Search Results (38)

Approximately 30% of tumors contain an activating mutation in the oncogene KRAS, leading to increased cell proliferation that often promotes non-small cell lung cancers, colorectal adenocarcinomas, pancreatic ductal adenocarcinomas (PDAC), and other cancers. Among the most common point mutations in KRAS is G12D, an example of an oncogenic sequence present in tumor cells but not normal cells. We developed peptide nucleic acid (PNA) oligomers that selectively bind KRAS G12D sequences and fused them with novel cell-penetrating peptide flanking regions (CPP-PNA-G12D) then evaluated them. Electrophoretic mobility shift assays demonstrated in vitro binding to and selectivity for KRAS G12D over wild-type KRAS and KRAS G12C. Cells and nuclei were able to uptake CPP-PNA-G12D at high efficiency as shown by fluorescent microscopy and flow cytometry. Cell viability assays showed a striking dose-response effect in on-target cells expressing KRAS G12D, while relatively sparing off-target cells expressing KRAS G12C. CPP-PNA-G12D constructs were effective against a panel of PDAC cell lines and in female Balb/c mice bearing patient-derived xenografts. These results show promise for an enhanced PNA-delivery peptide conjugate strategy as a potential therapeutic strategy to selectively target KRAS mutant cancer cells, with the potential to expand this technology to additional cancer-derived mutant oncogenes. Full article

KRAS G12C, long considered an undruggable oncogenic driver, has become one of the most consequential therapeutic targets in non-small cell lung cancer (NSCLC). The discovery of a cryptic binding pocket accessible in the GDP-bound state enabled covalent inhibitors—sotorasib and adagrasib—that have received regulatory approval for previously treated KRAS G12C-mutant NSCLC, with sotorasib demonstrating PFS and OS superiority over docetaxel in CodeBreaK 200 and adagrasib showing meaningful intracranial activity and a progression-free survival benefit over docetaxel in KRYSTAL-12. Yet response durability is limited by on-target switch-II pocket mutations, upstream RTK and SHP2-mediated bypass signaling, downstream MAPK and PI3K-AKT reactivation, phenotypic plasticity, and adverse modulation by co-occurring STK11, KEAP1, and TP53 alterations. Next-generation covalent inhibitors (divarasib, glecirasib, olomorasib), tri-complex RAS(ON) inhibitors (RMC-6291), pan-KRAS agents, and rationally designed combinations with EGFR, SHP2, SOS1, and PD-1 inhibitors are repositioning KRAS-directed therapy toward earlier lines of treatment. This review integrates the structural, signaling, and clinical biology of KRAS G12C with contemporary trial and real-world evidence to examine the emerging case for first-line KRAS G12C inhibition in genomically defined subsets of NSCLC. First-line use nonetheless remains investigational; platinum-based chemoimmunotherapy remains the standard of care outside of clinical trials, and a frontline indication will require confirmation from randomized phase III trials. Full article

Background/Objectives: Chronic lymphocytic leukemia (CLL) is characterized by clinical and biological heterogeneity shaped by intrinsic signaling programs and microenvironmental interactions. Established biomarkers, including IGHV mutation status and TP53 alterations, provide important clinical and molecular information, but do not fully capture the diversity of pathways that sustain leukemic cell fitness. Aberrant calcium signaling contributes to leukemic survival; however, the clinical relevance of Ca²⁺/calmodulin-dependent protein kinase kinase 2 (CaMKK2), a calcium-responsive kinase, has not been defined. This study evaluated CaMKK2 as a candidate marker associated with high-risk disease biology and a pathway of interest for further study. Methods: CaMKK2 expression was quantified in purified CD19⁺ CLL cells from a clinically annotated cohort balanced by immunoglobulin heavy chain variable region (IGHV) mutation status. Associations with time to treatment and overall survival were analyzed. Functional relevance was assessed by pharmacologic inhibition of CaMKK2 in primary CLL cells using metabolic (MTS) and apoptosis (Annexin V/PI) assays. Correlations between CaMKK2 expression and inhibitor sensitivity were determined. The impact of CaMKK2 inhibition on nurse-like cell (NLC) differentiation and macrophage-mediated leukemic support was evaluated in ex vivo culture systems. Results: Elevated CaMKK2 expression was enriched in IGHV-unmutated CLL and associated with shorter time to treatment and inferior overall survival. Pharmacological inhibition of CaMKK2 was associated with reduced primary CLL viability in a dose-dependent manner and increased Annexin V/PI-defined total cell death with sensitivity correlating with CaMKK2 expression levels. Inhibition also attenuated CD163⁺ macrophage polarization and impaired NLC-mediated support of leukemic cells. Conclusions: CaMKK2 expression is associated with IGHV-unmutated, high-risk CLL biology. Pharmacologic inhibition of CaMKK2 was associated with reduced leukemic cell viability and altered macrophage phenotypes in ex vivo systems. These findings are exploratory, derived from a limited cohort, and support further investigation into the role of CaMKK2 in CLL biology, but do not establish independent prognostic value or direct on-target causality. Full article

