Rearranged During Transfection Research Articles

Loss of tumor suppressor TMEM127 drives RET-mediated transformation through disrupted membrane dynamics

Internalization from the cell membrane and endosomal trafficking of receptor tyrosine kinases (RTKs) are important regulators of signaling in normal cells that can frequently be disrupted in cancer. The adrenal tumor pheochromocytoma (PCC) can be caused by activating mutations of the rearranged during transfection (RET) receptor tyrosine kinase, or inactivation of TMEM127, a transmembrane tumor suppressor implicated in trafficking of endosomal cargos. However, the role of aberrant receptor trafficking in PCC is not well understood. Here, we show that loss of TMEM127 causes wildtype RET protein accumulation on the cell surface, where increased receptor density facilitates constitutive ligand-independent activity and downstream signaling, driving cell proliferation. Loss of TMEM127 altered normal cell membrane organization and recruitment and stabilization of membrane protein complexes, impaired assembly, and maturation of clathrin-coated pits, and reduced internalization and degradation of cell surface RET. In addition to RTKs, TMEM127 depletion also promoted surface accumulation of several other transmembrane proteins, suggesting it may cause global defects in surface protein activity and function. Together, our data identify TMEM127 as an important determinant of membrane organization including membrane protein diffusability and protein complex assembly and provide a novel paradigm for oncogenesis in PCC where altered membrane dynamics promotes cell surface accumulation and constitutive activity of growth factor receptors to drive aberrant signaling and promote transformation.

Exploring the Impact of Hepatic Impairment on Pralsetinib Pharmacokinetics.

Pralsetinib is a kinase inhibitor indicated for the treatment of metastatic rearranged during transfection (RET) fusion-positive non-small cell lung cancer. Pralsetinib is primarily eliminated by the liver and hence hepatic impairment (HI) is likely alter its pharmacokinetics (PK). Mild HI has been shown to have minimal impact on the PK of pralsetinib. This hepatic impairment study aimed to determine the pralsetinib PK, safety and tolerability in subjects with moderate and severe HI, as defined by the Child-Pugh and National Cancer Institute Organ Dysfunction Working Group (NCI-ODWG) classification systems, in comparison to subjects with normal hepatic function. Based on the Child-Pugh classification, subjects with moderate and severe HI had similar systemic exposure (area under the plasma concentration time curve from time 0 to infinity [AUC0-∞]) to pralsetinib, with AUC0-∞ geometric mean ratios (GMR) of 1.12 and 0.858, respectively, compared to subjects with normal hepatic function. Results based on the NCI-ODWG classification criteria were comparable; the AUC0-∞ GMR were 1.22 and 0.858, respectively, for subjects with moderate and severe HI per NCI-ODWG versus those with normal hepatic function. These results suggested that moderate and severe hepatic impairment did not have a meaningful impact on the exposure to pralsetinib, thus not warranting a dose adjustment in this population.

Aberrant SOX10 and RET expressions in patients with Hirschsprung disease

BackgroundHSCR is a complex genetic disorder characterized by the absence of ganglion cells in the intestine, leading to a functional obstruction. It is due to a disruption of complex signaling pathways within the gene regulatory network (GRN) during the development of the enteric nervous system (ENS), including SRY-Box Transcription Factor 10 (SOX10) and REarranged during Transfection (RET). This study evaluated the expressions of SOX10 and RET in HSCR patients in Indonesia.MethodsTotal RNA of 19 HSCR ganglionic and aganglionic colons and 16 control colons were analyzed using quantitative real-time polymerase chain reaction for SOX10 and RET with GAPDH as the reference gene. Livak’s method (2−ΔΔCT) was used to determine the expression levels of SOX10 and RET.ResultsMost patients were males (68.4%), in the short aganglionosis segment (78.9%), and had undergone transanal endorectal pull-through (36.6%). There were significant upregulated SOX10 expressions in both ganglionic (2.84-fold) and aganglionic (3.72-fold) colon of HSCR patients compared to controls’ colon (ΔCT 5.21 ± 2.04 vs. 6.71 ± 1.90; p = 0.032; and ΔCT 4.82 ± 1.59 vs. 6.71 ± 1.90; p = 0.003; respectively). Interestingly, the RET expressions were significantly downregulated in both ganglionic (11.71-fold) and aganglionic (29.96-fold) colon of HSCR patients compared to controls’ colon (ΔCT 12.54 ± 2.21 vs. 8.99 ± 3.13; p = 0.0004; and ΔCT 13.90 ± 2.64 vs. 8.99 ± 3.13; p = 0.0001; respectively).ConclusionsOur study shows aberrant SOX10 and RET expressions in HSCR patients, implying the critical role of SOX10 and RET in the pathogenesis of HSCR, particularly in the Indonesian population. Our study further confirms the involvement of SOX10-RET within the GNR during the ENS development.

