Perspectives of carriers of X‐linked retinal diseases on genetic testing and gene therapy: A global survey

Female carriers of X‐linked inherited retinal diseases (IRDs) are burdened with potentially passing their disease‐causing variant to future generations, as well as exhibiting signs of retinal disease themselves. This study aimed to investigate carriers' experiences of genetic testing, emotions relating to having affected children, and their knowledge regarding genetic testing and gene therapy. An online survey was advertised to self‐identified carriers worldwide. Two hundred and twenty‐eight carriers completed the survey with mean age of 51 years (SD ± 15.0). A majority of respondents resided in the United States of America (51%), Australia (19%), and the United Kingdom (14%). Most carriers identified with feelings of guilt (70%), concern (91%), and anxiety (88%) for their child. Female carriers who had given birth to children had significantly greater gene therapy knowledge compared to carriers who had not (p < 0.05). Respondents agreed that their eyecare provider and general practitioner helped them understand their condition (63%), however, few carriers reported receiving psychological counselling (9%) or family planning advice (5%). Most respondents (78%) agreed that gene therapy should be available to carriers. This study emphasises the importance of providing appropriate counselling to female carriers and illustrates the motivation of many to participate in emerging treatment options, such as gene therapy.


| INTRODUCTION
X-linked inherited retinal diseases (IRDs) such as choroideremia and X-linked retinitis pigmentosa (XLRP) lead to progressive retinal degeneration in males (XY individuals) which typically results in severe visual impairment.Female carriers of X-linked IRDs (XX individuals) exhibit variable retinal phenotypes, ranging from near normal retinae to severe retinal degeneration (known as 'male-pattern' phenotype). 1 A general misconception is that female carriers are only mildly affected by the presence of a disease-causing variant in one of their X-chromosomes, 2 therefore, female X-linked IRD patients are largely overlooked 3 or incorrectly diagnosed. 4Improving genetic testing technologies and availability 5,6 has led to an increased awareness of the retinal phenotypes presented by female carriers, and the variable prognoses these clinical presentations portend.In addition to their own health journey, female carriers of X-linked IRDs are burdened with the possibility of passing their disease-causing variant to future generations.
Other studies have investigated the experiences and psychosocial impacts of X-linked disease on female carriers, but this has not been examined in depth in female X-linked IRD patients.In a prior study, Dunn et al (2008) reported on female carriers of haemophilia and their experiences of receiving a genetic diagnosis, 7 female carriers reported a sense of shock and grief, concern about having children, feeling blamed by their partner, and a 'prolonged sense of guilt'. 7A more recent study by Choi et al. distributed a survey to female carriers of varying X-linked diseases, including adrenoleukodystrophy (46.3%), blue cone monochromacy (7.4%) and chronic granulomatous disease (6.7%). 8The survey found that most of the women (54%) found out about their carrier status after having an affected child, even though a majority of respondents (83.2%) felt they already had relevant symptoms to their X-linked condition. 8wever, this study only had a single choroideremia carrier (0.7%) and no carriers of XLRP.
Understanding the perspectives of female carriers may aid in the counselling provided upon diagnosis, and potential inclusion into therapeutic clinical trials, such as gene therapy.Following diagnosis, knowledge of the risks associated with passing on the disease-causing variant may directly influence future family planning, 8 relationship with partners, 7 and interest in participating in future clinical trials.Previous studies have reported that severely affected female carriers of X-linked conditions often want to avoid having an affected child, 8 and carriers with affected male relatives usually want to avoid having an affected child compared to carriers without any personal experience of the condition. 9However, whether female carriers are receiving appropriate advice regarding family planning and reproductive options such as in vitro fertilisation with preimplantation genetic testing (IVF with PGT) remains unknown.Inadequate understanding of the negative emotions and experiences of female carriers hinders the support offered to carriers by healthcare professionals to cope with the emotional strain of the condition and the ability to make informed decisions about family planning.
In X-linked IRDs, there is also a time-sensitive need to learn more about female carriers.Choroideremia and RPGR-related retinitis pigmentosa are two of the leading candidates for gene therapy treatment, 10 and it will be important to determine whether a subset of female carriers-those with a 'male pattern' phenotype for example-will be suitable for inclusion in these upcoming trials.This study aimed to explore female carriers' genetic testing experiences, emotions relating to being a female carrier of an X-linked IRD (i.e., choroideremia and X-linked retinitis pigmentosa), and knowledge of genetic testing, genetic counselling, and gene therapy.

