•RAS is the most frequently mutated oncogene in human cancers, accounting for approximately 30% of mutations in all human cancers.•Despite playing a distinct role in tumorigenesis, various attempts ...to inhibit K-RAS directly in the past were unsuccessful.•Additionally, inhibiting downstream Kras signaling through approaches such as inhibiting RAF, MEK and ERK have been unsuccessful.•Recently, a binding pocket (S-IIP) has been identified in K-RAS G12C that can be targeted by covalent inhibitors.•The K-RAS G12C mutation is present in about 13% of lung adenocarcinoma and 3% of colorectal cancer cases. Several inhibitors of this specific mutation have been developed, with initial evidence of impressive clinical activity.•Other approaches including, SHP2, SOS1 and eIF4 inhibition, are being evaluated to abrogate tumor growth in K-RAS mutant cells.
RAS is the most frequently mutated oncogene in human cancers, with mutations in about 30% of all cancers. RAS exists in three different isoforms (K-RAS, H-RAS and N-RAS) with high sequence homology. K-RAS is the most commonly mutated RAS isoform. The Ras protein is a membrane bound protein with inherent GTPase activity and is activated by numerous extracellular stimuli, cycling between an inactive (GDP-bound) and active (GTP-bound) form. When bound to GTP, it is switched “on” and activates intracellular signaling pathways, critical for cell proliferation and angiogenesis. Mutated RAS is constitutively activated and persistently turned “on” thereby enhancing downstream signaling and leading to tumorigenesis. Various attempts to inhibit Kras in the past were unsuccessful. Recently, several small molecules (AMG510, MRTX849, JNJ-74699157, and LY3499446) have been developed to specifically target K-RAS G12C. Additionally, various other approaches including, SHP2, SOS1 and eIF4 inhibition, have been utilized to abrogate tumor growth in K-RAS mutant cells, resulting in a renewed interest in this pathway. In this review article, we provide an overview on the role of K-RAS in tumorigenesis, past approaches to inhibiting Kras, and current and future prospects for targeting Kras.
Abstract
Objective
Numerous scattered case studies continue to demonstrate a strong correlation between acquired KRAS mutations and epidermal growth factor receptor-tyrosine kinase inhibitor ...resistance in non-small cell lung cancer. However, the comprehensive understanding of the KRAS pathway following the failure of epidermal growth factor receptor-tyrosine kinase inhibitor therapy remains limited.
Methods
We conducted a retrospective evaluation of the next generation sequencing data from 323 patients with advanced non-small cell lung cancer and EGFR-activating mutations after experiencing progression with epidermal growth factor receptor-tyrosine kinase inhibitor therapy. Our analysis specifically focused on the acquired changes to the KRAS gene.
Results
Among the 323 patients with advanced non-small cell lung cancer and EGFR-activating mutations who experienced resistance to epidermal growth factor receptor-tyrosine kinase inhibitor therapy, 14 individuals (4.3%) developed resistance due to acquired KRAS alterations. Of these 14 patients, 10 cases (71.4%) were due to KRAS missense mutations, 1 case (7.2%) was due to KRAS gene fusion and 3 cases (21.4%) were due to KRAS amplification. Notably, we identified one newly demonstrated KRAS gene fusion (KRAS and LMNTD1), one KRAS G13D and one KRAS K117N. The emergence of acquired KRAS alterations was often accompanied by novel mutations and high tumor mutation burden, with TP53, CNKN2A, PIK3CA, MYC, STK11, CDK4, BRCA2 and ERBB2 being the most frequently observed concurrent mutations. The median progression-free survival and overall survival for the 14 patients were 5.2 and 7.3 months, respectively. Acquired KRAS missense variants were associated with significantly worse progression-free survival compared with other KRAS variant subtypes (P < 0.028).
Conclusions
This study provides significant evidence of the role of acquired KRAS variants in the development of resistance to epidermal growth factor receptor-tyrosine kinase inhibitor therapy. Our results contribute to the growing body of knowledge on the mutational profiles associated with resistance to epidermal growth factor receptor-tyrosine kinase inhibitor treatment. Furthermore, our study highlights the KRAS gene change as a significant mechanism of resistance to epidermal growth factor receptor-tyrosine kinase inhibitor therapy.
