Anaplastic lymphoma kinase (ALK) gene rearrangement is a known oncogenic driver in non-small cell lung cancer (NSCLC). ALK tyrosine kinase inhibitors (TKIs) have been clearly shown to produce excellent therapeutic effects and prolong survival in patients with this gene mutation. According to current treatment guidelines, ALK inhibitors are the first-line treatment of choice for ALK-positive advanced NSCLC patients. However, although ALK TKIs are very effective, there is still a small group of patients who do not achieve good treatment outcomes, developing resistance and tumor progression within 3 to 6 months of initial ALK TKI use. This is called primary resistance. Intrinsic resistance to ALK inhibition occurs when the best clinical response after first-generation and second/third-generation TKI treatment is disease progression. Approximately 5-7% of cases after crizotinib treatment, 9% after ceritinib treatment, and 25% after lorlatinib treatment show no response to treatment, and no specific ALK mutation has been found to explain the occurrence of primary resistance. Currently, many different resistance mechanisms are known, some of which are still ALK-related, while others are ALK-independent alternative survival pathways. However, most research focuses on acquired resistance, with very few studies on primary resistance, only a few case reports. Therefore, this study aims to explore the primary ALK TKI resistance mechanisms.

The investigators plan to explore the incidence and mechanisms of primary ALK TKI resistance in ALK-positive advanced NSCLC patients who develop primary resistance or rapid progression (within 3-6 months) during ALK inhibitor treatment by re-obtaining tumor samples for genetic analysis.

Lung cancer is the most common cause of cancer death in Taiwan and worldwide [1]. The traditional chemotherapy, platinum-doublet chemotherapy, provided limited survival benefit. In the recent decades, the major evolution in lung cancer is the molecular subtyping of NSCLC, especially lung adenocarcinoma, which has resulted in a paradigm shift in the management of patients in advanced disease. The target therapy provided better treatment response and longer overall survival. For example, patients with NSCLC harboring Epidermal growth factor receptor (EGFR) mutations can get remarkable benefit and longer survival from EGFR tyrosine kinase inhibitors (TKIs) treatment comparing with the traditional platinum-base chemotherapy [2].

Anaplastic lymphoma kinase (ALK) gene rearrangements define a distinct molecular subset of NSCLC, typically occurring in younger, non-smoking individuals with adenocarcinoma histology [3]. The discovery of ALK fusions, particularly EML4-ALK, revolutionized the treatment of these patients with the development of ALK TKIs. First-generation ALK inhibitors, such as crizotinib, demonstrated remarkable initial responses, significantly improving progression-free survival (PFS) compared to standard chemotherapy [4]. However, the emergence of resistance, both primary and acquired, remains a major clinical challenge.

Primary resistance, defined as the lack of initial response or early progression within the first few months of ALK TKI therapy, occurs in a subset of patients and represents a significant obstacle to optimal treatment outcomes [5]. Primary resistance to ALK inhibition occurs, even with advanced TKI therapies, when the best clinical outcome is disease progression. Notably, approximately 5-7% of patients following crizotinib, 9% following ceritinib, and 25% following lorlatinib demonstrated a lack of response, and in these cases, no identifiable ALK mutations were detected [6]. Unlike acquired resistance, which typically arises from secondary mutations within the ALK kinase domain or activation of bypass signaling pathways, the mechanisms underlying primary resistance are less well understood.

Several potential factors contribute to primary resistance. One key aspect involves inherent tumor heterogeneity. NSCLC tumors are complex ecosystems with diverse subclones, some of which may harbor pre-existing genetic alterations that confer resistance to ALK inhibition [7]. These alterations might include co-occurring mutations in genes involved in cell signaling pathways, such as KRAS, EGFR, or TP53, which can bypass ALK dependence [6]. Furthermore, variations in ALK fusion isoforms or copy number alterations may also influence initial drug sensitivity. For example, some less common ALK fusion variants might possess distinct structural properties that reduce their affinity for ALK TKIs.

Another potential mechanism involves the tumor microenvironment (TME). The TME, composed of stromal cells, immune cells, and extracellular matrix, plays a critical role in tumor growth and drug response [8]. Components of the TME, such as cancer-associated fibroblasts (CAFs) and immunosuppressive cells, can secrete growth factors and cytokines that activate alternative signaling pathways, thereby promoting resistance to ALK inhibition. Additionally, the TME can create a physical barrier that limits drug penetration and efficacy.

Furthermore, the complexity of ALK signaling networks and the potential for activation of bypass pathways contribute to primary resistance. For example, activation of the EGFR or MET signaling pathways can provide alternative growth signals, bypassing the need for ALK activation [9]. Understanding these bypass pathways is crucial for developing combination therapies that can overcome primary resistance.

Investigating primary resistance requires a comprehensive approach that integrates genomic, transcriptomic, and proteomic analyses. Next-generation sequencing (NGS) can identify pre-existing genetic alterations that confer resistance. Single-cell RNA sequencing can dissect tumor heterogeneity and identify resistant subclones. Proteomic profiling can reveal activation of bypass signaling pathways. Clinical studies analyzing pre-treatment tumor samples and correlating genomic and clinical data are essential for identifying predictive biomarkers of primary resistance.

In summary, primary resistance to ALK TKIs in NSCLC is a complex phenomenon driven by multiple factors, including tumor heterogeneity, TME interactions, pharmacokinetic variations, and activation of bypass signaling pathways. Further research is needed to elucidate the precise mechanisms underlying primary resistance and to develop strategies to overcome this clinical challenge, ultimately improving outcomes for patients with ALK-positive NSCLC.

Inclusion Criteria:

1. Age ≥ 18 years old.
2. Histologically or cytologically diagnosed NSCLC
3. ALK gene fusion detected at initial diagnosis of NSCLC
4. Patients exhibiting primary resistance to ALK-TKI therapy, defined as the absence of initial response or progression within 3 to 6 months of treatment initiation, were included.
5. Able and willing to provide written informed consent prior to performing any study related procedures and to comply with the study protocol.

Exclusion Criteria:

1. Unable to provide written inform consent
2. The patients had received other systemic treatments except ALK TKI.
3. Unable to undergo tumor biopsy or venipuncture

ALK-positive advanced NSCLC patients who develop primary resistance or rapid progression (within 3-6 months) during ALK inhibitor treatment.