Small-Molecule RAS Inhibitors as Anticancer Agents: Discovery,Development, and Mechanistic Studies

Mutations of RAS oncogenes are responsible for about 30% of all human cancer types, including pancreatic, lung, and colorectal cancers. While KRAS1 is a pseudogene, mutation of KRAS2 (commonly known as KRAS oncogene) is directly or indirectly associated with human cancers. Among the RAS family, KRAS is the most abundant oncogene related to uncontrolled cellular proliferation to generate solid tumors in many types of cancer such as pancreatic carcinoma (over 80%), colon carcinoma (40–50%), lung carcinoma (30–50%), and other types of cancer. Once described as ‘undruggable’, RAS proteins have become ‘druggable’, at least to a certain extent, due to the continuous efforts made during the past four decades. In this account, we discuss the chemistry and biology (wherever available) of the small-molecule inhibitors (synthetic, semi-synthetic, and natural) of KRAS proteins that were published in the past decades. Commercial drugs, as well as investigational molecules from preliminary stages to clinical trials, are categorized and discussed in this study. In summary, this study presents an in-depth discussion of RAS proteins, classifies the RAS superfamily, and describes the molecular mechanism of small-molecule RAS inhibitors     

KRAS-Mutant Non-Small-Cell Lung Cancer: From Past Efforts to Future Challenges

KRAS is the most frequently mutated oncogene identified in human cancers. Despite the numerous efforts to develop effective specific inhibitors against KRAS, this molecule has remained “undruggable” for decades. The development of direct KRAS inhibitors, such as sotorasib, the first FDA-approved drug targeting KRAS G12C, or adagrasib, was made possible with the discovery of a small pocket in the binding switch II region of KRAS G12C. However, a new challenge is represented by the necessity to overcome resistance mechanisms to KRAS inhibitors. Another area to be explored is the potential role of co-mutations in the selection of the treatment strategy, particularly in the setting of immune checkpoint inhibitors. The aim of this review was to analyze the state-of-the-art of KRAS mutations in non-small-cell lung cancer by describing the biological structure of KRAS and exploring the clinical relevance of KRAS as a prognostic and predictive biomarker. We reviewed the different treatment approaches, focusing on the novel therapeutic strategies for the treatment of KRAS-mutant lung cancers.     

Molecular Glues: Capable Protein-Binding Small Molecules That Can Change Protein–Protein Interactions and Interactomes for the Potential Treatment of Human Cancer and Neurodegenerative Diseases

Molecular glue (MG) compounds are a type of unique small molecule that can change the protein–protein interactions (PPIs) and interactomes by degrading, stabilizing, or activating the target protein after their binging. These small-molecule MGs are gradually being recognized for their potential application in treating human diseases, including cancer. Evidence suggests that small-molecule MG compounds could essentially target any proteins, which play critical roles in human disease etiology, where many of these protein targets were previously considered undruggable. Intriguingly, most MG compounds with high efficacy for cancer treatment can glue on and control multiple key protein targets. On the other hand, a single key protein target can also be glued by multiple MG compounds with distinct chemical structures. The high flexibility of MG–protein interaction profiles provides rich soil for the growth and development of small-molecule MG compounds that can be used as molecular tools to assist in unraveling disease mechanisms, and they can also facilitate drug development for the treatment of human disease, especially human cancer. In this review, we elucidate this concept by using various types of small-molecule MG compounds and their corresponding protein targets that have been documented in the literature.     

KRAS: A Druggable Target in Colon Cancer Patients

Mutations in KRAS are among the most frequent aberrations in cancer, including colon cancer. KRAS direct targeting is daunting due to KRAS protein resistance to small molecule inhibition. Moreover, its elevated affinity to cellular guanosine triphosphate (GTP) has made the design of specific drugs challenging. Indeed, KRAS was considered ‘undruggable’. KRASG12C is the most commonly mutated variant of KRAS in non-small cell lung cancer. Currently, the achievements obtained with covalent inhibitors of this variant have given the possibility to assess the best therapeutic approach to KRAS-driven tumors. Mutation-related biochemical assets and the tissue of origin are expected to influence responses to treatment. Further attempts to obtain mutant-specific KRAS (KRASG12C) switch-II covalent inhibitors are ongoing and the results are promising. Drugs targeted to block KRAS effector pathways could be combined with direct KRAS inhibitors, immunotherapy or T cell-targeting approaches in KRAS-mutant tumors. The development of valuable combination regimens will be essential against potential mechanisms of resistance that may arise during treatment.     