Unraveling the Promise of RET Inhibitors in Precision Cancer Therapy by Targeting RET Mutations.

Over the past decades, the role of rearranged during transfection (RET) alterations in tumorigenesis has been firmly established. RET kinase inhibition is an essential therapeutic target in patients with RET-altered cancers. In clinical practice, initial efficacy can be achieved in patients through the utilization of multikinase inhibitors (MKIs) with RET inhibitory activity. However, the effectiveness of these MKIs is impeded by the adverse events associated with off-target effects. Recently, many RET-selective inhibitors, characterized by heightened specificity and potency, have been developed, representing a substantial breakthrough in the field of RET precision oncology. This Perspective focuses on the contemporary understanding of RET mutations, recent advancements in next-generation RET inhibitors, and the challenges associated with resistance to RET inhibitors. It provides valuable insights for the development of next-generation MKIs and selective RET inhibitors.

Primary Resistance to RET Inhibition in a RET Fusion-Positive Pancreatic Neuroendocrine Carcinoma.

We present a 54-year-old White male with a diagnosis of stage IV pancreatic neuroendocrine carcinoma. Next-generation sequencing of the tumor/blood identified a complex tumor genome, which included a rearranged during transfection (RET) gene fusion. The patient initially received cytotoxic chemotherapy with a significant radiographic response. After 4 cycles of chemotherapy, the patient was transitioned to a clinical trial using selpercatinib, a RET inhibitor, as maintenance therapy. Unfortunately, our patient developed progression of disease at the first treatment monitoring scan. Our patient suffered primary resistance to RET-targeted therapy. Proposed mechanisms of resistance include intrinsic resistance of the nuclear receptor co-activator 4-RET fusion to RET inhibition, the RET fusion representing a passenger alteration to another tumorigenic driver pathway and/or decreased efficacy of RET inhibition after platinum-based chemotherapy. Our patient's clinical course highlights the fact that "actionable" genomic alterations do not always equate to patient benefit.

Successful treatment with selpercatinib after pralsetinib-related pneumonitis and intracranial failure in a patient with RET-rearranged nonsmall cell lung cancer.

Pralsetinib and selpercatinib are two highly potent and selective rearranged during transfection (RET) inhibitors that substantially improved the clinical outcome of patients with RET-rearranged non-small cell lung cancer. Treatment with one RET inhibitor after failure of the other is generally not recommended because of cross-resistance mechanisms. We report the case of a patient affected by metastatic RET-rearranged non-small cell lung cancer who experienced long-lasting disease control with pralsetinib. After 13 months from treatment start, the patient developed recurrent drug-related pneumonitis, requiring temporary interruptions and dose reductions and eventually failing to control the disease. Selpercatinib was then started as an off-label treatment, allowing both clinical and radiological intracranial disease control. Selpercatinib was well-tolerated at full dosage, and no pulmonary event occurred. In our case report, after pralsetinib dose reduction due to pulmonary toxicity, the therapeutic switch to selpercatinib allowed the patient to receive a full-dose treatment, eventually restoring disease control. Our case report and a few literature data suggest that switching from pralsetinib to selpercatinib may represent a therapeutic opportunity, especially for patients with brain metastases.

