# Ras Isoforms from Lab Benches to Lives--What Are We Missing and How Far Are We?
## 6 Future Recommendations for Devising Isoform-Specific Targeting
Researchers have developed Proteolysis Targeting Chimeras (PROTAC)-like degradation-resistant monobody called 12 VC1 that recognizes and noncovalently inhibited only active K-Ras mutants G12C and G12V [177]. Antibodies generally fail to detect mutant epitopes generated by cancer cells as they are present inside the cell. Single-chain variable fragments specific for these mutant peptides bound to HLA can be used to generate antibodies called mutation-associated neoantigen (MANAbodies). Previous studies have reported TCR-mimic antibody which targets K-Ras G12V/HLA-A*0201 [178]. This approach was further modified using antibody-drug conjugate (ADC) as TCRm-ADCs (2A5-MMAE and 2E8-MMAE) which showed higher specificity and lower toxicity [179]. Despite these signs of progress targeting K-Ras G12V remains a challenge unmet. Different K-Ras G12 mutant subtypes differently induct downstream signaling molecule and thus differ in their sensitivity to specific drugs. For example, G12C phosphorylates ERK to a higher level than G12D which is supported by the finding that MEK inhibitor selumetinib is more effective in K-Ras G12C tumors when compared to G12D tumors [180; 181]. Allele-specific mutation in K-Ras in different patient populations further adds complexity to finding an inhibitor and limits the monotherapeutic approach. Table 3 shows the various drugs/interventions currently in different phases of clinical trials for Ras isoforms. The biological differences due to the variability of K-Ras mutations are not well understood. The most common allele-specific mutation in K-Ras is K-Ras G12D and K-Ras-G12V. In pancreatic ductal adenocarcinoma (PDAC), G12D has low survivability whereas G12R has a higher probability of survival. In the case of non-small cell lung cancer (NSCLC), G12V or G12C mutant patients have a higher probability of survival when compared to other codon 12 mutations [182]. Also, K-Ras mutations co-occurring with mutations in tumor-suppressor genes may further lead to different biological behavior [183] and hence responsiveness to therapeutic agents. Comparative analysis of single and multiple mutations in mutant colorectal cancer (mCRC) shows K-Ras mutations in about 90% of patients. The most common mutation is present in exon 2 (92%), while 10% of the patients had multiple mutations. The median progression-free survival (PFS) and overall survival (O.S) was longer in patients with multiple mutations (22.7 months, 40.7 months) when compared to patients with single mutations (8.8 months, 12.8 months) [184] A detailed study on the clinical pathology and prognosis of the different K-Ras mutants shows that p.G12C and p.G12S variants are the most aggressive clinical subtypes in mCRC patients. A median follow-up performed 25.6 months post-diagnosis, shows the median survival was 7.3 months and 5 months in patients with K-Ras G12C and K-Ras G12S mutations respectively [185]. Patients with mutations in K-Ras exon 2 codons 12, 13 and N-Ras exon 2-4 are resistant to anti-EGFR treatment. The role of K-Ras in the negative prognosis of mCRC was observed in patients under different treatment regimens namely- irinotecan, oxaliplatin, and bevacizumab in various combinations. The lowest PFS, as well as OS, was observed in mCRC with K-Ras mutations. When compared to non-K-Ras mutated tumors, the PFS and OS of K-Ras G12C, K-Ras G13D were inferior. Although mutations in K-Ras G12V have a negative prognostic effect on PFS, K-Ras G12D and K-Ras G12V did not have any significant effect on OS [186]. In another study, the effect of K-Ras and N-Ras mutation on the prognosis of mCRC specifically in the case of liver-metastasis (LiM) and lung- metastasis (LuM) was assessed. K-Ras as well as N-Ras mutation affected the PFS and the OS of mCRC patients in the case of (LiM) but not in (LuM) [187]. In a subset of CRC patients who underwent hepatic resection, K-Ras mutations were negatively associated with OS and relapse-free survival (RFS) as well [188].