Gene Therapy Targeting p53 and KRAS for Colorectal Cancer Treatment: A Myth or the Way Forward?

Colorectal cancer (CRC) is the third most commonly diagnosed malignancy worldwide and is responsible as one of the main causes of mortality in both men and women. Despite massive efforts to raise public awareness on early screening and significant advancements in the treatment for CRC, the majority of cases are still being diagnosed at the advanced stage. This contributes to low survivability due to this cancer. CRC patients present various genetic changes and epigenetic modifications. The most common genetic alterations associated with CRC are p53 and KRAS mutations. Gene therapy targeting defect genes such as TP53 (tumor suppressor gene encodes for p53) and KRAS (oncogene) in CRC potentially serves as an alternative treatment avenue for the disease in addition to the standard therapy. For the last decade, significant developments have been seen in gene therapy for translational purposes in treating various cancers. This includes the development of vectors as delivery vehicles. Despite the optimism revolving around targeted gene therapy for cancer treatment, it also has various limitations, such as a lack of availability of related technology, high cost of the involved procedures, and ethical issues. This article will provide a review on the potentials and challenges of gene therapy targeting p53 and KRAS for the treatment of CRC.     

Natural Products of Marine Origin for the Treatment of Colorectal and Pancreatic Cancers: Mechanisms and Potential

Gastrointestinal cancer refers to malignancy of the accessory organs of digestion, and it includes colorectal cancer (CRC) and pancreatic cancer (PC). Worldwide, CRC is the second most common cancer among women and the third most common among men. PC has a poor prognosis and high mortality, with 5-year relative survival of approximately 11.5%. Conventional chemotherapy treatments for these cancers are limited due to severe side effects and the development of drug resistance. Therefore, there is an urgent need to develop new and safe drugs for effective treatment of PC and CRC. Historically, natural sources—plants in particular—have played a dominant role in traditional medicine used to treat a wide spectrum of diseases. In recent decades, marine natural products (MNPs) have shown great potential as drugs, but drug leads for treating various types of cancer, including CRC and PC, are scarce. To date, marine-based drugs have been used against leukemia, metastatic breast cancer, soft tissue sarcoma, and ovarian cancer. In this review, we summarized existing studies describing MNPs that were found to have an effect on CRC and PC, and we discussed the potential mechanisms of action of MNPs as well as future prospects for their use in treating these cancers.     

If Virchow and Ehrlich Had Dreamt Together: What the Future Holds for KRAS-Mutant Lung Cancer

Non-small-cell lung cancer (NSCLC) with Kirsten rat sarcoma (KRAS) mutations has notoriously challenged oncologists and researchers for three notable reasons: (1) the historical assumption that KRAS is “undruggable”, (2) the disease heterogeneity and (3) the shaping of the tumor microenvironment by KRAS downstream effector functions. Better insights into KRAS structural biochemistry allowed researchers to develop direct KRAS(G12C) inhibitors, which have shown early signs of clinical activity in NSCLC patients and have recently led to an FDA breakthrough designation for AMG-510. Following the approval of immune checkpoint inhibitors for PDL1-positive NSCLC, this could fuel yet another major paradigm shift in the treatment of advanced lung cancer. Here, we review advances in our understanding of the biology of direct KRAS inhibition and project future opportunities and challenges of dual KRAS and immune checkpoint inhibition. This strategy is supported by preclinical models which show that KRAS(G12C) inhibitors can turn some immunologically “cold” tumors into “hot” ones and therefore could benefit patients whose tumors harbor subtype-defining STK11/LKB1 co-mutations. Forty years after the discovery of KRAS as a transforming oncogene, we are on the verge of approval of the first KRAS-targeted drug combinations, thus therapeutically unifying Paul Ehrlich’s century-old “magic bullet” vision with Rudolf Virchow’s cancer inflammation theory.     

