2025 Theses Doctoral

Chromosomal instability as a driver of metastasis and cancer immune evasion in non-small cell lung cancer

Caprio, Lindsay A.

Up to 40% of all diagnosed brain metastases (BMs), a complication which portends an especially dismal prognosis despite recent therapeutic advances, originate from non-small cell lung cancers (NSCLC). NSCLCs which are characterized by co-occurring oncogenic KRAS and inactivating STK11 mutations, ~15% of all diagnosed cases, have an enhanced brain metastatic potential, and are particularly resistant to immune checkpoint blockade (ICB).

The salient biology of the treatment-naïve NSCLC BM ecosystem and tumor-intrinsic mechanisms by which STK11-mutant NSCLCs promote aggressive cancer behavior have both remained understudied. Here, we performed comprehensive, single nuclei RNA sequencing (snRNA-seq) coupled with whole genome sequencing to profile a cohort of NSCLC primary tumors (PTs) and BMs, validating our findings with extensive computational and matched patient tissue cohorts. We show that i) NSCLC BMs are enriched for computational and tissue surrogates of chromosomal instability (CIN) compared to PTs, ii) a pre-existing, CINhigh population which adopts neural-like features exists in NSCLC PTs and expands in BM samples, and iii), NSCLC BMs are characterized by a particularly immunosuppressive tumor microenvironment, altogether corroborating findings our group made in treatment-naïve melanoma BMs and in a pan-cancer BM analysis.

In our sequenced cohort, we found that tumors with STK11 mutation were particularly enriched for CIN. We experimentally confirmed this finding in vitro, showing that human and murine cancer cell lines with inactivating STK11-mutations, which encodes LKB1 protein, have higher rates of anaphase chromosome mis-segregation, and higher percentage of micronuclei formation than their LKB1-wild type counterparts. To facilitate the identification of putative CINhigh models, we developed an artificial-intelligence (AI) based tool, micronuclAI, to score cells grown in vitro for the presence of micronuclei. Through direct genetic modulation of CIN levels in LKB1-proficient and -deficient models, we show that CIN dictates type I IFN production in vitro and responses to ICB in vivo.

Additionally, we highlight that loss of LKB1 itself may promote CIN, with restoration of a functional allele leading to the rescue of immunogenic output and sensitivity to ICB. Finally, we show that functional relief of CIN through genetic knockout or pharmacological inhibition of cGAS, followed by re-engagement of STING, equilibrates the pathway towards its physiological function. Coupled with STING agonist or ICB therapy in vivo, cGAS-deficient tumors become overwhelmingly infiltrated by anti-tumor immune populations, leading to significant tumor eradication.

Overall, this work highlights cGAS inhibition in combination with ICB or STING agonists as an attractive therapeutic strategy for aggressive, chromosomally unstable NSCLC.