Microsecond Molecular Dynamics Simulation to Gain Insight Into the Binding of MRTX1133 and Trametinib With KRASG12D Mutant Protein for Drug Repurposing.

The Kirsten Rat Sarcoma (KRAS) G12D mutant protein is a primary driver of pancreatic ductal adenocarcinoma, necessitating the identification of targeted drug molecules. Repurposing of drugs quickly finds new uses, speeding treatment development. This study employs microsecond molecular dynamics simulations to unveil the binding mechanisms of the FDA-approved MEK inhibitor trametinib with KRASG12D, providing insights for potential drug repurposing. The binding of trametinib was compared with clinical trial drug MRTX1133, which demonstrates exceptional activity against KRASG12D, for better understanding of interaction mechanism of trametinib with KRASG12D. The resulting stable MRTX1133-KRASG12D complex reduces root mean square deviation (RMSD) values, in Switch I and II domains, highlighting its potential for inhibiting KRASG12D. MRTX1133's robust interaction with Tyr64 and disruption of Tyr96-Tyr71-Arg68 network showcase its ability to mitigate the effects of the G12D mutation. In contrast, trametinib employs a distinctive binding mechanism involving P-loop, Switch I and II residues. Extended simulations to 1 µs reveal sustained network interactions with Tyr32, Thr58, and GDP, suggesting a role of trametinib in maintaining KRASG12D in an inactive state and impede the further cell signaling. The decomposition binding free energy values illustrate amino acids' contributions to binding energy, elucidating ligand-protein interactions and molecular stability. The machine learning approach reveals that van der Waals interactions among the residues play vital role in complex stability and the potential amino acids involved in drug-receptor interactions of each complex. These details provide a molecular-level understanding of drug binding mechanisms, offering essential knowledge for further drug repurposing and potential drug discovery.

The KRAS G12D mutation increases the risk of unresectable recurrence of resectable colorectal liver-only metastasis.

PURPOSE: Unresectable recurrence is a critical predictor of outcomes for colorectal cancer patients. We attempted to identify the prognostic factors, especially for unresectable recurrence-free survival (URFS) as a new endpoint, in patients with resectable colorectal liver-only metastasis (CRLOM). METHODS: We investigated patients with resectable CRLOM, who underwent an R0 resection for both CRC and CRLOM between January, 2014 and March, 2019 at a single institution. The exclusion criteria were patients who received neoadjuvant treatment, the absence of data for genetic analyses, and the presence of multiple cancers, synchronous CRC, or familial adenomatous polyposis. The prognostic factors were examined retrospectively using data on pre-hepatectomy factors, including primary tumor molecular profiling results. RESULTS: We analyzed the data of 101 patients who underwent curative-intent surgery for CRLOM. Multivariate analysis revealed that KRAS G12D mutation-positivity (hazard ratio [HR]: 7.69; p < 0.01), RYR2 mutation-positivity (HR: 4.03; p < 0.01), and KRAS G12S mutation-positivity (HR: 3.96; p = 0.03), CA19-9 > 37 U/ml before hepatectomy (HR: 3.62; p < 0.01), and primary tumor pN2 stage (HR: 3.22; p = 0.03) were significant predictors of the URFS. CONCLUSIONS: This is the first study to show that specific KRAS and RYR2 mutations were associated with the URFS.

Structural perspectives on recent breakthrough efforts toward direct drugging of RAS and acquired resistance.

