Elexacaftor/Tezacaftor/Ivacaftor Efficacy in a Cohort of Italian Patients with CFTR Rare Mutations

Abstract

Background: Cystic Fibrosis is an inherited disorder caused by mutations in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene, encoding a chloride and bicarbonate channel widely expressed in epithelia. Loss of CFTR function leads to dehydration of the epithelium surface with thicker mucus secretions from tissues. The lungs, pancreas, liver, intestines, and sweat glands are the most common affected organs. However, pulmonary disease remains the main cause of morbidity and mortality. Fortunately, elexacaftor/tezacaftor/ivacaftor (ETI) therapy is showing unprecedented clinical benefits in patients with Cystic Fibrosis (CF) carrying at least one F508del mutation in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene. However, almost 35% of the CF population living in the Mediterranean area still lacks effective CFTR modulator therapies because of the elevated incidence of patients with (pw)CF harboring CFTR rare mutations (RMs), different from F508del. Methods: Twenty-three pwCF harboring RM including the N1303K underwent off-label ETI treatment for 6-12 months. Respiratory function in terms of FEV1 and FVC was measured after 3, 6, and 12 months of treatment. In addition, we analyzed sweat chloride concentration, body mass index (BMI), and quality of life before and after treatment. Possible adverse effects were recorded. Results: All patients included in this off-label program displayed a substantial improvement in respiratory function. In particular, patients carrying the N1303K mutation showed an improvement in FEV1 and FVC similar to that observed in subjects harboring the F508del mutation, although sweat chloride concentration was not significantly decreased. No severe adverse effect was reported. Conclusions: This study strengthens the clinical efficacy of ETI in pwCF harboring the N1303K and other CFTR rare variants. Since these CFTR RMs have not been approved for ETI therapy in Europe, this study may promote the inclusion of these variants in the list of CFTR mutations responsive to ETI.

1. Introduction

Cystic Fibrosis (CF) is caused by mutations in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene, encoding a chloride and bicarbonate channel widely expressed in human epithelia. CF-related lung disease, characterized by excessive mucus production, bronchial obstruction, infection, and chronic inflammation, remains the main cause of morbidity and mortality [1].

Although over 2000 genetic variants have been described for CFTR, the F508del variant accounts for almost 90% of total CFTR variants in northern Europe and north America. However, F508del is present only in 65–70% of patients with (pw)CF in the Mediterranean area [1]. In Italy, similarly to other Mediterranean areas, the incidence of CFTR rare mutations (RMs) is higher compared to northern Europe and north America. Data from the Italian Registry of Cystic Fibrosis (https://registroitalianofibrosicistica.it, accessed on 28 February 2025) indicate that N1303K (10.3%), G542X (8.7%), 2789+5G>A (5.5%), 2183AA>G (3.6%), 3849+10kbC>T (2.7%), and G85E (2.2%) are the most representative CFTR RM in Italy.

The European Medicines Agency (EMA) has approved elexacaftor/tezacaftor/ivacaftor (ETI) therapy for pwCF aged 2 and older carrying at least one F508del mutation in CFTR. The US Food and Drug Administration (FDA) approval includes an additional 177 responsive CFTR variants [2].

ETI treatment in eligible pwCF leads to a remarkable increase in lung function and body mass index (BMI), while also reducing hospitalizations for pulmonary exacerbations (PEx) [3]. However, many CFTR RMs still have no EMA or FDA approval for ETI therapy. Among these orphan variants, N1303K has shown a variable in vitro response to ETI [4,5], although recent reports identified promising clinical responses [6,7]. Despite the ETI-dependent improved respiratory function, it has been recently reported that sweat chloride concentration is not affected by the treatment in pwCF and N1303K variants. Similar to pwCF with the F508del mutation, mild or moderate adverse events have also been reported in patients with the N1303K variant undergoing ETI therapy [7]. The present study aims to evaluate the clinical response to ETI in a cohort of 23 Italian patients carrying CFTR RM for which this therapy still awaits EMA approval.

