Decoding Sex Difference in LKB1-mutant Lung Adenocarcinoma with Clinically Relevant Mouse Models Restricted; Files Only

Abstract

LKB1/STK11 is the second most mutated tumor suppressor gene in lung adenocarcinoma (LUAD), following TP53. Despite its well-established role in tumor progression and metastasis, existing mouse models of LKB1-mutant LUAD have not effectively recapitulated distant metastasis to clinically relevant organ sites. Here, we confirmed metastases to distant organs such as the liver in our KRAS^G12DLKB1^-/-Rosa-Luc genetically engineered mouse model (KLL-GEMM). Using the KLL-GEMM-derived cell line, WRJ388, we conducted in vivo selection to generate WRJ388-BM3, which exhibited robust distant metastasis capacity upon intracardiac injections in nude mice. In vitro characterization revealed the invasive phenotype of WRJ388-BM3 in the 3-D matrix.

 

In parallel, we observed that males were significantly more susceptible to tumor formation compared to female KLL-GEMMs, uniquely recapitulating the clinical sex disparity of LKB1-mutant LUAD. WRJ388-AD1, generated from in vivo selection process, was used to establish tumor colonization with various host backgrounds. NK cells emerged as key contributors to the observed sex difference, which was most pronounced in LKB1-mutant LUADs—likely due to their reduced MHC-I expression, rendering them susceptible to NK cell–mediated 'missing-self' recognition. Flow cytometry analysis of whole blood from syngeneic mice indicated a higher frequency of immature NK cells in females, consistent with trends observed in human data. In addition, baseline frequencies of immature NK cells were significantly higher in females that resisted tumor formation following challenge, compared to those that developed tumors, suggesting a potential association between NK cell maturation states and tumor outcome. Following tumor challenge, immunofluorescence analysis revealed elevated NK cell presence in female lungs, even in the presence of LKB1-mutant tumors. Consistently, gene set enrichment analysis of the TCGA-LUAD dataset demonstrated a stronger NK cell–mediated response in female LKB1-mutant patients, after adjusting for baseline sex differences using data from LKB1-wildtype LUAD patients.

 

Together, our findings establish a robust preclinical platform for modeling metastasis in LKB1-mutant LUAD and uncover a novel sex-biased immunological barrier to tumor development mediated by NK cells. These insights have significant implications for understanding tumor progression and for developing sex-informed therapeutic strategies in LKB1-mutant lung cancer.

Table of Contents

Chapter 1. Introduction 1

1.1.         Lung Cancer 2

1.2.         Heterogeneity of Cancer-driver Mutations in Lung Adenocarcinoma 2

1.3.         Metastasis of Lung Cancer 4

1.4.         STK11/LKB1 as a Tumor Suppressor in Lung Cancer 6

1.5.         Pre-clinical Models for Lung Cancers 9

1.6.         Sex Differences in Human Cancers including Lung Cancer 14

1.7.         Immune Response Mediated by Natural Killer Cells 16

1.8.         Dissertation Goals 20

1.9.         Reference 22

 

Chapter 2. A Novel Model for Distant Metastasis of LKB1-mutant Lung Adenocarcinoma 36

2.1.         Authors 37

2.2.         Abstract 38

2.3.         Introduction 39

2.4.         Method 41

2.5.         Results 44

2.6.         Discussion4 8

2.7.         Figure Legend 50

2.8.         Figures 53

2.9.         References 58

 

Chapter 3. Suppression of LKB1-mutant Lung Adenocarcinoma by Natural Killer Cells from Females60

3.1.         Authors 61

3.2.         Abstract 62

3.3.         Introduction 63

3.4.         Results 65

3.5.         Discussion 73

3.6.         Methods 75

3.7.         References 77

3.8.         Figure Legends 83

3.9.         Main Figures 90

3.10.      Supplementary Methods 95

3.11.      Supplementary Tables 102

3.12.      Supplemental Figure Legends 106

3.13.      Supplemental Figures 114

 

Chapter 4. General Discussion and Future Directions 125

3.1.         Distant metastasis in LKB1-mutant LUAD mouse model 127

3.2.         Establishment of WRJ388-AD1 cells 129

3.3.         NK-mediated suppression on LKB1-mutant lung cancers in female 130

3.4.         Antigen presentation reduced by LKB1 loss as a vulnerability 133

3.5.         Conclusion 134

3.6.         Future Directions 135

3.7.         References 138

About this Dissertation

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• Permission granted by the author to include this thesis or dissertation in this repository. All rights reserved by the author. Please contact the author for information regarding the reproduction and use of this thesis or dissertation.

School	Laney Graduate School
Department	Biological and Biomedical Sciences
Subfield / Discipline	Cancer Biology
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Health Sciences, Immunology Biology, Cell Health Sciences, Oncology
关键词	Mouse model LKB1-mutant lung adenocarcinoma Tumor suppressor Sex disparity Natural killer cells
Committee Chair / Thesis Advisor	Wei Zhou, Emory University
Committee Members	Gregory Lesinski, Emory University Frank Schneider, Emory University Sumin Kang, Emory University Adam Marcus, Emory University

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