| Study design
The survey was hosted on Qualtrics Survey Software (Qualtrics, Provo, UT) and REDCap 11 (hosted by the University of Melbourne), which are secure web applications for creating and handling online surveys.Participants indicated consent following the study description on the survey webpage.Carrier status was self-reported and not confirmed by any clinical and/or genetic testing.The survey was available in English only, therefore, respondents had to be English speaking.No age-limit was set.
The survey was distributed globally by clinicians, support groups and associations (see acknowledgements), and social media.Survey responses were collected between 2 December 2021 and 30 April 2023.The survey had three sections: (i) respondent demographics and genetic testing experiences; (ii) female carrier emotions; and (iii) knowledge relating to genetic testing, genetic counselling, and gene therapy (Supplementary Material 1).

| Demographics and genetic testing experiences
Data were collected on respondents' age, ethnicity, country of residence, and marital status.Respondents were asked their X-linked retinal condition (i.e., XLRP, choroideremia, X-linked retinoschisis, other), age of diagnosis, and details of genetic testing (if applicable, i.e., year of testing, mutated gene, motivation for testing).Respondents were also required to indicate whether they have any children and, if so, whether they knew they were a carrier prior to their first pregnancy.

| Carrier emotions
In this section, female carriers were asked about their thoughts and emotions relating to being a carrier of an IRD.Respondents answered nine questions regarding their emotions relating to their own vision, their child's vision, family planning, speaking to their child and partner about their carrier status, and regret for not having genetic testing prior to pregnancy.All questions used a 5-step Likert scale (from strongly agree to strongly disagree) or not applicable option.

| Female carrier knowledges and opinions
The final section comprised of 14 questions regarding genetic testing, genetic counselling, and gene therapy, which were taken from the larger Attitudes to Gene Therapy for the Eye (AGT-Eye) questionnaire by Mack et al. 12 Respondents selected their level of agreement using a 5-step Likert scale, from strongly agree to strongly disagree.

| Data analysis
Carrier emotions and knowledge sections (Sections 2 and 3) were collapsed to a three-point scale to summarise data collected (i.e., strongly agree and agree, and strongly disagree and disagree combined), and evaluated using their original scores (scored from 1 to 5).Scoring of select statements were reversed and total scores (%) were calculated to ensure a higher score in the female carrier knowledge section indicated high awareness of genetic testing, a benefit of genetic counselling services, and knowledge of gene therapy processes and outcomes.Statistical analyses were performed using GraphPad Prism version 9.0 (GraphPad Software, Boston, MA, USA).A Kruskal-Wallis test was used to compare the age of diagnosis of carriers between the United States of America (USA), Australia, and United Kingdom.A Mann-Whitney U test was used to determine whether giving birth to a child, having genetic testing, or genetic counselling affected knowledge of female carriers.

| RESULTS
Three-hundred and thirty-two responses were collected.After removal of ineligible responses (n = 57), 278 responses were available for analysis.Responses were deemed ineligible if they were not completed by a carrier of an X-linked IRD (based on responses to the carrier experience section), demographics section was blank, or majority of the survey was incomplete (i.e., none of the sections were completed).Respondent demographics are summarised in Table 1.The majority of respondents were from the USA (51%), Australia (20%), the United Kingdom (14%), and Canada (4%) (Figure 1).