The study identified 14 cases showing acquired KRAS alterations associated with resistance to epidermal growth factor receptor-tyrosine kinase inhibitor. These findings contribute to our comprehension of the mutational profiles linked to epidermal growth factor receptor-tyrosine kinase inhibitor resistance.
Zrenjski ravnik je kraška uravnava v severni Istri na Hrvaškem, ki ima na severnem in južnem robu obsežna območja ponornega kontaktnega krasa. Vodotoki pritekajo z eocenskih klastičnih kamnin in ...ponikajo v zakrasele karbonatne kamnine pretežno kredne starosti. Na območju smo opravili morfografsko, morfostrukturno in morfometrično analizo in nato morfogenetsko in morfodinamično interpretacijo. Zaključili smo, da je razvoj kontaktnega krasa potekal vsaj v treh različnih fazah. Najprej je deloval kot korozijska uravnava v plitvem krasu, kasneje je prišlo do tektonskega dviga in antecedentnega vrezovanja vodotokov v korozijsko uravnavo. Šele v zadnji fazi je prišlo do pretočitve površinskih tokov v podzemlje in oblikovanja kontaktnega krasa.
Due to groundbreaking developments and continuous progress, the treatment of advanced and metastatic non-small cell lung cancer (NSCLC) has become an exciting, but increasingly challenging task. This ...applies, in particular, to the subgroup of NSCLC with oncogenic driver alterations. While the treatment of epidermal growth factor receptor (EGFR)-mutated and anaplastic lymphoma kinase (ALK)-rearranged NSCLC with various tyrosine kinase inhibitors (TKIs) is well-established, new targets have been identified in the last few years and new TKIs introduced in clinical practice. Even for KRAS mutations, considered for a long time as an "un-targetable" alteration, promising new drugs are emerging. The detection and in-depth molecular analysis of resistance mechanisms has further fueled the development of new therapeutic strategies. The objective of this review is to give a comprehensive overview on the current landscape of targetable oncogenic alterations in NSCLC.
Ras GTPases are mutated at codons 12, 13, and 61, with different frequencies in KRas, HRas, and NRas and in a cancer-specific manner. The G13D mutant appears in 25% of KRas-driven colorectal cancers, ...while observed only rarely in HRas or NRas. Structures of Ras G13D in the three isoforms show an open active site, with adjustments to the D13 backbone torsion angles and with disconnected switch regions. KRas G13D has unique features that destabilize the nucleotide-binding pocket. In KRas G13D bound to GDP, A59 is placed in the Mg2+ binding site, as in the HRas-SOS complex. Structure and biochemistry are consistent with an intermediate level of KRas G13D bound to GTP, relative to wild-type and KRas G12D, observed in genetically engineered mouse models. The results explain in part the elevated frequency of the G13D mutant in KRas over the other isoforms of Ras.
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•Ras G13D proteins have open active sites with disconnected switches I and II•KRas G13D shows unique destabilization of the nucleotide-binding pocket•KRas G13D has attenuated oncogenic phenotype relative to KRas G12D•KRas G13D and KRas G12D are more sensitive to Erk than to Akt inhibition
Johnson et al. show that conformational states and biochemical properties of the KRas G13D oncogenic mutant in the context of isoform-specific residues unique to KRas lead to destabilization of the active site, consistent with its intermediate phenotype between wild-type KRas and KRas G12D in genetically engineered mice.
