrant Cancer Cell Proliferation and Resource for Precision Medicine Applications

The plasma membrane proteome resides at the interface between the extra- and intra-cellular environment and through its various roles in signal transduction, immune recognition, nutrient transport, and cell-cell and cell-matrix interactions plays an absolutely critical role in determining the fate of a cell. Our work was aimed at exploring the landscape of the cancer cell-membrane proteome responsible for sustaining uncontrolled cell proliferation, and its intrinsic resources for enabling detection and therapeutic interventions. SKBR3/HER2+ breast cancer cells were used as a model system and mass spectrometry for characterizing the proteome. The number of identified cell-membrane proteins exceeded 2,000, with ~1,300 being matched by two or more unique peptides. Classification in four major categories, i.e., proteins with receptor or enzymatic activity, CD antigens, transporters, and cell adhesion proteins, uncovered overlapping roles in biological processes that drive cell growth, apoptosis, differentiation, immune response, adhesion and migration, as well as capabilities for signaling crosstalk and alternate pathways for proliferation. The large number of tumor markers (>50) and putative drug targets (>100) exposed a vast potential for yet unexplored detection and targeting opportunities, whereas the presence of 15 antigen immunological markers enabled an assessment of epithelial, mesenchymal or stemness characteristics. Serum-starved cells displayed altered processes related to mitochondrial OXPHOS/ATP synthesis, protein folding and localization, while serum-treated cells exhibited attributes that support tissue invasion and metastasis. Altogether, our findings advance the understanding of the biological triggers that sustain aberrant cancer cell proliferation, survival and development of resistance to therapeutic drugs, and reveal the vast innate opportunities for guiding immunological profiling and precision medicine applications aimed at target selection or drug discovery.

cancer biology↗

bioRxiv2019

Preferential phosphatidylinositol 5-phosphate binding contributes to a destabilization of the VHS domain structure of Tom1

Tom1 transports endosomal ubiquitinated proteins that are targeted for degradation in the lysosomal pathway. Infection of eukaryotic cells by Shigella flexneri boosts oxygen consumption and promotes the synthesis of phosphatidylinositol-5-phosphate [PtdIns5P], which triggers Tom1 translocation to signaling endosomes. Removing Tom1 from its cargo trafficking function hinders protein degradation in the host and, simultaneously, enables bacterial survival. Tom1 preferentially binds PtdIns5P via its VHS domain, but the effects of a reducing environment as well as PtdIns5P on the domain structure and function are unknown. Thermal denaturation studies demonstrate that, under reducing conditions, the monomeric Tom1 VHS domain switches from a three-state to a two-state transition behavior. PtdIns5P reduced thermostability, interhelical contacts, and conformational compaction of Tom1 VHS, suggesting that the phosphoinositide destabilizes the protein domain. Destabilization of Tom1 VHS structure was also observed with other phospholipids. Isothermal calorimetry data analysis indicates that, unlike ubiquitin, Tom1 VHS endothermically binds to PtdIns5P through two noncooperative binding sites, with its acyl chains playing a relevant role in the interaction. Altogether, these findings provide mechanistic insights about the recognition of PtdIns5P by the VHS domain that may explain how Tom1, when in a different VHS domain conformational state, interacts with downstream effectors under S. flexneri infection.

biophysics↗