Targeting RET alterations in non-small cell lung cancer

Rearranged during transfection (RET) alterations, which lead to aberrant activation of the RET proto-oncogene, have been identified in various cancers. In non-small cell lung cancer (NSCLC), RET mutations often manifest as RET fusion genes and are observed in 1–2 % of patients with NSCLC. In recent years, selective RET inhibitors such as selpercatinib and pralsetinib, approved by the Food and Drug Administration (FDA) in 2020, have been part of the revolutionary changes in the treatment landscape for non-small cell lung cancer. While first-generation RET inhibitors have become part of the standard of care for RET-fusion positive NSCLC, a new challenge has emerged: acquired resistance to RET inhibitors. RET resistance is a complex phenomenon that can manifest as either on-target or off-target resistance. Numerous studies have been conducted to identify the mechanisms behind this resistance. This review provides an overview of the biology of RET in NSCLC, methods of RET testing, and a comprehensive analysis of the clinical outcomes associated with multikinase and selective RET inhibitors for NSCLC. Additionally, we will explore future perspectives for RET fusion-positive NSCLC, including ongoing trials and the challenges involved in overcoming resistance to RET inhibitors.

Current Status of Targeted Therapy for Biliary Tract Cancer in the Era of Precision Medicine.

First-line chemotherapy has been established for advanced biliary tract cancer (BTC). However, few treatment options are available as second-line treatment. Advances in comprehensive genomic analysis revealed that nearly half of patients with BTC harbor targetable genetic alterations such as fibroblast growth factor receptor (FGFR), isocitrate dehydrogenase (IDH), BRAF, human epidermal growth factor receptor 2 (HER2), microsatellite instability (MSI)-high, neurotrophic tropomyosin receptor kinase (NTRK), rearranged during transfection (RET), and poly (adenosine diphosphate-ribose) polymerase (PARP). This review summarizes currently available options in precision medicine and clinical trials for patients with advanced BTC.

Selpercatinib combination with the mitochondria-targeted antioxidant MitoQ effectively suppresses RET–mutant thyroid cancer

Genetic alternation of REarranged during Transfection (RET) that leads to constitutive RET activation is a crucial etiological factor for thyroid cancer. RET is known to regulate mitochondrial processes, although the underlying molecular mechanisms remain unclear. We previously showed that the multi-kinase inhibitors vandetanib and cabozantinib increase the mitochondrial membrane potential (Δψm) in RET-mutated thyroid tumor cells and that this effect can be exploited to increase mitochondrial enrichment of Δψm-sensitive agents in the tumor cells. In this study, we hypothesized that the RET-selective inhibitor, selpercatinib, can increase Δψm and, subsequently, tumor cell uptake of the mitochondria-targeted ubiquinone (MitoQ) to the level to break the mitochondrial homeostasis and induce lethal responses in RET-mutated thyroid tumor cells. We show that selpercatinib significantly increased Δψm, and its combination with MitoQ synergistically suppressed RET-mutated human thyroid tumor cells, which we validated using RET-targeted genetic approaches. Selpercatinib and MitoQ, in combination, also suppressed CCDC6-RET fusion cell line xenografts in mice and prolonged animal survival more effectively than single treatments of each agent. Moreover, we treated two patients with CCDC6-RET or RETM918T thyroid cancer, who could not take selpercatinib at regular doses due to adverse effects, with a dose-reduced selpercatinib and MitoQ combination. In response to this combination therapy, both patients showed tumor reduction. The quality of life of one patient significantly improved over a year until the tumor relapsed. This combination of selpercatinib with MitoQ may have therapeutic potential for patients with RET-mutated tumors and intolerant to regular selpercatinib doses.

Generating Potential RET-Specific Inhibitors Using a Novel LSTM Encoder-Decoder Model.

The receptor tyrosine kinase RET (rearranged during transfection) plays a vital role in various cell signaling pathways and is a critical factor in the development of the nervous system. Abnormal activation of the RET kinase can lead to several cancers, including thyroid cancer and non-small-cell lung cancer. However, most RET kinase inhibitors are multi-kinase inhibitors. Therefore, the development of an effective RET-specific inhibitor continues to present a significant challenge. To address this issue, we built a molecular generation model based on fragment-based drug design (FBDD) and a long short-term memory (LSTM) encoder-decoder structure to generate receptor-specific molecules with novel scaffolds. Remarkably, our model was trained with a molecular assembly accuracy of 98.4%. Leveraging the pre-trained model, we rapidly generated a RET-specific-candidate active-molecule library by transfer learning. Virtual screening based on our molecular generation model was performed, combined with molecular dynamics simulation and binding energy calculation, to discover specific RET inhibitors, and five novel molecules were selected. Further analyses indicated that two of these molecules have good binding affinities and synthesizability, exhibiting high selectivity. Overall, this investigation demonstrates the capacity of our model to generate novel receptor-specific molecules and provides a rapid method to discover potential drugs.