2. Materials and Methods

Between March 2022 and June 2024, 23 pwCF took part in this study (M:10, 43%; mean age 39.2 y ± 15.3, range 13–58): all carried only the CFTR RM, including N1303K (n = 11, 48%), and received off-label ETI treatment for 6–12 months at the University Hospitals of Verona, Ancona, or Treviso (Italy). All patients recruited were of Italian origin (Caucasian), except for #23 who was from Ghana. The recruitment of patients was based on these inclusion criteria: advanced lung disease (with ppFEV1 < 40 and/or under evaluation for lung transplantation), or clinical worsening due to recurrent PEx, not eligible for ETI according to regulatory criteria indicated by the Italian Medicines Agency (AIFA), bearing one CFTR variant described as responsive in vivo or ex vivo. ETI was administered at a dosage of 2 tablets containing 75 mg of ivacaftor, 50 mg of tezacaftor, and 100 mg of elexacaftor in the morning, and 1 tablet containing 150 mg of ivacaftor in the evening. The ethics committee of the University Hospital of Verona approved this study (approval code: n° 216 CET; approved on: 8 May 2024). Patients’ written consent was obtained. The following parameters were recorded after 3, 6, and 12 months of treatment: sweat chloride test; respiratory function, measured as predicted percent forced expiratory volume at 1st second (ppFEV1) and percent forced vital capacity (ppFVC); body mass index (BMI); number of hospitalizations; PEx rate; quality of life (CFQR-respiratory domain); and any adverse events. Data before and after treatment were compared by the Wilcoxon signed rank test. Results were considered statistically significant when p < 0.05. All analyses were performed using SigmaPlot version 14.0 (Systat, Palo Alto, CA, USA).

3. Results

Characteristics of all 23 pwCF (2 children, 21 adults) are shown in

Table 1. Individual data before and after 6 months of ETI treatment in pwCF harboring RMs.

ID	Age	Sex	CFTR mut# 1	CFTR mut# 2	PI/PS	Sweat Chloride Conc. (mmol/L)		ppFEV1		BMI
						Before	After	Before	After	Before	After
1	33	M	N1303K	2183AA>G	PI	119	102	36	48	21.6	22.3
2	35	M	N1303K	711+5G>A	PI	107	103	68	87	21.1	23.7
3	37	M	N1303K	1717-1G>A	PI	107	106	44	71	24.2	26.9
4	13	F	N1303K	1717-1G>A	PI	104	93	81	93	17.2	18.0
5	13	F	N1303K	2183AA>G	PI	87	84	82	87	17.3	18.1
6	32	M	N1303K	2183AA>G	PI	93	98	91	99	21.6	21.4
7	23	F	N1303K	2183AA>G	PI	89	108	14	21	14.0	16.2
8	48	M	N1303K	P205S *	PI	78	41	37	38	21.1	24.6
9	21	M	N1303K	G542X	PI	109	102	48	84	21.3	25.9
10	58	F	N1303K	R1162X	PI	103	94	44	55	20.1	20.4
11	23	F	N1303K	W1282X	PI	113	151	28	50	24.8	24.9
12	51	F	3849+10KbC>T	R1162X	PS	71	54	57	64	22.4	21.8
13	56	M	3849+10KbC>T	R1162X	PS	71	35	57	58	20.1	20.3
14	58	M	3849+10KbC>T	G542X	PS	92	13	43	45	23.9	24.5
15	45	F	2789+5G>A	R553X	PS	97	81	39	45	23.9	24.5
16	31	F	R347P *	W1282X	PI	84	44	38	56	20.4	22.2
17	56	F	G85E *	1584+18672bpA>G	PS	67	52	91	100	20.4	20.6
18	28	F	G85E *	621+1G>T	PI	100	68	34	36	18.3	19.1
19	42	F	W57G	A234D *	PS	104	14	62	65	19.5	21.0
20	51	F	2183AA>G	711+3A>G	PS	53	56	70	77	19.5	21.0
21	29	F	2183AA>G	711+5G>A	PS	96	73	28	36	16.8	17.3
22	52	M	2789+5G>A	711+5G>A	PS	125	92	-	-	23.2	24.4
23	33	M	A559T	A559T	PI	107	92	46	52	20.0	22.6

PI, pancreatic insufficiency; PS, pancreatic sufficiency; BMI, body mass index; * mutations already established as responsive only by FDA; patient 22 was not able to perform spirometry, because of cognitive impairment.

. Seven patients (30%) showed a severe decrease in lung function (ppFEV1 < 40%), with six of them (26%) requiring oxygen therapy. Two patients (9%) were awaiting lung transplantation.

Overall data for the entire cohort showed significant improvement in respiratory functional tests, including FEV1 (p < 0.001; +11.3 and +14.6 pp) and FVC (p < 0.001; +10.6 and +13.0 pp), after 6 and 12 months of ETI, respectively (Table 1).

Among the 23 patients enrolled, only 6 subjects (26%) reported mild to moderate adverse effects (

Table 2. Adverse effects reported in the study cohort.

ID	Treatment Duration (Months)	Adverse Effects (PEx Excluded)
1	12	skin erythema
2	12	None
3	12	None
4	12	None
5	12	None
6	12	None
7	12	None
8	11	None
9	12	None
10	12	abdominal swelling, short-term memory deficit, skin erythema
11	12	None
12	12	None
13	12	None
14	12	skin erythema
15	12	skin erythema, mild hypertension
16	12	short-term memory deficit, transient anomia
17	11	None
18	12	None
19	12	None
20	8	None
21	12	None
22	12	mild hypertension
23	12	None

PEx, pulmonary exacerbations.