The Kirsten rat sarcoma viral oncoprotein homolog (KRAS) is currently a primary focus of oncologists and translational scientists, driven by exciting results with KRAS-targeted therapies for non-small cell lung cancer (NSCLC) patients. While KRAS mutations continue to drive high cancer diagnosis and death, researchers have developed unique strategies to target KRAS variations. Having been investigated over the past 40 years and considered "undruggable" due to the lack of pharmacological binding pockets, recent breakthroughs and accelerated FDA approval of the first covalent inhibitors targeting KRASG12C, have largely sparked further drug development. Small molecule development has targeted the previously identified primary location alterations such as G12, G13, Q61, and expanded to address the emerging secondary mutations and acquired resistance. Of interest, the non-covalent KRASG12D targeting inhibitor MRTX-1133 has shown promising results in humanized pancreatic cancer mouse models and is seemingly making its way from bench to bedside. While this manuscript was under review a novel class of first covalent inhibitors specific for G12D was published, These so-called malolactones can crosslink both GDP and GTP bound forms of G12D. Inhibition of the latter state suppressed downstream signaling and cancer cell proliferation in vitro and in mouse xenografts. Moreover, a non-covalent pan-KRAS inhibitor, BI-2865, reduced tumor proliferation in cell lines and mouse models. Finally, the next generation of KRAS mutant-specific and pan-RAS tri-complex inhibitors have revolutionized RAS drug discovery. This review will give a structural biology perspective on the current generation of KRAS inhibitors through the lens of emerging secondary mutations and acquired resistance.

Anti-tumor efficacy of HRS-4642 and its potential combination with proteasome inhibition in KRAS G12D-mutant cancer.

KRAS G12D is the most frequently mutated oncogenic KRAS subtype in solid tumors and remains undruggable in clinical settings. Here, we developed a high affinity, selective, long-acting, and non-covalent KRAS G12D inhibitor, HRS-4642, with an affinity constant of 0.083 nM. HRS-4642 demonstrated robust efficacy against KRAS G12D-mutant cancers both in vitro and in vivo. Importantly, in a phase 1 clinical trial, HRS-4642 exhibited promising anti-tumor activity in the escalating dosing cohorts. Furthermore, the sensitization and resistance spectrum for HRS-4642 was deciphered through genome-wide CRISPR-Cas9 screening, which unveiled proteasome as a sensitization target. We further observed that the proteasome inhibitor, carfilzomib, improved the anti-tumor efficacy of HRS-4642. Additionally, HRS-4642, either as a single agent or in combination with carfilzomib, reshaped the tumor microenvironment toward an immune-permissive one. In summary, this study provides potential therapies for patients with KRAS G12D-mutant cancers, for whom effective treatments are currently lacking.

Pathways and mechanism of MRTX1133 binding to KRAS G12D elucidated by molecular dynamics simulations and Markov state models.

KRAS G12D is the most common oncogenic mutation identified in several types of cancer. Therefore, design of inhibitors targeting KRAS G12D represents a promising strategy for anticancer therapy. MRTX1133 is a highly potent inhibitor (approximate experiment Kd ≈ 0.0002 nM) of KRAS G12D and is currently in Phase 1/2 study, however, pathways of the compound binding to KRAS G12D has remained unknown, and the mechanism underlying the complicated dynamic process are challenging to capture experimentally, which hinder the structure-based anti-cancer drug design. Here, using MRTX1133 as a probe, unbiased molecular dynamics (MD) was used to simulate the process of MRTX1133 spontaneously binding to KRAS G12D. In six of 42 independent MD simulation (a total of 99 µs), MRTX1133 was observed to successfully associate with KRAS G12D. The kinetically metastable states refer to the potential pathways of MRTX1133 binding to KRAS G12D were revealed by Markov state models (MSM) analysis. Additionally, 8 key residues that are essential for MRTX1133 recognition and tight binding at the preferred low energy states were identified by MM/GBSA analysis. In sum, this study provides a new perspective on understanding the pathways and mechanism of MRTX1133 binding to KRAS G12D.

A novel Imidazo[1,2-a]pyridine derivative modulates active KRASG12D through off-like conformational shifts in switch-I and switch-II regions, mimicking inactive KRASG12D.