MET pathway alterations have evolved from a niche translational interest into one of the most clinically actionable axes in non-small cell lung cancer (NSCLC). Three biologically distinct lesions—MET exon 14 (METex14) skipping mutations, focal high-level MET amplification, and c-Met protein overexpression—are now individually targetable, each with its own diagnostic prerequisites and therapeutic class. Selective type Ib MET tyrosine kinase inhibitors (capmatinib, tepotinib) anchor first-line therapy for METex14, while next-generation agents and type II inhibitors are being developed to address on-target D1228 and Y1230 resistance mutations. In parallel, MET amplification has emerged as a leading mechanism of acquired resistance to osimertinib in EGFR-mutated NSCLC, with the SAVANNAH, SACHI, and INSIGHT 2 trials providing biomarker-guided combination strategies. The 2025 accelerated approval of telisotuzumab vedotin for c-Met-overexpressing tumors expanded the therapeutic armamentarium beyond kinase inhibition. Despite these advances, lineage plasticity, polyclonal bypass signaling, and inconsistent diagnostic thresholds for MET amplification continue to limit durable benefit. This review integrates the molecular biology, current clinical evidence, resistance mechanisms, and a proposed 2025 treatment algorithm for MET-altered NSCLC, with emphasis on the translational interface between mutation class, drug class, and emerging combinatorial approaches. As a narrative review, it synthesizes peer-reviewed literature and pivotal trial and regulatory data through early 2026, identified by structured searches of PubMed and major oncology congress proceedings, and prioritizes sources that link mutation class to drug class and resistance mechanism. Full article

Hematological malignancies, including multiple myeloma (MM), leukemia, and lymphoma, represent a major global health burden, accounting for approximately 6.6% of all cancer cases and contributing to significant mortality. The evolutionary conserved WNT/β-catenin signaling pathway is a critical regulator of normal hematopoietic stem cell homeostasis, and its dysregulation is a hallmark of various hematological malignancies. Aberrant activation through mutations, overexpression of ligands, or disruption of the destruction complex drives uncontrolled proliferation, impaired differentiation, and therapeutic resistance to therapy in acute and chronic leukemias, lymphomas, and multiple myeloma. Therapeutic interventions targeting this pathway, such as GSK-3 inhibitors, β-catenin antagonists, and small molecules like CWP291 and salinomycin, have demonstrated promising antitumor effects. Furthermore, combining WNT/β-catenin inhibition with targeted or epigenetic therapies, such as venetoclax and chidamide, can produce synergistic antitumor effects and overcome chemoresistance. Despite this potential, clinical translation is hampered by on-target toxicities in healthy tissues, pathway complexity, and a lack of predictive biomarkers. We conclude that the future of WNT-directed therapy lies in developing biomarker-selective agents, advanced drug delivery systems to improve specificity, and exploring novel combinations with immunotherapy to harness the anti-tumor immune response. Full article