In calmodulin (CaM)-rich environments, oncogenic KRAS plays a critical role in adenocarcinomas by promoting PI3K/Akt signaling. We previously proposed that at elevated calcium levels in cancer, CaM ...recruits PI3Kα to the membrane and extracts K-Ras4B from the membrane, organizing a K-Ras4B–CaM–PI3Kα ternary complex. CaM can thereby replace a missing receptor-tyrosine kinase signal to fully activate PI3Kα. Recent experimental data show that CaM selectively promotes K-Ras signaling but not of N-Ras or H-Ras. How CaM specifically targets K-Ras and how it extracts it from the membrane in KRAS-driven cancer is unclear. Obtaining detailed structural information for a CaM–K-Ras complex is still challenging. Here, using molecular dynamics simulations and fluorescence experiments, we observed that CaM preferentially binds unfolded K-Ras4B hypervariable regions (HVRs) and not α-helical HVRs. The interaction involved all three CaM domains including the central linker and both lobes. CaM specifically targeted the highly polybasic anchor region of the K-Ras4B HVR that stably wraps around CaM's acidic linker. The docking of the farnesyl group to the hydrophobic pockets located at both CaM lobes further enhanced CaM–HVR complex stability. Both CaM and K-Ras4B HVR are highly flexible molecules, suggesting that their interactions permit highly dynamic flexible-body motions. We, therefore, anticipate that the flexible-body interaction is required to extract K-Ras4B from the membrane, as conformational plasticity enables CaM to orient efficiently to the polybasic HVR anchor, which is partially diffused into the liquid-phase membrane. Our structural model of the CaM–K-Ras4B HVR association provides plausible clues to CaM's regulatory action in PI3Kα activation involving the ternary complex in cell proliferation signaling by oncogenic K-Ras.
Oncogenic KRas reprograms pancreatic ductal adenocarcinoma (PDAC) cells to states which are highly resistant to apoptosis. Thus, a major preclinical goal is to identify effective strategies for ...killing PDAC cells. Artesunate (ART) is an anti-malarial that specifically induces programmed cell death in different cancer cell types, in a manner initiated by reactive oxygen species (ROS)-generation. In this study we demonstrate that ART specifically induced ROS- and lysosomal iron-dependent cell death in PDAC cell lines. Highest cytotoxicity was obtained in PDAC cell lines with constitutively-active KRas, and ART did not affect non-neoplastic human pancreatic ductal epithelial (HPDE) cells. We determined that ART did not induce apoptosis or necroptosis. Instead, ART induced ferroptosis, a recently described mode of ROS- and iron-dependent programmed necrosis which can be activated in Ras-transformed cells. Co-treatment with the ferroptosis inhibitor ferrostatin-1 blocked ART-induced lipid peroxidation and cell death, and increased long-term cell survival and proliferation. Importantly, analysis of PDAC patient mRNA expression indicates a dependency on antioxidant homeostasis and increased sensitivity to free intracellular iron, both of which correlate with Ras-driven sensitivity to ferroptosis. Overall, our findings suggest that ART activation of ferroptosis is an effective, novel pathway for killing PDAC cells.
The Role of Autophagy in Cancer Santana-Codina, Naiara; Mancias, Joseph D; Kimmelman, Alec C
Annual review of cancer biology,
03/2017, Letnik:
1, Številka:
1
Journal Article
Recenzirano
Odprti dostop
Autophagy is a highly conserved and regulated process that targets proteins and damaged organelles for lysosomal degradation to maintain cell metabolism, genomic integrity, and cell survival. The ...role of autophagy in cancer is dynamic and depends, in part, on tumor type and stage. Although autophagy constrains tumor initiation in normal tissue, some tumors rely on autophagy for tumor promotion and maintenance. Studies in genetically engineered mouse models support the idea that autophagy can constrain tumor initiation by regulating DNA damage and oxidative stress. In established tumors, autophagy can also be required for tumor maintenance, allowing tumors to survive environmental stress and providing intermediates for cell metabolism. Autophagy can also be induced in response to chemotherapeutics, acting as a drug-resistance mechanism. Therefore, targeting autophagy is an attractive cancer therapeutic option currently undergoing validation in clinical trials.
Allele-specific KRAS inhibitors are an emerging class of cancer therapies. KRAS-mutant (KRASMUT) non-small-cell lung cancers (NSCLCs) exhibit heterogeneous outcomes, driven by differences in ...underlying biology shaped by co-mutations. In contrast to KRASG12C NSCLC, KRASG12D NSCLC is associated with low/never-smoking status and is largely uncharacterized.
