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PROGRESS TOWARDS INVESTIGATION OF THE IMPACT OF TARGETED INHIBITION OF POTENTIAL DNA-DAMAGE RESPONSE REGULATION OF TRANSLATION IN IRRADIATED NON-SMALL CELL LUNG CANCER CELL LINES

Johnston, Eleanor (2018) PROGRESS TOWARDS INVESTIGATION OF THE IMPACT OF TARGETED INHIBITION OF POTENTIAL DNA-DAMAGE RESPONSE REGULATION OF TRANSLATION IN IRRADIATED NON-SMALL CELL LUNG CANCER CELL LINES. Master's Thesis, University of Pittsburgh. (Unpublished)

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Abstract

ABSTRACT:
Lung cancer has a 5-year survival rate of 17% with current standard-of-care radiation and chemotherapies (1,2,4,8,9). Immune escape contributes to poor tumor clearance driving the development of therapies that promote anti-tumor immune responses (29-33). The cytotoxic T-lymphocyte response distinguishes cancerous cells by the presentation of mutated endogenous antigens on MHC-1 molecules (26,29-33). Radiation therapy has been shown to increase MHC-1 antigen presentation (22-23,30). It has been proposed that DNA-damage response signaling inhibits translation during radiation induced cell recovery resulting in a post-repair spike in translation and MHC-1 presentation (22-23,30). We hypothesize that ATM and ATR inhibition will disrupt the negative regulation of translation post-radiation to impact the rate of MHC-1 presentation. Increasing antigen presentation before cell recovery from radiation damage may increase the magnitude of mutant antigens to stimulate anti-tumor CD8+ T-cell activation.
This investigation attempted to monitor the impact of DNA-damage response inhibition on translation and subsequent MHC-1 presentation post-radiation at two levels. First, fluctuations in the intracellular peptide pool available for MHC-1 antigen loading was measured via the activity of Transporter associated with Antigen Processing (TAP) responsible for shuttling cytosolic peptides into the ER for MHC-1 antigen loading (20-2226). Fluorescence Recovery After Photobleaching (FRAP) of TAP1-mNeonGreen was used to calculate the lateral diffusion of TAP within the ER membrane as diffusion is inversely correlated to TAP activity. Second, MHC-1 cell surface presentation was measured via flow cytometry.

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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Johnston, Eleanoremj22@pitt.eduemj22
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Committee ChairRomero, Guillermoggr@pitt.eduggr
Thesis AdvisorBakkenist, Christopherbakkenistcj@upmc.educjb38
Committee MemberO'Sullivan, Rodderickrjo@pitt.edurjo
Date: 11 September 2018
Date Type: Publication
Defense Date: 3 August 2018
Approval Date: 11 September 2018
Submission Date: 7 August 2018
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Number of Pages: 48
Institution: University of Pittsburgh
Schools and Programs: School of Medicine > Molecular Pharmacology
Degree: MS - Master of Science
Thesis Type: Master's Thesis
Refereed: Yes
Uncontrolled Keywords: DNA-damage response, NSCLC, Radiation therapy, MHC-1 presentation, Cap-dependent translation
Date Deposited: 11 Sep 2018 18:34
Last Modified: 11 Sep 2018 18:34
URI: http://d-scholarship.pitt.edu/id/eprint/35209

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