Hyperinsulinemia and Its Pivotal Role in Aging,Obesity, Type 2 Diabetes,Cardiovascular Disease and Cancer

For many years, the dogma has been that insulin resistance precedes the development of hyperinsulinemia. However, recent data suggest a reverse order and place hyperinsulinemia mechanistically upstream of insulin resistance. Genetic background, consumption of the “modern” Western diet and over-nutrition may increase insulin secretion, decrease insulin pulses and/or reduce hepatic insulin clearance, thereby causing hyperinsulinemia. Hyperinsulinemia disturbs the balance of the insulin–GH–IGF axis and shifts the insulin : GH ratio towards insulin and away from GH. This insulin–GH shift promotes energy storage and lipid synthesis and hinders lipid breakdown, resulting in obesity due to higher fat accumulation and lower energy expenditure. Hyperinsulinemia is an important etiological factor in the development of metabolic syndrome, type 2 diabetes, cardiovascular disease, cancer and premature mortality. It has been further hypothesized that nutritionally driven insulin exposure controls the rate of mammalian aging. Interventions that normalize/reduce plasma insulin concentrations might play a key role in the prevention and treatment of age-related decline, obesity, type 2 diabetes, cardiovascular disease and cancer. Caloric restriction, increasing hepatic insulin clearance and maximizing insulin sensitivity are at present the three main strategies available for managing hyperinsulinemia. This may slow down age-related physiological decline and prevent age-related diseases. Drugs that reduce insulin (hyper) secretion, normalize pulsatile insulin secretion and/or increase hepatic insulin clearance may also have the potential to prevent or delay the progression of hyperinsulinemia-mediated diseases. Future research should focus on new strategies to minimize hyperinsulinemia at an early stage, aiming at successfully preventing and treating hyperinsulinemia-mediated diseases.     

Past and Future Strategies to Inhibit Membrane Localization of the KRAS Oncogene

KRAS is one of the most studied oncogenes. It is well known that KRAS undergoes post-translational modifications at its C-terminal end. These modifications are essential for its membrane location and activity. Despite significant efforts made in the past three decades to target the mechanisms involved in its membrane localization, no therapies have been approved and taken into the clinic. However, many studies have recently reintroduced interest in the development of KRAS inhibitors, either by directly targeting KRAS or indirectly through the inhibition of critical steps involved in post-translational KRAS modifications. In this review, we summarize the approaches that have been applied over the years to inhibit the membrane localization of KRAS in cancer and propose a new anti-KRAS strategy that could be used in clinic.     

Overview of Ferroptosis and Synthetic Lethality Strategies

Ferroptosis, a term first proposed in 2012, is iron-dependent, non-apoptotic regulatory cell death induced by erastin. Ferroptosis was originally discovered during synthetic lethal screening for drugs sensitive to RAS mutant cells, and is closely related to synthetic lethality. Ferroptosis sensitizes cancer stem cells and tumors that undergo epithelial−mesenchymal transition and are resistant to anticancer drugs or targeted therapy. Therefore, ferroptosis-inducing molecules are attractive new research targets. In contrast, synthetic lethal strategies approach mechanisms and genetic abnormalities that cannot be directly targeted by conventional therapeutic strategies, such as RAS mutations, hypoxia, and abnormalities in the metabolic environment. They also target the environment and conditions specific to malignant cells, have a low toxicity to normal cells, and can be used in combination with known drugs to produce new ones. However, the concept of synthetic lethality has not been widely adopted with ferroptosis. In this review, we surveyed the literature on ferroptosis-related factors and synthetic lethality to examine the potential therapeutic targets in ferroptosis-related molecules, focusing on factors related to synthetic lethality, discovery methods, clinical application stages, and issues in drug discovery.     