Exposure‐Response Relationships for Pralsetinib in Patients with RET‐Altered Thyroid Cancer or RET Fusion‐Positive Nonsmall Cell Lung Cancer

AbstractPralsetinib is a highly potent oral kinase inhibitor of oncogenic RET (rearranged during transfection) fusions and mutations. Pralsetinib received approval from the United States Food and Drug Administration for the treatment of patients with metastatic RET fusion‐positive non‐small cell lung cancer (NSCLC), and received accelerated approval for the treatment of patients with RET fusion‐positive thyroid cancer. Exposure–response (ER) analyses of efficacy were performed separately in patients with thyroid cancer and in patients with NSCLC, but data for all patients were pooled for the safety analysis. ER models were developed with time‐varying exposure; the effect of covariates was also examined. For patients with NSCLC, a higher starting dose was associated with improved progression‐free survival (PFS), but this improvement did not correlate with a higher exposure overall. Significant covariates included sex and baseline Eastern Cooperative Oncology Group (ECOG) score. For patients with thyroid cancer, a higher exposure was associated with improved PFS. Significant covariates included prior systemic cancer therapy and ECOG score. For safety, higher exposure was associated with a greater risk of grade ≥3 anemia, pneumonia, and lymphopenia. Patients with an ECOG score of ≥1 had an increased risk of grade ≥3 pneumonia. Non‐White patients had a lower risk of grade ≥3 lymphopenia. ER analysis revealed that higher pralsetinib exposure was associated with improved PFS in thyroid cancer, but not in NSCLC. However, a higher starting dose (ie, 400 vs ≤300 mg daily) was correlated with better PFS for all indications. Higher exposure was also associated with an increased risk of grade ≥3 adverse events (AEs); however, the overall incidence of these events was acceptably low (≤20%). This analysis supports the use of a 400 mg starting dose of pralsetinib, allowing for dose reduction in the event of AEs.

[Precision medicine in endocrinology exemplified by medullary thyroid cancer].

Medullary thyroid cancer (MTC) is a prime example for precision medicine in endocrinology and underlines the immediate benefits of basic, translational and healthcare research for patients with a rare disease in clinical . A mutation in the rearranged during transfection (RET) proto-oncogene that codes for a transmembrane receptor protein tyrosine kinase, leads to constitutive activation of the kinase, which is the decisive pathomechanism for the disease. The MTC occurs in a sporadic (somatic RET mutation) or hereditary form (RET germline mutation, multiple endocrine neoplasia types 2 and 3). For germline mutation carriers the timing of preventive thyroidectomy depends on the RET genotype. For advanced metastasized RET-mutant MTC, selective RET kinase inhibitors are available, which are currently considered to be game changers in the treatment. Based on the specific tumor marker calcitonin, MTC can be identified at an early stage during the differential diagnosis of thyroid nodules. The preoperative calcitonin level even enables statements on the degree of dissemination of the disease and on the probability of a cure through surgery. A new development is the consideration of desmoplasia as a histopathological biomarker for the metastatic potential of a MTC, which could possibly modify the operative approach as well as the future MTC nomenclature. Furthermore, the postoperative calcitonin level and the calcitonin doubling time are highly valid prognostic markers for tumor burden and biological aggressiveness of MTC and therefore decisive for patient follow-up. Biochemical, molecular and histological markers enable a risk-adapted surgical treatment and together with new targeted systemic treatments have contributed to a paradigm shift in the diagnostics, prognosis and treatment of MTC in recent years. Endocrine precision medicine for MTC therefore enabled a change from the previous purely symptom-oriented to a modern preventive and individualized treatment.

Enhancer-activated RET confers protection against oxidative stress to KMT2A-rearranged acute myeloid leukemia.