KRASG12D are the most prevalent oncogenic mutations and a promising target for solid tumor therapies. However, its inhibition exhibits tremendous challenge due to the necessity of high binding affinity to obviate the need for covalent binders. Here we report the evidence of a novel class of Imidazo[1,2-a]pyridine derivative as potentially significant novel inhibitors of KRASG12D, discovered through extensive ligand-based screening against 2-[(2R)-piperidin-2-yl]-1H-indole, an important scaffold for KRASG12D inhibition via switch-I/II (S-I/II) pocket. The proposed compounds exhibited similar binding affinities and overlapped pose configurations to 2-[(2R)-piperidin-2-yl]-1H-indole, serving as a reliable starting point for drug discovery. Comparative free energy profiles demonstrated that C4 [2-methyl-3-((5-phenyl-1H-1,2,4-triazol-3-yl)methyl)imidazo[1,2-a]pyridine] effectively shifted the protein to a stable low-energy conformation via a prominent transition state. The conformational changes across the transition revealed the conformational shift of switch-I and II to a previously known off-like conformation of inactive KRASG12D with rmsd of 0.91 Å. These conformations were even more prominent than the privileged scaffold 2-[(2R)-piperidin-2-yl]-1H-indole. The representative structure overlay of C4 and another X-ray crystallography solved BI-2852 bound inactive KRASG12D revealed that Switch-I and II exhibited off-like conformations. The cumulative variance across the first eigenvalue that accounted for 57 % of the collective variance validated this on-to-off transition. In addition, the relative interaction of C4 binding showed consistent patterns with BI-2852. Taken together, our results support the inhibitory activity of [2-methyl-3-((5-phenyl-1H-1,2,4-triazol-3-yl)methyl)imidazo[1,2-a]pyridine] by shifting active KRASG12D to an inactive conformation.

In Silico Prediction of New Inhibitors for Kirsten Rat Sarcoma G12D Cancer Drug Target Using Machine Learning-Based Virtual Screening, Molecular Docking, and Molecular Dynamic Simulation Approaches.

Single-point mutations in the Kirsten rat sarcoma (KRAS) viral proto-oncogene are the most common cause of human cancer. In humans, oncogenic KRAS mutations are responsible for about 30% of lung, pancreatic, and colon cancers. One of the predominant mutant KRAS G12D variants is responsible for pancreatic cancer and is an attractive drug target. At the time of writing, no Food and Drug Administration (FDA) approved drugs are available for the KRAS G12D mutant. So, there is a need to develop an effective drug for KRAS G12D. The process of finding new drugs is expensive and time-consuming. On the other hand, in silico drug designing methodologies are cost-effective and less time-consuming. Herein, we employed machine learning algorithms such as K-nearest neighbor (KNN), support vector machine (SVM), and random forest (RF) for the identification of new inhibitors against the KRAS G12D mutant. A total of 82 hits were predicted as active against the KRAS G12D mutant. The active hits were docked into the active site of the KRAS G12D mutant. Furthermore, to evaluate the stability of the compounds with a good docking score, the top two complexes and the standard complex (MRTX-1133) were subjected to 200 ns MD simulation. The top two hits revealed high stability as compared to the standard compound. The binding energy of the top two hits was good as compared to the standard compound. Our identified hits have the potential to inhibit the KRAS G12D mutation and can help combat cancer. To the best of our knowledge, this is the first study in which machine-learning-based virtual screening, molecular docking, and molecular dynamics simulation were carried out for the identification of new promising inhibitors for the KRAS G12D mutant.

Discovery, optimization and biological activity evaluation of genipin derivatives as potential KRAS G12D inhibitors.

A series of genipin derivatives were designed and synthesized as potential inhibitors targeted KRAS G12D mutation. The majority of these compounds demonstrated potential antiproliferative effects against KRAS G12D mutant tumor cells (CT26 and A427). Notably, seven compounds exhibited the anticancer effects with IC50 values ranging from 7.06 to 9.21 µM in CT26 (KRASG12D) and A427 (KRASG12D) cells and effectively suppressed the colony formation of CT26 cells. One representative compound SK12 was selected for further investigation into biological activity and action mechanisms. SK12 markedly induced apoptosis in CT26 cells in a concentration-dependent manner. Moreover, SK12 elevated the levels of reactive oxygen species (ROS) in tumor cells and exhibited a modulatory effect on the KRAS signaling pathway, thereby inhibiting the activation of downstream phosphorylated proteins. The binding affinity of SK12 to KRAS G12D protein was further confirmed by the surface plasmon resonance (SPR) assay with a binding KD of 157 µM. SK12 also exhibited notable anticancer efficacy in a nude mice tumor model. The relative tumor proliferation rate (T/C) of the experimental group (50 mg/kg) was 31.04 % (P < 0.05), while maintaining a commendable safety profile.