Moving beyond PARP Inhibition: Current State and Future Perspectives in Breast Cancer

Breast cancer is the most frequent and lethal tumor in women and finding the best therapeutic strategy for each patient is an important challenge. PARP inhibitors (PARPis) are the first, clinically approved drugs designed to exploit synthetic lethality in tumors harboring BRCA1/2 mutations. Recent evidence indicates that PARPis have the potential to be used both in monotherapy and combination strategies in breast cancer treatment. In this review, we show the mechanism of action of PARPis and discuss the latest clinical applications in different breast cancer treatment settings, including the use as neoadjuvant and adjuvant approaches. Furthermore, as a class, PARPis show many similarities but also certain critical differences which can have essential clinical implications. Finally, we report the current knowledge about the resistance mechanisms to PARPis. A systematic PubMed search, using the entry terms “PARP inhibitors” and “breast cancer”, was performed to identify all published clinical trials (Phase I-II-III) and ongoing trials (ClinicalTrials.gov), that have been reported and discussed in this review.     

Small-Molecule Modulation of Protein Lipidation: From Chemical Probes to Therapeutics

Protein lipidation is a widespread modification that regulates protein subcellular localization, structure and function. Dysregulation of protein lipidation has been implicated in various human diseases, including neurological disorders, infectious diseases and cancers. Thus lipid-modifying enzymes and their substrate proteins are emerging as attractive drug targets. The development of small-molecule modulators of protein lipidation has remarkably impacted our understanding of lipid-modification biology and potential therapeutics. In this review, we summarize recent progress in small-molecule targeting of protein lipidation and highlight therapeutic opportunities.     

The Role of EREG/EGFR Pathway in Tumor Progression

Aberrant activation of the epidermal growth factor receptor (EGFR/ERBB1) by erythroblastic leukemia viral oncogene homolog (ERBB) ligands contributes to various tumor malignancies, including lung cancer and colorectal cancer (CRC). Epiregulin (EREG) is one of the EGFR ligands and is low expressed in most normal tissues. Elevated EREG in various cancers mainly activates EGFR signaling pathways and promotes cancer progression. Notably, a higher EREG expression level in CRC with wild-type Kirsten rat sarcoma viral oncogene homolog (KRAS) is related to better efficacy of therapeutic treatment. By contrast, the resistance of anti-EGFR therapy in CRC was driven by low EREG expression, aberrant genetic mutation and signal pathway alterations. Additionally, EREG overexpression in non-small cell lung cancer (NSCLC) is anticipated to be a therapeutic target for EGFR-tyrosine kinase inhibitor (EGFR-TKI). However, recent findings indicate that EREG derived from macrophages promotes NSCLC cell resistance to EGFR-TKI treatment. The emerging events of EREG-mediated tumor promotion signals are generated by autocrine and paracrine loops that arise from tumor epithelial cells, fibroblasts, and macrophages in the tumor microenvironment (TME). The TME is a crucial element for the development of various cancer types and drug resistance. The regulation of EREG/EGFR pathways depends on distinct oncogenic driver mutations and cell contexts that allows specific pharmacological targeting alone or combinational treatment for tailored therapy. Novel strategies targeting EREG/EGFR, tumor-associated macrophages, and alternative activation oncoproteins are under development or undergoing clinical trials. In this review, we summarize the clinical outcomes of EREG expression and the interaction of this ligand in the TME. The EREG/EGFR pathway may be a potential target and may be combined with other driver mutation targets to combat specific cancers.     

Telomere Targeting Approaches in Cancer: Beyond Length Maintenance

Telomeres are crucial structures that preserve genome stability. Their progressive erosion over numerous DNA duplications determines the senescence of cells and organisms. As telomere length homeostasis is critical for cancer development, nowadays, telomere maintenance mechanisms are established targets in cancer treatment. Besides telomere elongation, telomere dysfunction impinges on intracellular signaling pathways, in particular DNA damage signaling and repair, affecting cancer cell survival and proliferation. This review summarizes and discusses recent findings in anticancer drug development targeting different “telosome” components.     

Computationally Empowered Workflow Identifies Novel Covalent Allosteric Binders for KRASG12C

Due to its frequent mutations in multiple lethal cancers, KRAS is one of the most-studied anticancer targets nowadays. Since the discovery of the druggable allosteric binding site containing a G12C mutation, KRAS^G12C has been the focus of attention in oncology research. We report here a computationally driven approach aimed at identifying novel and selective KRAS^G12C covalent inhibitors. The workflow involved initial enumeration of virtual molecules tailored for the KRAS allosteric binding site. Tools such as pharmacophore modeling, docking, and free-energy perturbations were deployed to prioritize the compounds with the best profiles. The synthesized naphthyridinone scaffold showed the ability to react with G12C and inhibit KRAS^G12C. Analogues were prepared to establish structure-activity relationships, while molecular dynamics simulations and crystallization of the inhibitor-KRAS^G12C complex highlighted an unprecedented binding mode.     

