Frequency of KRAS p.Gly12Cys Mutation in Brazilian Patients With Lung Cancer

KRAS is one of the most frequently mutated genes in NSCLC. The frequency of KRAS mutations varies among distinct populations, accounting for approximately 25% in Whites and , 10% in East Asians. KRAS driver mutations are mostly located in codons 12 and 13, and the most frequent one is the p.Gly12Cys (c.34G.T) mutation. In lung cancer, KRAS mutations are associated with smokers and with a more aggressive phenotype. Efforts have been made in the past decade for rendering KRAS mutations susceptible to targeting. However, until lately, KRASmutated tumors were, unfortunately, undruggable.


INTRODUCTION
Lung cancer is the deadliest cancer worldwide, and in Brazil. 1,2 In the past decade, targeted therapies have revolutionized the clinical management of lung cancer, particularly in non-small-cell lung cancer (NSCLC) subtype. [3][4][5][6][7][8] The most successful examples of targeted therapies are the EGFR and ALK inhibitors, used for EGFRmutated and ALK-translocated tumors, respectively. 9,10 KRAS is one of the most frequently mutated genes in NSCLC. The frequency of KRAS mutations varies among distinct populations, accounting for approximately 25% in Whites and , 10% in East Asians. 11 KRAS driver mutations are mostly located in codons 12 and 13, and the most frequent one is the p.Gly12Cys (c.34G.T) mutation. [12][13][14][15][16][17][18] In lung cancer, KRAS mutations are associated with smokers and with a more aggressive phenotype. 12,[19][20][21][22] Efforts have been made in the past decade for rendering KRAS mutations susceptible to targeting. 23 However, until lately, KRASmutated tumors were, unfortunately, undruggable. 10 Recently, the agents AMG-510 (sotorasib, Amgen, Thousand Oaks, CA) and MRTX849 (adagrasib, Mirati Therapeutics, San Diego, CA) were developed to target the KRAS p.Gly12Cys mutation. 24,25 These specific inhibitors locked KRAS p.Gly12Cys mutation in an inactive state, hampering the oncogenic signals and allowed the normal function of remained wild-type KRAS. [24][25][26] In a phase I study, 32.2% (19 out of 59) of sotorasib-treated patients presented with objective response, and 88.1% (52 out of 59) presented with the disease control. 26 In a phase I and II study, 94% (17 out of 18) adagrasib-treated lung patients presented with disease control, and objective response was not yet available (KRYSTAL-1 study; ClinicalTrials.gov identifier: NCT03785249). 25 The frequency of KRAS p.Gly12Cys in admixture NSCLC populations remains scarce. Herein, we report the frequency of the KRAS p.Gly12Cys mutation in a series of 844 Brazilian NSCLC cases, followed by the data gathered from Brazil's previously reported studies.

METHODS
This retrospective study included 844 patients diagnosed with NSCLC. Seven hundred fifty-four patients were diagnosed at Barretos Cancer Hospital (BCH), and 90 patients were diagnosed at Bacchi Laboratory. Tobacco exposure, performance status, and overall survival data were provided for a subset of patients (BCH). This study was approved by the local IRB (Project no. 630/2012), and all procedures were performed following the Helsinki Declaration.
KRAS mutational status was evaluated from FFPE tumor tissue using different methodologies. The cases diagnosed at Barretos Cancer Hospital from 2014 to 2017 (n = 319) were genotyped by polymerase chain reaction followed by direct Sanger sequencing, and from 2018 to 2020 (n = 435) was assessed by next-generation sequencing, using the TruSight Tumor 15 (Illumina Waltham, MA) as reported by our group. 12,27,28 The cases diagnosed at Bacchi Laboratory were analyzed by qPCR TaqMan-MGB allelic discrimination assay (n = 67) and by FoundationOne (n = 23) between 2018 and 2020. 29,30 Genetic ancestry was analyzed in a subset of patients from Barretos Cancer Hospital (n = 660 out of 844), as previously described. 12 For statistical analysis, the percentage was used to describe categorical variables, and medians were used to describe continuous variables. Fisher's exact test and χ 2 test were used for the association between KRAS mutations and the clinicopathogic data. The log-rank test and the Kaplan-Meier curves were used to analyze patients' overall survival. The Cox regression method was used to investigate the association of clinicopathologic data to the outcome (death). All tests were made in the software IBM SPSS Statistics version 22 with a limit of statistical significance of 0.05.
KRAS mutation status was further associated with clinicopathologic and ancestry features (Table 1). Significant associations were found between the presence of KRAS mutations and smoking status, ECOG PS at diagnosis, and Native American ancestry (Table 1). Patients harboring KRAS mutation had worse overall survival than wild-type patients ( Fig 1B). Besides, smoking and higher ECOG PS at diagnosis were significantly associated with higher risk of death by multivariate Cox regression analysis (P , .0001 and P , .0001, respectively).

DISCUSSION
The KRAS p.Gly12Cys mutation became a new target for personalized therapy with the sotorasib and adagrasib. 23,25,26,31 Our study analyzed the frequency of p.Gly12Cys mutation in the Brazilian NSCLC population. We observed that 25% of the 3,247 cases were KRAS-mutated, and the most common variant was the p.Gly12Cys, present in 285 (9%) of the cases. Currently, expanded access is available for Brazilian patients and also for patients around the world, since both are non-US Food and Drug Administration-approved drugs. Once US Food and Drug Administration approves any of these drugs-sotorasib and adagrasib-compassionate drug use may be the option for obtaining access for Brazilian patients.
In our study, the presence of KRAS mutations was associated with smoking status (current or quitter) and worse overall survival. These data are in agreement with the literature. 12,[19][20][21][22] A recent review reported that KRAS mutations are present in 18%-32% of lung adenocarcinoma, 12.8% of large cell carcinoma, 10% of adenosquamous carcinomas, and 1.6%-7.1% of squamous-cell carcinomas in White patients. 32 Moreover, African-American patients with NSCLC are more frequently identified with KRAS mutations than White patients. 32 The frequency of KRAS mutations in Western populations with lung adenocarcinoma is about 26% and about 6% in the squamous-cell carcinoma population. 33 In Asian patients, the frequency of KRAS mutations is 11.2% of patients with NSCLC. 33 According to The Cancer Genome Atlas, KRAS mutations are found in 33% of lung adenocarcinoma. 34  The role of genetic ancestry in KRAS mutational status in NSCLC is poorly explored. A recent metadata analysis showed that KRAS mutations were more present in White and Black NSCLC patient groups than in Asian. 36 In a previous study, our group reported that KRAS mutations were associated with low Asian genetic ancestry background. 12 In the current study, using a panel of genetic ancestry markers, these findings were not confirmed in a multivariate analysis. Therefore, further studies using an admixture of populations are needed to clarify this important issue.
In conclusion, we showed that approximately 10% of Brazilian patients with NSCLC harbor the KRAS p.Gly12Cys variant and are therefore potentially responsive to the new anti-KRAS agents.