Clinicopathologic and genomic information were collected from patients with NSCLCs harboring a KRAS mutation at the Dana-Farber Cancer Institute (DFCI), Memorial Sloan Kettering Cancer Center, MD Anderson Cancer Center, and Imperial College of London. Multiplexed immunofluorescence for CK7, programmed cell death protein 1 (PD-1), programmed death-ligand 1 (PD-L1), Foxp3, and CD8 was carried out on a subset of samples with available tissue at the DFCI. Clinical outcomes to PD-(L)1 inhibition ± chemotherapy were analyzed according to KRAS mutation subtype.
Of 2327 patients with KRAS-mutated (KRASMUT) NSCLC, 15% (n = 354) harbored KRASG12D. Compared to KRASnon-G12D NSCLC, KRASG12D NSCLC had a lower pack-year (py) smoking history (median 22.5 py versus 30.0 py, P < 0.0001) and was enriched in never smokers (22% versus 5%, P < 0.0001). KRASG12D had lower PD-L1 tumor proportion score (TPS) (median 1% versus 5%, P < 0.01) and lower tumor mutation burden (TMB) compared to KRASnon-G12D (median 8.4 versus 9.9 mt/Mb, P < 0.0001). Of the samples which underwent multiplexed immunofluorescence, KRASG12D had lower intratumoral and total CD8+PD1+ T cells (P < 0.05). Among 850 patients with advanced KRASMUT NSCLC who received PD-(L)1-based therapies, KRASG12D was associated with a worse objective response rate (ORR) (15.8% versus 28.4%, P = 0.03), progression-free survival (PFS) hazard ratio (HR) 1.51, 95% confidence interval (CI) 1.45-2.00, P = 0.003, and overall survival (OS; HR 1.45, 1.05-1.99, P = 0.02) to PD-(L)1 inhibition alone but not to chemo-immunotherapy combinations ORR 30.6% versus 35.7%, P = 0.51; PFS HR 1.28 (95%CI 0.92-1.77), P = 0.13; OS HR 1.36 (95%CI 0.95-1.96), P = 0.09 compared to KRASnon-G12D.
KRASG12D lung cancers harbor distinct clinical, genomic, and immunologic features compared to other KRAS-mutated lung cancers and worse outcomes to PD-(L)1 blockade. Drug development for KRASG12D lung cancers will have to take these differences into account.
•Among KRAS NSCLC, KRASG12D NSCLC defines a subset with a lower pack-year smoking history, lower PD-L1 TPS, and lower TMB.•The smoking history among KRASG12D NSCLC is bimodal—those with a greater smoking history have higher PD-L1 TPS and TMB.•KRASG12D NSCLC is associated with diminished infiltration of CD8+ T cells.•Patients with KRASG12D NSCLC experience worse clinical outcomes to PD-(L)1 inhibition compared to KRASnon-G12D NSCLC.•KRASG12D NSCLC may respond better to chemotherapy added to PD-1 blockade, especially with a low smoking history/unknown PD-L1/TMB.
Mutant KRAS-induced tumorigenesis is highly involved in the progression of pancreatic, lung, and breast cancer. Comparatively, KRAS G12D and KRAS G12C are the most frequent mutations that promote ...cancer progression and aggressiveness. Although KRAS mutant inhibitors exhibit significant therapeutic potential, day by day, they are becoming resistant among patients. Multi-epitope based cancer vaccines are a promising alternative strategy that induces an immune response against tumor antigens. In the present study, we have designed, constructed, and validated a novel multi-epitope vaccine construct against KRAS G12D and G12C mutants using reverse vaccinology and immunoinformatics approaches. In addition, the vaccine construct was structurally refined and showed significant physiochemical properties, and could induce an immune response. Furthermore, the optimized vaccine construct was cloned into a pET‑28a (+) expression vector through in silico cloning. Conclusively, the multi-epitope vaccine construct is structurally stable, soluble, antigenic, non‑allergic, and non‑toxic. Further, it has to be studied in in vitro and in vivo to evaluate its therapeutic efficacy against KRAS-mutated cancers in the near future.
Graphical abstract