Thrombospondin-1 in Urological Cancer: Pathological Role,Clinical Significance,and Therapeutic Prospects

Angiogenesis is an important process for tumor growth and progression of various solid tumors including urological cancers. Thrombospondins (TSPs), especially TSP-1, are representative “anti”-angiogenic molecules and many studies have clarified their pathological role and clinical significance in vivo and in vitro. In fact, TSP-1 expression is associated with clinicopathological features and prognosis in many types of cancers. However, TSP-1 is a multi-functional protein and its biological activities vary according to the specific tumor environments. Consequently, there is no general agreement on its cancer-related function in urological cancers, and detailed information regarding regulative mechanisms is essential for a better understanding of its therapeutic effects and prognostic values. Various “suppressor genes” and “oncogenes” are known to be regulators and TSP-1-related factors under physiological and pathological conditions. In addition, various types of fragments derived from TSP-1 exist in a given tissue microenvironment and TSP-1 derived-peptides have specific activities. However, a detailed pathological function in human cancer tissues is not still understood. This review will focus on the pathological roles and clinical significance of TSP-1 in urological cancers, including prostate cancer, renal cell carcinoma, and urothelial cancer. In addition, special attention is paid to TSP-1-derived peptide and TSP-1-based therapy for malignancies.     

Roles of microRNAs in Gastrointestinal Cancer Stem Cell Resistance and Therapeutic Development

Resistance to cancer treatment is one of the major challenges currently faced when treating gastrointestinal (GI) cancers. A major contributing factor to this resistance is the presence of cancer stem cells (CSCs) in GI cancers (e.g., colorectal, pancreatic, gastric, liver cancer). Non-coding RNAs, such as microRNAs (miRNAs), have been found to regulate several key targets that are responsible for cancer stemness, and function as oncogenic miRNAs (oncomiRs) or tumor suppressor miRNAs. As a result, several miRNAs have been found to alter, or be altered by, the expression of CSC-defining markers and their related pathways. These miRNAs can be utilized to affect stemness in multiple ways, including directly targeting CSCs and enhancing the efficacy of cancer therapeutics. This review highlights current studies regarding the roles of miRNAs in GI CSCs, and efforts towards the development of cancer therapeutics.     

KRAS Affects Adipogenic Differentiation by Regulating Autophagy and MAPK Activation in 3T3-L1 and C2C12 Cells

Kirsten rat sarcoma 2 viral oncogene homolog (Kras) is a proto-oncogene that encodes the small GTPase transductor protein KRAS, which has previously been found to promote cytokine secretion, cell survival, and chemotaxis. However, its effects on preadipocyte differentiation and lipid accumulation are unclear. In this study, the effects of KRAS inhibition on proliferation, autophagy, and adipogenic differentiation as well as its potential mechanisms were analyzed in the 3T3-L1 and C2C12 cell lines. The results showed that KRAS was localized mainly in the nuclei of 3T3-L1 and C2C12 cells. Inhibition of KRAS altered mammalian target of rapamycin (Mtor), proliferating cell nuclear antigen (Pcna), Myc, peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer binding protein beta (C/ebp-β), diacylglycerol O-acyltransferase 1 (Dgat1), and stearoyl-coenzyme A desaturase 1 (Scd1) expression, thereby reducing cell proliferation capacity while inducing autophagy, enhancing differentiation of 3T3-L1 and C2C12 cells into mature adipocytes, and increasing adipogenesis and the capacity to store lipids. Moreover, during differentiation, KRAS inhibition reduced the levels of extracellular regulated protein kinases (ERK), c-Jun N-terminal kinase (JNK), p38, and phosphatidylinositol 3 kinase (PI3K) activation. These results show that KRAS has unique regulatory effects on cell proliferation, autophagy, adipogenic differentiation, and lipid accumulation.