| Genetic testing and counselling experiences
Over half (56%) female carriers had genetic testing and 38% had genetic counselling.Most respondents self-reported their motivation for undertaking genetic testing was because they had affected relatives and wanted to know if they were a carrier (57%) or wanted to confirm their relative's diagnosis (36%).One in five carriers had genetic testing because they had symptoms suggestive of an IRD (20%) or were interested in family planning (18%).
Most carriers sought genetic counselling to understand their condition (57%) and the risk of having a child with an IRD (39%) (Table 2).These goals were met, with most respondents reporting that the genetic counsellor covered these topics (69% for understanding the condition and 48% for family planning).Nearly half of the respondents (42%) thought their genetic counsellor was 'very helpful' or 'extremely helpful'.Most carriers (63%) reported that their clinician (i.e., optometrist, ophthalmologist, and/or healthcare provider) helped them understand their condition and 26% reported that their clinician helped them with understanding the risk of having a child with an IRD (Table 2).However, one in four carriers reported that their clinician was no help at all.
Of the 223 carriers who had given birth to children, only 39% were diagnosed (clinically and/or genetically) as a carrier prior to pregnancy.One in four carriers (23%) reported that their decision to have children was affected 'a fair amount' or 'a lot' by their carrier status.Most responses were received from the USA, Australia, and the United Kingdom, therefore, experiences of female carriers from these countries were compared.There was a statistically significant difference in age of onset between the USA and Australia, with Australian women on average diagnosed 10 years later than those in the USA (median 37 years vs. 27 years, p = 0.03).The median age of diagnosis in the United Kingdom was in between the two, at 30 years, which was not statistically significant to USA or Australia ( p > 0.99 and p = 0.48, respectively).
Women in the USA (55%) and Australia (55%) had lower rates of genetic testing, compared to the United Kingdom (69%).Similarly, women in the USA (36%) and Australia (31%) had lower rates of genetic counselling, compared to the United Kingdom (46%).Female carriers in Australia (18%) and the United Kingdom (26%) were less likely to know about their carrier status prior to pregnancy, compared to the USA (40%).

| Female carrier emotions
The second section of the survey explored female carriers' emotions relating to their vision, their child's vision, and telling their partner and child about their carrier status (Figure 2).Values in  respondents felt anxious (89%), guilty (71%), and concerned (91%) for their child.Over half of the carriers (52%) also agreed they felt anxious about their own vision.Although most respondents (64%) said they did not feel nervous telling their partner of their carrier status, there were mixed feelings about telling their child (41% were not nervous, while 39% were).Half of the respondents (51%) indicated regret for not having genetic testing prior to pregnancy.

| Female carrier knowledge
Following removal of ineligible or incomplete results, the final section was completed by 262 respondents.These questions captured the knowledge of carriers about genetic testing, genetic counselling, and gene therapy (Table 3).Most respondents disagreed that genetic testing was only required for family planning (69%), and most had been told about genetic testing by a healthcare professional (54%).A strong finding was that most respondents were confident in the data privacy arrangements in place, and were not fearful of losing their privacy if they undertook genetic testing (74%).Gaps in knowledge of ocular gene therapy were apparent, with only 39% aware that the treatment was not administered through the blood stream.However, the majority of carriers agreed that gene therapy may slow down the progression of the disease (65%) and understood that ocular gene therapy does not change the risk of passing the mutation to their children (60%).Of interest, most respondents agreed that gene therapy should be available for carriers of IRDs (78%).
A sub analysis was performed to determine factors promoting improved knowledge in female carriers of X-linked IRDs (Table 4).
Genetic testing knowledge was significantly greater for female carriers who had not given birth to children, had genetic testing, and genetic counselling ( p < 0.05, p < 0.0001, and p < 0.001, respectively).As expected, genetic counselling knowledge score was higher in female carriers who had genetic testing and genetic counselling ( p < 0.001 and p < 0.0001, respectively).However, gene therapy knowledge only improved in female carriers who had given birth to children ( p < 0.05).

| DISCUSSION
This is the first global survey exploring the psychosocial effects of female carriers of X-linked IRDs.Considering the relatively large number of responses obtained (n = 278), we believe our survey accurately represents the perspectives of female carriers of X-linked IRDs, particularly choroideremia and X-linked retinitis pigmentosa.