Impact of KRASG12D subtype and concurrent pathogenic mutations on advanced non-small cell lung cancer outcomes / Impacto del subtipo KRASG12D y mutaciones patogénicas concurrentes en los resultados del cáncer de pulmón de células no pequeñas avanzado

Purpose: Mutations in the Kirsten rat sarcoma viral (KRAS) oncogene constitute a significant driver of lung adenocarcinoma, present in 10–40% of patients, which exhibit heterogeneous clinical outcomes, mainly driven by concurrent genetic alterations. However, characterization of KRAS mutational subtypes and their impact on clinical outcomes in Latin America is limited. Methods: A cohort study was conducted at the National Cancer Institute (INCan) of Mexico. Individuals with advance-staged of adenocarcinoma and KRAS mutations, detected by next-generation sequencing, having undergone at least one line of therapy were included for analysis. Clinical and pathological characteristics were retrieved from institutional database from June 2014 to March 2023. Results: KRAS was identified in fifty-four (15.6%) of 346 patients, among which 50 cases were included for analysis. KRASG12D (n = 16, 32%) and KRASG12C (n = 16, 32%) represented the most prevalent subtypes. KRASG12D mutations were associated with female (p = 0.018), never smokers (p = 0.108), and concurrences with EGFR (25.0% vs. 17.6%, p = 0.124) and CDKN2A (18.8% vs. 14.7%, p = 0.157). KRASG12D patients showed a better ORR (66.6% vs. 30.0%; OR 4.66, 95% CI 1.23–17.60, p = 0.023) and on multivariate analysis was significantly associated with better PFS (HR 0.36, 95% CI 0.16–0.80; p = 0.012) and OS (HR 0.24, 95% CI 0.08–0.70; p = 0.009). Conclusions: To our knowledge, this study represents the first effort to comprehensively characterize the molecular heterogeneity of KRAS-mutant NSCLC in Latin American patients. Our data reinforce the current view that KRAS-mutated NSCLC is not a single oncogene-driven disease and emphasizes the prognostic impact of diverse molecular profiles in this genomically defined subset of NSCLC. Further validation is warranted in larger multicenter Latin American cohorts to confirm our findings.(AU)

Design, synthesis, and evaluation of purine and pyrimidine-based KRAS G12D inhibitors: Towards potential anticancer therapy.

Oncogenic RAS mutations, commonly observed in human tumors, affect approximately 30% of cancer cases and pose a significant challenge for effective cancer treatment. Current strategies to inhibit the KRAS G12D mutation have shown limited success, emphasizing the urgent need for new therapeutic approaches. In this study, we designed and synthesized several purine and pyrimidine analogs as inhibitors for the KRAS G12D mutation. Our synthesized compounds demonstrated potent anticancer activity against cell lines with the KRAS G12D mutation, effectively impeding their growth. They also exhibited low toxicity in normal cells, indicating their selective action against cancer cells harboring the KRAS G12D mutation. Notably, the lead compound, PU1-1 induced the programmed cell death of KRAS G12D-mutated cells and reduced the levels of active KRAS and its downstream signaling proteins. Moreover, PU1-1 significantly shrunk the tumor size in a pancreatic xenograft model induced by the KRAS G12D mutation, further validating its potential as a therapeutic agent. These findings highlight the potential of purine-based KRAS G12D inhibitors as candidates for targeted cancer therapy. However, further exploration and optimization of these compounds are essential to meet the unmet clinical needs of patients with KRAS-mutant cancers.

Clinicopathologic features, concurrent genomic alterations, and clinical outcomes of patients with KRAS G12D mutations in resected lung adenocarcinoma.

