The primary cause of cancer-related deaths in triple-negative breast cancer (TNBC) is the progression to distant sites from the mammary gland. TNBC is an aggressive disease that lacks hormone and growth factor receptors; thus has no targeted therapies, is highly metastatic, and has a poor prognosis. Focal adhesion kinase (FAK) signaling promotes cell invasion and migration with overexpression correlating with metastatic breast cancer. High levels of inflammatory marker nitric oxide synthase 2 (NOS2) are found in TNBC tissue samples compared to normal mammary epithelial cells. Inflammation and the progression of TNBC and metastasis are critical characteristics of the TNBC tumor microenvironment (TME). Therefore, understanding how NOS2 regulates cell invasion and migration will help elevate the progression of TNBC in metastasis. We will use 4T1, a syngeneic mouse model of TNBC cells, with wildtype (WT) NOS2 or without the NOS2 gene knocked out (KO) with CRISPR/Cas9. We hypothesize that NOS2-mediated NO in TNBC cells regulates FAKs for cell adhesion and migration. We will measure FAKs and invasion in our 4T1 cell lines (NOS2 WT, KO1, and KO2) using microscopy techniques, scratch or wound healing assays, and PET track-etched membrane invasion assays. These experiments aim to identify the role of NOS2-derived nitric oxide (NO) during the migration and progression of TNBC.

High-risk human papillomaviruses (HPVs) are the cause of almost all cervical cancers and a substantial fraction of anogenital and oropharyngeal cancers. HPV16 is responsible for the growing number of oral cancers. HPV infects epithelial tissues and its viral lifecycle exhibits both an early and late stage that coincides with host cell differentiation from basal layer epithelial cells to upper layer differentiated cells. To explore the switch that ensures the completion of HPV’s lifecycle, viral L1 (the major capsid protein) was chosen as a marker. L1 expression is differentiation-dependent. The early stage of the viral cycle modifies signaling pathways, including epidermal growth factor and immune response, which further enables the proliferative state of the cell followed by a signal for late gene expression L1. The L1 marker is expressed in only a subset of infected cells, which poses a significant challenge for exploring HPVs full lifecycle. The research focuses on establishing methods to induce the L1 late viral production in monolayer keratinocytes. According to previous research, transforming growth factor beta (TGF-beta1) activates differentiation. The effect of TGF-beta1 on HPV16-positive oral cells is being tested to see if it activates L1 as a method to consistently increase late expression in HPV-positive cells. The expression of L1 marker will be measured at the RNA-level using RT-qPCR and western blot will be used to confirm protein-level expression. Successful induction of L1 in monolayer allows us to test viral lifecycle by indicating which cells have entered the productive stage of infection.

Androgen receptor (AR) is the primary driver of prostate cancer. Patients with metastases are treated with androgen deprivation therapy (ADT). Currently, ADT is the only way to block AR signaling. However, some patients treated with ADT relapse, resulting in a more advanced metastatic castration-resistant prostate cancer (mCRPC). AR in mCRPC uses cell adhesion-mediated drug-resistant pathways (CAM-DR) to evade cell death. Laminin integrin alpha-6 beta-1 is a driver of CAM-DR and a direct transcriptional target of AR. Transcriptional targeting activates pro-survival signaling by alpha-6 beta-1. Alternative therapies are needed to target AR for mCRPC patients responding poorly to ADT. Alternative therapies could also be used in conjunction with existing therapies, providing more opportunities to patients and increasing efficacy of treatment. The survival mechanism of integrin alpha-6 beta-1 can be targeted by HYD1 (kikmviswkg), a synthetic D-amino acid tumor adhesion peptide. It is hypothesized that HYD1 stabilizes and enriches integrin alpha-6 beta-1 at the cell surface and prevents normal protein internalization and signal transduction, thus impacting the normal function and activity of integrin alpha-6 beta-1. Despite this hypothesis, the effect of androgen stimulation on integrin expression is unknown. This present study utilizes immunoblotting to determine the effect of androgen treatment on the binding of integrin alpha-6 beta-1. Metastatic CRPC cell lines C4-2 and 22RVI will undergo HYD1 treatment and western blotting will be used to determine the impact of androgen stimulation on integrin alpha-6 beta-1 expression.

Studies have shown a link between greater rates and risks of breast cancer in women that work night shifts, such that this type of work has been labeled a probable carcinogen. It has been suggested that the disruption of circadian genes- PER and CRY - could be partly responsible for this increased breast cancer risk. The suprachiasmatic nucleus (SCN) in the brain is the central circadian pacemaker; however, how and if Triple-negative breast cancer (TNBC) cells, an aggressive type of breast cancer that is characterized by the absence of estrogen receptors (ER), progesterone receptors (PR), as well as HER-2 growth factor receptors, express particular circadian clock genes when disconnected from the “master clock” is not extensively researched nor understood. The aim of this study was to answer the following questions: Do triple-negative breast cancer cells (TNBCs) express circadian genes? Do TNBCs exhibit rhythmic circadian gene expression in vitro? Are TNBCs and the circadian genes responsive to external red-light exposure at a wavelength shown to alter mitochondrial function? Cells were cultured in vitro, exposed or not exposed to red light, and then harvested at 6-hour time intervals for 24 hours. The harvested cells endured RNA isolation, followed by cDNA synthesis and PCR to amplify circadian genes PER1, PER2, PER3, CRY1, CRY2. The expression level was compared to constitutive reference genes to determine whether circadian rhythmicity was present in TNBC cells. The first trial has shown evidence for wavelength of light to impact the extent or lack thereof, of circadian rhythmicity.