Background: Alpelisib plus fulvestrant improves outcomes in PIK3CA-mutated, hormone receptor-positive, HER2-negative metastatic breast cancer. However, on-target hyperglycemia often leads to dose modification or discontinuation. We aimed primarily to determine whether evening alpelisib after a ≥5 h fast with low-carbohydrate guidance reduces severe hyperglycemia versus standard morning dosing, and secondarily, to assess time to first grade 3–4 hyperglycemia, efficacy, and quality of life (QoL). Methods: ITACA was an open-label, randomized, phase IIb trial in three Croatian centers. Patients progressing on endocrine therapy were randomized 1:1 to evening alpelisib 300 mg after a ≥5 h fast with low-carbohydrate guidance or standard morning alpelisib, both with fulvestrant. The primary endpoint was the exposure-adjusted incidence rate (EAIR) of first grade 3–4 hyperglycemia within 90 days or 30 days post-discontinuation. Secondary endpoints were time to first grade 3–4 hyperglycemia, efficacy, and QoL. Results: Forty-two patients were randomized (21 per arm). Median age was 60 vs. 63 years in the evening vs. morning arms. In the safety set, EAIR of first grade 3–4 hyperglycemia was 378 vs. 742 per 100 person-years (11/21 vs. 14/20 patients with ≥1 event, unadjusted IRR 0.51, 95% CI 0.23–1.12). Adjusted Poisson models favored evening dosing. Analyses suggested delayed onset (median 73 vs. 9.5 days), with no detriment in efficacy or QoL. Conclusions: Evening alpelisib preceded by fasting and low-carbohydrate guidance may improve metabolic tolerability without compromising efficacy or QoL. These findings support evaluation in a larger trial incorporating prospective metabolic adherence and pharmacokinetic assessments. Full article

Background: Point mutations in mitochondrial DNA (mtDNA) cause a range of neurometabolic disorders that currently have no curative treatments. The m.8993T>G mutation in the Homo sapiens MT-ATP6 gene leads to neurogenic muscle weakness, ataxia, and retinitis pigmentosa (NARP) when heteroplasmy exceeds approximately 70%. Methods: We engineered a split DddA-derived cytosine base editor (DdCBE), each half fused to programmable TALE DNA-binding domains and a mitochondrial targeting sequence, to correct the m.8993T>G mutation in patient-derived induced pluripotent stem cells (iPSCs). Seven days after plasmid delivery, deep amplicon sequencing showed 35 ± 3% on-target C•G→T•A conversion at position 8993, reducing mutant heteroplasmy from 80 ± 2% to 45 ± 3% with less than 0.5% editing at ten predicted off-target loci. Results: Edited cells exhibited a 25% increase in basal oxygen consumption rate, a 50% improvement in ATP-linked respiration, and a 2.3-fold restoration of ATP synthase activity. Directed neural differentiation yielded 85 ± 2% Nestin-positive progenitors compared to 60 ± 2% in unedited controls. Conclusions: Edits remained stable over 30 days in culture. These results establish mitochondrial base editing as a precise and durable strategy to ameliorate biochemical and cellular defects in NARP patient cells. Full article

CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR-associated protein) nucleases enable precise genome editing, but off-target cleavage remains a critical challenge. Here, we report the development of MAD7_HF, a high-fidelity variant of the MAD7 nuclease engineered through a bacterial screening system leveraging the DNA gyrase-targeting toxic gene ccdB. This system couples survival to efficient on-target cleavage and minimal off-target activity, mimicking the transient action required for high-precision editing. Through iterative selection and sequencing validation, we identified MAD7_HF, harboring three substitutions (R187C, S350T, K1019N) that enhanced discrimination between on- and off-target sites. In Escherichia coli assays, MAD7_HF exhibited a >20-fold reduction in off-target cleavage across multiple mismatch contexts while maintaining on-target efficiency comparable to wild-type MAD7. Structural modeling revealed that these mutations stabilize the guide RNA-DNA hybrid at on-target sites and weaken interactions with mismatched sequences. This work establishes a high-throughput bacterial screening strategy that allows the identification of Cas12a variants with improved specificity at a given target site, providing a useful framework for future efforts to develop precision genome-editing tools. Full article