BACKGROUND: In light of the ongoing clinical development of KRAS G12D-specific inhibitors, we sought to investigate the clinicopathologic, co-occurring genomic features and outcomes of patients with KRAS G12D-mutant lung adenocarcinoma. METHODS: 3828 patients with completely resected primary lung adenocarcinomas were examined for KRAS mutations between 2008 and 2020. The association between KRAS G12D and clinicopathologic features, molecular profiles, and outcomes was investigated. RESULTS: 65 patients (1.7%) with KRAS G12D-mutant lung adenocarcinoma were identified. KRAS G12D mutation was more frequent in males, former/current smokers, radiologic solid tumors, and invasive mucinous adenocarcinoma. TP53 and STK11 were the two most frequent concomitant mutations in the KRAS G12D group. KRAS G12D mutation did not appear to be a prognostic factor in resected stage I-III lung adenocarcinomas, while KRAS non-G12D mutation was related to worse survival, especially in stage I tumors. KRAS G12D mutations were associated with positive but low (1-49%) PD-L1 expression compared to negative (<1%), while KRAS non-G12D mutation was associated with high PD-L1 expression (≥50%). TP53 co-mutation indicated higher PD-L1 expression, while STK11 co-mutation had a negligible impact on PD-L1 expression. Furthermore, data mining of MSK datasets from cBioPortal revealed that KRAS G12D and SKT11 co-mutation were associated with a diminished response to immunotherapy. CONCLUSIONS: KRAS G12D-mutant lung adenocarcinoma harbored unique clinicopathologic and genomic characteristics. Despite not being prognostic in resected lung adenocarcinoma, KRAS G12D might be a valuable biomarker in combination with certain co-mutations for identifying relevant subgroups of patients that could eventually influence treatment regimens.

Gluing GAP to RAS Mutants: A New Approach to an Old Problem in Cancer Drug Development.

Mutated genes may lead to cancer development in numerous tissues. While more than 600 cancer-causing genes are known today, some of the most widespread mutations are connected to the RAS gene; RAS mutations are found in approximately 25% of all human tumors. Specifically, KRAS mutations are involved in the three most lethal cancers in the U.S., namely pancreatic ductal adenocarcinoma, colorectal adenocarcinoma, and lung adenocarcinoma. These cancers are among the most difficult to treat, and they are frequently excluded from chemotherapeutic attacks as hopeless cases. The mutated KRAS proteins have specific three-dimensional conformations, which perturb functional interaction with the GAP protein on the GAP-RAS complex surface, leading to a signaling cascade and uncontrolled cell growth. Here, we describe a gluing docking method for finding small molecules that bind to both the GAP and the mutated KRAS molecules. These small molecules glue together the GAP and the mutated KRAS molecules and may serve as new cancer drugs for the most lethal, most difficult-to-treat, carcinomas. As a proof of concept, we identify two new, drug-like small molecules with the new method; these compounds specifically inhibit the growth of the PANC-1 cell line with KRAS mutation G12D in vitro and in vivo. Importantly, the two new compounds show significantly lower IC50 and higher specificity against the G12D KRAS mutant human pancreatic cancer cell line PANC-1, as compared to the recently described selective G12D KRAS inhibitor MRTX-1133.

Early elevations of RAS protein level and activity are critical for the development of PDAC in the context of inflammation.

The KRASG12D mutation was believed to be locked in a GTP-bound form, rendering it fully active. However, recent studies have indicated that the presence of mutant KRAS alone is insufficient; it requires additional activation through inflammatory stimuli to effectively drive the development of pancreatic ductal adenocarcinoma (PDAC). It remains unclear to what extent RAS activation occurs during the development of PDAC in the context of inflammation. Here, in a mouse model with the concurrent expression of KrasG12D/+ and inflammation mediator IKK2 in pancreatic acinar cells, we showed that, compared to KRASG12D alone, the cooperative interaction between KRASG12D and IKK2 rapidly elevated both the protein level and activity of KRASG12D and NRAS in a short term. This high level was sustained throughout the rest phase of PDAC development. These results suggest that inflammation not only rapidly augments the activity but also the protein abundance, leading to an enhanced total amount of GTP-bound RAS (KRASG12D and NRAS) in the early stage. Notably, while KRASG12D could be further activated by IKK2, not all KRASG12D proteins were in the GTP-bound state. Overall, our findings suggest that although KRASG12D is not fully active in the context of inflammation, concurrent increases in both the protein level and activity of KRASG12D as well as NRAS at the early stage by inflammation contribute to the rise in total GTP-bound RAS.