Ectopic activation of rearranged during transfection (RET) has been reported to facilitate lineage differentiation and cell proliferation in different cytogenetic subtypes of acute myeloid leukemia (AML). Herein, we demonstrate that RET is significantly (p < 0.01) upregulated in AML subtypes containing rearrangements of the lysine methyltransferase 2A gene (KMT2A), commonly referred to as KMT2A-rearranged (KMT2A-r) AML. Integrating multi-epigenomics data, we show that the KMT2A-MLLT3 fusion induces the development of CCCTC-binding (CTCF)-guided de novo extrusion enhancer loop to upregulate RET expression in KMT2A-r AML. Based on the finding that RET expression is tightly correlated with the selective chromatin remodeler and mediator (MED) proteins, we used a small-molecule inhibitor having dual inhibition against RET and MED12-associated cyclin-dependent kinase 8 (CDK8) in KMT2A-r AML cells. Dual inhibition of RET and CDK8 restricted cell proliferation by producing multimodal oxidative stress responses in treated cells. Our data suggest that epigenetically enhanced RET protects KMT2A-r AML cells from oxidative stresses, which could be exploited as a potential therapeutic strategy.

Impact of the quality of resected thyroid cancer tissue sample on next-generation sequencing testing.

Activating rearranged during transfection (RET) proto-oncogene alterations can be identified using next-generation sequencing (NGS) of tumor DNA/RNA. We assessed factors associated with NGS (Oncomine Dx Target Test [ODxTT]) success for resected thyroid cancer (TC) specimens, including sample age, processing conditions, and DNA/RNA quality. TC samples were from three Japanese hospitals, with sample age <1-<10 years, fixative 10%/15% neutralized buffered formalin (NBF), and fixation time ≤48 h/>48 h-≤72 h. NGS success rate was defined as the percentage of samples returning validated NGS results (RET fusion-positive/negative [RNA] or RET mutation-positive/negative [DNA], detected using ODxTT). DNA/RNA quality was assessed with indexes based on electrophoresis (DNA/RNA integrity number, DV200 ) and quantitative polymerase chain reaction (DNA/RNA integrity score [ddCq/ΔCq]). NGS success rate (N = 202) was 90%/93% (DNA/RNA) overall, 98%-100% (DNA and RNA) for samples <3 years old, and 91% (DNA and RNA) for samples ≥3-<5 years old fixed in 10% NBF for ≤48 h. Multivariate logistic regression analysis identified ddCq and ΔCq as significant predictors of DNA and RNA NGS success rates, respectively. Quality assessment of nucleic acid extracted from archival tissue samples is important for achieving high NGS success rates in clinical practice, especially for samples ≥3 years old.

Real-world clinical profile, RET mutation testing, treatments, and PROs for MTC in Europe.

This study aimed to describe real-world patient and physician characteristics, rearranged during transfection (RET) mutation testing and results, treatment patterns, and patient-reported outcomes (PROs) in advanced or metastatic medullary thyroid cancer (aMTC) across five populous European countries. Cross-sectional physician and patient surveys were used to collect quantitative and qualitative data in France, Germany, Italy, Spain, and the UK from July to December 2020, prior to the introduction of selective RET inhibitors in Europe. Physicians completed patient record forms and a survey about their specialty and practice site. Patients were asked to provide PRO data using four validated instruments, including the EuroQol 5 Dimension (EQ-5D) questionnaire. The physician-reported sample included 275 patients with aMTC, including 79 patients with RET mutation-positive disease; median age was 60 and 56 years, respectively. Overall, 75% were tested for RET mutation (35% germline only, 21% somatic only, 44% both). Common physician-cited barriers to RET mutation testing included high cost, difficulty accessing latest tests, and time delay for results. First-line systemic therapy (most commonly vandetanib or cabozantinib) was prescribed for 69% of patients overall and 82% of the RET mutation-positive subgroup. Second-line therapy was prescribed for 12% of patients who received first-line therapy; most patients remained on first-line therapy at data capture. PROs revealed substantial disease/treatment burden. Patients with aMTC report substantial disease/treatment burden. Outcomes could be improved by identifying patients eligible for treatment with selective RET inhibitors through more optimal RET mutation testing.

