EGF+61 A>G polymorphism does not predict response to first-generation EGFR tyrosine kinase inhibitors in lung cancer patients

Introduction

Epidermal growth factor (EGF) and its receptor (EGFR) play a key role in the pathogenesis of several tumors, including non-small cell lung cancer (NSCLC).^{1, 2} EGFR is a critical therapeutic target, and the presence of EGFR mutations has previously successfully guided clinical management in NSCLC.^{3, 4} EGFR mutations located at the tyrosine kinase domain, sensitizes NSCLC to treatment with anti-EGFR tyrosine kinase inhibitors (TKIs), such as erlotinib and gefitinib.^3-6 However, most TKI-treated patients will experience disease progression due to resistance mechanisms, such as the presence of EGFR resistance mutations.⁷

A single nucleotide polymorphism (SNP) in the 5′ untranslated region of the EGF gene (EGF+61 A>G — rs4444903) has been associated with increased levels of EGF and consequently is a risk for cancer development of distinct tumors.² In lung cancer, conflicting results have been reported in different populations.^8-11 Importantly, in advanced rectal cancer, EGF+61 A>G SNP predicted a complete pathologic response to cetuximab, an anti-EGFR monoclonal antibody.¹²

Although cetuximab and TKIs are two different forms of therapies directed at EGFR, they both result in blocking signal transduction.¹³ Thus, we hypothesized that EGF+61 A>G SNP might be associated with a response to first-generation TKIs in lung cancer. Thereafter, we investigated the association between EGF+61 A>G genotype and TKI response in a Brazilian series comprised of 111 TKI-treated lung adenocarcinoma patients harboring EGFR-sensitizing mutations.

Methods

Study population

This was a retrospective and observational study, which included patients with NSCLC, diagnosed between 2001 and 2017 at Barretos Cancer Hospital. The present study was approved by the Barretos Cancer Hospital IRB (Project No. 630/2012), and all methods were performed following the Helsinki declaration.

From a series of lung adenocarcinoma cases (n = 870), 183 patients presented with EGFR mutations and 127 out of 183 patients were TKI-treated (Fig 1). The response to TKI-treatment was determined using RECIST 1.1, and the clinical response was categorized as partial response (PR) for overall response rate (ORR), stable disease (SD) for disease control rate (DCR), or disease progression (DP). NSCLC patients harboring EGFR resistance mutations (eg, T790M) are not currently eligible for erlotinib, and gefitinib treatment; thus, these patients (n = 7) were not evaluated (Fig 1). Nine patients were excluded due to inconclusive genotyping results (Fig 1). The demographic and clinicopathological data from eligible patients (n = 111) were obtained by reviewing the medical records (Table 1).

Table 1. Demographic and clinicopathological features of EGFR-mutated TKI-treated NSCLC patients enrolled in the study (n = 111)

Characteristics		n = 111	Frequencies
Sex	Male	47	42.3%
	Female	64	57.7%
Age	Median (SD)	61.75 (11.48)
	≤63 years	58	52.3%
	>63 years	53	47.7%
Self-reported skin color	White	96	86.5%
	Other	15	13.5%
Smoking status	Never	65	58.6%
	Current	17	15.3%
	Former	28	25.2%
	NA	1	0.9%
ECOG PS	0–1	78	70.3%
	2	20	18.0%
	3–4	8	7.2%
	NA	5	4.5%
Weight loss in the last six months prior to diagnosis	>10%	18	16.2%
	<10%	46	41.4%
	None	39	35.1%
	NA	8	7.3%
TNM stage	I/II/III	12	10.8%
	IV	99	89.2%
Histological subtype	Acinar	42	37.8%
	Mucinous	1	0.9%
	Lepidic	8	7.3%
	Papillary	10	9.0%
	Solid	10	9.0%
	Others	7	6.3%
	Not defined	33	29.7%
Central nervous system metastasis	Yes	32	28.8%
	No	79	71.2%
Current status	Alive	24	21.6%
	Dead due to cancer	80	72.1%
	Dead due to comorbidity	1	0.9%
	NA	6	5.4%
Response to TKI treatment	Partial response	78	70.3%
	Stable disease	18	16.2%
	Disease progression	15	13.5%
EGFR mutation according to exon location	Exon 19	69	62.2%
	Exon 20	3	2.7%
	Exon 21	25	22.5%
	Exon 18 and exon 21	2	1.8%
	Exon 19 and exon 20	9	8.1%
	Exon 20 and exon 21	3	2.7%

• NA, data unavailable; SD, standard deviation.