Impact of KRASG12D subtype and concurrent pathogenic mutations on advanced non-small cell lung cancer outcomes.

PURPOSE: Mutations in the Kirsten rat sarcoma viral (KRAS) oncogene constitute a significant driver of lung adenocarcinoma, present in 10-40% of patients, which exhibit heterogeneous clinical outcomes, mainly driven by concurrent genetic alterations. However, characterization of KRAS mutational subtypes and their impact on clinical outcomes in Latin America is limited. METHODS: A cohort study was conducted at the National Cancer Institute (INCan) of Mexico. Individuals with advance-staged of adenocarcinoma and KRAS mutations, detected by next-generation sequencing, having undergone at least one line of therapy were included for analysis. Clinical and pathological characteristics were retrieved from institutional database from June 2014 to March 2023. RESULTS: KRAS was identified in fifty-four (15.6%) of 346 patients, among which 50 cases were included for analysis. KRASG12D (n = 16, 32%) and KRASG12C (n = 16, 32%) represented the most prevalent subtypes. KRASG12D mutations were associated with female (p = 0.018), never smokers (p = 0.108), and concurrences with EGFR (25.0% vs. 17.6%, p = 0.124) and CDKN2A (18.8% vs. 14.7%, p = 0.157). KRASG12D patients showed a better ORR (66.6% vs. 30.0%; OR 4.66, 95% CI 1.23-17.60, p = 0.023) and on multivariate analysis was significantly associated with better PFS (HR 0.36, 95% CI 0.16-0.80; p = 0.012) and OS (HR 0.24, 95% CI 0.08-0.70; p = 0.009). CONCLUSIONS: To our knowledge, this study represents the first effort to comprehensively characterize the molecular heterogeneity of KRAS-mutant NSCLC in Latin American patients. Our data reinforce the current view that KRAS-mutated NSCLC is not a single oncogene-driven disease and emphasizes the prognostic impact of diverse molecular profiles in this genomically defined subset of NSCLC. Further validation is warranted in larger multicenter Latin American cohorts to confirm our findings.

Alveolar type I cells can give rise to KRAS-induced lung adenocarcinoma.

Lung adenocarcinoma (LUAD) is the most prevalent subtype of lung cancer and presents clinically with a high degree of biological heterogeneity and distinct clinical outcomes. The current paradigm of LUAD etiology posits alveolar epithelial type II (AT2) cells as the primary cell of origin, while the role of AT1 cells in LUAD oncogenesis remains unknown. Here, we examine oncogenic transformation in mouse Gram-domain containing 2 (Gramd2)+ AT1 cells via oncogenic KRASG12D. Activation of KRASG12D in AT1 cells induces multifocal LUAD, primarily of papillary histology. Furthermore, KRT8+ intermediate cell states were observed in both AT2- and AT1-derived LUAD, but SCGB3A2+, another intermediate cell marker, was primarily associated with AT1 cells, suggesting different mechanisms of tumor evolution. Collectively, our study reveals that Gramd2+ AT1 cells can serve as a cell of origin for LUAD and suggests that distinct subtypes of LUAD based on cell of origin be considered in the development of therapeutics.

Salvianolic acid F suppresses KRAS-dependent lung cancer cell growth through the PI3K/AKT signaling pathway.