Selpercatinib: A narrative drug review

Rearranged during transfection (RET) alteration promotes oncogenesis in a few cancers. RET mutation positivity is seen in approximately 70% of medullary thyroid cancers, around 30% of differentiated papillary thyroid cancers, and 1-2% of non-small-cell lung cancers (NSCLC). To write this narrative drug review, we searched various websites like the United States Food and Drug Administration, PubMed, Google Scholar, UpToDate, and recently published papers in various international conferences using the search terms “RET,” “RET alteration,” “Retevmo,” “RET inhibitors,” and “selpercatinib.” We shortlisted 31 articles published between January 1980 and January 2024. We discuss the history, mechanism of action, resistance, pharmacodynamics, pharmacokinetics, dosing, toxicity, pivotal trials, and indications of selpercatinib. Selective RET inhibitors like selpercatinib are indicated in the treatment of RET-altered NSCLC and thyroid cancer.

GDF-15 Inhibits ADP-Induced Human Platelet Aggregation through the GFRAL/RET Signaling Complex.

Growth differentiation factor-15 (GDF-15) is proposed to be strongly associated with several cardiovascular diseases, such as heart failure and atherosclerosis. Moreover, some recent studies have reported an association between GDF-15 and platelet activation. In this study, we isolated peripheral blood platelets from healthy volunteers and evaluated the effect of GDF-15 on adenosine diphosphate (ADP)-induced platelet activation using the platelet aggregation assay. Subsequently, we detected the expression of GDF-15-related receptors on platelets, including the epidermal growth factor receptor (EGFR), human epidermal growth factor receptor 2 (HER2), human epidermal growth factor receptor 3 (HER3), transforming growth factor-beta receptor I (TGF-βRI), transforming growth factor-beta receptor II (TGF-βRII), glial-cell-line-derived neurotrophic factor family receptor α-like (GFRAL), and those rearranged during transfection (RET). Then, we screened for GDF-15 receptors using the GDF-15-related receptor microarray comprising these recombinant proteins. We also performed the immunoprecipitation assay to investigate the interaction between GDF-15 and the receptors on platelets. For the further exploration of signaling pathways, we investigated the effects of GDF-15 on the extracellular signal-regulated kinase (ERK), protein kinase B (AKT), and Janus kinase 2 (JAK2) pathways. We also investigated the effects of GDF-15 on the ERK and AKT pathways and platelet aggregation in the presence or absence of RET agonists or inhibition. Our study revealed that GDF-15 can dose-independently inhibit ADP-induced human platelet aggregation and that the binding partner of GDF-15 on platelets is GFRAL. We also found that GDF-15 inhibits ADP-induced AKT and ERK activation in platelets. Meanwhile, our results revealed that the inhibitory effects of GDF-15 can be mediated by the GFRAL/RET complex. These findings reveal the novel inhibitory mechanism of ADP-induced platelet activation by GDF-15.

Selective RET Inhibitors (SRIs) in Cancer: A Journey from Multi-Kinase Inhibitors to the Next Generation of SRIs.

RET is a receptor tyrosine kinase that plays an important role in the development of neurons and kidneys. The gene encoding the rearranged-during-transfection (RET) receptor tyrosine kinase was first discovered in the 1980s. Activating RET mutations and rearrangements have since been identified as actionable drivers of oncogenesis in numerous cancer types and are most prevalent in thyroid and non-small-cell lung cancer. Following the modest success of repurposed RET-active multikinase inhibitors, the first selective RET inhibitors (SRIs), selpercatinib and pralsetinib, received regulatory approval in 2020. Now, thousands of patients with RET-altered cancers have benefited from first-generation SRIs, with impressive deep and durable responses. However, following prolonged treatment with these SRIs, a number of acquired on-target resistance mutations have been identified together with other non-RET-dependent resistance mechanisms. Today, the focus is on how we can further evolve and improve the treatment of RET-altered tumors with next-generation SRIs, and a number of candidate drugs are in development. The ideal next-generation SRIs will be active against on-target acquired resistance alterations, including those that emerge in the CNS, and will have improved safety and tolerability relative to first-generation SRIs. In this review, we will provide an update on these candidates and their potential to meet the unmet clinical need for patients who progress on first-generation SRIs.

Modeling RET-Rearranged Non-Small Cell Lung Cancer (NSCLC): Generation of Lung Progenitor Cells (LPCs) from Patient-Derived Induced Pluripotent Stem Cells (iPSCs).

