Semester

Summer

Date of Graduation

2021

Document Type

Dissertation

Degree Type

PhD

College

School of Medicine

Department

Not Listed

Committee Chair

Lori Hazlehurst

Committee Member

Scott Weed

Committee Member

Michael Craig

Committee Member

Anne Cress

Committee Member

Werner Geldenhuys

Abstract

Multiple myeloma (MM) is a hematological malignancy characterized by the neoplastic proliferation of the plasma cells. MM is a relatively rare cancer that accounts for about 1.8% of all cancers and is the second most common hematologic malignancies, and despite the advancement from untreatable to treatable malignancy, it is yet incurable. Calcium ions (Ca2+) play an important role as second messengers in regulating a plethora of physiological and pathological processes, hence cytoplasmic Ca2+ is tightly regulated with strict spatial and temporal control to initiate, maintain, and terminate appropriate signaling pathways and phenotypes including cellular proliferation, cell cycle control, migration, gene expression, muscle contraction, metabolism, and cell death. Multiple Ca2+ pumps and channels exist in the cell to tightly regulate cytoplasmic Ca2+ levels. The plasma membrane Ca2+ transport ATPase (PMCA), and the Na+/Ca2+ exchanger (NCX) are present on the plasma membrane, the mitochondrial Ca2+ uniporter (MCU) are located on the mitochondrial membrane, and the sarcoplasmic reticulum/endoplasmic reticulum Ca2+-ATPase (SERCA) pump, the 1,4,5-triphosphate receptor (IP3R) and ryanodine receptor (RyR) channels are expressed on SE/ER membranes, all these channels work seamlessly to regulate and redistribute Ca2+ levels among various cellular compartments. The major Ca2+ regulatory pathway in non-excitable cells is the Store-Operated Ca2+ Entry Pathway (SOCE) and one its major contributors is TRPC1. TRPC1 is a member of the TRP protein superfamily and a potential modulator of store-operated Ca2+ entry (SOCE) pathways. While TRPC1 is ubiquitously expressed in most tissues, its dysregulated activity may contribute to the hallmarks of various types of cancers, including breast cancer, pancreatic cancer, glioblastoma multiforme, lung cancer, hepatic cancer, multiple myeloma and thyroid cancer. Our group has developed a novel cyclic peptide referred to as MTI-101 for the treatment of MM. MTI-101 is derived from a 10 D-amino acids peptide HYD1, which was discovered to block cancer cell adhesion to extracellular matrix. It was previously reported that acquired resistance to HYD-1 correlated with repression of genes involved in store operated Ca2+ entry (SOCE): PLCβ, SERCA, ITPR3, and TRPC1 expression. We sought to determine the role of TRPC1 heteromers in mediating MTI-101 induced cationic flux. Our data indicate that consistent with activation of TRPC heteromers, MTI-101 treatment induced Ca2+ and Na+ influx. However, replacing extracellular Na+ with NMDG did not reduce MTI-101-induced cell death. In contrast, decreasing extracellular Ca2+ reduced both MTI-101-induced Ca2+ influx as well as cell death. The causative role of TRPC heteromers was established by suppressing STIM1, TRPC1, TRPC4 or TRPC5 function both pharmacologically and by siRNA, resulting in a reduction in MTI-101-induced Ca2+ influx. Mechanistically MTI-101 treatment induces trafficking of TRPC1 to the membrane and co-immunoprecipitation studies indicate that MTI-101 treatment induces a TRPC1-STIM1 complex. Moreover, treatment with calpeptin inhibited MTI-101-induced Ca2+ influx and cell death indicating a role of calpain in the mechanism of MTI-101-induced cytotoxicity. Finally, components of the SOCE pathway were found to be poor prognostic indicators among MM patients, suggesting that this pathway is attractive for the treatment of MM.

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