Non-small-cell lung cancer (NSCLC) represents the most common type of lung cancer. The majority of patients with lung cancer characterized by activating mutations in the epidermal growth factor receptor (EGFR), benefit from therapies entailing tyrosine kinase inhibitors (TKIs). In this regard, osimertinib, a third-generation EGFR TKI, has greatly improved the outcome for patients with EGFR-mutated lung cancer. The AURA and FLAURA trials displayed the superiority of the third-generation TKI in both first- and second-line settings, making it the drug of choice for treating patients with EGFR-mutated lung cancer. Unfortunately, the onset of resistance is almost inevitable. On-target mechanisms of resistance include new mutations (e.g., C797S) in the kinase domain of EGFR, while among the off-target mechanisms, amplification of MET or HER2, mutations in downstream signaling molecules, oncogenic fusions, and phenotypic changes (e.g., EMT) have been described. This review focuses on the strategies that are currently being investigated, in preclinical and clinical settings, to overcome resistance to osimertinib, including the use of fourth-generation TKIs, PROTACs, bispecific antibodies, and ADCs, as monotherapy and as part of combination therapies. Full article

A subset of ALK+ non-small cell lung cancer (NSCLC) patients relapse on ALK inhibitor (ALKi) treatment due to on-target resistance mutations affecting the tyrosine kinase domain. Objective: In this study, we investigated the presence of minor resistant clones in pre-treatment tissue samples and assessed their predictive value for subsequent resistance mechanisms. Methods: Using the highly sensitive digital droplet (dd)PCR technique, we analyzed 40 tissue samples obtained from 17 patients who had developed on-target resistance mutations after receiving ALKi between 2013 and 2022. We focused on 10 on-target ALKi resistant mutations identified in our patient cohort. Results: Fifteen ALKi resistance mutations were detected in 13 samples from 11/17 patients. Among these, four mutations were observed as resistance mutations in follow-up biopsies taken after first or subsequent lines of ALKi. Comparison of the test results from two subsequent biopsies, before and directly after therapy, revealed presence of the resistance mutation identified upon relapse in the pre-treatment sample of three cases that were all taken from the same tumor location. In six cases taken from different tumor locations, the resistant mutations were not found in the pre-treatment sample. Conclusions: By using the highly sensitive ddPCR approach, we detected minor clones with on-target resistant mutations in both treatment-naive and relapse biopsies from ALK-positive NSCLC patients. The predictive value of these mutations as the potential resistance-causing mechanism was limited to relapses occurring at the same tumor location as the pre-treatment sample. Full article

Non-small-cell lung cancer (NSCLC) is a leading cause of cancer-related deaths worldwide, with epidermal growth factor receptor (EGFR) mutations present in a substantial proportion of patients. Third-generation EGFR tyrosine kinase inhibitors (EGFR TKI), exemplified by osimertinib, have dramatically improved outcomes by effectively targeting the T790M mutation—a primary driver of acquired resistance to earlier-generation EGFR TKI. Despite these successes, resistance to third-generation EGFR TKIs inevitably emerges. Mechanisms include on-target mutations such as C797S, activation of alternative pathways like MET amplification, histologic transformations, and intricate tumor microenvironment (TME) alterations. These resistance pathways are compounded by challenges in tolerability, adverse events, and tumor heterogeneity. In light of these hurdles, this review examines the evolving landscape of combination therapies designed to enhance or prolong the effectiveness of third-generation EGFR TKIs. We explore key strategies that pair osimertinib with radiotherapy, anti-angiogenic agents, immune checkpoint inhibitors, and other molecularly targeted drugs, and we discuss the biological rationale, preclinical evidence, and clinical trial data supporting these approaches. Emphasis is placed on how these combinations may circumvent diverse resistance mechanisms, improve survival, and maintain a favorable safety profile. By integrating the latest findings, this review aims to guide clinicians and researchers toward more individualized and durable treatment options, ultimately enhancing both survival and quality of life for patients with EGFR-mutated NSCLC. Full article