REarranged during Transfection (RET) oncogenic rearrangements can occur in 1-2% of lung adenocarcinomas. While RET-driven NSCLC models have been developed using various approaches, no model based on patient-derived induced pluripotent stem cells (iPSCs) has yet been described. Patient-derived iPSCs hold great promise for disease modeling and drug screening. However, generating iPSCs with specific oncogenic drivers, like RET rearrangements, presents challenges due to reprogramming efficiency and genotypic variability within tumors. To address this issue, we aimed to generate lung progenitor cells (LPCs) from patient-derived iPSCs carrying the mutation RETC634Y, commonly associated with medullary thyroid carcinoma. Additionally, we established a RETC634Y knock-in iPSC model to validate the effect of this oncogenic mutation during LPC differentiation. We successfully generated LPCs from RETC634Y iPSCs using a 16-day protocol and detected an overexpression of cancer-associated markers as compared to control iPSCs. Transcriptomic analysis revealed a distinct signature of NSCLC tumor repression, suggesting a lung multilineage lung dedifferentiation, along with an upregulated signature associated with RETC634Y mutation, potentially linked to poor NSCLC prognosis. These findings were validated using the RETC634Y knock-in iPSC model, highlighting key cancerous targets such as PROM2 and C1QTNF6, known to be associated with poor prognostic outcomes. Furthermore, the LPCs derived from RETC634Y iPSCs exhibited a positive response to the RET inhibitor pralsetinib, evidenced by the downregulation of the cancer markers. This study provides a novel patient-derived off-the-shelf iPSC model of RET-driven NSCLC, paving the way for exploring the molecular mechanisms involved in RET-driven NSCLC to study disease progression and to uncover potential therapeutic targets.

Clinical characteristics and progression of pre-/minimally invasive lung adenocarcinoma harboring ALK or RET rearrangements: a retrospective cohort study.

Patients harboring anaplastic lymphoma kinase (ALK) or rearranged during transfection (RET) rearrangements are usually diagnosed at a relatively late stage with nodal and distant metastasis, and rapid progression course of ALK/RET fusion-positive lung cancer were well-known. However, clinical characteristics and course of pre-/minimally invasive lung adenocarcinoma harboring ALK or RET fusions are poorly described. Identifying patients with gene fusions at early stage may offer surgical options that could cure those patients. We retrospectively included patients with surgically resected pre-/minimally invasive lung adenocarcinomas harboring epidermal growth factor receptor (EGFR) mutations or ALK/RET rearrangements, and further compared the patient clinical characteristics, nodule natural course, and survival outcomes. Radiological characteristics including ground-glass component, cystic airspace, pleural attachment, etc. were specially assessed for this study. EGFR (exons 18-22) was detected by Sanger sequencing and quantitative real-time polymerase chain reaction (qRT-PCR) was used to analyze the ALK/RET rearrangements. Lung cancer-specific survival (LCSS), relapse-free survival (RFS), and overall survival (OS) were all evaluated. Of 238 patients with pre-/minimally invasive lung adenocarcinomas, 226 patients had EGFR mutations, 7 patients had ALK fusions, and 5 patients had RET fusions. Average age at surgery was 45.3 years for ALK/RET-positive group and 52.6 years for EGFR-positive group (P=0.049). Radiologically, among the 12 patients with ALK/RET fusions, the majority of lesions (10/12) manifested as mixed ground-glass opacities (mGGOs), which was significantly more prevalent when compared with patients with EGFR mutations (83.4% vs. 24.3%, P<0.001). Moreover, a substantial proportion of cystic airspace was found in ALK/RET-positive group but not in EGFR-positive group (66.7% vs. 14.2%, P<0.001). Among four patients with ALK/RET fusions undergoing surveillance over 1 year before surgery, two of them developed rapid radiologic progression. The 5-year LCSS and RFS were 100%, 100% for ALK/RET-positive group, and 100%, 100% for EGFR-positive group, respectively. ALK/RET-positive pre-/minimally invasive lung adenocarcinomas were mostly characterized as mGGOs with cystic airspace developing rapid nodule progression, and no recurrence occurred during long-term follow-up after resection. This provides insights into proper curative surgery timing in the management of patients with gene fusions. However, these findings must be treated with caution and validated in future multi-center studies with larger sample size.