| Female carriers' genetic testing experiences
According to the National Society of Genetic Counsellors, 'genetic counseling is the process of helping people understand and adapt to the medical, psychological and familial implications of genetic contributions to disease'. 13Respondents of our survey generally indicated a positive experience with genetic counselling, with good alignment between their goals and the help that they received.However, only 19% of our respondents indicated receiving emotional support from their genetic counsellor.Although several studies report high overall satisfaction with genetic counselling, [14][15][16] genetic counselling may not lesson an individual's worries, simply by being 'provided with new information' about their condition. 14In particular, a study by Charles et al. investigated satisfaction of African American women who were at risk of BRCA1-and BRCA2-associated breast and ovarian cancer.
Whilst a majority of respondents (98%) were satisfied with genetic counselling, only 26% reported that their worries lessened and were able to cope better (22%). 14Austin et al. suggested conceptualising genetic counselling as a time-limited psychotherapeutic encounter to target educational and psychosocial aspects of receiving a genetic diagnosis. 17Such an approach by genetic counsellors may equip carriers to 'understand and adapt to the medical, psychological, and familial implications of genetic contributions to disease'. 13male carriers had different experiences of receiving their diagnosis and genetic testing depending on their country of residence.
Female carriers in the USA were diagnosed significantly younger (27 years) compared to carriers in Australia (37 years, p = 0.03).This may explain why female carriers were more likely to know about their T A B L E 4 Sub-analysis of female carriers' knowledge relating to genetic testing, genetic counselling, and gene therapy.Note: Median knowledge score as a percentage [% (interquartile range)] of female carriers depending on 1. whether they had given birth to any children, 2. previously had genetic testing, or 3. had genetic counselling.Bold represent statistically significant differences in knowledge score between respondents answering yes/no to each sub-analysis question.
carrier status prior to pregnancy in the USA compared to Australia (40% vs. 18%, respectively).This is important information when considering access to IVF and family planning counselling and supports the need for earlier diagnosis in Australia.However, both countries had similar overall rates of genetic testing (55% in both countries) and genetic counselling (USA: 36%; Australia: 31%).The USA is one of the leading countries for gene therapy clinical trials for X-linked RP and choroideremia. 1 The interest in developing therapeutic treatment for these conditions and larger population of people with IRDs in the USA, 18 compared to Australia 19 may lead to earlier diagnosis of female carriers through the clinical care of affected male relatives.
Experiences with clinicians (i.e., ophthalmologists, optometrists, and other healthcare providers) showed the most discrepancy between care providers in this study.A potential area for improved patient care would include better counselling and referral pathways for the provision of appropriate reproductive guidance to female carriers of IRDs.The majority of the female carriers reported that their clinician helped them understand their condition (63%), however, fewer received support for their emotions (9%), family planning (5%), and learning about IVF (5%).Similarly, in the study by Choi et al., only 10% of carriers of a wide range of X-linked conditions reported they had sufficient access to knowledgeable healthcare providers exploring symptoms and/or the risks associated with being a carrier. 8Insufficient counselling was also evident through an audit of a private tertiary ophthalmology practice in Australia, whereby approximately 70% of clinical records did not document any counselling provided to patients with an IRD. 20Whether this is due to clinical record documentation inaccuracies or is indicative of an area for improvement for clinicians remains unknown.However, considering that 25% of carriers reported that their clinician was no help to them, there is a demand for better counselling practices by clinicians.

| Emotions of female carriers
Similar to previous studies of other X-linked conditions, 7,8 our respondents reported anxiety, guilt, and concern associated with being a carrier and potentially passing the disease-causing variant to their children.However, interestingly, most respondents (64%) did not feel nervous about telling their partner of their carrier status.On the contrary, Dunn et al. reported that carriers identified with a fear of being blamed by their partners for passing down their disease-causing variant to their child. 7The results in the current study may be related to the fact that 61% of carriers with children were not diagnosed prior to pregnancy.Furthermore, their own guilt towards their children, may potentially overpower their consideration for their partner's emotions.Of the women who were aware of their carrier status prior to pregnancy (n = 85), only 14 indicated that it would have affected their decision to have children, while most respondents said it would not have affected (n = 61) or might have affected their decision (n = 10).
Similar to affected individuals of other ocular conditions, visuallyimpaired female carriers may also present with multiple psychological impacts. 21This will include fears about the uncertainty and the future, as well as a change in identity.These states of mind require dedicated psychological support to address them.