Results

The demographic and clinicopathological characteristics of the 111 eligible patients (Fig 1) are detailed in Table 1. The median age was 61.75 years (SD ± 11.48), 47 were males and 64 females, most patients were never smokers (n = 65), and the majority were diagnosed at disease stage IV (n = 99) (Table 1). Notably, most patients harboring EGFR-sensitizing mutations presented a partial response as the best TKI response (n = 78, 70.3%; Table 1), and none of the patients presented with a complete response (CR) (Table 1). The association between response to TKI treatment and demographic and clinicopathological characteristics from TKI-treated NSCLC patients harboring EGFR-sensitizing mutations (n = 111) was also assessed, and no association was revealed (Table S1).

EGF+61 A>G genotypic frequencies were 31.5% (n = 35) for AA, 49.6% (n = 55) for AG and 18.9% (n = 21) for GG. The allelic distribution was 56.3% for the A allele and 47.7% for the G allele (Table S2). The present series (n = 111) was in Hardy-Weinberg equilibrium, according to EGF+61 A>G polymorphism (P = 0.941).

The patients were grouped into three categories according to response to treatment (PR vs. SD vs. DP) and three genotypic models to assess the association between EGF+61 A>G polymorphism and response to TKI treatment. There was an association between response to TKI treatment and EGF+61 A>G polymorphism for a codominant and recessive model (P = 0.046 and P = 0.023, respectively; Table 2). No association was observed between response to TKI treatment and the dominant model (P = 0.601, Table 2). In addition, no association was observed between response to TKI treatment and allele frequencies (P = 0.163) (Table 2).

Since EGF+61 A>G genotypes (codominant and recessive model) were associated with response to TKI treatment, we assessed whether any clinical parameter could be influenced by this result as a confounder variable and a multivariate analysis (multinomial analysis) was conducted adjusted for age, TNM, and PS-ECOG. We observed that in the codominant model, AG genotype was independently associated with SD (P = 0.01; OR = 0.08; 95% CI: 0.01–0.60). Likewise, we observed that in the recessive model, AG + GG genotypes were independently associated with SD (P = 0.02; OR = 0.12; 95% CI: 0.20–0.72) (Table 3).

In addition, we evaluated the association between EGF+61 A>G genotypes and post-TKI PFS and OS. No association was observed between EGF+61 A>G codominant model (AA vs. AG vs. GG; P = 0.562), recessive model (AA vs. AG + GG; P = 0.709) and dominant model (AA +AG vs. GG; P = 0.401) and post-TKI PFS (Fig S1A–C, respectively). Likewise, no association was observed between EGF+61 A>G codominant model (AA vs. AG vs. GG; P = 0.493), recessive model (AA vs. AG

+GG; P = 0.318) and dominant model (AA + AG vs. GG; P = 0.342) and OS (Fig S1–F, respectively). Thus, the results show that the EGF+61 A>G polymorphism does not influence disease outcomes, both PFS and OS from NSCLC TKI-treated patients harboring EGFR-sensitizing mutations.

Discussion

Molecular therapies have revolutionized clinical management in NSCLC patients, with particular emphasis on EGFR-TKIs.^{3-5, 15-17} Herein, we report the impact of EGF+61 A>G genotype over ORR and DCR to first-generation TKIs (erlotinib and gefitinib) on 111 EGFR-mutated Brazilian lung adenocarcinoma patients.

The EGF+61 polymorphism (AG and AG + GG) was associated with stable disease in both univariate and multivariate in NSCLC patients treated with first-generation TKIs harboring EGFR-sensitizing mutations. Nevertheless, the SNP was not associated with ORR or disease outcome (progression-free and overall survival). It is well known that EGFR-sensitizing mutations can predict a response to first-generation TKIs, and TK-treated EGFR-mutated patients can benefit from a substantial improvement in progression-free and overall survival.^{6, 17} However, most patients will experience disease progression due to acquired resistance mechanisms.⁷ Currently, there is an effective third-generation TKI, osimertinib, for patients harboring EGFR-resistance mutations.¹⁸ Nonetheless, TKIs are considered high-cost drugs for low-middle income countries, and only first-generation TKIs (erlotinib/gefitinib) are supported by the Brazilian public health system for EGFR-mutated NSCLC patients. Thus, a biomarker for predicting both ORR and DCR in patients treated with first-generation TKIs harboring EGFR-sensitizing mutations would be valuable when financial resources are limited.

In conclusion, the present study revealed that the EGF+61 polymorphism (AG and AG + GG) was independently associated with stable disease, although the identification of this SNP cannot predict ORR, DCR or disease outcome in patients treated with first-generation TKIs harboring EGFR-sensitizing mutations.

Disclosure

The authors declare that they have nothing to disclose.