). Skin erythema was the most representative adverse effect, affecting three patients (13%). Mild hypertension and short-term memory deficit were reported in two patients (9%) each, whereas abdominal swelling and transient anomia were shown by one patient each.

BMI significantly increased by +1.0 and +2.1 points after 6 and 12 months, respectively, compared to baseline. Considering a period of 12 months of pre-treatment and the same period after starting ETI, the PEx rate decreased by 50%. One patient with the G85E variant discontinued oxygen therapy, whereas three pwCF with the N1303K variant showed a 50% reduction in oxygen supplementation after 3 months of ETI. Interestingly, patients with N1303K undergoing ETI treatment for 12 months showed similar clinical outcomes to pwCF with F508del [3]. Both sub-cohorts of patients with N1303K and other RMs showed a significantly improved respiratory function, BMI, and quality of life score after 3 months of therapy. While the sweat chloride test results of patients with the N1303K variant were unchanged after 12 months of therapy, other pwCF carrying RMs showed a significantly decreased (p = 0.008, −32 mmol/L) chloride concentration (Figure 1).

4. Discussion

Although the efficacy of ETI in improving respiratory function and nutritional status in pwCF with eligible CFTR mutations, particularly the F508del variant, has already been widely established, many other mutations are still in need of a therapeutic option. Here, we reported the efficacy of ETI on a cohort of pwCF harboring CFTR RM for which there is still no approved treatment in Europe, including the N1303K variant. The results showed a significant increase in respiratory function and nutritional status upon ETI treatment, with the clinical response being comparable to that of F508del CFTR [8]. In addition, the PEx rate decreased by almost 50% after therapy. Although respiratory function significantly improved after ETI treatment in patients harboring the N1303K variant, their sweat chloride concentration did not change, except for a patient heterozygous for the P205S variant, already included in the FDA list of responding mutations [2]. These data are consistent with two recently reported studies conducted in the USA and France [6,7], raising doubts as to the actual predictive role of sweat chloride concentration on respiratory function.

We observed a higher increase in ppFEV1 in pwCF harboring the N1303K variant (means of +13.4 and +15.8 pp after 6 and 12 months of treatment, respectively) compared with other patients with CFTR RM. Our results are therefore ameliorative compared with a recent study conducted on a US cohort of pwCF harboring the N1303K mutation [7]. It should be noted that pwCF with N1303K reported pancreas insufficiency (PI), whereas most of the remaining patients (75%) exhibited a pancreatic sufficient (PS) condition. This suggests that the severity of the CF phenotype (i.e., PS/PI condition) seems irrelevant to the response to ETI.

However, the improvement in CFQ-R respiratory domain score for patients with N1303K (means of +25.2 and +15.4 points after 6 and 12 months of treatment, respectively) was similar to that observed in the US prospective, multicenter, open-label, single-arm trial [7].

In the present study, pwCF with CFTR RMs undergoing ETI treatment reported only mild or moderate adverse effects, including mild hypertension, abdominal swelling, short-term memory deficit, and skin erythema.

Like F508del, N1303K is a class II CFTR mutation impairing the transport of unfolded protein to the plasma membrane [5]. However, N1303K has also been described as a gating mutation, impairing ATP-dependent chloride efflux from the channel [9]. Although the actual mechanism of CFTR correction sustained by ETI is still not fully known, class II mutations may be the ones predominantly affected by this therapy. Importantly, the poor efficacy of ETI in ameliorating the sweat chloride test in N1303K mutants contrasts with the major improvement it achieves in pwCF harboring the F508del variant. This discrepancy might be at least partially explained by the aforementioned complexity of the N1303K variant. Other CFTR rare variants, including G85E [10], A234D [11], 3849+10KbC>T, 711+3A>G, R347P [12], and A559T [13], have recently proved responsive to ETI both in vitro and ex vivo, using cellular models and organoids. Here, we demonstrate that pwCF with these CFTR RMs can clinically benefit from ETI. Therefore, this study confirms and extends previous reported data on the efficacy of ETI in terms of respiratory function, regardless of the class of CFTR genetic variants, except for the nonsense mutations.

Including these RM among those recognized as responsive to ETI could increase the percentage of Italian subjects covered by therapy from 66% to 82%. This would mean an overall reduction in hospitalization of at least 50% for these patients, thus reducing both direct and indirect costs for the Italian health system.

In conclusion, although this study presents some limitations, such as the small number of patients recruited and heterogeneity of CFTR RM, it strengthens the case for extending ETI approval in Europe to other CFTR mutations, including the N1303K variant. In addition, compared with previously published studies, this work provides long-term outcomes on ETI efficacy in pwCF harboring different RM.