Background: Anaplastic lymphoma kinase (ALK) plays a role in the development of lymphoma, lung cancer and neuroblastoma. While tyrosine kinase inhibitors (TKIs) have improved treatment outcomes, relapse remains a challenge due to on-target mutations and off-target resistance mechanisms. ALK-positive (ALK+) tumors can evade the immune system, partly through tumor-associated macrophages (TAMs) that facilitate immune escape. Cancer cells use “don’t eat me” signals (DEMs), such as CD47, to resist TAMs-mediated phagocytosis. TKIs may upregulate pro-phagocytic stimuli (i.e., calreticulin, CALR), suggesting a potential therapeutic benefit in combining TKIs with an anti-CD47 monoclonal antibody (mAb). However, the impact of this combination on both TKIs-sensitive and resistant ALK+ tumors requires further investigation. Methods: A panel of TKIs-sensitive and resistant ALK+ cancer subtypes was assessed for CALR and CD47 expression over time using flow cytometry. Flow cytometry co-culture and fluorescent microscopy assays were employed to evaluate phagocytosis under various treatment conditions. Results: ALK inhibitors increased CALR expression in both TKIs-sensitive and off-target resistant ALK+ cancer cells. Prolonged TKIs exposure also led to CD47 upregulation. The combination of ALK inhibitors and anti-CD47 mAb significantly enhanced phagocytosis compared to anti-CD47 alone, as confirmed by flow cytometry and fluorescent microscopy. Conclusions: Anti-CD47 mAb can quench DEMs while exposing pro-phagocytic signals, promoting tumor cell phagocytosis. ALK inhibitors induced immunogenic cell damage by upregulating CALR in both sensitive and off-target resistant tumors. Continuous TKIs exposure in off-target resistant settings also resulted in the upregulation of CD47 over time. Combining TKIs with a CD47 blockade may offer therapeutic benefits in ALK+ cancers, especially in overcoming off-target resistance where TKIs alone are less effective. Full article

The mammarenavirus matrix Z protein plays critical roles in virus assembly and cell egress. Meanwhile, heterotrimer complexes of a stable signal peptide (SSP) together with glycoprotein subunits GP1 and GP2, generated via co-and post-translational processing of the surface glycoprotein precursor GPC, form the spikes that decorate the virion surface and mediate virus cell entry via receptor-mediated endocytosis. The Z protein and the SSP undergo N-terminal myristoylation by host cell N-myristoyltransferases (NMT1 and NMT2), and G2A mutations that prevent myristoylation of Z or SSP have been shown to affect the Z-mediated virus budding and GP2-mediated fusion activity that is required to complete the virus cell entry process. In the present work, we present evidence that the validated on-target specific pan-NMT inhibitor DDD85646 exerts a potent antiviral activity against the prototypic mammarenavirus lymphocytic choriomeningitis virus (LCMV) that correlates with reduced Z budding activity and GP2-mediated fusion activity as well as with proteasome-mediated degradation of the Z protein. The potent anti-mammarenaviral activity of DDD85646 was also observed with the hemorrhagic-fever-causing Junin (JUNV) and Lassa (LASV) mammarenaviruses. Our results support the exploration of NMT inhibition as a broad-spectrum antiviral against human pathogenic mammarenaviruses. Full article

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. Full article

Specific signalling thresholds of the WNT/β-catenin pathway affect embryogenesis and tissue homeostasis in the adult, with mutations in this pathway frequently occurring in cancer. Excessive WNT/β-catenin activity inhibits murine anterior development associated with embryonic lethality and accounts for the driver event in 80% of human colorectal cancers. Uncontrolled WNT/β-catenin signalling arises primarily from impairment mutation in the tumour suppressor gene APC that otherwise prevents prolonged stabilisation of β-catenin. Surprisingly, no inhibitor compounds for WNT/β-catenin signalling have reached clinical use in part owing to the lack of specific in vivo assays that discriminate between on-target activities and dose-limiting toxicities. Here, we present a simple in vivo assay with a binary outcome whereby the administration of candidate compounds to pregnant and phenotypically normal Apc^flox/flox mice can rescue in utero death of Apc^min/flox mutant conceptus without subsequent post-mortem assessment of WNT/β-catenin signalling. Indeed, the phenotypic plasticity of born Apc^min/flox conceptus enables future refinement of our assay to potentially enable dosage finding and cross-compound comparisons. Thus, we show for the first time the suitability of endogenous WNT/β-catenin signalling during embryonic development to provide an unambiguous and sensitive mammalian in vivo model to assess the efficacy and bioavailability of potential WNT/β-catenin antagonists. Full article