| Female carriers' knowledge
Female carriers of IRDs in our study demonstrated similar knowledge of gene therapy, when compared to previously published data from people with IRDs. 22In particular, the two groups scored similarly on the statement: 'gene therapy slows down disease progression' (IRDs vs. carriers: 69.3% vs. 65% agreement, respectively).However, a difference between female carriers and people with IRDs were found for the statement that 'Having gene therapy does not mean a person will not pass on an eye condition to any children they may have in the future'.60% of female carriers knew that this was an incorrect statement compared to only 38% of people with an IRD. 22rthermore, it was evident that female carriers who had given birth to a child had significantly greater knowledge of gene therapy, suggesting this knowledge may not have been obtained through genetic testing or genetic counselling processes (Table 4), but rather through independent research or their affected child's care.As expected, genetic testing and genetic counselling in the current cohort was successful in providing relevant information regarding genetic testing and counselling processes, as evident by the knowledge scores obtained in these respective areas.
Few carriers had been told about current research studies by their genetic counsellors (23%) or clinicians (13%), suggesting that their therapeutic knowledge was obtained by independent research and/or advice provided to their male relatives.If future work showed that female carriers could be considered for inclusion in clinical trials, education about these options would be essential.
It appeared that the carriers were keen to see these options arise-with a large proportion agreeing that gene therapy should be available to them.Therefore, researchers should consider including visually symptomatic female carriers in clinical trials of emerging therapeutic options.This will require further data on the phenotypes and prognosis of affected female carriers, to determine who would be the best candidates for intervention.

| CONCLUSION
As our knowledge of the physical and psychological impacts of being a female carrier of X-linked IRDs increase, 1 it is important that both researchers and clinicians are aware of the full spectrum of support required.
Healthcare professionals should be equipped with the knowledge of the variable disease severity seen in female carriers of X-linked IRDs, and be prepared to offer appropriate medical and vision care.
However, it is also essential that female carriers are provided with the appropriate genetic counselling, family planning and psychological support, as highlighted in this study.Future studies could assess clinical practice patterns for care and counselling provided to female carriers of X-linked IRDs, in the aim of developing gold standard guidelines.
Our study demonstrated that female carriers experience feelings of guilt, concern, and anxiety for their child's vision and their own vision, particularly as they receive limited information.Appropriate counselling will promote informed decision-making regarding expectations, disease prognosis, and participation in future studies.We advocate for improved integration of clinical care between eyecare professionals and geneticists/genetic counsellors for comprehensive coverage of all the issues arising from receiving a carrier diagnosis.
Finally, a significant finding from this study was that 78% of the respondents felt that female carriers should have access to emerging gene therapy options.Whilst future work is required to identify the best candidates for these interventional trials, this data supports the inclusion of these women in such opportunities.

T A B L E 1
Respondent demographics.

Figure 2
Figure2represent responses by female carriers who the statement was applicable for.The researchers acknowledge there may be multiple reasons that would make each statement unapplicable for carriers.Therefore, respondents who indicated the statement was not applicable to them was not included in the calculation out of 100.Majority of Female carriers' emotions relating to their carrier status.Percentage values indicate level of agreement with the statements for applicable respondents.Number of respondents each statement applied for is indicated in brackets.n, number of respondents; NA, not applicable.[Colour figure can be viewed at wileyonlinelibrary.com]

4. 4 |
LimitationsThe current study relied on the self-reported diagnosis of female carriers of X-linked IRDs.Future studies should consider distribution of the survey to clinically and genetically confirmed carriers by clinicians, geneticists, genetic counsellors, and/or genetic testing facilities.The current survey was only available in English, which may have also limited responses obtained from countries where English is not the primary language.Translating the current survey to other languages may increase responses from countries with high prevalence of choroideremia and/or X-linked RP.The current study findings also do not represent the perspectives of female carriers of X-linked retinoschisis (XLRS) due to the low response rate (n = 3) obtained, and therefore future work should incorporate additional responses from this cohort.Female carriers of XLRS do not exhibit hallmark retinal findings such as those seen in carriers of XLRP or choroideremia, and any rare retinal signs detected are nonspecific abnormalities seen in healthy individuals, therefore, female carriers of XLRS are difficult to identify.23Additionally, they are usually at lower risk of vision loss than carriers of choroideremia and X-linked RP.This survey was disseminated through the organisations Foundation Fighting Blindness, the Choroideremia Research Foundation, Retina International and Retina Australia.We did not receive responses from people with all forms of X-linked IRD.In future studies, it would be useful to target recruitment to ensure enrolment of carriers of the rarer X-linked IRDs, such as blue cone monochromacy and congenital stationary night blindness.
Total (n = 278) Abbreviations: CHM, choroideremia; XLRP, X-linked retinitis pigmentosa; XLRS, X-linked retinoschisis.a Data presented as median [range] years, due to small sample size for this condition.b Mutated gene percentage calculated from 183 respondents, remaining 95 responses were intentionally left blank.