BACKGROUND: KRAS mutation is a common driver of NSCLC, and there is a high proportion of lung cancer patients with KRAS G12C and G12D mutation. KRAS was previously considered an "undruggable" target, but the first KRAS G12C mutation-targeted drug AMG510, entered the market in 2021. However, treatments for G12D mutant tumors remain to be discovered. Salvianolic acid F (SalF), a monomer derived from the traditional Chinese medicine Salvia miltiorrhiza (SM), and KRAS had high binding affinity, especially for KRAS G12D. There is an urgent need to investigate effective and safe novel targeted therapies against KRAS G12D-driven NSCLC. METHODS: To evaluate the anticancer effect of SalF, we used KRAS-overexpressing lung cancer cells in vitro, a subcutaneous transplant tumor model, and KRAS G12D mice model in vivo. Then, the binding effect of SalF and KRAS was investigated using molecular docking, proteolytic assays and protein thermal shift assays. More critically, the PI3K/AKT signaling pathway in the lung was investigated utilizing RT-qPCR and Western Blotting. RESULTS: This is the first study to evaluate the anticancer effect of SalF on KRAS-overexpressing lung cancer cells or KRAS G12D lung tumors in vivo. We demonstrated that SalF inhibits OE-KRAS A549 cell migration, proliferation and promotes apoptosis in vitro. In addition, we used a subcutaneous transplant tumor model to show that SalF suppresses the growth of lung cancer cells in vivo. Interestingly, our group found that SalF was strongly bound to G12D and could decrease the stability and promoted the degradation of the KRAS G12D mutant through molecular docking, proteolytic assays and protein thermal shift assays. Further research demonstrated that in the KrasG12D mice model, after SalF treatment, the number and size of mouse lung tumors were significantly reduced. More importantly, SalF can promote apoptosis by inhibiting downstream PI3K/AKT signaling pathway activation. CONCLUSION: SalF activated apoptosis signaling pathways, suppressed anti-apoptotic genes, and inhibited lung cancer cell growth. These datas suggested that SalF could effectively inhibit the growth of lung tumors with KRAS G12D mutation. SalF may be a novel inhibitor against KRAS G12D, providing a strong theoretical basis for the clinical treatment of lung cancer with KRAS mutations.

Reverse vaccinology and immunoinformatics approaches to design multi-epitope based vaccine against oncogenic KRAS.

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 pET­28a (+) expression vector through in silico cloning. Conclusively, the multi-epitope vaccine construct is structurally stable, soluble, antigenic, non­allergic, and non­toxic. 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.

Next batter up! Targeting cancers with KRAS-G12D mutations.

KRAS is the most frequently mutated oncogene in cancer. Activating mutations in codon 12, especially G12D, have the highest prevalence across a range of carcinomas and adenocarcinomas. With inhibitors to KRAS-G12D now entering clinical trials, understanding the biology of KRAS-G12D cancers, and identifying biomarkers that predict therapeutic response is crucial. In this Review, we discuss the genomics and biology of KRAS-G12D adenocarcinomas, including histological features, transcriptional landscape, the immune microenvironment, and how these factors influence response to therapy. Moreover, we explore potential therapeutic strategies using novel G12D inhibitors, leveraging knowledge gained from clinical trials using G12C inhibitors.

Elimination of oncogenic KRAS in genetic mouse models eradicates pancreatic cancer by inducing FAS-dependent apoptosis by CD8+ T cells.

Oncogenic KRASG12D (KRAS∗) is critical for the initiation and maintenance of pancreatic ductal adenocarcinoma (PDAC) and is a known repressor of tumor immunity. Conditional elimination of KRAS∗ in genetic mouse models of PDAC leads to the reactivation of FAS, CD8+ T cell-mediated apoptosis, and complete eradication of tumors. KRAS∗ elimination recruits activated CD4+ and CD8+ T cells and promotes the activation of antigen-presenting cells. Mechanistically, KRAS∗-mediated immune evasion involves the epigenetic regulation of Fas death receptor in cancer cells, via methylation of its promoter region. Furthermore, analysis of human RNA sequencing identifies that high KRAS expression in PDAC tumors shows a lower proportion of CD8+ T cells and demonstrates shorter survival compared with tumors with low KRAS expression. This study highlights the role of CD8+ T cells in the eradication of PDAC following KRAS∗ elimination and provides a rationale for the combination of KRAS∗ targeting with immunotherapy to control PDAC.